Clinical Immunology_ Principles and Practice ( PDFDrive )

CHaPtEr 22 Phagocyte Deficiencies 331


in the cytosol cleave the ester linkage, trapping the fluorescent lactate dehydrogenase should be monitored simultaneously. The
probe in the cytosol. The labeled neutrophils are added to each release of azurophilic granules can be assessed by determination
well and incubated in the absence or presence of phorbol myristate of β-glucuronidase activity. Supernatant fluids or cell extracts
acetate (PMA) to promote adherence through activated integrins. obtained from stimulated neutrophils are incubated with
At the end of the incubation, the wells are washed to remove 4-methylumbelliferyl-β-d-glucuronide. Alternatively, myeloper-
nonadherent cells. The fluorescence of each well is determined oxidase can be determined by using commercially available
with a fluorescent microplate reader and compared with the enzyme-linked immunoassays. CD63 is also found in the
fluorescence of a control well with a fixed number of fluorescent membrane of azurophilic granules and migrates to the neutrophil
cells. Under control conditions, fewer than 10% of the neutrophils surface after stimulation with fMLF in the presence of cytochalasin
adhere to plastic or to plastic coated with fetal bovine serum. b. The release of specific granules can be assessed by determination
Slightly more neutrophil adherence is observed on wells coated of lactoferrin levels using an enzyme-linked immunoassay. The
with fibrinogen. Treatment of normal neutrophils with PMA carcinoembryonic antigen CD66b (formerly CD67) is found on
for 30 minutes results in the adherence of 100% of the neutrophils the neutrophil surface and in the specific granules, and its
under all conditions. This adherence is abnormal in patients expression on the surface of the neutrophils is increased after
with LAD. Neutrophils isolated from patients with typical LAD-1 stimulation with fMLF or LPS. Detection of the constituents of
generally exhibit markedly reduced adherence under both secretory granules can be assessed by flow cytometric analysis
unstimulated and PMA conditions. of the change in expression of surface proteins, such as adhesion
molecules, and cytochrome b 558 of the NADPH oxidase.
Neutrophil Chemotaxis
Neutrophil chemotaxis in vivo can be evaluated by using skin Generation of Reactive Oxygen Species
−
windows. Skin blisters are gently raised on the volar surface of The release of reactive oxygen intermediates, such as O 2 and
the forearm using a vacuum pump and a blister device, with H 2 O 2 , is an important component of the neutrophil bactericidal
little hemorrhage or vascular damage. The roof of the blister is machinery. Neutrophils isolated from patients with CGD are
removed, and the exposed dermis is bathed with autologous unable to generate superoxide, leading to their oxygen-dependent
−
serum with the use of a skin window chamber. In 24 hours, bactericidal defect. The production of O 2 can be detected by
−
exudative neutrophils accumulate in the autologous serum bathing using the reduction of cytochrome c. Because O 2 causes a
the skin lesion. The skin chamber provides a mechanism for one-to-one stoichiometric reduction of ferricytochrome c to
characterizing the immune cells as well as the soluble immune ferrocytochrome c, the resultant increase in the absorption
mediators that accumulate in the autologous serum during the spectrum at 550 nM can be used to quantitate the production of
−
evolution of the inflammatory response. Chemotaxis in vitro is O 2 . Superoxide dismutase is added to an identical tube to control
generally measured by using a Boyden chamber. The Boyden for the nonspecific reduction of cytochrome c. However, since
−
chamber includes three components: a lower (chemoattractant) cytochrome is not permeable to the cells, the detection of O 2 is
chamber, a nitrocellulose or polycarbonate filter layer, and an limited to that released into the extracellular milieu. Neutrophils
−
upper cell chamber. The lower compartments of the Boyden isolated from patients with CGD produce little O 2 in response
–8
chamber are filled with a chemoattractant, such as fMLF; 10 to PMA in 10 minutes. However, some patients with forms of
M) or IL-8 (10 ng/mL). Recently an improved rapid fluorescence- CGD associated with residual superoxide production have low,
−
based measure of neutrophil chemotaxis, which uses a new 96-well but detectable, O 2 production in 60 minutes. Neutrophils isolated
disposable chemotaxis chamber that can be read in a fluorescence from X-linked heterozygous carriers of CGD can yield a full
−
microplate reader, has been developed. spectrum of O 2 production, whereas neutrophils from autosomal
recessive carriers of CGD generally yield a normal response.
Expression of Surface Antigens Although the detection of O 2 by reduction of cytochrome c is
−
The expression of cell surface antigens on neutrophils relies on useful in the diagnosis of patients with CGD, it cannot be used in
labeled mAbs analyzed by flow cytometry. The panel may include the diagnosis of carriers because of the wide spectrum of responses
the β 2 integrins (CD11a, CD11b, CD11c, and CD18), selectins that result from the degree of X-chromosome lyonization.
(CD62L), Fcγ receptors I, II, and III (CD64, CD32 and CD16), The extracellular release of H 2 O 2 can be measured by using
leukosialin (CD43), the common leukocyte antigen (CD45), and horseradish peroxidase–induced oxidation of either phenol red
markers for the specific granules (CD67), and azurophilic granules or Amplex red. PMN suspensions in the presence of horseradish
(CD63). The expression of surface antigens can be used to assess peroxidase and one of the chromophores are exposed to either
the responsiveness of neutrophils to particular ligands, such as PMA or buffer alone. Changes in optical density of phenol red
fMLF and lipopolysaccharide (LPS). at 600 nm can be determined with a standard microplate reader.
Amplex red is a much more sensitive fluorescent chromophore,
Neutrophil Degranulation and H 2 O 2 -dependent changes in fluorescence can be determined
The proteases, acid hydrolases, and inflammatory mediators with a fluorescence microplate reader.
−
released from storage granules in neutrophils can mediate bacterial The NBT test is a qualitative assay of O 2 production. Either
killing, tissue damage, healing, and immune regulation. Lactoferrin whole blood or isolated neutrophils are mixed with NBT in a
from specific granules can chelate iron, resulting in a bactericidal chamber slide and stimulated with PMA for 15–30 minutes at
or bacteriostatic effect. Stimulation of neutrophils with various 37°C. The slide is counterstained with 0.1% safranin and examined
secretagogues can release granular enzymes into the extracellular under a microscope. The NBT test yields a visual record of the
fluid. Treatment of the neutrophils with cytochalasin b (5 µg/ reduction of the NBT dye to the insoluble, blue-black deposits
mL) disrupts microfilament assembly and facilitates the release of formazan. Normal neutrophils, but not neutrophils from
of both specific and azurophilic enzymes. To differentiate patients with CGD, reduce the yellow dye to black-brown-blue
degranulation from cell lysis, release of the cytosolic enzyme aggregates in the cells. The NBT test can be used to diagnose

332 Part two Host Defense Mechanisms and Inflammation


X-linked carriers of CGD but cannot differentiate autosomal oN tHE HorIZoN
carriers from normal subjects.
An alternative to the NBT test is a flow cytometric assay, in • Early recognition and molecular diagnosis of all phagocyte defects,
which the dye, dihydrorhodamine-1,2,3 is used. Neutrophils are leading to prophylactic antimicrobial treatment, where indicated
loaded with the nonfluorescent dye and then stimulated with • Improvement in bone marrow transplantation technology to allow for
early, safe, successful, fertility-preserving transplantation in all cases
PMA for 15 minutes at 37°C. The H 2 O 2 produced oxidizes the • Understanding the mechanisms of the hepatic complications of chronic
dye and results in increased fluorescence, detectable with a flow granulomatous disease (CGD) that correlate with mortality
cytometer. Catalase is added to prevent cell-to-cell diffusion of • Characterizing the pathways that converge on signal transducer and
H 2 O 2 . Since dye is localized to the cytoplasm, and catalase is activator of transcription 3 (STAT3) signaling causing the complex
present in the extracellular fluid, the dihydrorhodamine-1,2,3 somatic and immune disorder, hyper-IgE recurrent infection syndrome
assay detects the intracellular production of reactive oxygen (HIES).
metabolites. Stimulation of normal neutrophils with PMA results
in a two-log shift in the fluorescence intensity. Neutrophils from ACKNOWLEDGMENT
an X-linked carrier of CGD exhibit mosaicism with a negatively
stained (abnormal) population and a brightly stained positive This work was supported by the intramural program of the
population. Neutrophils from a patient with X-linked CGD that National Institute of Allergy and Infectious Diseases.
lack gp91 phox express little increase in fluorescence, whereas
neutrophils from a patient with a deficiency in p47 phox exhibit a Please check your eBook at https://expertconsult.inkling.com/
slight increase in fluorescence. The major advantages of the for self-assessment questions. See inside cover for registration
dihydrorhodamine-1,2,3 assay are the sensitivity, the signal-to- details.
noise ratio, and the ease of counting a large number of cells.
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CHaPtEr 22 Phagocyte Deficiencies 333.e1


MUL t IPLE-CH o ICE QUES t I o NS

1. A 12-year-old boy with X-linked chronic granulomatous A. Tissue neutropenia leading to severe bacterial infection
disease (CGD) presents to your office with progressive fever, B. Excessive interferon (IFN)-γ production
dyspnea, and hypoxia a week after a Halloween hayride at a C. Poor root maintenance by defective macrophages
local farm. His chest X-ray shows extensive bilateral military D. Excessive interleukin (IL)-17 production
infiltrates. The MOST likely diagnosis and management: E. Severe tooth decay caused by impaired neutrophil trafficking
A. Burkholderia cepacia complex pneumonia; initiate 3. GATA2 deficiency is a complex disease with manifestations
antibacterials involving hematopoietic and lymphatic development. It is
B. Nocardia spp. pneumonia; initiate antibacterials often identified in adolescence or adulthood with cytopenias
C. Serratia marcescens pneumonia; initiate antibacterials and/or infections. Signs and symptoms that should lead to
D. Mulch pneumonitis; initiate antifungals and corticosteroids early consideration of GATA2 deficiency include:
E. Staphylococcus aureus bacteremia; initiate antibacterials A. Human papilloma virus (HPV) infection, mycobacterial
and obtain echocardiogram
infection, monocytopenia
2. Leukocyte adhesion defect-1 (LAD-1) leads to low numbers B. Respiratory syncytial virus (RSV) infection, staphylococcal
of neutrophils in the tissues (tissue neutropenia) and recurrent pneumonia, monocytosis
severe infections. Progressive severe periodontitis is charac- C. Influenza infection, Aspergillus infection, thrombocytopenia
teristic, with most patients losing all their teeth by late D. Adenovirus infection, Bordetella infection, anemia
adolescence. The immunological mechanism behind the E. Cytomegalovirus (CMV) infection, streptococcal infection,
periodontal disease is: lymphocytosis

23









Mast Cells, Basophils, and Mastocytosis



Joshua Boyce, Laura Fanning







Mast cells and basophils are functionally related effector cells of committed precursors that migrate to their ultimate tissue destina-
hematopoietic origin that are implicated in allergy, type 2 immune tions before acquiring the morphological and biochemical
3
responses to parasites, and innate immunity. Both cell types characteristics of mature mast cells. Moreover, circulating
derive from bone marrow progenitors and express high-affinity basophils are short-lived and terminally differentiated, whereas
immunoglobulin E (IgE) Fc receptors (FcεRI) on their surface. mast cells can survive for long periods, and retain the capacity
Both release histamine and generate a variety of other inflam- for proliferation and substantial inducible gene expression. This
matory mediators in response to both IgE- and non–IgE-mediated likely explains the high degree of heterogeneity among mast cell
activation. However, there are important differences between subpopulations.
mast cells and basophils in terms of their developmental origins, In mice, committed mast cell progenitors (MCPs) can be
+
hi,
morphology, distribution, and mechanisms and responses to identified cytofluorographically as a Kit (CD117) FcεRI , β7
+
1
stimulation (Table 23.1, Fig. 23.1). This chapter will deal with integrin mononuclear cell population. A similar cell population
4
the functional characteristics of both cell types in host defense was recently identified in human blood. In ex vivo colony forming
+
+
+
and disease. assays, both mouse and human CD117 , FcεRI , β7 integrin cells
give rise preferentially to mast cells. In studies of cells sorted from
various organs BALB/c mice, committed basophil precursors
KEY CONCEPTS (BaPs) were found exclusively in bone marrow, whereas committed
Origins of Basophils and Mast Cells mast cell progenitors were most abundant in the small intestine.
2
This study identified a splenic population of cells capable of
• Mast cells and basophils represent distinct hematopoietic lineages. giving rise to both mast cells and basophils with ex vivo culture
• Basophils are granulocytes that mature in the bone marrow, circulate
in blood, and can be recruited into peripheral tissues at sites of (bipotent basophil–mast cell precursor [BMCP]). However, neither
inflammatory responses. mature mast cells nor mast cell progenitors show significant
• Mast cell progenitors arise in bone marrow, but mast cells mature in overlap in gene expression profiles with basophils in either mice
4,5
peripheral tissues. or humans. Thus despite some functional similarities, the bulk
• Mast cells are present throughout connective tissues and mucosal of evidence suggests that the developmental pathways of mast cells
surfaces and are particularly abundant near epithelial surfaces exposed and basophils diverge early in hematopoiesis and that the two
to the environment.
• CD117 and its ligand stem cell factor (SCF) are critical for mast cell cell types likely serve correspondingly very different functions.
development and survival. CD117 and its ligand, stem cell factor (SCF), are absolutely
• Basophil development is promoted in the context of T H 2 inflammatory required for mast cell development. SCF exists as both a
responses and regulated by cytokines, including interleukin-3 (IL-3) membrane-bound protein expressed by fibroblasts and other
and thymic stromal lymphopoietin (TSLP). stromal cells, as well as a soluble form. Mice with mutations in
sh
v
sh
d
the loci for CD117 (W/W or W /W mice) or SCF (Sl/Sl mice)
lack mast cells, among other hematopoietic and nonhematopoietic
DEVELOPMENT AND DISTRIBUTION defects. Although loss-of-function mutations in CD117 or SCF
1
OF MAST CELLS have not been identified in humans, gain-of-function mutations
6
in CD117 are associated with systemic mastocytosis, suggesting
Mast Cell Development and Survival that the SCF-KIT axis is important for human mast cell develop-
Mast cells are evolutionarily ancient, predating the emergence ment as well. Although SCF can stimulate mast cell development
of adaptive immunity. A mast cell–like cell containing histamine from either mouse or human progenitor cells in vitro, mouse
and heparin has been identified in Styela plicata, a urochordate mast cells can be derived in vitro from unfractionated bone
1
species that first appeared around 500 million years ago. Studies marrow cells by using interleukin-3 (IL-3) or thymic stromal
7
in rodents indicate that mast cells derive from hematopoietic lymphopoietin (TSLP) as the sole exogenous growth factors.
stem cell (HSC)–derived granulocyte–monocyte progenitor cells Mice lacking IL-3 have wild-type levels of mast cells under
(GMPs) in bone marrow, which can give rise to basophils and steady-state conditions but show diminished capacity to develop
2
other lineages. In mice, the transcription factors C/EBPα and a reactive mast cell hyperplasia in the intestine in response to
8
MITF interact antagonistically and specify basophil versus mast helminth infections. TSLP knock-out mice display modest
7
cell fate, respectively, in GMPs. Basophils undergo full differentia- steady-state reductions in mast cell numbers in all organs studied.
tion in bone marrow, whereas mast cells differentiate from Additionally, transgenic overexpression of IL-9 results in marked
335

336 ParT TwO Host Defense Mechanisms and Inflammation



TABLE 23.1 Natural History, Major Mediators, and Surface Membrane Structures of Human
Mast Cells and Basophils
Characteristic Basophils Mast Cells
Natural History
Origin of precursor cells Bone marrow Bone marrow
Site of maturation Bone marrow Peripheral tissues (a few in bone marrow)
Mature cells in the circulation Yes (usually <1% of blood leukocytes) No
Mature cells recruited into tissues Yes (during immunological, inflammatory No
from circulation responses)
Mature cells normally residing in No (not detectable by microscopy) Yes
connective tissues
Proliferative ability of morphologically None reported Yes (in certain circumstances)
mature cells
Life span Days (like other granulocytes) Weeks to months (based on studies in rodents)

Mediators
Major mediators stored preformed in Histamine, chondroitin sulfates-glucuronidase, Histamine, heparin and/or chondroitin sulfates,
cytoplasm elastase, cathepsin G-like enzyme, eosinophil chymase and/or tryptase, many acid hydrolases,
cationic protein, major basic protein, Charcot- cathepsin G, carboxypeptidase A
Leyden crystal protein, eosinophil peroxidase,
some positive for tryptase, chymase,
carboxypeptidase A
Major lipid mediators produced upon LTC 4 , PAF PGD 2 , LTC 4 , LTB 4 , PAF, TXA 2
appropriate activation
Cytokines and growth factors released IL-2, IL-3, IL-4, IL-5, IL-6, IL-9, IL-13, IL-15, TSLP IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-8, IL-9, IL-10, IL-11,
upon appropriate activation IL-13, IL-15, IL-16, IL-17A, IL-18, IL-22, IL-24, IL-25,
IL-33, TNF-α, SCF, NGF, TGF-β, FGF-2, VEGF, IFN-α,
GM-CSF, TSLP, amphiregulin
Chemokines produced CCL3, CCL4, CCL12, CXCL2 CCL1, CCL2, CCL3, CCL4, CCL5, CCL7, CCL8, CCL13,
CCL17, CCL18, CCL19, CCL20, CCL22, CCL23,
CCL25, CCL28, CXCL2, CXCL3, CXCL4, CXCL5,
CXCL7, CXCL8, CXCL10, CXCL14, CXCL16, CXCL17,
XCL1, CX3CL1
Surface Structures
Ig receptors FcεRI, FcγRII (CDw32), FcγRIIB, FcγRIIIA FcεRI, FcγRI, FcγRIIA, FcγRIIB, FcγRIIIA
Cytokine/growth factor receptors IL-1RIIb (CD121b), IL-2R (CD25), IL-3R, IL-4Rα, KIT (SCF receptor), IL-3R, IL-4Rα, IL5Rα, IL-6R, IL-18R,
IL-5Rα, IL-18R, IL-33R, TSLPR, GM-CSFRα, TrkA IL-33R, IFN-γRα, TGF-β type-I and type-II β receptors,
TrkA, T1/ST2/IL-1R4
Chemokine receptors CCR1, CCR2, CCR3, CCR5, CXCR1, CXCR2, CCR1, CCR3, CCR4, CCR5, CXCR1, CXCR2, CXCR3,
CXCR4 CXCR4, CX3CR1
Other GPCRs CRTH2, C3aR, C5aR EP3, BLT1, BLT2, cysLT1, PAF receptor, C3aR, C5aR
LT, leukotriene; PG, prostaglandin; PAF, platelet-activating factor; TXA 2, thromboxane A 2; IL, interleukin; TSLP, thymic stromal lymphopoietin; SCF, stem cell factor; NGF, nerve
growth factor; TGF-β, transforming growth factor-β; FGF-2, fibroblast growth factor-2; VEGF, vascular endothelial growth factor; IFN-α, interferon-α; GM-CSF, granulocyte
macrophage–colony-stimulating factor; CCL, chemokine (C-C motif) ligand; CXCL, chemokine (C-X-C motif) ligand; XCL, chemokine (C motif) ligand; CX3CL, chemokine (C-X3-C
motif) ligand; TrkA, tropomyosin receptor kinase A; CCR, C-C chemokine receptor; CXCR, C-X-C chemokine receptor; CX3CR, C-X3-C chemokine receptor; GPCRs, G protein–
coupled receptors; CRTH2, chemoattractant receptor-homologous molecule expressed on Th2 cells; C3aR, complement component 3a receptor; C5aR, complement component 5a
receptor.





9
hyperplasia of mast cells in multiple organs and tissues. However, strong induction of leukotriene C 4 synthase (LTC 4 S) expression.
v
sh
sh
the profound reduction of mast cells in W/W , W /W , and Sl/ Thus cytokines other than SCF are likely to play context-specific
Sld mice argue that none of these cytokines can substitute for roles in mast cell development and may help to account for
the absence of SCF or CD117. the substantial heterogeneity of anatomically and functionally
In contrast to mice, SCF is absolutely required to derive distinct mast cell subpopulations (see below). Tissue mast cells
mast cells in vitro from human bone marrow, fetal liver, or cord vary widely in their capacity for cysLT generation, which may
blood progenitors. IL-3 cannot induce mast cell development reflect cytokine-induced modification of their effector phenotype
from human hematopoietic progenitor cells, although it does in vivo.
can enhance survival of human cord blood mast cells (as do
10
IL-4, IL-5, or IL-6). Human intestinal mast cells proliferate in Homing, Distribution, and Heterogeneity of Mast Cells
response to IL-4 when it is provided in combination with SCF. Mature mast cells are distributed throughout connective tissues
Notably, IL-4 primes human cord blood and/or intestinal mast and mucosal surfaces, often adjacent to blood or lymphatic vessels
cells for FcεRI-dependent degranulation, cytokine generation, or peripheral nerves. Much of the current understanding of mast
11
and cysteinyl leukotriene (cysLT) generation, the latter reflecting cell tissue homing comes from mouse models. MCPs home

CHaPTEr 23 Mast Cells, Basophils, and Mastocytosis 337


carboxypeptidase A3 (CPA3). Accordingly, the anatomically
distinct mouse mast cell subpopulations express very different
profiles of proteases (and correspondingly different GAG composi-
tions), dependent on the microenvironment. 16
In contrast to the mouse, human mast cells express only a
single mast cell–specific chymase gene and thus exhibit less
heterogeneity compared with their rodent counterparts. Human
mast cells can express up to four tryptase genes (α, β, γ and δ),
and CPA3. All human mast cells express tryptase β, and some
also express chymase, CPA3, and cathepsin G. Human mast cells
expressing tryptase in the absence (MC T ) or presence (MC TC )
FIG 23.1 Ultrastructures of Mast Cell and Basophil. A basophil of chymase, CPA3, and cathepsin G differ in anatomical distribu-
(B) adjacent to a mast cell (MC) in the ileal submucosa of a tion, with MC T dominating in normal lung, intestinal mucosa,
patient with Crohn disease. The basophil exhibits a bilobed and bronchial and nasal epithelium and MC TC dominating in
16
nucleus (solid arrows) whose chromatin is strikingly condensed skin, heart, uterus, and deep connective tissues. These distribu-
beneath the nuclear membrane. The basophil surface is relatively tions can change with disease and inflammation. For example,
smooth with a few blunt processes (arrowhead). The mast cell the fraction of MC TC among bronchial submucosal mast cells
17
nucleus is larger and its chromatin less condensed than that of increases markedly with disease severity in asthma, as do the
18
the basophil. The mast cell’s granules are smaller, more numer- numbers of MC TC infiltrating the airway smooth muscle. MC TC s
19
ous, and more variable in shape and content than those of the accumulate selectively in atherosclerotic plaques. In allergic
basophil. The mast cell surface has numerous elongated, thin rhinitis and eosinophilic esophagitis, MC T selectively accumulate
folds (curved arrows). (Original magnification ~ × 9000). (From in the inflamed mucosal surfaces. Thus while the extent and
Dvorak AM, Monahan RA, Osage JE, Dickersin GR. Crohn’s defining features of mast cell heterogeneity differ between human
disease: transmission electron microscopical studies. Hum Pathol and rodent mast cells, there is considerable lineage plasticity in
1980;11:606–619, with permission from Ann M. Dvorak.) both species that reflect both tissue localization and context.
One of the most strongly conserved aspects of mast cell
heterogeneity is the dependency, or lack thereof, on T cell–derived
accessory growth factors. Athymic nude mice altogether lack
constitutively to the mouse small intestine, requiring both α 4 β 7 MMCs in the intestinal mucosa despite the presence of wild-type
20
12
integrin, as well as the chemokine receptor CXCR2. Although levels of CTMCs in the same organ. MC T are lacking in the
the lungs of naïve mice contain few mast cells, MCP recruitment intestines of humans with T-cell immunodeficiencies, but MC TC
21
can be experimentally induced in response to allergen sensitization numbers are unaffected. The differential requirement of mast
and challenge. In this context, MCP recruitment to the lung is cell subsets for T-cell help likely reflects the capacity of T cell–
controlled by α 4 β 7 as well as α 4 β 1 and its counterligand, vascular derived growth factors to synergize with SCF to promote mast
12
cell adhesion molecule-1 (VCAM-1). To date, no studies have cell proliferation and/or survival. This may be functionally
addressed the requirements for homing or recruitment of MCPs important, since T cell–driven expansion of mast cells in the
to skin, the central nervous system, or other characteristic destina- intestine is essential for the elimination of certain helminthic
tions. Cultured human mast cells express the chemokine receptors infections. Moreover, the capacity of IL-4 and IL-9 to alter
13
CCR3 and CXCR4 and respond chemotactically to their cor- expression of key mast cell effectors suggests that effector systems
responding ligands. Mast cells localizing to asthmatic airway can be modulated by T cells to suit the needs of the host during
smooth muscle express CXCR3, and lung mast cell migration immune responses. 9,11
can be induced by airway smooth muscle cultures via activation
14
of CXCR3. However, definitive information regarding the DEVELOPMENT AND DISTRIBUTION
homing requirements for human mast cells in vivo is lacking OF BASOPHILS
because of technical constraints.
Once in the tissues, MCPs give rise to anatomically and Basophils are the rarest of granulocytes in mammals (<1% of
biochemically distinct subpopulations of mast cells. Rodent mast peripheral blood leukocytes) and take their name from the
cell subsets are often referred to as mucosal mast cells (MMCs) basophilic granules in their cytoplasm. The scarcity and short
and connective tissue mast cells (CTMCs), respectively, based on life span (~2.5 days in mice) of basophils hindered the study of
differences in anatomical distribution and staining characteristics. their functions until recently, but there is now a growing apprecia-
Early studies of Kanakura et al. used adoptive transfers to tion for their roles in various immune processes that are distinct
demonstrate that both MMCs and CTMCs arose in vivo from from the mast cell. These include FcεRI-dependent antigen capture
15
a single population of progenitor cells. The differences between and a potent and rapid capacity to generate and secrete IL-4
MMCs and CTMCs in staining characteristics relate to the relative and IL-13.
content of heparin versus chondroitin sulfate glycosaminoglycans Basophils are myeloid cells that originate and develop in bone
(GAGs) bound to the serglycin core of mast cell granules. These marrow. Like mast cells, they develop from GMPs, which become
GAGs impart the basophilic staining characteristics of mast cell BMCPs or BaPs, each of which has the capacity to develop into a
2,3
granules and are essential for the storage and stability of highly mature basophil. The basophil lineage commitment is regulated
charged serine proteases and histamine that comprise the major by the transcription factors GATA-2 and C/EBPα. Unlike mast
preformed products of mast cells. The mouse genome encodes cells, basophils exit bone marrow only after they have fully
four mast cell–restricted tryptases and several chymases (at matured. Basophil development is promoted in the context
least five of which can be expressed by mast cells), as well as of T-helper cell-2 (Th2) inflammatory responses. Cytokines,

338 ParT TwO Host Defense Mechanisms and Inflammation


including IL-3 and the epithelial cell–derived cytokine TSLP, KEY CONCEPTS
regulate basophil development as well. 3
Ex vivo developmental studies with human progenitor cells Mast Cell and Basophil Mediators
have suggested a common origin for basophils and eosinophils. • Mast cells and basophils produce distinct but overlapping panels of
Denburg et al. showed that a fraction of clonal colonies arising mediators with diverse biological effects.
from human peripheral blood progenitor cells contained both • Some mast cell and basophil mediators are preformed and stored in
eosinophils and basophils, with some cells showing an intermedi- cytoplasmic granules (e.g., histamine, proteoglycans, proteases). These
22
ate phenotype with granules of both lineages. Saito et al. showed can be released rapidly (seconds to minutes) upon cellular activation
and degranulation.
that recombinant IL-3 stimulated the growth of both eosinophils • Mast cells and basophils produce lipid mediators (e.g., PGD 2 , LTC 4 ),
23
and basophils from cord blood progenitors. Human basophils which are derived from arachidonic acid and are newly synthesized
express eosinophil cationic protein, major basic protein, Charcot- upon cell activation.
Leyden crystal protein, and eosinophil peroxidase, all of which • Mast cells and basophils can transcribe and secrete many cytokines,
24
are eosinophil granule markers, consistent with a common chemokines, and growth factors (see Table 23.1).
origin of both cell types. A recent transcriptional profiling study
confirmed that mouse blood basophils are more similar to
eosinophils in their gene expression pattern than to any other
5
cell type. The shared lineage pathway for basophils and eosino- are among the oldest antiallergy medications, and their efficacy
phils explains why blood basophil counts are frequently elevated supports the role of histamine in mediating the clinical signs
in conditions associated with increased blood eosinophil counts. and symptoms of rhinitis, allergic conjunctivitis, and urticaria.
Basophil recruitment to the tissues is likely governed by a Proteases, including tryptases, chymase, and CPA3, comprise
combination of protein and lipid chemoattractants. Human the major protein component of mast cell secretory granules.
basophils express CCR1, CCR2, CCR3, CCR5, CXCR1, and CXCR2 Tryptases are the most abundant proteases in the human mast
and respond ex vivo to the corresponding ligands. Basophils cell and are found in basophils as well. Human tryptases are
strongly express chemoattractant receptor homologue expressed encoded by several different alleles, including α, β, γ, and δ. As
by Th2 cells (CRTH2), a specific receptor for prostaglandin D 2 noted previously, only a single mast cell–specific chymase gene
(PGD 2 ). PGD 2 is an abundant product of activated mast cells, exists in humans. CPA3 is expressed strongly by mast cells and
1,16
and CCR3 ligands (CCL11, CCL24, CCL26) are abundantly weakly by basophils. Tryptase β can activate protease-activated
1
expressed by structural cells in allergic inflammation. Thus all receptors, inducing fibroblast proliferation and collagen secretion.
of these are good candidates to account for the accumulation Both human tryptase β and its murine orthologue, mouse mast
of basophils that is frequently observed in rhinitis and asthma, cell protease 6, can induce neutrophil recruitment in models of
and that is characteristic of allergen-induced late-phase responses. bacterial sepsis, suggesting prominent functions in innate
immunity. Human mast cell chymase converts angiotensin 1 to
BIOLOGICAL MEDIATORS PRODUCED BY MAST angiotensin 2, suggesting a potential role in the control of blood
CELLS AND BASOPHILS pressure and cardiovascular homeostasis. CPA3 can neutralize
25
endothelin 1 and the snake venom protein sarafotoxin. These
vasoprotective and antivenom functions are consistent with a
Mast cells and basophils can elaborate a broad array of potent role for resident cutaneous and perivascular mast cells vascular
biologically active mediators (see Table 23.1). Some of these homeostasis and host defense. The functions of the mast cell
products are stored preformed in cytoplasmic granules (e.g., proteases are reviewed comprehensively elsewhere. 16,26
histamine, proteases, proteoglycans, certain cytokines), and others
are synthesized de novo upon activation of the cell (e.g., products Newly Synthesized Mediators
of arachidonic acid oxidation via the cyclooxygenase [COX] or Both mast cells and basophils utilize membrane phospholipid-
lipoxygenase pathways, platelet-activating factor [PAF], cytokines, derived arachidonic acid to synthesize eicosanoid mediators and
19
chemokines, growth factors). These mediators enable mast cells can use the resultant lysophospholipids to generate PAF. These
and basophils to orchestrate various functions in inflammation mediators are released within 15 minutes of activation. Both
and host defense. Some, such as histamine and cysLT, are targets mast cells and basophils express 5-LO and LTC 4 S, permitting
of the best-established drugs for the treatment of allergic diseases, them to generate LTC 4 , the parent cysLT, from endogenous
validating the pathogenetic importance of mast cells, basophils, arachidonic acid. Mast cells, but not basophils, can generate
and their associated effector systems. smaller quantities of the dihydroxy leukotriene, LTB 4 , which is
a potent neutrophil chemoattractant. LTC 4 is converted extracel-
Preformed Mediators lularly to LTD 4 (the most potent known bronchoconstrictor)
Mediators that are stored in secretory granules can be rapidly and then to the stable metabolite LTE 4 . 3,19 CysLTs are abundant
(seconds to minutes) released into the extracellular environment in allergic inflammation, increasing microvascular permeability
upon stimulation. These mediators include histamine, proteo- and causing bronchoconstriction. Mouse models suggest they
glycans, and (in the case of mast cells) proteases. 19 play important roles in amplifying Th2 responses through several
Histamine is stored in the granules of mast cells and, to a cellular targets. There are three cysLT receptors, respectively
27
lesser extent, basophils. Histamine is released within minutes termed CysLT 1 R, CysLT 2 R, and GPR99. Both CysLT 1 R antagonists
of cell activation and mediates increased vascular permeability, and 5-LO inhibitors show efficacy in asthma, supporting the
smooth muscle contraction, mucus production, and increased pathogenic role of the cysLT in this disease.
heart rate and cardiac output, gastric acid secretion, itching, and In contrast to basophils, mast cells generate substantial quanti-
sneezing. These actions are mediated via four types of histamine ties of PGD 2 , a product of successive metabolism of arachidonic
receptors (H1–4) on target cells. 1,3,16 H1 receptor antagonists acid by COX-1 or COX-2 and hematopoietic PGD 2 synthase, an

CHaPTEr 23 Mast Cells, Basophils, and Mastocytosis 339


enzyme that is expressed more strongly by mast cells than any chains. Human hematopoietic cells other than mast cells and
other cell type. PGD 2 binds to two specific receptors, termed basophils (e.g., eosinophils and dendritic cells [DCs]) express
DP 1 and CRTH2 (also known as DP 2 ). Once released, PGD 2 FcεRI in the absence of the β chain. The β chain amplifies signal-
elicits vasodilation (via DP 1 ) and bronchoconstriction (via the ing through the receptor, and the tetrameric form on mast cells
29
actions of a putative metabolite at the T prostanoid receptor). and basophils is thus fully functional. Activation occurs when
Additionally, PGD 2 is potently chemotactic for basophils, adjacent FcεRI receptors, occupied by receptor-bound IgE, are
eosinophils, Th2 cells, and group 2 innate lymphoid cells (ILC2), cross-linked by multivalent antigen, stimulating degranulation
19
each of which express CRTH2. PGD 2 induces cytokine generation and release of preformed mediators as well as de novo production
(IL-4, IL-5, IL-13) from both human Th2 cells and ILC2 and and release of additional mediators described above. 1,3,19,29 The
could, therefore, amplify type 2 inflammation in a polyclonal signaling cascades that are initiated by FcεRI cross-linking are
fashion. There are several CRTH2-selective antagonists in clinical reviewed in detail elsewhere. 30
development for the treatment of asthma and other allergic At the ultrastructural level, stimulation of sensitized human
diseases. basophils with specific antigen provokes fusion of the membranes
enveloping individual cytoplasmic granules with the plasma
Cytokines, Chemokines, and Growth Factors membrane (Fig. 23.2). This results in the release of the granules’
Basophils and mast cells produce a wide variety of cytokines contents via multiple narrow communications between single
and chemokines that recruit and activate other cells (see Table granules and the cell surface. IgE-dependent degranulation of
23.1). Mast cells generate tumor necrosis factor-α (TNF-α), IL-6, human lung mast cells results in the fusion of granule membranes
and IL-1β, which is consistent with their sentinel role in innate with the plasma membrane (Fig. 23.3). However, in this cell type
antimicrobial immunity (Chapter 3). Mast cells can generate the first ultrastructural changes detectable in the stimulated cells
the Th2 cytokines IL-5 and IL-13, a functional property that is are granule swelling, followed by fusion of individual granule
induced when the cells are exposed to IL-4. TNF-α can be stored membranes forming interconnecting chains of swollen granules;
in granules constitutively as well as produced de novo in response histamine release is initiated by the opening of these channels
19
to activation. Activated mouse and human mast cells express to the exterior through multiple narrow points of fusion with
a large range of both CC and CXC chemokines, likely reflecting the plasma membrane. 31
their role in facilitating the recruitment of bloodborne effector
cells. A recent study demonstrated that mast cell progenitors in the Non–IgE-Mediated Activation
intestinal mucosa are an important source of IL-9 and IL-13 in a Mast cells and basophils can be activated through a variety of
28
model of food allergy. Basophils can secrete large quantities of non–IgE-mediated mechanisms. Both mast cells and basophils
IL-4 (and IL-13) rapidly, contributing to Th2 immune responses express receptors for complement fragments C3a and C5a and
in a variety of settings. These observations suggest that basophils release histamine in response to these complement components.
are more specialized than mast cells for a role in type 2 immunity. The C5a receptor (CD88), in particular, is expressed in human
lung on MC TC s, and elevated C5a concentrations have been
MECHANISMS OF ACTIVATION OF MAST CELLS measured in induced sputum from subjects with asthma. Mast
2,19
AND BASOPHILS cells and basophils can be activated by microbial constituents
via Toll-like receptors (TLRs), many of which are expressed on
their surface and internal membranes. 19,29 Both cell types respond
to the barrier-derived cytokines IL-33 and TSLP, both of which
KEY CONCEPTS drive Th2-type immune responses. Each cytokine can directly
Mechanisms of Mast Cell and Basophil Activation activate mast cells and, in some cases, amplify IgE-mediated
29
responses. In contrast to the complement fragment-dependent
• Mast cells and basophils have cell-surface high-affinity immunoglobulin activation, IL-33 and TSLP fail to induce degranulation but are
E (IgE) receptors (FcεRI) that are tetrameric and thus fully functional, powerful inducers of cytokine production, particularly when
unlike the trimeric FcεRI found on other cells (e.g., dendritic cells they are provided together. Thus non–IgE-dependent activation
[DCs] and eosinophils).
• IgE-mediated activation of mast cells and basophils occurs when mechanisms tend to elicit a response from mast cells that is
multivalent antigen cross-links IgE antibodies bound to adjacent FcεRI more compartmentalized than FcεRI-mediated activation.
receptors, resulting in degranulation and de novo mediator production A variety of drugs, such as opiates and muscle relaxants, can
and release. directly activate mast cells. 19,29 Recently, the murine counterpart
• Mast cells and basophils can be activated by non–IgE-mediated of the human G protein-coupled receptor, MAS-related G
mechanisms (anaphylatoxins, Toll-like receptor [TLR] ligands, certain protein-coupled receptor X2 [MRGPRX2], was found to be
cytokines, drugs, venoms, etc.). necessary to trigger mast cell degranulation in vivo response to
• Some types of non–IgE-mediated activation cause a more compart-
mentalized activation (e.g., cytokine production without degranulation a range of basically charged drugs, as well as compound 48/80,
32
caused by activation of mast cells or basophils by interleukin-33 [IL-33] a classic secretagogue for mast cells. The potential that certain
and thymic stromal lymphopoietin [TSLP]). drugs possess in their ability to mimic components of innate
immunity to trigger mast cells represents a major conceptual
FcεRI-Mediated Activation (and Inhibition of step forward toward understanding drug-induced pseudoallergic
reactions that do not involve specific IgE.
IgE-Dependent Activation) Basophil activation is promoted by IL-3, and other cytokines,
Both mast cells and basophils express the high-affinity IgE including IL-18 and IL-33, can activate basophils and enhance
3
receptor, FcεRI. The FcεRI expressed on these cells is unique in their effector functions. Basophils can be directly activated by
3
that it has a tetrameric structure consisting of a single IgE-binding protease allergens, including the house dust mite protease Derp1.
αα chain, a single β chain, and two identical disulfide-linked γ Finally, both mast cells and basophils respond to PAF with

340 ParT TwO Host Defense Mechanisms and Inflammation


























A




B
FIG 23.2 (A) Transmission electron micrograph of a human basophil in a preparation of peripheral
blood leukocytes obtained by separation over Ficoll-Hypaque. All of the cytoplasmic granules
(some indicated by solid arrows) contain particulate electron-dense material. N, nucleus. (Original
magnification ~ × 19 800). (B) A human basophil 2 minutes after exposure to antigen in vitro.
The cell exhibits extrusion of granules from six separate sites on the plasma membrane (small
arrows). At this time after cell stimulation, particle-filled granules retain their shape and characteristic
structure even after exposure to extracellular milieu. Cationized ferritin coats the cell surface and
enters culs-de-sac that contain exteriorized granules. The cell exhibits no fully intracytoplasmic
typical basophilic granules, but one of the smaller kind of granules (curved arrow) can be observed
in the perinuclear region. N, nucleus. (Original magnification ~ × 19 200). (From Dvorak AM,
Newball HH, Dvorak HF, Lichtenstein LM. Antigen-induced IgE-mediated degranulation of human
basophils. Lab Invest 1980;43:126–139, with permission from Nature Publishing Group Ltd.)


activation and histamine release, and PAF concentrations are
33
elevated in the plasma after anaphylactic reactions. These Allergic Disease
findings suggest a mechanism by which endogenous lipid Anaphylaxis
metabolism can magnify the physiological effects of mast cell Anaphylaxis is an acute, life-threatening, multisystem reaction
activation in anaphylaxis. that results from the precipitous release of mast cell and basophil
mediators into the circulation (Chapter 42). These mediators cause
MAST CELLS AND BASOPHILS IN DISEASE AND the cardinal manifestations of hypotension, bronchoconstriction,
HOST DEFENSE intestinal colic, urticaria, and, in some instances, disseminated
intravascular coagulation. Anaphylactic reactions can be triggered
by foods, insect stings, and medications, among other factors.
Anaphylaxis, like mast cell and/or basophil activation, can be
CLINICaL rELEVaNCE immunological or nonimmunological.
Mast Cells and Basophils in Health and Disease The classic mechanism of anaphylaxis in human disease is
IgE mediated, with an interaction between antigen and antigen-
• Mast cells and basophils are primary effector cells in atopic disorders specific IgE bound to FcεR1 on mast cells and/or basophils. In
(anaphylaxis, asthma, allergic rhinitis, atopic dermatitis). this instance, anaphylaxis can be considered a severe form of
• Antigen-specific, immunoglobulin E (IgE)-mediated mast cell activation type I hypersensitivity. Production of histamine and leukotrienes
results in an immediate clinical reaction and, in some cases, contributes
to a late-phase reaction. in response to IgE-mediated activation of both mast cells and
• Medications that block mast cell and basophil mediators are mainstays basophils induces smooth muscle contraction and contributes
3
of treatment for atopic diseases. to the symptoms and physiology of systemic anaphylaxis. Besides
• Mast cells and basophils contribute to host defense against parasites the classically described IgE-mediated type of anaphylaxis, IgG-
and bacterial pathogens. driven, PAF-mediated anaphylaxis has also been described in
• Mast cells may play a role in other human diseases, including athero- mouse models. Mast cells are dispensable for this type of ana-
sclerosis, autoimmune diseases, and burns and in responses to venom
toxins. phylaxis, whereas basophils (among other lineages) have been
3
reported as PAF-producing effector cells. PAF levels are elevated

CHaPTEr 23 Mast Cells, Basophils, and Mastocytosis 341























B








A
FIG 23.3 (A) Mast cell purified from human lung. The cell contains many cytoplasmic granules
with scroll-like substructural elements (solid arrows) and eight large non–membrane-bound lipid
bodies (open arrows). The plasma membrane has prominent folds. N, nucleus. (Original magnification
~ × 12 900). (B) An isolated human lung mast cell 10 minutes after exposure to anti-IgE in vitro.
Some degranulation channels (C), formed by fusion of membranes surrounding individual cytoplasmic
granules, contain altered granule matrix; others (EC) are empty. Cationized ferritin stains the
plasma membrane and the membranes of some empty degranulation channels (EC). The membranes
lining other channels (C) are unstained. A few unaltered scroll-containing granules (solid arrows)
remain. Numerous lipid bodies are also present (open arrows). The cytoplasm contains prominent
filaments. N, nucleus. (Original magnification ~ × 9 100). (From Galli SJ, Dvorak AM, Dvorak HF.
Prog Allergy 1984;34:1, with permission from Ann M. Dvorak.)


19
in the serum of patients with severe peanut anaphylaxis, suggesting prostanoids. The involvement of mast cells in early- and late-
that PAF may serve to amplify the physiological manifestations phase responses is supported by earlier studies using mast cell
of IgE-dependent anaphylaxis. Immune complex–mediated stabilizing cromone drugs, as well as anti-IgE.
complement activation has been proposed as a mechanism Immunohistological studies provide additional support for
for some anaphylactic reactions to drugs. Nonimmunological the role of mast cells in asthma. Bronchial biopsies from subjects
anaphylaxis can be caused by complement activation in the with asthma exhibit mast cells localizing to the airway smooth
absence of immune complexes or by direct activation of mast muscle bundles, a region generally lacking mast cells in biopsies
34
cells, often by drugs, such as opiates or vancomycin. As noted from healthy controls and from subjects with nonasthmatic
19
above, some of these (particularly muscle relaxants) may result eosinophilic bronchitis. Smooth muscle mast cell density cor-
from drugs that activate MRGPRX2 or related receptors. 32 related strongly with disease severity and with hyperresponsiveness
35
to methacholine, a cardinal feature of asthma. Notably, these
Asthma increases were observed irrespectively of atopic status of the study
Asthma is a chronic inflammatory disease of the airways character- subjects. In biopsy samples from subjects with severe asthma,
ized by variable airflow obstruction and airway hyperresponsive- mast cells exhibit substantially higher proportions with positive
ness (AHR) (Chapter 41). The most direct evidence for a immunostaining for chymase than in specimens from those
mechanistic contribution of mast cells to allergic asthma comes with mild asthma. Thus both the distribution and phenotype
from allergen challenge studies. In sensitized individuals, allergen of mast cells is altered in the airways of patients with severe
challenge causes an increase in mast cell granule contents in asthma. The mechanisms responsible for these alterations, and
bronchoalveolar lavage (BAL) fluid, including histamine, tryptase, the factors responsible for driving mast cell activation in severe
and cysLTs. About 60% of patients having an immediate, early- disease, particularly in individuals without atopy remain to be
phase asthmatic response will have a late-phase reaction beginning determined.
3–5 hours later, peaking around 8 hours, and resolving in about Mast cells may play an especially important role in aspirin-
24 hours. Mast cells contribute to both the early-phase and the exacerbated respiratory disease (AERD), a distinctive disorder
late-phase asthmatic response, via the release of histamine and in which asthma, severe rhinosinusitis, and nasal polyposis are
production of proinflammatory cytokines, leukotrienes, and associated with idiosyncratic respiratory reactions to aspirin and

342 ParT TwO Host Defense Mechanisms and Inflammation


other drugs that inhibit COX-1. Although tissue mast cell burden around sites of parasitical infestation. Subsequent studies have
in AERD does not exceed that seen in aspirin-tolerant control demonstrated a role for mast cells in mouse models of helminth
patients with asthma, baseline levels of serum tryptase and urinary infection in the gut as well as in parasitical skin infections. 37
PGD 2 metabolites are higher, potentially reflecting a loss of Production of IL-4 and IL-13 by mast cells and basophils
homeostatic PGE 2 from the surrounding tissue. These values may play an important role in early parasitical infections, leading
increase markedly with aspirin challenges, and treatment with to IgE production, secretion of effector molecules, activation of
cromolyn or nedocromil blocks the characteristic reactions, smooth muscle cells, and increased mucus production, among
36
1
supporting their mast cell dependency. Notably, a mouse model other effects. Furthermore, although expulsion of hookworm
suggests that mast cell activation in response to aspirin challenge during primary infection in mice depends on mast cells, basophils
36
depends on IL-33 rather than the allergen. It is tempting to are necessary to expel hookworms during secondary infection,
1
speculate than innate cytokines could account for mast cell possibly because of activation by helminth-specific antibodies.
activation in other nonatopic disorders. Intestinal helminth infections typically elicit a T cell-dependent
There is growing evidence that basophils play a role in the expansion of mast cells and basophils in the involved mucosal
8
pathogenesis of asthma as well. Basophils are enriched in post- site. In that regard, mast cells and basophils can be viewed as
mortem human lung tissue from patients who have died as a components of the effector arm of adaptive immunity.
result of asthma as well as in bronchial biopsy specimens of Mast cells are important in the generation of immune
1,3
patients with asthma. In mouse models of allergic airway responses to bacterial pathogens. For example, mast cell deficient
inflammation, basophils have been shown to promote Th2 mice have increased mortality in a model of Escherichia coli–
3
cytokine responses and pathological airway inflammation. The induced peritonitis. Mast cells have shown protective roles in K.
specific functions of basophils relative to asthma are not yet pneumoniae lung infections and Pseudomonas skin infections in
37
known, though it seems likely that they could contribute Th2 mice. Mice lacking mast cells as a result of loss-of-function Kit
cytokines and cysLTs. and SCF mutations are significantly more susceptible to lethal
gram-negative sepsis than are congenic mast cell–sufficient
Allergic Rhinitis controls. Reconstitution of the tissues with wild-type mast cells
Allergic rhinitis is an inflammatory disease of the nasal mucosa restores protective immunity. Both proteases and TNF-α are
that occurs as a reaction to airborne allergens (Chapter 41). linked to the protection from bacterial infections conveyed by
Increased numbers of activated MC T mast cells are found in the mast cells.
nasal epithelium of patients with allergic rhinitis, and these
numbers increase further with allergy provocation challenge or Mast Cells in Other Diseases
34
during seasonal disease exacerbations. Basophils are present Mast cells may play a role in other disease processes as well.
3
in nasal washes from patients with allergic rhinitis. In addition, Increased numbers of mast cells have long been noted in human
nasal challenges with allergens lead to the release of histamine atherosclerotic lesions compared with healthy control tissue.
and cysLTs, which are preformed mediators of both mast cells Mast cell functions within these plaques are not completely
and basophils. 34 understood, although mouse models suggest that atherosclerotic
changes are promoted by release of proinflammatory cytokines
Atopic Dermatitis by mast cells. Mouse models of autoimmunity, including
38
40
39
Atopic dermatitis (AD) is a chronic, relapsing inflammatory skin autoantibody-mediated arthritis and multiple sclerosis, have
condition that is commonly diagnosed in young children and demonstrated a pathological role for mast cells. Skin mast cells
often precedes the development of allergic rhinitis and asthma are activated rapidly following thermal trauma, and the release
(Chapter 44). The pathogenesis of AD remains incompletely of mast cell proteases is crucial for development of the inflam-
understood but is likely multifactorial with contributions from matory response in burn lesions. 41
skin barrier dysfunction, innate and adaptive immune dysregula- Recent work in mice has shown that mast cells protect from
tion, and environmental and genetic factors. Increased numbers the detrimental effects of arthropod or reptile venoms via the
of mast cells can be seen in skin lesions from human AD and degradation of venom toxins by mast cell proteases. Furthermore,
mouse AD models, but their role in the pathophysiology of the acquired Th2 immune responses associated with IgE production
disease is not known. 34 can protect mice from challenge with potentially lethal doses of
Basophils have been identified in lesional skin of patients venoms from honeybee or the Russell viper. 42
with AD, but their precise role in human disease remains unclear.
In murine models of AD-like skin disease, significant TSLP- Mast Cell Roles in Normal Physiology
dependent basophilia has been noted in lesional skin. TSLP is In addition to their immune functions, mast cells are thought
also associated with human AD, so it is possible that TSLP-induced to have normal physiological functions as well. Data from mouse
basophil populations could play a role in human disease as well. 1,3 studies suggest a role for mast cells in intestinal barrier function
43
and homeostatic intestinal epithelial migration. Human uterine
Mast Cells and Basophils in Immunity tissue has increased density of mast cells during pregnancy, where
44
Although mast cells are found throughout most tissues, they they may mediate contractility, in addition to contributing to
are notably located at interfaces with the environment, such as local immunity.
skin and the mucosal surfaces. This suggests a potential role in
pathogen recognition and host defense. Additionally, mast cells MASTOCYTOSIS AND OTHER
have the capacity to increase in numbers during mucosal inflam- MAST CELL DISORDERS
matory responses, suggesting that their effector capabilities can be
magnified to suit the context. The first associations of mast cells A variety of human disorders are associated with excessive
with host defense included observations of mast cells clustering numbers of mast cells, increased activation of mast cells, or both.

CHaPTEr 23 Mast Cells, Basophils, and Mastocytosis 343


Mastocytosis can be limited to skin or have widespread systemic Skin findings may be seen in both CM and SM, and the most
effects. Mast cell disorders cause a broad range of symptoms common skin-related symptom in patients with mastocytosis is
that can mimic a variety of other conditions, often making pruritus. Accumulations of mast cells in skin can present as a
diagnosis a challenge. variety of lesions. Urticaria pigmentosa (UP), also called macu-
lopapular cutaneous mastocytosis (MPCM), is the most common
Epidemiology skin manifestation of mastocytosis in both adults and children.
Mastocytosis can occur at any age, although it tends to have a Lesions of UP appear as small, tan to reddish-brown macules
more benign and transient course in children and often is limited or slightly elevated papules (Fig. 23.5A–C). UP is present more
to cutaneous manifestations. In a Polish case series of 100 children commonly in the less aggressive forms of SM.
with cutaneous mastocytosis (CM), 73% had onset of the disease Diffuse cutaneous mastocytosis (DCM) is characterized by
45
by age 6 months and 94% within the first year of life. Most edema and increased skin thickness, with or without a yellowish-
cases of systemic mastocytosis (SM) are diagnosed in middle brown coloration. DCM is rare, accounting for 1–3% of CM.
age. The exact prevalence of mastocytosis is unknown, but Cutaneous mastocytomas are well-demarcated flat or slightly
an estimate from a recent Danish population-based study is elevated lesions that may have a yellow or red-brown coloration.
46
approximately 1 : 10 000. With rare exceptions, mastocytosis Mastocytomas can be solitary or multiple and are typically 2–5 cm
does not appear to be inherited. in size. The least frequent form of CM (<1% of cases) is telan-
giectasia macularis eruptive perstans (TMEP). TMEP typically
Pathogenesis presents in adulthood and consists of tan/brown macules with
The manifestations of mastocytosis can result from mast cell telangiectasias.
mediator release (both chronic and episodic) and from excessive Lesions of UP, DCM, and cutaneous mastocytomas display
accumulation of mast cells in one or more tissues. Although the the Darier sign: localized urtication and redness of lesions fol-
molecular pathogenesis is incompletely understood, mastocytosis lowing rubbing, scratching, or stroking of skin. Mastocytomas
is frequently associated with somatic gain-of-function mutations can also precipitate severe systemic symptoms when rubbed. In
in KIT (CD117). The most common of these mutations is both CM and SM, mast cell mediator release can occur either
47
Asp816Val, or D816V. KIT-activating mutations lead to SCF- chronically or episodically, resulting in a variety of symptoms,
independent activation. Mast cells are thought to accumulate in including flushing, pruritus, shortness of breath, nausea, vomiting,
tissues because of clonal expansion and apoptotic defects of abdominal pain, diarrhea, hypotension, syncope, fatigue, and
KIT-mutated mast cells in mastocytosis. 6 headache. Anaphylaxis can be seen in both CM and SM. Interest-
ingly, urticaria and angioedema are uncommon in mastocytosis.
Clinical Features Gastrointestinal (GI) symptoms are particularly common in
The clinical signs and symptoms of mastocytosis are varied SM, with diarrhea and abdominal pain reported in up to 80%
48
and can affect many different organ systems (Fig. 23.4). Clinical of patients with SM. Neuropsychiatric symptoms, including
features can be broadly categorized as cutaneous, related to mast depression, irritability, increased somnolence, and problems with
cell mediator release, or resulting from organ infiltration by mast memory and concentration, are often observed in mastocytosis,
49
cells. although the mechanisms are not well-understood. Osteopenia
and osteoporosis are observed in a subset of patients with SM.
This may be attributed to the effects of mast cell mediators,
Mediator release syndrome Mast cell infiltration
including histamine, tryptase, heparin, IL-6, TNF-α, and trans-
General General forming growth factor-β (TGF-β), on bone turnover. 50
• Fatigue • Lymph node Mast cell mediator release can be triggered by a variety of
• Weight loss enlargement
factors, including medications (including nonsteroidal antiinflam-
Central nervous system matory drugs [NSAIDs], opiates, muscle relaxants, and contrast
• Headache Skin media), alcohol, surgical or endoscopic procedures, infections
• Altered cognitive function • Mastocytoma
• Urticaria pigmentosa (viral, bacterial, parasitical), physical factors (exercise, friction,
Skin • Diffuse cutaneous extremes of temperature), and emotional stress. Patients with
• Pruritus mastocytosis mastocytosis are particularly susceptible to anaphylaxis during
• Urtication • Telangiectasia macularis
eruptiva perstans allergic reactions to Hymenoptera stings. Severe allergic reactions
Lungs to Hymenoptera stings should prompt an evaluation for masto-
• Bronchoconstriction
Abdomen cytosis, and patients with mastocytosis should be offered venom
Cardiovascular • Hepatosplenomegaly immunotherapy. 51
• Flush • Ascites Infiltration and/or proliferation of mast cells in organs other
• Syncope • Impaired liver function
• Hypotension • Malabsorption than the skin distinguish SM from CM. The organ systems most
• Tachycardia • Diarrhea often affected in SM include bone marrow, GI tract, lymph nodes,
• Weight loss
Abdomen liver, spleen, and cortical bone. The most frequent hematological
52
• Abdominal pain abnormality in SM is anemia, and eosinophilia is also common.
• Peptic ulcer disease Bones The lymph nodes and spleen are commonly infiltrated in all
• Gastric hypersecretion • Bone marrow lesions
• Diarrhea • Hematologic disease subtypes of SM, manifesting as lymphadenopathy or splenomegaly
• Vomiting (e.g., leukemia, in many patients.
• Nausea lymphoma)
• Skeletal lesions Classification
Bones (osteoporosis,
• Bone remodeling pathologic fractures) The World Health Organization (WHO) has defined seven variants
FIG 23.4 Clinical manifestations of mastocytosis. of mastocytosis (Table 23.2). CM is limited to the skin and more

344 ParT TwO Host Defense Mechanisms and Inflammation







































A B


















C D
FIG 23.5 Cutaneous Mastocytosis and Histopathology. Urticaria pigmentosa is the most common
form of cutaneous mastocytosis. In childhood, lesions are disseminated and consist of well-
demarcated hyperpigmented macules (e.g., arrows) (A). In adults, lesions may be numerous with
less well-demarcated brownish-red macules and papules (B). Histopathology of cutaneous
mastocytosis shows many mast cells containing abundant tryptase immunoreactive cytoplasmic
granules in the papillary dermis (antitryptase, AA1 clone, Dako; original magnification ~ × 400)
(C). Bone marrow pathology of indolent mastocytosis is characterized by paratrabecular lymphoid
nodule containing small, well-differentiated lymphoid cells around substantial numbers of fusiform
cells with prominent granules in the tryptase stain (arrow) (antitryptase, AA1 clone, Dako; original
magnification ~ × 250) (D). (Courtesy of Cem Akin.)



commonly found in children. SM can include findings in a variety The term monoclonal mast cell activation syndrome (MMAS)
of organ systems, with or without skin involvement, and is defined refers to the condition in patients who have clinical symptoms
53
by a set of major and minor diagnostic criteria (Table 23.3). consistent with mast cell activation and meet one or two of the
The different categories of SM are distinguished by clinical signs minor criteria for SM but do not fully meet the diagnostic criteria.
and symptoms and/or pathological findings, including mast cell The term mast cell activation syndrome (MCAS) describes patients
52
burden and involvement of non–mast cell lineages, and have presenting with clinical symptoms of mast cell activation affecting
different prognostic implications, as described below. at least two organ systems, but not fulfilling the criteria for

CHaPTEr 23 Mast Cells, Basophils, and Mastocytosis 345



TABLE 23.2 world Health Organization Most of the diagnostic criteria for mastocytosis (see Table
(wHO) Classification of Mastocytosis 23.3) depend on bone marrow histopathological findings. Bone
marrow biopsy is therefore an essential part of the diagnostic
Cutaneous mastocytosis process. It should be considered for adult patients with typical
Indolent systemic mastocytosis skin lesions and as a confirmatory test in patients with elevated
Systemic mastocytosis with an associated clonal hematologic non–
mast cell lineage disease tryptase levels. Typical findings on bone marrow biopsy in
Aggressive systemic mastocytosis mastocytosis include multifocal dense mast cell aggregates (>15
Mast cell leukemia mast cells per cluster) and abnormal morphological features of
Mast cell sarcoma mast cells (Fig. 23.5D). Such features include spindle shapes,
Extracutaneous mastocytoma bi- or multilobed nuclei, and expression of CD2 or CD25. 52
The differential diagnosis of SM includes a variety of disorders
with similar clinical manifestations, such as MMAS and MCAS
(described above), hereditary/acquired angioedema, carcinoid
TABLE 23.3 Diagnosis of Systemic syndrome, and pheochromocytoma.
Mastocytosis Based on world Health
Organization (wHO) Criteria a Treatment
Major Criterion
Multifocal, dense infiltrates of mast cells (≥ 15 mast cells in THEraPEUTIC PrINCIPLES
aggregates) detected in sections of bone marrow and/or other Approach to Treatment of Mastocytosis
extracutaneous organ(s)
• Avoidance of symptomatic triggers
Minor Criteria • Control of symptoms
1. More than 25% of mast cells in biopsy sections of bone marrow or • Antihistamines (H 1 and H 2 receptor antagonists)
other extracutaneous organs are spindle shaped or display atypical • CysLT receptor antagonists
morphology, or, of all mast cells in bone marrow aspirate smears, > • Aspirin
25 % are immature or atypical • Disodium cromoglycate
2. Detection of an activating point mutation at codon 816 of c-KIT in • Epinephrine
blood, bone marrow, or another extracutaneous organ • Management of osteoporosis
3. Mast cells in the bone marrow, blood, or other extracutaneous • Cytoreductive therapy for aggressive disease
organs express CD2 and/or CD25 in addition to normal mast cell • Imatinib
markers • Other tyrosine kinase inhibitors
4. Serum total tryptase persistently exceeds 20 ng/mL • Management of associated hematological disorders
a One major and one minor, or three minor, criteria are needed for the diagnosis of
systemic mastocytosis. There is no cure for mastocytosis, and treatment is focused on
Data from Horny HP, Metcalfe DD, Bennett JM, et al. Mastocytosis In: Swerdlow SH, control of symptoms. All patients with mastocytosis should avoid
Campo E, Harris NL, Jaffe ES, Pileri SA, Stein H, Thiele J, Vardiman JW, editors.
WHO classification of tumors of hematopoietic and lymphoid tissues. Lyon, France: exposures to symptomatic triggers. In addition, patients should
IARC Press; 2008: p. 54–63. have epinephrine autoinjectors available at all times to treat
possible anaphylaxis. Pharmacological treatment of mastocytosis
includes drugs that fall into three main categories: (i) those that
54
anaphylaxis or for MMAS. Patients who have anaphylaxis of block or interfere with mast cell mediators, (ii) those that prevent
unknown etiology are more properly diagnosed with idiopathic mast cell activation, and (iii) those that affect mast cell survival
anaphylaxis. and proliferation.
Type 1 histamine (H 1 )-receptor antagonists are not only
Diagnosis particularly useful for the control of cutaneous symptoms,
For all patients with mastocytosis, the diagnostic process begins including pruritus and flushing, but may also aid in the manage-
with a thorough history and physical examination. The diagnosis ment of headache and neuropsychiatric symptoms (headache,
of CM is made based on the appearance of characteristic skin depression, cognitive impairment). Type 2 histamine (H 2 )-receptor
lesions and may be confirmed with skin biopsy showing mast antagonists are used in the treatment of GI symptoms, such as
cell infiltrates in the skin. In adults with cutaneous findings, abdominal pain, cramping, diarrhea, heartburn, nausea, and
systemic disease must be ruled out. vomiting. CysLT receptor antagonists are used in conjunction
The diagnostic process for patients with suspected mast cell with antihistamines as a second-line treatment for cutaneous
disease typically includes measurement of serum tryptase. If the symptoms. Aspirin may be helpful for some patients with flushing,
tryptase is found to be elevated during an anaphylactic episode particularly those with high levels of urinary prostaglandin D 2
or other symptomatic event, the measurement should be repeated or 11β-PGF 2α but must be used with caution, as NSAIDs may
at least 24 hours after symptoms have resolved. If basal tryptase trigger anaphylaxis in some patients. 52
52
is elevated (>20 ng/mL), there is a high probability of SM. Disodium cromoglycate (cromolyn) is a weak in vivo inhibitor
Another blood test that may be useful, particularly in patients of mast cell activation but is used orally to reduce GI and
52
with borderline tryptase levels or an indeterminate clinical neuropsychiatric symptoms. Certain H 1 -receptor antagonists,
presentation, is a screening test for the D816V mutation in including ketotifen and rupatadine, are reported to block mast
peripheral blood leukocytes. Both elevated tryptase and the D816V cell activation.
mutation are minor criteria for mastocytosis. Other blood tests Osteoporosis in patients with SM is typically treated in the
that should be performed to evaluate for specific organ involve- same manner as in patients in the general population. Bisphos-
ment include complete blood count and liver function tests. phonate drugs are the first line of treatment, along with

346 ParT TwO Host Defense Mechanisms and Inflammation


appropriate intake of calcium and vitamin D. Interferon-α There is a great need for better understanding of molecular
(IFN-α) has been reported to be helpful as a second-line agent mechanisms of mast cell–related human diseases for improved
in refractory disease. 52 diagnosis and treatment in the long term. For most atopic diseases
Cytoreductive therapy has a role in the treatment of aggressive and for mastocytosis, the current standard of care consists of
systemic mastocytosis (ASM) and mast cell leukemia, although avoidance of triggers and symptomatic therapies aimed at media-
59
data from large clinical trials are lacking. Imatinib, a tyrosine tor blockade. Drugs that safely and selectively suppress the
kinase inhibitor, has been effective in reducing mast cell burden pathogenic functions of mast cells could substantially improve
in only a small subset of patients who lack the D816V mutation quality of life in patients suffering from common conditions,
in KIT. Although clinical responses to other tyrosine kinase such as asthma, as well as rare ones, such as mastocytosis. At
52
inhibitors have been largely disappointing, a very recent trial the same time, the critical functions for mast cells in innate
of midostaurin, which is active on D816V, showed substantial immunity identified in mouse models require investigation in
55
promise in patients with advanced SM. More studies are needed humans. Whether mast cell effector functions can be selectively
to determine those patients most likely to benefit. exploited to enhance immune responses and vaccine efficacy is
Additional treatment of patients with SM and associated clonal an area of active investigation.
hematological non–mast cell lineage disease (AHNMD) depends
on the associated hematological disorder. The general approach Please check your eBook at https://expertconsult.inkling.com/
is to treat AHNMD as if SM were not present and to treat the for self-assessment questions. See inside cover for registration
SM as if the AHNMD were not present, while closely monitoring details.
patients for mast cell activation symptoms during therapy. 56
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CHaPTEr 23 Mast Cells, Basophils, and Mastocytosis 347.e1


MULTIPLE-CHOICE QUESTIONS

1. Which of the following is TRUE of the development of mast 3. Mast cells and basophils can be activated by which of the
cells and basophils? following mechanisms?
A. Mast cells and basophils both develop and mature fully A. immunoglobulin (Ig)E cross-linking
in bone marrow. B. Platelet-activating factor
B. Mast cells originate in bone marrow but mature in periph- C. Binding of complement components C3a or C5a
eral tissues; basophils originate and mature fully in bone D. All of the above
marrow. 4. Which of the following is TRUE of the diagnosis of potential
C. Mast cells originate and mature fully in bone marrow; mastocytosis?
basophils originate in bone marrow but mature in periph- A. A child with an isolated mastocytoma requires a bone
eral tissues. marrow biopsy to rule out systemic mastocytosis.
D. Mast cells and basophils both originate in bone marrow B. An adult with urticaria pigmentosa and an elevated serum
and mature in peripheral tissues.
tryptase level requires a bone marrow biopsy to rule out
2. Which of the following mast cell mediators is preformed and systemic mastocytosis.
stored in granules? C. Elevated serum tryptase level during an anaphylactic
A. Interleukin (IL)-6 reaction is a diagnostic criterion for systemic mastocytosis.
D. Detection of a D816V mutation in c-KIT is not useful in
B. LTC 4
C. Tryptase making a diagnosis of systemic mastocytosis.
D. IL-4

24









Eosinophils and Eosinophilia



Anna Kovalszki, Peter F. Weller








Eosinophils are terminally differentiated, bone marrow–derived governing this homing of eosinophils to mucosal tissues are
granulocytes that normally circulate in blood in low numbers and not fully known, the chemokine eotaxin-1 is involved in the
tend to localize in those tissues with mucosal epithelial surfaces. homing of eosinophils to the GI tract, but not to the respira-
2
Increased blood or tissue eosinophils occur in helminth parasite tory tract. Eosinophils live longer compared with neutrophils
infections, as well as in allergic diseases and a variety of other, and probably persist in tissues for several weeks. They are
often idiopathic, conditions. Conventionally, the major focus principally tissue-dwelling cells: as demonstrated in rodents, for
on eosinophils has been delineating the “effector” functions every eosinophil present in the circulation, there are 300–500
of these end-stage granulocytes, including what roles these in tissues.
cells play as helminthotoxic effector cells and the contribution
they make to the immunopathogenesis of allergic diseases. Eosinophil Adherence Mechanisms
More recent findings have indicated that eosinophil functions The transit of eosinophils from bone marrow, through the circula-
1-4
are considerably more extensive. Eosinophils contain stores tion, and into tissues is governed, in part, by multiple adherence
of multiple, preformed cytokines; engage in cognate cell–cell molecules expressed on eosinophils (Fig. 24.1). As for other
interactions with other cell types, including lymphocytes; and leukocytes, recruitment of eosinophils into tissue sites of inflam-
have roles in varied host immune and inflammatory responses mation utilizes combinatorial interactions involving specific
not conventionally marked by quantitatively extensive eosinophil adhesion molecules (via their expression and altered affinity
infiltration. 4 states) that mediate cellular interactions with the vascular
endothelium and actions of chemoattractant molecules. Eosino-
phils express several adhesion molecules that they broadly share
PRODUCTION AND DISTRIBUTION with other leukocytes. These adhesion molecules mediate their
OF EOSINOPHILS initial rolling and subsequent adherence to endothelial cells.
Similar to neutrophils, eosinophils can adhere via CD11/CD18
Eosinophilopoiesis heterodimeric β 2 integrins to the intercellular adhesion molecule-1
The development of eosinophils in bone marrow can be elicited and -2 (ICAM-1 and ICAM-2). Likewise, specific sialoglycopro-
by three cytokines. Granulocyte macrophage–colony-stimulating teins mediate adherence between eosinophils and endothelial
factor (GM-CSF), interleukin (IL)-3, and IL-5 all promote E- and P-selectins. Unlike neutrophils, but similar to lymphocytes,
eosinophilopoiesis. In contrast to IL-3 and GM-CSF, which also eosinophils are able to bind to vascular cell adhesion molecule-1
promote the development of other lineages, IL-5 uniquely (VCAM-1). Eosinophils express two α 4 integrins, very late activa-
promotes the development and terminal differentiation of tion antigen-4 (VLA-4, α 4 β 1 ) and α 4 β 7 , which bind to VCAM-1.
5
eosinophils. IL-5 is produced by type 2 innate lymphoid cells, Moreover, α 4 β 7 binds to the mucosal addressin cell adhesion
CD8 T cells, natural killer (NK) cells, and other leukocytes, molecule (MAdCAM). The β 2 integrin αdβ 2 , which binds ICAM-3
including eosinophils themselves. IL-5 is a defining cytokine and is expressed on other leukocytes, is an additional integrin
product of T-helper cell-2 (Th2) CD4 T cells. The production that mediates eosinophil adhesion to VCAM-1. Enhanced expres-
of IL-5 by Th2 lymphocytes accounts for the eosinophilia sion of VCAM-1 on the vascular endothelium, as elicitable by
accompanying T-helper 2 (Th2) cell–mediated immune responses IL-4 or IL-13 stimulation, may contribute to the localization of
characteristic of helminth infections and allergic diseases. eosinophils in some tissue sites of inflammation.
Eosinophilopoiesis develops over about a week. Retained In addition to mediating interactions with the endothelium,
in bone marrow is a pool of mature eosinophils. IL-5, alone eosinophil adherence molecules, by their interactions with
and in concert with the chemokine eotaxin-1 (CCL11), rapidly extracellular matrix components, modulate the activity of
releases this pool of mature eosinophils into the circulation to eosinophils that have exited the bloodstream. Eosinophil VLA-6,
acutely increase blood eosinophilia and facilitate recruitment of α 6 β 1 , binds laminin. Both α 4 β 1 and α 4 β 7 interact with specific
1
eosinophils to sites of inflammation. Blood eosinophils circu- domains of tissue fibronectin, and these interactions can enhance
late with a half-life of about 8–18 hours. Eosinophils leave the eosinophil functional responses. Eosinophils express CD44
circulation and localize in tissues, especially those with mucosal (PGP-1), which binds hyaluronic acid. Siglec-8, a sialic acid–
interfaces with the outside world, such as the gastrointestinal binding immunoglobulin-like lectin, is expressed on eosinophils
(GI) and lower genitourinary tracts. Although the mechanisms and binds sialoglycoconjugates. 6

349

350 Part two Host Defense Mechanisms and Inflammation


Extracellular Eosinophil Endothelial
matrix molecules adherence molecules adherence molecules
Integrins Integrins

β )
Laminin VLA-6 (α 6 1 CD11b/CD18 (Mac-1) ICAM-1
ICAM-1
CD11a/CD18 (LFA-1) ICAM-2
ICAM-3
β
VLA-4 (α β ) α d 2
4 1
Fibronectin VLA-4 (α β ) VCAM-1
4 1
β
4 7
α 4 7 α β MadCAM
Sialoglycoproteins Sialoglycoproteins Selectins
Sialyl-Lewis X E-selectin
Hyaluronic acid PGP-1 PSGL-1 P-selectin
Siglec-8 Selectin
L-selectin GlyCAM-1, CD34
Sialoglycoconjugates
FIG 24.1 Adherence Mechanisms Utilized by Human Eosinophils to Bind to Vascular Endothelial
Cells and the Extracellular Matrix Molecules. ICAM, intercellular adhesion molecule; VCAM,
vascular cell adhesion molecule; MAdCAM, mucosal addressin cell adhesion molecule; VLA, very
late activation antigen.


KEY CoNCEPtS
Actions of Eosinophilopoietic Cytokines
Interleukin (IL)-3, Granulocyte Macrophage–
Colony-Stimulating Factor (GM-CSF), IL-5
Promote eosinophil development and maturation in bone marrow
(IL-5).
Release a pool of mature eosinophils from bone marrow (IL-5).
Sustain the viability and antagonize apoptosis of mature eosinophils,
enhance multiple effector responses of mature eosinophils.





Eosinophil Chemoattractants
Mobilization of eosinophils into tissues is governed by receptor-
mediated chemoattractant stimuli. Chemoattractants promote
the directed migration of eosinophils and may enhance the FIG 24.2 Transmission Electron Micrograph of a Human
adhesion of eosinophils to vascular endothelium and their Eosinophil. The numerous cytoplasmic specific granules contain
subsequent migration through the endothelium. Many compounds the electron-dense crystalline cores that are unique to eosinophils.
have been identified as eosinophil chemoattractants, including In addition, lipid bodies are visible as globular, uniformly dark
humoral immune mediators, such as platelet-activating factor structures. (Original magnification × 11,180.) (Courtesy of Dr.
(PAF) and the complement anaphylatoxins C5a and C3a; certain Ann M. Dvorak, Beth Israel Deaconess Medical Center, Harvard
cytokines; and several chemokines, most notably the eotaxins. Medical School, Boston.)
None of these is specific solely for eosinophils, but eotaxin-1,
1
eotaxin-2, and eotaxin-3 exhibit the most restricted specificity.
Eotaxins signal through CCR3 chemokine receptors that are large, cytoplasmic “specific” granules that are morphologically
expressed on eosinophils as well as basophils, some Th2 cells, distinct because of their unique content of crystalloid cores.
and some mast cells. Thus recruitment of eosinophils to sites Crystalloid cores are recognizable by transmission electron
of immunological reactions is governed by their response to microscopy and usually appear electron dense (see Fig. 24.2).
chemoattractants that facilitate intravascular emigration and The cores and surrounding matrices of specific granules contain
direct migration of extravascular eosinophils, as well as by the cationic proteins that account for the tinctorial staining of granules
functional states of eosinophil adherence molecules and the with eosin. Eosinophils at sites of inflammation can exhibit
differential expression of endothelial cell adherence ligands. morphological changes in their specific granules, including
loss of either matrix or core components from within intact
STRUCTURE OF EOSINOPHILS granules, compatible with the extracellular release of granule
constituents.
Human eosinophils, unlike neutrophils, usually have a bilobed Lipid bodies, cytoplasmic structures distinct from granules
nucleus (Fig. 24.2). Defining attributes of eosinophils are their (see Fig. 24.2), are roughly globular in shape and range in size

CHaPtEr 24 Eosinophils and Eosinophilia 351


from minute to the size of specific granules. Lipid bodies are induced for expression on eosinophils in sites of inflammation.
dissolved by common alcohol-based hematological stains but In addition, eosinophils can express CD40, CD154 (CD40 ligand),
are preserved and stain darkly with osmium fixation. Lipid bodies CD153 (CD30 ligand), CD28 (B7–2), and CD86. 4
lack a delimiting membrane but contain internal membranes Eosinophils express receptors for several lipid media-
that are often obscured by overlying lipid. Lipid bodies are found tors, including PAF and leukotriene B 4 (LTB 4 ), which are
in neutrophils and other cells, especially in association with chemoattractants for eosinophils and stimulate eosinophil
inflammation; but eosinophils typically contain more lipid bodies degranulation and respiratory burst activity. Eosinophils also
compared with neutrophils. Lipid body formation in eosinophils have receptors for prostaglandins D 2 and E 2 and for cysteinyl
is rapidly inducible within minutes. In eosinophils, key enzymes leukotrienes.
involved in eicosanoid formation, including prostaglandin H
synthase, the 5- and 15-lipoxygenases, and leukotriene (LT) C 4 CONSTITUENTS OF EOSINOPHILS
synthase, localize at lipid bodies; and lipid bodies are sites of
eicosanoid synthesis. 7 Eosinophil-specific granules contain preformed proteins that
include both specific cationic proteins and stores of diverse
CELL-SURFACE RECEPTORS AND PROTEINS cytokines and chemokines.
Eosinophil receptors for immunoglobulins include those for Cationic Granule Proteins
immunoglobulin G (IgG), IgE, and IgA. The receptor for IgG Eosinophil granule cationic proteins have been extensively
on eosinophils is principally the low-affinity FcγγRII (CD32), studied because of their abundance in the granules and their
whereas neutrophils have FcγRII and FcγRIII (CD16). capacity to exert multiple effects on host cells and microbial
Eosinophil IgE receptors include the high-affinity IgE receptor targets. Major basic protein (MBP), named for its quantitative
FcεRI, typically found on basophils and mast cells, as well as predominance within the granule and its markedly cationic
FcεRII, the low-affinity IgE receptor, such as CD23, found on (basic) isoelectric point of about 11, is a 13.8- to 14-kilodalton
lymphocytes, monocytes, and antigen-presenting cells (APCs). (kDa) protein. A homolog of MBP that is somewhat smaller
Although FcεRI α-chain protein is present within eosinophils, (13.4 kDa) and less basic (pI 8.7) has been identified. MBP
its surface expression can be low or undetectable. Engagement lacks enzymatic activity and probably exerts its varied effects
of eosinophil FcεRI does not elicit exocytotic degranulation, as via its markedly cationic nature. MBP was found to be toxic to
it does on basophils and mast cells. FcεRI may participate in both airway epithelium and helminths and to have antibacterial
IgE-mediated antigen uptake by antigen-presenting eosinophils. effects. 8
Eosinophil expression of IgE receptors is notable because IgE A second granule protein is eosinophil peroxidase (EPO), an
levels and eosinophil numbers frequently increase concomitantly enzyme distinct from neutrophil myeloperoxidase. Cationic EPO
in helminth infections as well as allergic diseases. (pI 10.8) uses hydrogen peroxide and halide ions to form
Eosinophils express FcαRI (CD89), which binds secretory hypohalous acids, which are toxic for parasites, bacteria, and
IgA more potently than other forms of IgA. Engagement of FcαRI tumor and host cells. EPO utilizes bromide in preference to
triggers eosinophil release of granule proteins. With the char- chloride and is even more active with a pseudohalide, thiocyanate,
acteristic localization of eosinophils to mucosal surfaces of the to generate oxidant products, including hypobromous and
respiratory, GI, and genitourinary tracts, this IgA receptor enables hypothiocyanous acids.
eosinophils to engage secretory IgA present at these mucosal Two additional granule proteins are eosinophil cationic protein
sites. (ECP) (18 kDa, pI 10.8) and eosinophil-derived neurotoxin
Eosinophils have receptors for complement components, (EDN) (18–19 kDa, pI 8.9). EDN, never demonstrated to be
including C1q (CR1), C3b/C4b (CR1), iC3b (CR3), C3a, and neurotoxic for humans, is so named only because, after it is
C5a. Both C3a and C5a are eosinophil chemoattractants and injected intracerebrally into test rabbits, it elicits a characteristic
stimulate the production of oxygen radicals by eosinophils. neuropathological response. Both ECP and EDN are ribonucleases
Eosinophils express several receptors for chemokines. CCR1 is (RNases). EDN expresses 100 times more RNase activity compared
a receptor for MIP-1α, MCP-3, and RANTES, whereas CCR3 is with ECP, although their toxic effects on bacterial, parasitic, and
a receptor for eotaxin-1, eotaxin-2, eotaxin-3, MCP-3, and mammalian target cells are not simply a result of their RNase
RANTES. Eosinophils express CXCR4 and respond to the ligand catalytic activities.
for this receptor, stromal cell–derived factor 1α. Within the specific granule, MBP is localized to the crystalloid
Mature eosinophils, like their immature precursors, express core and to tubulovesicular structures within and arising from
8
receptors for the three cytokines, GM-CSF, IL-3, and IL-5, which the granules, whereas ECP, EDN, and EPO are localized in the
promote eosinophilopoiesis and stimulate the functioning of matrix of the granule around the core (see Fig. 24.2). MBP is
mature eosinophils. In addition, eosinophils have receptors for also found in low amounts (~3% of eosinophil levels) in basophils,
a broad range of other cytokines, including IL-1α, IL-2, IL-4, but whether this reflects endocytosis or endogenous synthesis
interferon (IFN)-α and IFN-γ, tumor necrosis factor (TNF)-α, is not known. Uptake of MBP and EPO into mast cells is known
stem cell factor (c-KIT), and IL-16 (which signals via CD4 on to occur via endocytosis. Small amounts of EDN and ECP are
eosinophils). Thus eosinophils are subject to stimulation by many present in neutrophils, and as neutrophils contain messenger
cytokines, although little is understood about how many of them RNA (mRNA) transcripts for these, EDN and ECP are likely
affect eosinophil functioning in vivo. synthesized by neutrophils. Nevertheless, eosinophils are the
Of pertinence to interactions between eosinophils and B and dominant source of these four cationic proteins. The properties
T lymphocytes, eosinophils can express several relevant plasma of these proteins and their numerous biological effects have been
9
membrane proteins. Class II major histocompatibility complex reviewed, as these proteins have major effects not only in the
(MHC) proteins, generally absent on blood eosinophils, are potential role of eosinophils in host defense against helminth

352 Part two Host Defense Mechanisms and Inflammation


parasites but also in contributing to tissue dysfunction and damage be elicited without other attributes by mediators and mechanisms
in eosinophil-related allergic and other diseases. that remain to be delineated.
Cytokines and Chemokines MECHANISMS OF EOSINOPHIL DEGRANULATION
Eosinophils are capable of elaborating at least four dozen diverse
cytokines and chemokines, and studies continue to identify more As eosinophil granules contain four major cationic proteins and
cytokines released by eosinophils. The potential activities of a multitude of preformed cytokines and chemokines, the processes
eosinophil-derived cytokines are extensive. Eosinophil-derived by which eosinophils mobilize these granule constituents for
cytokines include those with autocrine growth factor activities their extracellular release are important in understanding the
for eosinophils and those with potential roles in acute and chronic regulated functioning of eosinophils. Unlike mast cells or basophils
inflammatory responses. A notable feature of eosinophils as a that undergo acute exocytotic degranulation in response to
source of cytokines is that they contain stores of these cytokines cross-linking of their high-affinity Fcε receptors, an analogous
10
preformed within eosinophil granules and secretory vesicles. mechanism to elicit comparable exocytotic degranulation of
Thus in contrast to most lymphocytes, which must be induced fluid-phase eosinophils has not been identified. Cross-linking
to synthesize de novo cytokines destined for release, eosinophils of eosinophil IgG or IgA Fc receptors in vitro can stimulate
can immediately release preformed cytokine and chemokine release of eosinophil cationic proteins, but this rapid FcR-mediated
proteins into the surrounding milieu. The local, rapid release of acute “degranulation” process is cytolytic for eosinophils. In
eosinophil-derived cytokines in tissues to effect adjacent cells contrast, observations of eosinophils on the surfaces of large
could, and has been shown to, readily induce responses in varied nonphagocytosable multicellular helminth parasites do provide
cell types. evidence that eosinophils can degranulate by exocytosis to
Eosinophils synthesize the three growth factor cytokines wholesale release granule contents on the surfaces of target
GM-CSF, IL-3 and IL-5, which promote eosinophil survival, parasites.
antagonize eosinophil apoptosis, and enhance eosinophil effec- An alternative mechanism of mobilizing granule contents for
tor responses. Other cytokines elaborated by human eosinophils secretion that eosinophils utilize is a process of vesicular transport-
that may have activities in acute and chronic inflammatory mediated “piecemeal” degranulation. Electron microscopic
responses include IL-1α, IL-6, IL-8, IFN-γ, TNF-α, and MIP- observations of lesional eosinophils provided evidence that
1α. Human eosinophils can elaborate other various “growth” eosinophil granule contents were mobilized in vivo by selective
factors, including transforming growth factor (TGF)- α, incorporation into small vesicles that traffic to the cell surface
TGF-β 1 , vascular endothelial growth factor, platelet-derived and release these granule contents. By this process, there may
growth factor (PDGF)-β, heparin-binding epidermal growth be agonist-elicited selective secretion of certain eosinophil-derived
11
factor, and a proliferation-inducing ligand (APRIL). These cytokines. Ultrastructural studies have demonstrated that
cytokines have roles in contributing to epithelial hyperplasia secretory vesicles arise from granules and transport cytokines,
11
and fibrosis, as well as other immune homeostatic activities. such as IL-4. Insights into the selectivity and mechanisms of
In addition, eosinophils are recognized as sources of specific differential cytokine secretion have been gained by the finding,
cytokines and chemokines capable of stimulating or inhibiting at least for IL-4, that a receptor for IL-4 mediates the transport
12
lymphocyte responses, including IL-2, IL-4, IL-10, IL-12, IL-16, of IL-4 from granules and within secretory vesicles. How this
RANTES, and TGF-β 1 . Of note, eosinophil cytokines include process of vesicular transport is regulated and functions to
those associated with Th2 (IL-4, IL-5, IL-13), Th1 (IL-12, IFN- selectively mobilize specific eosinophil granule–derived cytokines
γ), and T-regulatory (IL-10, TGF-β) responses, emphasizing the or cationic proteins remains under investigation.
diverse immunoregulatory potentials for eosinophil-secreted In addition to regulated release of granule contents from
cytokines. 10 viable eosinophils, a common, but often overlooked, occurrence
is the lysis of eosinophils. Both cutaneous and pulmonary biopsy
ACTIVATED EOSINOPHILS specimens of eosinophil-associated diseases contain free, extracel-
lular, but still membrane-bound, eosinophil granules. These free
A well-recognized attribute of eosinophils is that, in conjunc- extracellular granules express cytokine, chemokine, and cysteinyl
tion with eosinophilic diseases, some blood and tissue eosino- leukotriene receptors and are secretion competent even outside of
phils may exhibit various alterations, indicating that these cells intact eosinophils. 13,14 Cytolytic stimuli elicit both the release of
have been “activated.” These changes include increased meta- nuclear DNA to form extracellular DNA “traps” and the release
bolic activity, diminished density (“hypodense”), enhanced of free secretion-competent eosinophil granules in humans. 15
LTC 4 formation, and morphological alterations, including
cytoplasmic vacuolization, alterations in granule numbers and FUNCTIONS OF EOSINOPHILS
size, and losses within specific granules of MBP-containing
cores or matrices. Activated eosinophils may exhibit enhanced Conventional considerations of the roles that eosinophils may
plasma membrane expression of some proteins, including play have been guided by quantitative considerations so that
CD69, human leukocyte antigen–D related (HLA-DR), and those diseases characteristically marked by more prominent
CD25. eosinophilia have occasioned the most interest. Thus studies
Features associated with in vivo “activated” eosinophils can have focused on the “effector” roles eosinophils play in host
be elicited, in part, by exposing eosinophils to specific stimuli, defense against helminth infections and in the immunopatho-
including GM-CSF, IL-3, and IL-5. In addition, interactions with genesis of allergic and other eosinophilic diseases. Additional
extracellular matrix components can further contribute to roles for eosinophils must be considered in immune or inflam-
eosinophil activation. Eosinophil “activation,” however, is not a matory responses not conventionally recognized to contain
singularly binary process, and some attributes of activation can abundant eosinophils. 4

CHaPtEr 24 Eosinophils and Eosinophilia 353



ROLES IN HOST DEFENSE OTHER EOSINOPHIL FUNCTIONS
Because the host response to infections with multicellular hel- Other potential functions for the eosinophil are not fully defined.
minth parasites is characteristically associated with eosinophilia, In addition to the acute release of lipid, peptide, and cytokine
it is often believed that eosinophils evolved to have a role in mediators of inflammation, eosinophils probably contribute to
killing helminths, especially during their larval stages. Indeed, chronic inflammation, including the development of fibrosis.
in vitro eosinophils can kill numerous helminths, organisms too Eosinophils can be a major source of the fibrosis-promoting
large to be phagocytosed. Eosinophils adhere to the parasite and cytokine TGF-β. Additional roles of eosinophils in modulating
deposit eosinophil granule contents onto its surface. Cell products extracellular matrix deposition and remodeling are suggested
that can contribute to parasite death include MBP, ECP, EDN, by studies of normal wound healing. During dermal wound
and EPO. healing, eosinophils infiltrate into wound sites and sequentially
16
As reviewed earlier, the helminthotoxic roles of eosinophils express TGF-α early and TGF-β 1 later during wound healing.
in vivo are less certain in humans and rodents. In eosinophil- These findings suggest that eosinophils may contribute to the
depleted mice, the intensities of primary and secondary infections more chronic subepithelial airway fibrosis characteristic of chronic
with some helminths have not been greater than in eosinophilic asthma.
mice, nor have IL-5 transgenic mice exhibited increased resistance Additional functions for eosinophils are indicated by the
to infection with some helminth species. Moreover, schistosome findings that they may be induced to express class II MHC proteins
20
infections in two lines of eosinophil-ablated mice have shown and can function as APCs. Blood eosinophils lack HLA-DR
no differences in measures of infection compared with normal expression, but eosinophils recovered from the airways 48 hours
17
mice. Nevertheless, murine studies need to be interpreted with after segmental antigen challenge have been shown to express
caution. Many experimental infections involve introducing HLA-DR. Cytokines, including GM-CSF, IL-3, IL-4, and IFN-γ,
helminth infections that are often host species–restricted into induce eosinophil HLA-DR expression. Both murine and human
unnatural host mice, in which innate immune responses may eosinophils can function as HLA-DR–dependent MHC-restricted
be prominent. Natural human infections are usually a consequence APCs in stimulating the proliferation of T cells. In vivo, murine
of repeated exposures, during which acquired, rather than innate, eosinophils can process exogenous antigens in the airways, traffic
immunity becomes prominent. Thus eosinophil functions as to regional lymph nodes, and function as antigen-specific APCs
helminthotoxic cells in vivo remain unclear. Eosinophils might to stimulate responses of CD4 T cells. 21
have alternative functions in host responses to helminths, includ- Eosinophils, that normally become resident in submucosal
ing functioning as APCs and even favoring the survival of and less prominently in other tissues, undoubtedly participate
Trichinella larvae in muscles. 18,19 in ongoing homeostatic immune responses at these sites. Some
of these responses are mediated by cytokines secreted by eosino-
ROLES IN DISEASE PATHOGENESIS phils, including IL-6 and APRIL to stimulate plasma cells
22
development and IL-4 to activate macrophages in fat tissue
23
The abilities of eosinophils to release biologically active lipids and effect glucose metabolism. Further investigations will help
as paracrine mediators of inflammation and to release preformed delineate eosinophil’s functional roles and interactions with other
cationic and cytokine granule constituents enable eosinophils cells, so that the scope of eosinophil functions will probably
to contribute to the immunopathogenesis of various diseases, extend beyond its currently more defined role as an effector cell
1
including asthma. Eosinophils form several classes of biologically contributing to allergic inflammation.
active lipids. Eosinophils may liberate PAF, whose diverse activities
can be mediated either directly or by stimulating other cells to EOSINOPHILIA AND EOSINOPHILIC DISORDERS
release leukotrienes, prostaglandins, and complement peptides.
Stimulated eosinophils release LTC 4 . LTD 4 and LTE 4 are formed Diverse infectious, allergic, neoplastic and idiopathic disease
from LTC 4 by the sequential enzymatic removal of glutamic acid processes can be associated with increased blood and/or tissue
and glycine from its tripeptide glutathione side chain. LTC 4 and eosinophil numbers. Blood eosinophilia, present when eosino-
especially LTD 4 have bronchoconstrictor activities, constrict phil numbers are in excess of their usual level of <450/µL of
terminal arterioles, dilate venules, and stimulate airway mucus blood, may be intermittently, modestly, or (less frequently)
secretion. Thus eosinophils are a potential source of two major markedly increased. Blood eosinophil numbers are not neces-
types of mediator lipids, the sulfidopeptide leukotrienes and sarily indicative of the extent of eosinophil involvement in
PAF. affected tissues.
Oxidants released by eosinophils, including superoxide anion, Some patients with sustained blood eosinophilia can develop
hydroxyl radical, and singlet oxygen, as well as EPO-catalyzed organ damage, especially cardiac damage. This cardiac involvement
hypothiocyanous acid and other hypohalous acids, have the can include the formation of intraventricular thrombi and
potential to damage host tissues. endomyocardial fibrosis with secondary mitral or tricuspid
Released eosinophil granule proteins are immunologically regurgitation (Fig. 24.3). Such damage can complicate the
detectable in fluids, including blood, sputum, and synovial fluids, sustained eosinophilia of hypereosinophilic syndromes and has
and in tissues, including the respiratory and GI tracts, skin, and been noted with eosinophilias accompanying other diseases,
heart, in association with various eosinophil-related diseases. including eosinophilia with carcinomas, lymphomas, GM-CSF,
The eosinophil cationic proteins, including MBP, ECP, and EPO, or IL-2 administration, drug reactions, and parasitic infections.
can damage various cell types. Thus extracellular release of Most patients with eosinophilia, however, develop no evidence
eosinophil granule proteins, by degranulation or cytolysis of of endomyocardial damage. Conversely, cardiac disease can rarely
eosinophils, could contribute to local tissue damage by causing present in patients without known eosinophilia. The pathogenesis
dysfunction and damage to adjacent cells. of eosinophil-mediated cardiac damage involves both usually

354 Part two Host Defense Mechanisms and Inflammation


INFECTIOUS DISEASES ASSOCIATED
WITH EOSINOPHILIA

Eosinophilia is encountered only with specific infectious diseases.
With active bacterial or viral infections, eosinopenia is charac-
teristic. This suppression of blood eosinophils is, in part, caused
by heightened endogenous corticosteroid production as well as
by inflammatory mediators released during these infections. This
suppression of eosinophilia, with either serious bacterial infections
or marked inflammation, accounts for the absence of otherwise
expected eosinophilia in some patients with helminth infections,
24
including those with hyperinfection strongyloidiasis. As a general
clinical guideline, patients with a febrile illness and an increased
or even normal blood eosinophilia are not likely to have common
bacterial or viral infections, unless they have adrenal insufficiency
FIG 24.3 Eosinophil Endomyocardial Disease. A large thrombus or a confounding medication-elicited eosinophilia.
is present in the apex of the left ventricle and the chordae Helminth Parasites
tendineae are entrapped, leading to severe mitral valve
regurgitation. Helminth parasites are multicellular metazoan organisms—the
“worm” parasites. Infections with diverse helminths elicit
24
eosinophilia (Chapter 31). Although eosinophilia may provide
a hematological clue to the presence of helminth infection, the
heightened numbers of eosinophils and some activating events, absence of blood eosinophilia does not exclude such infections.
as yet ill-defined, that promote eosinophil-mediated tissue damage. The eosinophilic response to helminths is determined both by
Cardiac damage progresses through three stages. In the first the host’s immune response and by the parasite, including its
stage, there is damage to the endocardium and infiltration of distribution, migration, and development within the infected
the myocardium with eosinophils and lymphocytes, with host. The level of eosinophilia tends to parallel the magnitude
eosinophil degranulation and myocardial necrosis. Elevated and extent of tissue invasion by helminth larvae or adults. In
plasma levels of troponin can be a sensitive assay of early several helminth infections, the migration of infecting larvae or
eosinophil-mediated cardiac damage. A similar acute eosinophilic subsequent developmental stages through tissues is greatest early
myocarditis can develop with drug hypersensitivity reactions in infection, and hence the magnitude of the elicited eosinophilia
and may be more fulminant. The first stage is frequently clinically will be the greatest in these early phases. In established infections,
occult, although subungual splinter hemorrhages may be local eosinophil infiltration will often be present around helminths
prominent. Elevations of serum troponins as a measure of within tissues, without significant blood eosinophilia. Eosinophilia
myocardial injury should be evaluated. Echocardiography usually may be absent in those helminth infections that are well contained
detects no abnormalities at this stage, although cardiac magnetic within tissues (e.g., intact echinococcal cysts) or are solely
resonance imaging (MRI) is evolving as a technique to potentially intraluminal within the intestinal tract (e.g., Ascaris, tapeworms).
detect cardiac involvement at an earlier stage. Uncommonly, In some established infections, increases in blood eosinophilia
death due to acute progressive cardiac disease can occur. Cor- may be episodic. Intermittent leakage of cyst fluids from echi-
ticosteroid therapy during the acute stage may help control and nococcal cysts can transiently stimulate increases in blood
prevent the evolution of myocardial fibrosis. eosinophilia and also cause symptoms attributable to allergic or
The second stage of heart disease, the formation of thrombi anaphylactic reactions (urticaria, bronchospasm). For tissue-
along the damaged endocardium, affects either or both ventricles dwelling helminths, increases in eosinophilia may occur principally
and occasionally the atrium. Outflow tracts near the aortic and in association with migration of adult parasites, as in loiasis and
pulmonic valves are usually spared. These thrombi can embolize gnathostomiasis.
to the brain and elsewhere. Finally, in the fibrotic stage, progressive Helminth infections more likely to elicit prolonged hypereo-
scarring leads to entrapment of chordae tendineae with the sinophilia in adults include filarial and hookworm infections
24
development of mitral and/or tricuspid valve regurgitation and and strongyloidiasis (Table 24.1). Trichinellosis can elicit an
to endomyocardial fibrosis, producing a restrictive cardiomy- acute hypereosinophilia. Strongyloides stercoralis infection, difficult
opathy. Echocardiography and MRI are valuable in detecting to diagnose solely by stool examinations, is especially important
intracardiac thrombi and the manifestations of fibrosis. Patients to exclude, not only because it elicits modest to even marked
with sustained eosinophilia should be monitored by using eosinophilia but also because, unlike other helminths, it can
echocardiography and serum troponin assays for evidence of develop into a disseminated, often fatal, disease (hyperinfection
24
cardiac disease. In an older series of patients referred to the syndrome) in patients given immunosuppressive corticosteroids.
National Institutes of Health (NIH), much of the mortality among Enzyme-linked immunosorbent assay (ELISA) serology has proved
these patients with hypereosinophilia was attributable to end-stage valuable in detecting strongyloidiasis and should be obtained
congestive heart failure. In contemporary times, earlier recognition for patients with eosinophilia likely to receive corticosteroids.
of cardiac involvement, mitral valve replacement with biopros- Some tissue- or blood-dwelling helminths that are not diagnosable
theses and additional therapeutic options for hypereosinophilic by stool examinations but that can cause marked eosinophilia
syndromes (see below) have largely minimized the morbidity require diagnostic examination of blood or biopsied tissues or
24
and mortality attributable to end-stage eosinophilic endomyo- specific serological tests. Infections with these organisms include
cardial disease. filarial infections, trichinellosis, and visceral larva migrans. In

CHaPtEr 24 Eosinophils and Eosinophilia 355



TABLE 24.1 Parasitical Diseases Capable invasive pathogen. (ii) Coccidioidomycosis, following primary
of Causing Marked (>3000/µL) or Long- infection, especially in conjunction with erythema nodosum,
Standing Eosinophilia and at times with progressive disseminated disease, can elicit
blood eosinophilia and may cause an eosinophilic meningitis.
Helminth Chronic (iii) Basidiobolomycosis infection can also be associated with
Infection Hypereosinophilia Eosinophilia eosinophilia. 24
Angiostrongyliasis costaricensis
Ascariasis + During early lung Human Immunodeficiency Virus and
migration Retroviral Infections
Hookworm infection + During early lung + Common cause of Eosinophilia can uncommonly accompany human immunode-
migration low-grade ficiency virus (HIV) infections for several reasons. First, leukopenia
eosinophilia may increase eosinophil percentages without reflecting true
Strongyloidiasis Uncommonly + Self-perpetuating,
may last > 50 eosinophilia. Second, adverse reactions to medications may elicit
years eosinophilia. Third, patients with acquired immunodeficiency
Trichinellosis + With heavy infections syndrome (AIDS) who develop adrenal insufficiency as a result
Visceral larva migrans + Principally in children of cytomegalovirus and other infections may exhibit eosinophilia
Gnathostomiasis as a consequence. In addition, modest, and uncommonly marked,
Cysticercosis eosinophilia and eosinophilic pustular folliculitis can be observed
Echinococcosis + May be episodic in some patients with HIV infection. Eosinophilia more com-
24
with cyst fluid 24
leakage monly develops with HTLV-1 infections.
Filariases:
Tropical pulmonary + + ALLERGIC DISEASES ASSOCIATED
eosinophilia WITH EOSINOPHILIA
Loiasis + Especially in +
expatriates Among the noninfectious diseases associated with eosinophilia
Onchocerciasis + +
Flukes: (Table 24.2) are allergic diseases, notably those mediated by
Schistosomiasis + During early infection + IgE-dependent mechanisms. In these diseases, including allergic
in nonimmune rhinitis, conjunctivitis, and asthma, eosinophils are present in
patients involved tissues as well as often being increased in blood.
Fascioliasis + During early infection +
Clonorchiasis + During early infection +
Paragonimiasis + During early infection + MYELOPROLIFERATIVE AND NEOPLASTIC DISEASE
Fasciolopsiasis + During early infection +
Eosinophilia can occur with specific neoplastic diseases, as well
Adapted from Wilson ME, Weller PF. Eosinophilia. In: Guerrant RL, Walker DH, as in some disorders of uncertain etiology, including some
Weller PF, eds. Tropical Infectious Diseases: Principles, Pathogens and Practice, 3rd hypereosinophilic syndromes.
ed. Philadelphia, Penn.: Churchill Livingstone; 2011:943.
Hypereosinophilic Syndromes
A syndrome previously termed idiopathic hypereosinophilic
children, owing to their propensity for geophagous pica and syndrome is not a single entity but rather a constellation of
ingestion of dirt contaminated by dog ascarid eggs, visceral larva leukoproliferative disorders characterized by sustained overpro-
migrans caused by Toxocara canis is a potential etiology for duction of eosinophils. The three original diagnostic criteria for
sustained eosinophilia. ELISA serological testing can evaluate this syndrome were eosinophilia in excess of 1500/µL of blood
this possibility. persisting for longer than 6 months; lack of an identifiable
parasitic, allergic, or other etiology for eosinophilia; and signs
Other Infections: Protozoa and Fungi and symptoms of organ involvement. In contemporary practice,
Infections with single-celled protozoan parasites do not char- if eosinophilia is sustained over a month and the other criteria
acteristically elicit eosinophilia. This is true of all intestinal, are reliably met before the 6-month time frame, a diagnosis can
blood-, and tissue-infecting protozoa, with three exceptions. Two be made and treatment promptly initiated. Moreover, in recent
intestinal protozoans, Dientamoeba fragilis and Isospora belli, years there has been increasing recognition that hypereosinophilic
can at times be associated with low-grade eosinophilia. Hence, syndromes (HES), even at times without evident organ damage,
in patients with symptoms of enteric infection and eosinophilia, encompass a spectrum of disorders, and progress has been made
diagnostic trophozoites of D. fragilis or oocysts of I. belli should in identifying the underlying defects in some of these (Fig. 24.4). 25
be sought in stool examinations. Fecal examinations for I. belli
oocysts must be specifically requested, as they are not usually
detected in routine stool ova and parasite examinations. Other
enteric protozoa do not elicit eosinophilia and, if detected in
stool examinations, should not be accepted as causes of eosino- KEY CoNCEPtS
philia. Sarcocystis, a myositis producing protozoan, can elicit Hypereosinophilic Syndromes
modest eosinophilia.
Three fungal diseases can be associated with eosinophilia. (i) Eosinophilia sustained in excess of 1500/µL.
Aspergillosis is accompanied by eosinophilia only in the form Absence of allergic, parasitic, or other etiologies for eosinophilia.
Usually evidence of organ involvement.
of allergic bronchopulmonary aspergillosis, not when it is an

356 Part two Host Defense Mechanisms and Inflammation



TABLE 24.2 Eosinophil-associated Diseases and Disorders
allergic or atopic Diseases
Asthma
Allergic rhinitis
Eosinophilic esophagitis
Atopic dermatitis
Allergic urticaria
Nasal polyps
Myeloproliferative and Neoplastic Disorders
Hypereosinophilic syndromes: myeloproliferative, lymphoproliferative, and others
Leukemia
Lymphoma- and tumor-associated
Mastocytosis

Pulmonary Syndromes
Parasite-induced eosinophilic lung diseases:
Loeffler syndrome: patchy migratory infiltrates, resolving over weeks, seen with transpulmonary migration of helminth parasites, especially Ascaris
Tropical pulmonary eosinophilia: miliary lesions and fibrosis; heightened immune response causing one form of lymphatic filariasis; increased
immunoglobulin E (IgE) and antifilarial antibodies
Pulmonary parenchymal invasion: paragonimiasis
Heavy hematogenous seeding with helminths: trichinellosis, schistosomiasis, larva migrans
Allergic bronchopulmonary aspergillosis
Chronic eosinophilic pneumonia: dense peripheral infiltrates, fever; progressive, blood eosinophilia may be absent; steroid responsive
Acute eosinophilic pneumonia: acute presentation diagnosed by bronchoalveolar lavage or biopsy
Eosinophilic granulomatosis with polyangiitis (Churg–Strauss syndrome) vasculitis: small- and medium-sized arteries; granulomas, necrosis; asthma often
antecedent; extrapulmonary (e.g., neurological, cutaneous, cardiac, or gastrointestinal) involvement likely
Drug- and toxin-induced eosinophilic lung diseases
Other: hypereosinophilic syndromes, neoplasia, bronchocentric granulomatosis

Skin and Subcutaneous Diseases
Skin diseases: atopic dermatitis, blistering diseases, including bullous pemphigoid, urticarias, drug reactions
Diseases of pregnancy: pruritic urticarial papules and plaques syndrome, herpes gestationis
Eosinophilic pustular folliculitis
Eosinophilic cellulitis (Wells syndrome)
Kimura disease and angiolymphoid hyperplasia with eosinophilia
Shulman syndrome (eosinophilic fasciitis)
Episodic angioedema with eosinophilia: recurrent periodic episodes with fever, angioedema, and secondary weight gain; may be long-standing without
untoward cardiac dysfunction
Gastrointestinal Disorders
Eosinophilic gastroenteritides
Inflammatory bowel disease: eosinophils in lesions; occasionally blood eosinophilia with ulcerative colitis
rheumatologic Diseases
Vasculitis: Eosinophilic granulomatosis with polyangiitis (Churg–Strauss) and cutaneous necrotizing eosinophilic vasculitis
Immunological reactions
Medication-related eosinophilias
Immunodeficiency diseases: Job’s syndrome and Omenn syndrome
Transplant rejections
Endocrine
Hypoadrenalism: Addison disease, adrenal hemorrhage, hypopituitarism
other Causes of Eosinophilia
Atheromatous cholesterol embolization
Hereditary
Serosal surface irritation, including peritoneal dialysis and pleural eosinophilia
Adapted from Weller PF. Eosinophilia and eosinophil-related disorders. In: Adkinson NF, Jr., Yunginger JW, Busse WW, et al., eds. Allergy: Principles and Practice, 6th ed.
Philadelphia, Penn.: Mosby; 2003:1105.

CHaPtEr 24 Eosinophils and Eosinophilia 357



Hypereosinophilic Syndromes (HESs)


Eosinophils > 1,500/mm 3
Persistent eosinophilia and/or end-organ damage/dysfunction
Exclusion of secondary causes of eosinophilia




Myeloproliferative variant Lymphocytic
variant Familial Undefined Overlap* Associated**



Myeloproliferative PDGFRA Chronic Clonal lymphocyte Family history of EGID eosinophilic CSS Systemic
- etiology unknown -associated HES eosinophilic population by documented pneumonia, mastocytosis,
leukemia flow cytometry persistent eosinophilia inflammatory
or eosinophilia of myalgia syndrome, bowel disease,
PCR analysis of unknown cause and other organ- sarcoidosis, HIV,
T-cell receptor usage restricted and other disorders
eosinophilic disorders
F/P negative and F/P positive by Demonstrable
clonal eosinophilia by RT - PCR cytogenetic
HUMARA*** or FISH abnormalities
or and/or blasts
≥4 of the following: on peripheral Benign Complex Episodic
smear
Dysplastic eosinophils on
peripheral smear Asymptomatic Symptomatic with Cyclical angioedema
Serum B12>1000 pg/ml with no evidence organ dysfunction and eosinophilia
Anemia and/or of organ involvement but does not meet
thrombocytopenia criteria for
Hepatosplenomegaly myeloproliferative or
Bone marrow cellularity >80% lymphocytic variant
Spindle-shaped mast cells
Myelofibrosis
FIG 24.4 Classification of Hypereosinophilic Syndromes Based on a Workshop Summary
Report. Specific syndromes discussed at the workshop are indicated in bold. *Incomplete criteria,
apparent restriction to specific tissues/organs. †Peripheral eosinophilia, >1500/mm in association
3
with a defined diagnosis. ‡ Presence of the FLPL1/PDGFRA (F/P) mutation. § Clonality analysis
based on the digestion of genomic DNA with methylation-sensitive restriction enzymes followed
by polymerase chain reaction (PCR) amplification of the CAG repeat at the human androgen
receptor gene (HUMARA) locus at the X chromosome. CSS, Churg-Strauss syndrome (now called
eosinophilic granulomatosis with polyangiitis); EGID, eosinophil gastrointestinal diseases; FISH,
fluorescence in situ hybridization. (From Klion AD, Bochner BS, Gleich GJ, et al. Approaches
to the treatment of hypereosinophilic syndromes: a workshop summary report. J Allergy Clin
Immunol 2006; 117:1294, with permission from the American Academy of Allergy, Asthma and
Immunology.)



26
Some patients with HES, termed myeloproliferative variants the first line of therapy for FIP1LI/PDGFRA-positive HES. For
of HES, exhibit features common to myeloproliferative disorders, patients with any evidence of cardiac involvement, including
including elevated vitamin B 12 and lactate dehydrogenase (LDH) elevated troponin levels, it is recommended that glucocorticoids
levels, splenomegaly, cytogenetic abnormalities, myelofibrosis, be administered along with initiation of imatinib therapy. Other
anemia, myeloid dysplasia, and often elevated serum level of patients with eosinophilia without F/P mutations have also
mast cell tryptase. In many patients with myeloproliferative HES, responded to imatinib, indicating that other receptor tyrosine
the molecular defect has been identified as a chromosome 4 kinase mutations can underlie some of these CEL/myeloproliferative
27
deletion that yields a fusion gene encoding a FIP1LI/PDGFRA forms of HES. The presence of more than four myeloproliferative
(PDGF-α receptor) (F/P) protein that constitutively expresses features commonly seen in mutation-positive disease, predicted
receptor kinase activity. This fusion gene can be diagnostically response in those without known mutations. Some of these
evaluated by reverse transcription–polymerase chain reaction features included dysplastic eosinophils, vitamin B 12 level >1000
(RT-PCR) or fluorescence in situ hybridization (FISH) (Chapter picograms per milliliter (pg/mL), tryptase level ≥12 nanograms
96). Importantly, the majority of patients with this fusion muta- per milliliter (ng/mL), anemia/thrombocytopenia, hypercellular
tion, which constitutes a form of chronic eosinophilic leukemia marrow, and spindled mast cells, reticulin fibrosis, and dysplastic
28
(CEL), respond to therapy with imatinib, which is considered megakaryocytes on bone marrow biopsy. In addition, clonal

358 Part two Host Defense Mechanisms and Inflammation


abnormalities in the eosinophil lineage have been reported in a secondary eosinophil-mediated cardiac damage, for reasons that
few patients (see Fig. 24.4). are not known.
Another variant form of HES is a lymphoproliferative form
−
+
resulting from clonal expansions of lymphocytes, often CD4 CD3 Pulmonary Eosinophilias
29
Th2-like lymphocytes, which elaborate IL-5. These aberrant T Blood eosinophilia can infrequently accompany pleural fluid
cells can be sought by flow cytometry or T-cell receptor (TCR) eosinophilia, a nonspecific response seen with various disorders,
analysis. These patients, who may have elevated IgE levels, usually including trauma and repeated thoracenteses. In addition, several
do not develop eosinophilic endomyocardial disease but are at pulmonary parenchymal disorders can be associated with
risk for developing T-cell lymphomas. 29 eosinophilia (see Table 24.2). 32
In addition to these recognized variants, there are a substantial Helminth parasites are responsible for four forms of eosino-
number of patients with HES for whom the etiologies of the philic lung disease. 24,32 The first form, Loeffler syndrome, is marked
26
eosinophilia remain unknown. Some such patients develop no by blood eosinophilia, eosinophilic patchy pulmonary infiltrates
signs or symptoms of disease and can be monitored without that appear and resolve over weeks, and, at times, bronchospasm,
30
therapy. For those who require therapy, including those with and is typically caused by those helminth parasites (Ascaris
29
lymphoproliferative variants, glucocorticosteroids are the lumbricoides/Ascaris suum), and less commonly hookworm and
26
mainstay of treatment. With glucocorticoid therapy, partial or Strongyloides that migrate through the lungs early in their
24
complete remission of eosinophilia within 1 month has been developmental lifecycle. Stool examinations are not helpful, as
26
reported to occur in 85% of patients. Second-line agents include the pulmonary response is elicited by infecting larval forms
26
hydroxyurea and IFN-α. The neutralizing anti-IL-5 monoclonal months before productive egg-laying from later adult stages
antibody (mAb), mepolizumab, has beneficial and steroid-sparing begins in the intestines. Diagnosis is made on epidemiological
effects in those with FIP1L1-PDGFRA negative hypereosinophilic grounds. 24
31
syndromes, but it is approved by the US Food and Drug The second form of helminth-induced lung disease is the
Administration (FDA) for treating severe eosinophilic phenotype syndrome of tropical pulmonary eosinophilia, which develops
asthma but not yet approved for hypereosinophilic syndromes. in a minority of patients infected with lymphatic-dwelling filarial
23
Anti-CD52 mAb (alemtuzumab) and allogeneic hematopoietic species. This syndrome is characterized by marked blood
cell transplantation have been used for particularly severe and eosinophilia, a paroxysmal nonproductive cough, wheezing,
refractory HES. occasional weight loss, lymphadenopathy, and low-grade fevers.
In contrast to older reports, with earlier diagnosis and On chest X-rays, increased bronchovesicular markings, diffuse
therapy and with more varied and targeted therapeutic options, interstitial lesions 1–3 mm in diameter or mottled opacities,
morbidity and, particularly, mortality in HES syndromes have usually more prominent in lower lung fields, are common. Patients
been reduced. have markedly increased numbers of blood and alveolar eosino-
phils, and elevations in both total serum IgE and antifilarial
Eosinophilia With Tumors or Leukemias antibodies.
The F/P-positive and related chromosomal fusion gene mediated A third form of helminth-induced lung disease is caused by
27
myeloproliferative variants of HES are forms of CEL. Eosino- helminths that invade the pulmonary parenchyma, notably lung
philia is a characteristic of the M4Eo subtype of acute myeloid flukes that cause paragonimiasis.
leukemia, having the common M4 characteristic of chromosomal The fourth form of lung disease is caused by larger than usual
16 abnormalities. Other forms of eosinophilic leukemia, often numbers of helminth organisms that are carried hematogenously
with specific cytogenetic and molecular genetic abnormalities, into the lungs. Examples include schistosomiasis, trichinellosis,
27
have been recognized. Eosinophilia may accompany chronic and larva migrans.
myelogenous leukemia (often with basophilia) but is uncommon Bronchopulmonary aspergillosis constitutes another type of
with acute lymphoblastic leukemia. Eosinophilia may be observed eosinophil-associated pulmonary disease. Two forms of idiopathic
32
in some patients with lymphoma, including Hodgkin disease, eosinophilic pneumonia are recognized. In chronic eosinophilic
especially the nodular sclerosing form, T-cell lymphoblastic pneumonia, patients may exhibit peripheral pulmonary infiltrates
lymphoma, and adult T-cell leukemia/lymphoma. A small propor- that can extend across lobar fissures. Chronic eosinophilic
tion of patients with carcinomas, especially of mucin-producing pneumonia is of unknown etiology and is responsive to
epithelial cell origin, have associated blood and tissue eosinophilia. corticosteroids but is prone to relapse. An acute form of eosinophilic
Eosinophilia may accompany angioimmunoblastic lymphade- pneumonia, which manifests as fever, pulmonary infiltrates, and
nopathy, mycosis fungoides, Sézary syndrome, and lymphomatoid respiratory insufficiency, is diagnosable by finding eosinophils
papulosis. Eosinophilia occurs in about 20% of patients with in bronchoalveolar lavage (BAL) fluids or on lung biopsy. Acute
systemic mastocytosis and may be the presenting finding in the eosinophilic pneumonia, which often follows new exposures to
absence of cutaneous manifestations. cigarette or other smoke or dusts, responds to corticosteroid
treatment and does not relapse.
ORGAN SYSTEM INVOLVEMENT The major vasculitis associated with eosinophilia is eosinophilic
AND EOSINOPHILIA granulomatosis with polyangiitis (EGPA, formerly called Churg-
33
Strauss syndrome) (Chapter 58). Late-onset asthma, eosinophilia,
Eosinophilic syndromes limited to specific organs, such as and at times transient pulmonary infiltrates antedate the develop-
eosinophilic pneumonias or eosinophilic GI disorders (EGIDs; ment of systemic vasculitis in about half the cases. Pulmonary
Chapter 46), characteristically do not extend beyond their own involvement is seen in almost all patients, and pulmonary
target organ, and hence lack the multiplicity of organ involvement infiltrates occur in three-quarters of them. Nasal and sinus
often found with non–organ-specific hypereosinophilic syn- involvement is common. Corticosteroids, anti-IgE mAb, or
dromes. They also do not have the predilection to develop anticysteinyl leukotriene agent therapies for asthma may mask

CHaPtEr 24 Eosinophils and Eosinophilia 359


the evolution of EGPA. Neurological, cutaneous, cardiac, and Immunological Disorders
33
GI organ involvement is common. Cardiac involvement includes
pericarditis and small vessel cardiac vasculitis, and, much less CLINICaL PEarLS
commonly, endomyocardial thrombosis and fibrosis. Eosinophilia and Drug Reactions
Diverse drugs are capable of eliciting pulmonary eosinophilia.
More commonly implicated medications include nonsteroidal Drug reactions Examples
antiinflammatory drugs (NSAIDs) and antimicrobial medica- Interstitial nephritis Semisynthetic penicillins,
tions. Likewise, toxic agents, including those from occupational cephalosporins
exposure, can be responsible for pulmonary eosinophilia. Each of Pulmonary infiltrates Nitrofurantoin, sulfas, nonsteroidal
antiinflammatory drugs (NSAIDs)
these reactions has a defined etiological stimulus and hence differs Pleuropulmonary Dantrolene
from idiopathic and other eosinophilic diseases, but the clinical Hepatitis Semisynthetic penicillins, tetracyclines
presentation of drug- and toxin-elicited pulmonary eosinophilias Hypersensitivity vasculitis Allopurinol, phenytoin
can resemble other forms of pulmonary eosinophilia, including Asthma, nasal polyps Aspirin
acute or chronic eosinophilic pneumonia. Eosinophilia–myalgia L-Tryptophan contaminant syndrome
Asymptomatic Ampicillin, penicillins, cephalosporins
Skin and Subcutaneous Diseases Cytokine-mediated Granulocyte macrophage–colony-
stimulating factor (GM-CSF),
A number of cutaneous diseases can be associated with heightened interleukin (IL)-2
34
blood eosinophils, including atopic dermatitis, blistering dis- DRESS (drug reaction with Minocycline, allopurinol, anticonvulsants
orders including bullous pemphigoid, drug reactions, and two eosinophilia and systemic
diseases associated with pregnancy: (i) herpes gestationis and symptoms)
(ii) the syndrome of pruritic urticarial papules and plaques of Adapted from Weller PF. Eosinophilia and eosinophil-related
pregnancy. Eosinophilic pustular folliculitis is seen mostly in disorders. In: Adkinson NF, Jr., Yunginger JW, Busse WW, et al.,
patients with HIV infections and in those treated for hematological eds. Allergy: Principles and Practice, 6th ed. Philadelphia, Penn.:
malignancies or after bone marrow transplantation. In patients Mosby; 2003:1105.
with cutaneous involvement and eosinophilia, angiolymphoid
hyperplasia with eosinophilia and Kimura disease, eosinophilic
cellulitis (Wells syndrome), eosinophilic fasciitis, and eosinophilic Adverse reactions to medications are a common cause of eosino-
pustular folliculitis can be differentiated on the basis of histo- philia. Although often considered as hypersensitivity reactions,
pathology of biopsied lesions. Another syndrome, episodic in most instances of drug-associated eosinophilia, the mecha-
angioedema with eosinophilia, is characterized by recurring nism leading to eosinophilia is not understood. Eosinophilia
episodes of angioedema, urticaria, fever, and marked blood may develop without other manifestations of adverse drug reac-
eosinophilia. This syndrome responds to glucocorticosteroid tions, such as rashes or drug fevers. In addition, drug-induced
therapy. eosinophilia may be associated with distinct clinicopathological
patterns in which eosinophilia accompanies drug-induced
Gastrointestinal Diseases diseases that are characteristically limited to specific organs
EGIDs (Chapter 46), including eosinophilic esophagitis, eosino- with or without associated blood eosinophilia. When organ
philic gastroenteritis, and eosinophilic colitis, represent a het- dysfunction develops, cessation of drug administration is neces-
erogeneous collection of disorders in which there may be sary. Drug-induced interstitial nephritis may be accompanied by
eosinophilic infiltration of the mucosa, the muscle layer or the blood eosinophilia, and eosinophils may be detectable in urine.
serosa, the last of which can lead to eosinophilic ascites. Peripheral Unlike granulocyte–colony-stimulating factor (G-CSF) therapy,
blood eosinophilia may occur in EGIDs, although with eosino- therapy with GM-CSF can lead to prominent blood and tissue
philic esophagitis, peripheral blood eosinophil counts are often eosinophilia. Administration of IL-2 or of IL-2-stimulated lym-
normal. Eosinophils are present in the lesions of collagenous phocytes can be followed by the development of eosinophilia,
colitis and ulcerative colitis, but blood eosinophilia is usually most likely as a result of stimulated production of IL-5. Reac-
absent. GI eosinophilia elicited by intestinal helminths and tions to medications, often anticonvulsants, minocycline, and
eosinophilic enterocolitis as a result of hypersensitivity reactions allopurinol, can elicit DRESS (drug reaction with eosinophilia
34
to medications must be excluded in patients with these diseases and systemic symptoms). In addition to cutaneous eruptions,
who have tissue eosinophilia. fever, lymphadenopathy, hepatitis, nephritis, atypical lympho-
cytosis, GI tract involvement, and eosinophilia are common
Rheumatological Disorders but variable elements of this drug-induced syndrome, which
Of the various forms of vasculitis, only two are commonly can be fatal. The triggering medication must be stopped, and
associated with eosinophilia. The principal eosinophil-related corticosteroids are often administered.
vasculitis is EGPA, formerly called Churg-Strauss syndrome (as Some primary immunodeficiency syndromes are associated
35
discussed above; and in Chapter 58). Cutaneous necrotizing with eosinophilia. Hyper-IgE syndrome is characterized by
eosinophilic vasculitis with hypocomplementemia and eosino- recurrent staphylococcal abscesses of the skin, lungs, and other
philia is a distinct vasculitis of small dermal vessels that are sites; pruritic dermatitis; hyperimmunoglobulinemia E; and
extensively infiltrated with eosinophils. This form of vasculitis eosinophilia of blood, sputum, and tissues. Eosinophilia is
may occur in patients with connective tissue diseases. In addition, characteristic of Omenn syndrome, combined immunodeficiency
eosinophilia may uncommonly accompany rheumatoid arthritis with hypereosinophilia (Chapter 35).
itself but more commonly results from adverse reactions to Infiltration of eosinophils accompanies rejection of lung,
treatment medications (including NSAIDs, gold, and tetracyclines) kidney, and liver allografts. Tissue and blood eosinophilia occur
or concomitant vasculitis. early in the rejection process, and eosinophil counts and

360 Part two Host Defense Mechanisms and Inflammation


eosinophil granule protein levels (in urine, BAL fluids, and blood, sputum, or tissue examinations, as well as results of
24
involved allograft tissues) have correlated with prognosis, severity, serological tests. The duration and magnitude of the eosinophilia
and response to rejection therapy. may suggest some entities, especially if it is prolonged or markedly
elevated (see Table 24.1). Other causes of eosinophilia that are
Endocrine Diseases amenable to treatment include eosinophilia secondary to medica-
The loss of endogenous adrenoglucocorticosteroid production tions, for which cessation of the offending drug may be indicated
in Addison disease, adrenal hemorrhage, or hypopituitarism can if the eosinophilia is accompanied by organ damage. Likewise,
cause increased blood eosinophilia, although usually not more if eosinophilia is secondary to glucocorticosteroid deficiency,
than mild to moderate. diagnostic testing can corroborate this deficiency and lead to
the administration of replacement corticosteroids and consequent
Other Causes of Eosinophilia resolution of the eosinophilia.
The syndrome of atheromatous cholesterol embolization is at Because allergic diseases usually are associated with at least
times associated with hypocomplementemia, eosinophilia, and some degree of eosinophilia, clinical and laboratory evidence of
eosinophiluria. Rarely, cases of hereditary eosinophilia among such disease should be sought. If the eosinophilia is not attribut-
family members have been recognized. Irritation of serosal surfaces able to allergic diseases, parasitic infections, medications, or
can be associated with eosinophilia, and related diseases can steroid deficiency, further evaluation will be guided by whether
include Dressler syndrome; eosinophilic pleural effusions; the patient has evidence of organ disease and, if so, which organs
peritoneal and, at times, blood eosinophilia developing during are involved (see Table 24.2). This is germane, for instance, in
chronic peritoneal dialysis; and perhaps the eosinophilia that defining whether the patient has a distinct eosinophilic pulmonary,
follows abdominal irradiation. GI, or cutaneous syndrome. Bone marrow examinations in most
patients with eosinophilia are not usually informative, revealing
only evidence of enhanced eosinophilopoiesis; but bone marrow
should be examined if there is suspicion of a hematological
tHEraPEUtIC PrINCIPLES malignancy or myeloproliferative disorder. For patients with
Therapy of Specific Eosinophilic Diseases sustained eosinophilia who meet the criteria for HES, diagnostic
testing should aim to identify which variant form of HES the
Eosinophil-associated Diseases with Identifiable Etiologies patient may have, in which case, bone marrow examination is
Parasitic infections Treat causative parasite often needed (see Fig. 24.4).
Drug-reaction related Terminate eliciting medication
eosinophilias oN tHE HorIZoN
Adrenal insufficiency Corticosteroid replacement therapy
Allergic/atopic diseases Varied, may include topical or inhaled Identify the causes of those hypereosinophilic syndromes for which
corticosteroids etiologies currently remain unknown.
Delineation of the signaling mechanisms that govern the agonist-specific,
Distinct Eosinophilic Syndromes Involving Specific organs differential secretion of cytokines that are preformed and stored within
Eosinophilic pulmonary diseases: eosinophil granules and secretory vesicles.
Acute eosinophilic pneumonia Corticosteroids Continue to evaluate the therapeutic efficacies of anticytokine therapeutics,
Chronic eosinophilic pneumonia Corticosteroids, interferon-α including anti–interleukin (IL)-5 neutralizing antibodies, in the treatment
Eosinophilic granulomatosis with Corticosteroids, interferon-α of the varied forms of eosinophilic diseases.
polyangiitis Identify biomarkers that are predictive of eosinophil-mediated tissue
damage and that can be used for clinical diagnostic and therapeutic
Hypereosinophilic Syndromes monitoring tools.
F/P-positive myeloproliferative Imatinib
Lymphoproliferative and other Corticosteroids, interferon-α,
hydroxyurea, anti–interleukin (IL)-5 Please check your eBook at https://expertconsult.inkling.com/
monoclonal antibody (mAb), other for self-assessment questions. See inside cover for registration
details.

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2011;12:151–9.

CHaPtEr 24 Eosinophils and Eosinophilia 361.e1


MUL t IPLE-CH o ICE QUES t I o NS

1. Which of the following may NOT contribute to or elicit 3. Which three cytokines promote eosinophil development in
eosinophilia? bone marrow and sustain eosinophil viability?
+
−
A. CD3 , CD4 aberrant lymphocyte subsets A. Transforming growth factor (TGF)-β, interleukin (IL)-2,
B. FIP1L1-PDGFRα chromosomal gene rearrangement IL-4
C. Giardia lamblia infection B. IL-7, IL-2, IL-13
D. Strongyloides stercoralis infection C. Granulocyte macrophage–colony-stimulating factor (GM-
CSF), IL-3, IL-5
2. A 45-year-old male who has had eosinophilia of between
3
2500 and 5000/ mm documented for the past 6 months has D. Interferon (INF)-γ, IL-8, IL-10
been referred to you. Your evaluation could include:
A. Flow cytometry for lymphocyte phenotyping
B. Serum immunoglobulin E (IgE)
C. Serum troponin
D. Strongyloides enzyme-linked immunosorbent assay (ELISA)
serology
E. All of the above

25









Host Defenses to Viruses



Barry T. Rouse, Scott N. Mueller







Viruses as obligate intracellular parasites require their host to rhinoviruses, coronaviruses, measles virus, varicella-zoster virus
replicate them and to facilitate their spread to others. In humans, (VZV), and respiratory syncytial virus (RSV) are often spread
most clinically relevant infections were derived from other by aerosol transmission, as well as person-to-person contact.
animals, and this process continues. Recent examples include Many of the herpes viruses target the skin or the mucosae, such
human immunodeficiency virus (HIV), Ebola virus, severe acute as herpes simplex virus (HSV) and VZV. HSV, in particular, can
respiratory syndrome (SARS) virus, and Zika virus. Viral infections infect the oral and genital mucosae, the eye, and skin through
are rarely lethal, even if they are highly cytolytic to individual small cuts and abrasions. Other herpes viruses, such as Epstein-
cells. Mortality commonly occurs when viruses jump species, Barr virus (EBV) and cytomegalovirus (CMV), target mucosae.
when the virus undergoes a major antigenic change (i.e., influenza CMV can also spread vertically from mother to baby or rarely
viruses), or when host immunity is compromised. HIV (Chapter via blood transfusions. Human papillomavirus (HPV) targets
39) represents one of the most dramatic human examples of an skin and mucosae and causes warts and may transform cells,
exotic virus that kills its host. However, HIV kills slowly, providing inducing cancers, such as cervical cancer. Some viruses, such as
ample time to spread to new hosts and an effective strategy for West Nile virus, Dengue virus, Semliki forest virus, and Zika
persistence in the species. Death or dire consequences following virus, can enter through skin via insect vectors. HIV and hepatitis
virus infection in mammals with inadequate immunity are well B virus (HBV) are commonly spread via sexual contact. HIV,
illustrated by observations that fetuses or neonates, especially if HBV, and hepatitis C virus (HCV) can also infect humans by
deprived of passive immunity, succumb to many agents well direct entry into the bloodstream via transfusions or contaminated
tolerated by healthy adults. The science of viral immunology needles.
seeks to understand mechanisms of virus–host interactions with Most human viruses replicate only in certain target tissues,
a view to applying this knowledge to the design of effective this being mainly the consequence of viral receptor distribution.
vaccines and immunomodulators that control virus infections. Many viruses use two receptors, such as the use of the CD4
These objectives are facilitated by an increasing wealth of coreceptor and the chemokine receptor CCR5 on T cells by HIV.
immunological techniques, an expanding array of genetically After attachment to a cellular receptor, viruses may fuse with
manipulated animal models, and an abundance of high through- the cell membrane or be endocytosed and then gain entry into
put technologies, which generate data that can be subjected to the cytoplasm or nucleus by fusing with the vesicular membrane
complex computational analysis. Such analyses can yield signatures (enveloped viruses, such as HSV and HIV), or translocate across
indicative of optimal immunogenicity and vaccine efficacy or the cell membrane or induce lysis of the endocytic vesicle once
failure and can explain the variable outcome of infections in in the cytoplasm (nonenveloped viruses, such as Norwalk virus
1
individual hosts. In most situations, defense against viruses and poliovirus). Viruses then utilize host cell machinery and
involves multiple immune components, and the impact of a specialized virally encoded proteins to replicate rapidly within
single mechanism varies greatly according to the method by the cell. Once they have multiplied within the cell, many viruses
which individual viruses enter, replicate, and spread within the induce cytolysis to facilitate release of new infectious virions
host. In this chapter, we highlight the principal means by which (e.g., poxviruses, poliovirus, and herpes viruses). Other viruses
the host achieves immunity after infection by viruses. Table 25.1 are released from infected cells by budding through the cell
presents an overview. membrane in the absence of cell death (e.g., HIV and influenza
virus). Having entered the body, however, viruses encounter
VIRAL ENTRY AND INFECTION numerous innate defenses and activate the components of adaptive
immunity. The latter usually assures that clinical disease, if not
Access to target tissues presents numerous obstacles for entry infection, will not become evident. Successful exploitation of
and infection by most human viruses. Most effective of these these defenses through the use of vaccines (Chapter 90) remains
are the mechanical barriers provided by skin and the mucosal a central challenge for many human viruses, particularly those
surfaces, as well as the chemically hostile environment of the that cause chronic infections, such as HIV and HCV. 2
gut (Fig. 25.1). A number of common human viral pathogens
enter through the gastrointestinal tract, including rotavirus, INNATE IMMUNITY TO VIRUSES
enteric adenoviruses, and hepatitis A virus (HAV). These are
usually spread via person-to-person contact or contaminated Viral infection induces an extensive array of defense mechanisms
food and water. Respiratory infections caused by influenza viruses, in the host. Innate defenses come into play to block or inhibit

365

366 Part tHrEE Host Defenses to Infectious Agents



TABLE 25.1 Viral Infections and Immunity TABLE 25.2 Sensors of Viral Infection
Viral Event Obstacles time Course toll-Like receptors (tLrs)
TLR3 dsRNA, MCMV, VSV, LCMV, HSV, EBV
Transmission Mechanical and chemical 0 TLR7 and TLR8 ssRNA, Influenza virus, HIV, VSV
barriers TLR9 dsDNA, HSV, MCMV
Infection and Innate immunity 0 → TLR2 MV hemagglutinin protein, HSV, HCMV
replication TLR4 MMTV envelope protein, RSV
Infection stopped or Viral antigens transported Within 24 hours
spreads to lymphoid tissues rIG-I-Like Helicases (rLHs)
Infection controlled Specific antibodies and 4–10 days
cell-mediated immunity RIG-I Influenza virus, VSV, HCV, JEV, MV,
Sterile immunity Immune memory 14 days to years RSV, Sendai virus, EBV
Viral persistence if Immune disruption or Weeks to years MDA-5 Poly(I:C), MV, Sendai virus, VSV,
infection not evasion MCMV, Picornaviruses
controlled
NOD-Like receptors (NLrs)
NLRP3 Influenza virus, Sendai virus,
Adenovirus, Vaccinia virus
NOD2 Influenza virus, VSV, RSV
Ocular infection Other sensors
• HSV Gastrointestinal tract AIM2 Vaccinia virus, MCMV
• Adenoviruses • Rotavirus ZBP1 (DAI) Cytosolic dsDNA, HSV
• Adenoviruses IFI16 Cytosolic dsDNA, HSV
Respiratory tract • Hepatitis A virus cGAS Cytosolic dsDNA, HSV
• Influenza virus • Caliciviruses
• RSV AIM2, absent in melanoma-2; IFI16, Gamma-interferon-inducible protein Ifi-16; cGAS,
• Rhinoviruses cyclic GMP-AMP Synthase; ZBP1, Z-DNA-binding protein 1; DAI, DNA-dependent
• Coronaviruses activator of IFN; dsRNA, double-strand RNA; EBV, Epstein-Barr virus; HCMV, human
• Adenoviruses cytomegalovirus; HCV, hepatitis C virus; HIV, human immunodeficiency virus;
parainfluenza virus HSV, herpes simplex virus 1/2; JEV, Japanese encephalitis virus; LCMV, lymphocytic
• VZV choriomeningitis virus; MCMV, murine cytomegalovirus; MDA-5, melanoma
• Measles virus Genitourinary tract differentiation-associated gene; MMTV, mouse mammary tumor virus; MV, measles
• HSV virus; NLR, NOD-like receptor; RLH, RIG-I-like helicase; RSV, respiratory syncytial virus;
• HIV
Skin entry and infection • HBV ssRNA, single-strand RNA; TLR, Toll-like receptor; VSV, vesicular stomatitis virus.
• HSV • CMV
• Human papillomavirus • Human papillomavirus
• West Nile virus
FIG 25.1 Common Routes of Entry and Infection for Human double-stranded RNA (TLR 3 and TLR7/8, respectively) or
Viral Pathogens. CMV, cytomegalovirus; HBV, hepatitis B virus; double-stranded DNA (TLR9).
HIV, human immunodeficiency virus; HSV, herpes simplex virus; The RLHs retinoic acid-inducible gene I (RIG-I) and
RSV, respiratory syncytial virus; VZV, varicella-zoster virus. melanoma differentiation-associated gene (MDA-5) mediate
cytoplasmic recognition of viral nucleic acids. These activate
mitochondrial antiviral signaling (MAVS) proteins to stimulate
initial infection, to protect cells from infection, or to eliminate IFN-I production and activate inflammasomes, which are
virus-infected cells. Innate mechanisms occur well before the molecular complexes that facilitate the activation of caspases
4
effectors of adaptive immunity become active, but they are critical and induce the production of proinflammatory IL-1β and IL-18.
for the initiation of adaptive immunity via the elicitation of NLRs are a second class of cytosolic sensors of PAMPs that activate
inflammation that promotes immune cell activation. The innate inflammasomes via the adapter protein ASC. These include the
immune defenses are initiated via pattern recognition receptors NLRP (or NALP), NOD, and IPAF/NAIP receptors. Three major
(PRRs), which recognize pathogen-associated molecular patterns inflammasomes have been shown to be involved in antiviral
3
(PAMPs) (Chapter 3). These include transmembrane receptors immunity: the NLRP3 inflammasome, the RIG-I inflammasome,
of the Toll-like receptor (TLR) family, two families of intracel- and the AIM2 inflammasome. 3
lular receptors including the NOD-like receptors (NLRs) and the The innate defense system consists of multiple cellular
RIG-I–like helicases (RLHs), as well as the sensor molecule absent components and many specialized proteins. The longest known
in melanoma-2 (AIM2). Additionally, the molecules cyclic gua- and best-studied antiviral proteins are the α/β IFNs, which act
nosine monophosphate–adenosine monophosphate (GMP-AMP) by binding to the type I IFN receptor and result in the transcrip-
synthase (cGAS), DDX41, IFI16, and Z-DNA–binding protein tion of more than 100 IFN-stimulated genes. One consequence
1 (ZBP1) can sense cytosolic DNA (Table 25.2). These cellular of this “antiviral state” is the inhibition of cell protein synthesis
5
sensors promote the expression of interleukin-1 (IL-1) and and the prevention of viral replication. Multiple leukocyte subsets
IL-18, type I (α/β) interferon (IFN-I), and a variety of IFN- are involved in innate defense, including macrophages, DCs,
stimulated genes and inflammatory cytokines, and chemokines. neutrophils, natural killer (NK) cells, natural killer T cells (NKT
TLRs are cell surface or endosomal membrane–bound proteins cells), and γδT cells. Furthermore, tissue cells, including fibroblasts,
expressed by numerous cells, including dendritic cells (DCs), epithelial cells, and endothelial cells, express PRRs and respond
macrophages, lymphocytes, and parenchymal cells. Expression to viral infection via the production of innate cytokines, including
of TLRs is largely inducible in most cell types, although some IFN-I and IL-1. IFN-I is a critical link between the innate and
(TLR7/8/9) are constitutively expressed at high levels by special- adaptive immune system, via activation of DCs and T cells, as
6
ized plasmacytoid DCs for rapid IFN production. Different TLR well as protecting T cells from NK cell-mediated attack. IFN-Is
molecules recognize specific viral products, such as single- and can also activate NK cells and induce other cytokines that promote

CHaPtEr 25 Host Defenses to Viruses 367



KEY CONCEPtS PRRs; the cytokines IFN-I, IL-I, IL-33, and IL-12; and phagocytes,
Major Antiviral Innate Defense Mechanisms including macrophages, monocytes, and DCs) serve to shape
the nature and effectiveness of the subsequent adaptive response
Acting to block infection: to viral pathogens. For instance, DCs require innate signals, such
Natural antibodies as IFN-I and IL-12, for maturation and optimal T-cell activation.
+
Complement components Furthermore, CD8 T cells responding to viruses need IFN-I
Some cytokines and chemokines and IL-33 signals for expansion and memory formation. Thus
Acting to protect cells from infection: both the magnitude and the type of innate response induced by
Interferon-α/β virus infection have a marked influence on the generation of
Interferon-γ (IL-γ)
IL-1, IL-18 adaptive immune responses.
Acting to destroy or inhibit virus-infected cells:
Natural killer (NK) cells ADAPTIVE IMMUNITY TO VIRUSES
Natural killer T cells (NKT cells)
Macrophages Innate immunity generally only slows, rather than stops, viral
Neutrophils infection, allowing time for the adaptive immune response to
γδ T cells
Nitric oxide begin. The two major divisions of adaptive immunity, antibody-
Involved in regulating antiviral inflammatory response: mediated and T-cell–mediated, are mainly directed at different
ILs-1, 6, 10, 12, 18, 23, 33 targets. Antibodies usually function by binding to free viral
Transforming growth factor (TGF)-β particles and, in so doing, block infection of the host cell (Chapter
Chemokines (CCL2, 3, 4, 5) 15). In contrast, T cells act principally by recognizing and destroy-
ing virus-infected cells or by orchestrating an inflammatory
response that includes several antiviral components (Chapters
NK responses, such as IFN-γ and IL-12. NK cells produce 16, 17). As all viruses replicate within cells and many can spread
proinflammatory cytokines; they can kill infected cells and interact directly between cells without reentering the extracellular environ-
with DCs, and are an important component of innate defense ment, resolution of infection is reliant more on T-cell function
against viruses. NK cells can protect against some herpes viruses, than on antibody function. However, broadly neutralizing antiviral
which downregulate major histocompatibility complex (MHC) antibodies have the potential to be effective therapies against
expression in the cells they infect. NK cells are also important many different human infections, including HIV, influenza viruses,
in resistance to mouse and human CMV and possibly to HIV, and Ebola virus. Recent advances have allowed researchers to
7
influenza virus, and Ebola virus. NK cells have also recently isolate and identify human monoclonal antibodies (mAbs) against
12
been shown to possess traits of adaptive immunity and, like T these and other pathogens, offering promise of new therapies
8
and B cells, can form populations of memory cells. NK cells as well as significant insight for vaccine design. Antiviral antibodies
are regulated by an array of activating and inhibitory receptors, are also very important as an immunoprotective barrier against
whose expression and function are just beginning to be under- reinfection. It is the presence of antibodies at portals of entry—
stood. Uninfected cells are usually protected from NK cell cytolysis most often mucosal surfaces—that is of particular relevance to
13
as they deliver negative signals, such as high expression of MHC influenza, HSV, and HIV infections. Yet, how to generate vaccines
molecules. In contrast, virus-infected cells are killed either because that induce optimal antibody responses, including broadly
they deliver positive signals or because they lack adequate MHC- neutralizing antibodies, remains an important unsolved problem.
negative signals. NK cells may also control excessive immune Initiation of adaptive immunity is closely dependent on early
+
responses to viruses by killing CD4 T cells and indirectly regulat- innate mechanisms that activate antigen-presenting cells (APCs),
ing cytotoxic T lymphocyte (CTL) responses. NKT cells may principally subsets of DCs. APCs and lymphocytes are drawn
provide some antigen-specific innate immune protection against into lymphoid tissues by chemokine and cytokine signals and
certain viruses, such as influenza virus. 9 are retained there for a few days to facilitate effective intercellular
Several classes of innate host proteins function in antiviral interactions. The architecture of the secondary lymphoid tissues
defense. These include natural antibodies, which may play a role supports the coordinated interactions among the cells of the
14
in defense against some viral infections, as well as pentraxins adaptive immune system through a network of supportive
10
15
and complement proteins. Some viruses may be directly stromal cells and local chemokine gradients (Chapter 2). The
inactivated by complement activation or be destroyed by phago- induction events occur in lymph nodes draining an infection
cytic cells that bind and ingest complement-bound virions. Several site or in the spleen if virus enters the bloodstream. The passage
proinflammatory cytokines and chemokines induced by virus of viral antigens to lymph nodes usually occurs in DCs. Some
infection also play key roles in defense. Foremost among these viruses are able to compromise the function of APCs, such as
is IL-1 and other members of the IL-1 family, including IL-18 HSV and measles virus, which can inhibit DC maturation.
11
and IL-33. These cytokines influence both innate and adaptive B-cell activation occurs following antigen encounter in the
immune cells and play critical roles in antiviral defense. Other B-cell follicles, and possibly the T-cell zones, in the spleen or
antiviral cytokines are produced early following infection, such lymph nodes. Some activated B cells become short-lived plasma
as TNF-α, IFN-γ, IL-12, IL-6, and chemokines, such as MIP-1α. cells, whereas others move to the edges of the B-cell follicles and
+
In particular, IL-12 is a potent inducer of IFN-γ from NK cells. interact with antigen-specific helper CD4 T cells via presentation
Inflammatory chemokines may also play an important role in of antigenic peptides on B-cell MHC class II molecules. These
innate antiviral defense by orchestrating macrophage, neutrophil, Bcl6-dependent CD4 T follicular helper (Tfh) cells are specialized
DC, and NK cell responses at the site of infection. Not only are for providing help for B-cell responses and are needed to promote
16
these components of innate immunity involved in mediating and regulate B-cell responses. Activated B cells initiate germinal
initial protection against viruses; several components (e.g., the center (GC) reactions with the help of CD4 Tfh cells, ensuring

368 Part tHrEE Host Defenses to Infectious Agents


somatic hypermutation and affinity maturation for the selection Primary infection Recall infection
of high-affinity, antibody-producing, long-lived plasma cells, as Expansion Contraction Memory
17
well as memory B cells. At the molecular level, upregulation
of the transcription factors Blimp-1, XBP-1, and IRF-4 dictates
plasma cell formation, whereas Pax-5 expression delineates B
cells destined for GC reactions and the memory B-cell lineage.
Antibody binding to epitopes expressed by native proteins at
the surface of free virions usually blocks viral attachment or
penetration of target cells. Sometimes the consequence is viral Antigen - specific T cells
lysis (with complement proteins also involved), opsonization,
or sensitization for destruction by Fc receptor–bearing cells that
mediate antibody-dependent cellular cytotoxicity (ADCC).
Occasionally, however, Fc receptor binding of antibody-bound
virus may facilitate infection and result in more severe tissue Time
damage. This occurs in Dengue fever and may happen in some A
instances in HIV infection. The antibody involved in the protec-
tion of mucosal surfaces in humans is predominantly secretory Naive Effector 90–95% death
immunoglobulin A (IgA), but serum-derived IgG may also be T EM T CM
13
protective, particularly in such sites as the vaginal mucosa.
Both antibody isotypes act mainly to block infection of epithelial
cells, although in some instances, the antibody may transport hi hi
antigen from within the body across epithelial cells to the outside. CD62L hi CD62L lo CD62L lo lo CD62L hi
CCR7
CCR7
Mucosal antibody persists for a much shorter period compared CD69 lo CCR7 lo CCR7 lo CD69 lo
CD69
with serum antibody, which explains, in part, why immunity to T RM
mucosal pathogens is usually of much shorter duration compared CD62L lo
with immunity to systemic viral infections. CCR7 lo
CD69 hi
B
KEY CONCEPtS FIG 25.2 Expansion/Contraction/Memory Phases of Adaptive
Immunity and Memory Cell Subsets. (A) Dynamics of primary
Antiviral T- and B-Cell Immunity and secondary (recall) T-cell responses to viral infection. Both
primary and recall T-cell responses undergo expansion and
Effector recognized
Systems Molecules Control Mechanisms contraction phases, followed by stable immune memory. Recall
responses induce a larger effector pool and reduced contraction
Antibody Surface proteins or Neutralization of virus, further boosting the memory pool. (B) Effector and memory
virions opsonization, or T-cell differentiation. Antigen stimulation expands effector cells,
destruction of infected
cells by ADCC most of which die during the contraction phase. Effector memory
Antibody + Surface proteins Infected cell destruction T (T EM ) cells that are formed gradually convert to central memory
complement expressed on by ADCC or T (T CM ) cells over time, with corresponding changes in surface
infected cells complement-mediated marker expression. Some effector T cells develop into resident
lysis memory T (T RM ) cells that persist in the tissues and do not
Mucosal Surface proteins or Viral neutralization, reenter the circulation.
antibody virions opsonization, and
(IgA) transcytosis
CD4 T cells Viral peptides Antiviral cytokine and
+
(10–20 mers) chemokine Like B-cell responses, T-cell responses to viral infections also
presented on MHC production; help for begin within lymphoid tissues. Specific CD8 CTL precursors
+
+
class II surface, CD8 T-cell and B-cell recognize antigen in the context of MHC class I–peptide antigen
internal or responses; killing +
nonstructural infected cells; complexes on DCs. The CD8 T cells become activated, proliferate,
proteins presented regulatory functions to and differentiate into effectors. Expansion of these naïve antigen-
by APCs reduce specific precursors is considerable, often exceeding 10 000-fold,
immunopathology and results in an effector population that can account for 40%
+
CD8 T cells Viral peptides Killing infected cells or or more of a host’s total CD8 T-cell population (Fig. 25.2).
+
(8–10 mers) purging virus without Various factors, including antigen and APCs, costimulatory
presented on MHC cell death; antiviral
class I surface, cytokine and molecules (e.g., CD28 and 4–1BB), and inflammatory cytokines
internal, or chemokine production (e.g., IFN-I and IL-12) are required to program the development
+
nonstructural of functional effector lymphocytes. In some infections, CD4
proteins presented T-cell help is also important to prime robust CTL responses via
on infected cells or signals, including CD40 that are delivered to DCs. Activated
18
by cross-presentation CTL effectors then exit lymphoid organs and access almost all
ADCC, antibody-dependent cellular cytotoxicity; APC, antigen- body locations via the bloodstream. However, effectors do not
presenting cell; IgA, immunoglobulin A; MHC, major histocompatibility stay activated for long once the virus is cleared, and approximately
complex. 95% die by a process termed activation-induced cell death.

CHaPtEr 25 Host Defenses to Viruses 369


Following this contraction phase, the remaining cells differentiate tumor necrosis factor [TNF]-α, IL-2) and Th17-producing cells
into memory cells, which remain as a more or less stable popula- (IL-17a and IL-22). A third effector subset, Th2 cells producing
tion in the host for many years. They represent an expanded (IL-4, IL-5, and IL-13), also participates in inflammatory reactions,
pool of CTL precursors that can be activated upon secondary although in the case of viruses, these are usually more tissue
encounter with antigen and provide enhanced protection upon damaging than protective. This situation can occur in responses
reinfection with the same virus (see next section). Although to RSV infection. Regulatory T cells (Tregs) are a further subset
+
much of our knowledge of T-cell responses to viruses has been of CD4 T cells of particular importance, since these cells largely
obtained from murine studies, it is increasingly clear that the act to regulate the function of effector subsets and, in so doing,
22
fundamental principles are the same or similar in humans. 19 influence the severity and duration of inflammatory reactions
+
T-cell immunity against a particular virus involves both CD4 (Chapter 18). Tregs produce antiinflammatory cytokines, such
+
+
and CD8 T-cell subsets that recognize peptides derived from as IL-10 and TGF-β, and can be distinguished from other CD4
viral antigens bound to surface MHC proteins (class II and class subsets by their expression of a unique transcription factor FoxP3.
+
I, respectively) (Chapters 5, 6). Complexes of viral peptides bound The balance of CD4 T-cell subset representation in response
to MHC class II proteins are generated by APCs from scavenged to a virus infection is critical. In situations where responses
and processed virus-infected cells or viral particles. Antigen–MHC become overtly tissue damaging and chronic, the balance favors
class I complexes are expressed on the surface of infected cells, effector subsets. In such situations, changing the balance to favor
and antigen can also be transferred to APCs from infected cells Tregs can result in diminished lesions.
by a process known as cross-presentation. Recent experiments in
mice have also demonstrated a role for transfer of antigen between IMMUNOLOGICAL MEMORY
DCs as they migrate from infected tissues to the lymphoid tissues.
Multiple subsets of DCs exist and specialize somewhat in antigen Immunological memory is a cardinal feature of adaptive immunity.
20
presentation on MHC-I or MHC-II. During the process of The goal of vaccinology is to induce long-lived immunological
activation, T cells can receive signals from multiple DC types in memory to protect against reinfection (Chapter 90). Following
+
a temporally controlled sequence that coordinates CD4 and infection with certain viruses, memory can be exceptionally
+
18
CD8 T-cell interactions. Use of MHC class I and class II tet- long-lived, potentially for the life of the host (e.g., yellow fever
+
+
ramers to directly visualize antigen-specific CD8 and CD4 T-cell and smallpox viruses). 19,23 Memory is defined by the persistence
responses, respectively, has demonstrated the significant size of of specific lymphocytes and antibody-producing plasma cells
T-cell responses to viruses, such that the majority of the activated rather than that of antigen to induce continuous lymphocyte
T cells seen at the peak of the response are virus-specific. activation. Humoral memory to viruses involves long-lived plasma
CTLs function by recognizing virus-infected cells and killing cells in bone marrow, which provide a continuous low-level source
them; this often involves perforins and cytotoxic granules contain- of serum antibody. This maintenance of humoral immunity also
ing granzymes. Effector CTLs can also induce death in target involves a population of homeostatically maintained memory B
cells following engagement of the Fas ligand on the CTL with cells, which may be required to maintain stable numbers of
Fas on target cells. Both pathways lead to apoptosis of the target long-lived plasma cells over time. The pool of memory T cells
cell, involving the degradation of nucleic acids, including those is regulated by low-level homeostatic division controlled by the
+
+
of the virus. Alternatively, CD8 T cells also mediate defense cytokines IL-7 and IL-15. For memory CD8 T cells, IL-7 is primar-
through the release of various cytokines after antigen recognition. ily important for survival, whereas IL-15 is crucial for low-level
Some of the cytokines and chemokines most highly produced proliferation to maintain the size of the memory T-cell pool.
by CTLs include IFN-γ, TNF-α, lymphotoxin-α, and RANTES
(CCL5) (Chapters 9, 10). These cytokines can have multiple KEY CONCEPtS
antiviral effects on infected cells and on the cells around them, Principles of Antiviral Immunity
including purging of virus from infected cells without killing
the cells. This is particularly important for such viruses as HSV, Many human viral infections are successfully controlled by the immune
which infects nonrejuvenating cells, such as nerve cells. system.
+
CD4 T cells are involved in antiviral defense as well as being Certain emerging viruses may overwhelm the immune system and cause
severe morbidity and mortality.
modulators of inflammatory reactions to viruses. Multiple Other viruses have developed mechanisms to overwhelm or evade the
+
functional subsets of CD4 T cells are recognized based largely immune system and persist.
on the types of cytokines produced when they recognize antigen. Individuals with defects in innate or adaptive immunity demonstrate
+
+
CD4 T cells are more broadly reactive than CD8 T cells; they more severe viral infections.
recognize larger peptides processed from viral proteins and are T-cell immunity is more important for control than are antibodies in many
+
restricted by MHC class II. These CD4 T cells participate in viral infections.
antiviral immunity in several ways. They can act as helper cells Antibodies are important to minimize reinfection, particularly at mucosal
sites.
for the development of high-affinity antibody responses and for Immune memory is often sufficient to prevent secondary disease, although
+
+
more functional CD8 T-cell responses. 16,21 Additionally, CD4 not in all viral infections.
T cells act as effectors and orchestrate inflammatory reactions, Tissue-specific immune memory may be important to rapidly protect
which either serve a protective function or, in some cases, become against reinfection at peripheral sites (e.g., skin and mucosae).
prolonged causing chronic tissue damage (Chapter 16). The latter
can happen in HCV-mediated hepatitis and HSV-mediated Immunological memory is defined by a pool of antigen-specific
+
stromal keratitis. Occasionally, CD4 T cells can mediate direct cells whose increased frequency enables rapid control of viral
+
cytotoxicity, but they are less effective than CD8 T cells. The reinfection (see Fig. 25.2). IL-7Rα-expressing effector T cells are
+
principal subsets of CD4 T cells involved in inflammatory the precursors of this memory pool. This population of cells,
reactions are T helper-1 (Th1) cells (producing mainly IFN-γ, which constitutes about 5–10% of the effector pool, preferentially

370 Part tHrEE Host Defenses to Infectious Agents


survives the contraction phase and gradually differentiates into T RM cells can trigger enhanced early inflammation to drive local
24
a stable memory population. Upon reinfection, these memory immunity. This is in contrast to T EM cells, which continue to
cells can be rapidly activated and, by virtue of their increased migrate through nonlymphoid tissues, rather than being seques-
+
frequency, mediate more rapid clearance of the viral pathogen. tered in peripheral tissues, and also differs from the CD8 and
+
Moreover, repeated stimulation of memory cells via multiple CD4 T CM , which migrate largely through lymphoid organs (spleen
infections with the same virus, or prime-boost vaccine regimes, and lymph nodes). These differences may define the physiological
further increases the size of the antigen-specific memory T-cell raison d'être for these memory T-cell subsets, highlighting that
25
pool. Restimulation also affects the activation status and tissue measurement of memory T cells in human peripheral blood is
distribution of memory T cells, which may enhance protection a poor representation of the total-body memory T cell pool.
from viral infection in mucosal and other tissues. T RM cells can be detected in tissues by using markers, such
Experiments in humans and mice have demonstrated that as CD69 and CD103, although these are imperfect identifiers,
memory T cells are heterogeneous. Memory T cells were divided including in human tissues. T RM cells in different anatomical
into effector memory (T EM ) and central memory (T CM ) subsets, locations share a common genetic signature and require common
defined by expression of two surface molecules involved in T-cell transcription factors for their formation. Yet, these cells also
lo
24
lo
migration: CD62L and CCR7. The CD62L CCR7 T EM subset adopt unique gene expression that is imprinted by the tissue
is found primarily in nonlymphoid tissues and the spleen, whereas environment, and presumably imparts specialized functions on
hi
hi
the CD62L CCR7 T CM subset is largely present in lymph nodes T RM cells in each location. However, memory in certain peripheral
and the spleen. The current model predicts that effector T cells tissues, such as lungs, appears to wane over time, suggesting that
form the T EM subset and that these cells gradually convert to a memory T cells may not persist in sufficient numbers in this
T CM phenotype over time (Fig. 25.2B). Although the conditions site. This rationalizes a need for vaccines that induce optimal
that control the rate of this conversion are unknown, it is likely numbers of memory T cells in tissues as well as blood.
that the amounts of antigen and inflammatory signals received
during the effector phase greatly influence this. It has also been
+
shown that CD4 T-cell help is required for the generation of IMMUNE EVASION AND IMMUNITY TO CHRONIC
+
long-lived memory CD8 T cells, via interactions with DCs. 21 VIRAL INFECTIONS
Studies suggest that T CM are capable of mounting stronger
proliferative responses following reinfection. Tissue-specific Many, if not all, viruses employ immune blunting or delay tactics
homing of T EM cells permits them to enter sites of potential viral to circumvent aspects of the immune system, allowing them
27
infection, such as skin and mucosae. However, we now know time to replicate further or escape detection (Table 25.3). One
that many memory T cells found at sites of previous viral infec- such mechanism may involve killing or infecting APCs. Viruses
26
tions take up long-term residence in tissues. This includes skin, may also delay or prevent apoptosis induced by CTLs within
+
intestines, lungs, the liver, and the brain. These resident memory infected cells. Other viral evasion measures aimed at the CD8 T
T cells (T RM cells) are sequestered from the circulation and provide cell–mediated antiviral defense system inhibit antigen processing,
rapid protection against viruses, such as HSV, in skin, where thereby minimizing effector CTL induction. To escape CTL killing,
they localize with a unique dendritic morphology and undergo many viruses also downregulate the MHC molecules on the
slow surveillance of the tissue (Fig. 25.3). Notably, activation of surface of infected cells. In addition, viruses may produce various
mimics or modulators/inhibitors of cytokines, chemokines, or
Epidermis
TABLE 25.3 Mechanisms and Examples of
Viral Immune Evasion
Mechanism Example
Interference with viral antigen HSV (ICP47), EBV (EBNA-1), HIV
processing and presentation (Nef, Tat), HPV (E5), CMV (UL6)
Evasion of NK cell function HIV (Nef), EBV (EBNA-1), CMV
(UL40, UL18)
+
+
CD8 T RM Basement CD8 T RM Inhibition of cell apoptosis Adenovirus (RID complex and
membrane E1B), HIV (Nef), EBV (BHRF-1)
Destruction of T cells HIV
Interference with antiviral EBV (IL-10 homologue),
+
CD4 T
CD4 T EM EM cytokines and chemokines CMV(US28 chemokine receptor
+
homologue), vaccinia virus
(IL-18-binding protein), HIV (Tat
chemokine activity)
Dermis
Inhibition of complement action HSV, pox viruses
+
+
FIG 25.3 Unique Subsets of Memory CD8 and CD4 T Cells Inhibition of DC maturation HSV, vaccinia virus
Reside Within Peripheral Tissues, at Sites of Previous Viral Frequent antigenic variation Influenza virus, HIV
Infection, and Provide Rapid Protection Against Reinfection. Infection of immune privileged Measles virus, VZV and HSV
(neurons)
site
+
Resident memory CD8 T cells (T RM ) remain localized in the Immune exhaustion HIV, HCV, HBV
epidermis in skin after herpes simplex virus (HSV) infection.
Resident memory CD4 T (T EM ) cells continue to migrate through CMV, cytomegalovirus; DC, dendritic cell; EBV, Epstein–Barr virus; HBV, hepatitis B
+
the dermal layers of skin, with access to blood and lymphoid virus; HCV, hepatitis C virus; HIV, human immunodeficiency virus; HPV, human
papillomavirus; HSV, herpes simplex virus; IL-18, interleukin-18; NK, natural killer; RID,
tissues. receptor internalization and degradation; VZV, varicella-zoster virus.

CHaPtEr 25 Host Defenses to Viruses 371


other components of the immune system or their receptors. exhausted T cells. These studies implicated multiple inhibitory
Viruses also resort to antigenic hypervariability to escape antibody receptors as a potential therapeutic targets, and although combina-
or T-cell recognition. This can occur during transmission from tions of these checkpoint inhibitor blockade therapies are proving
31
host to host (e.g., influenza virus), or within hosts during chronic highly beneficial to the treatment of certain cancers, similarly
infection through the generation of viral escape mutants. The efficacious responses have yet to be demonstrated during chronic
latter is particularly important for HIV and HCV infections. virus infection.
The success of many viral pathogens rests in their ability to
subvert the host immune response. The most successful human
viruses can escape the immune system and persist for the life OUTCOMES OF VIRUS INFECTION: IMMUNITY
28
of the host. Two well-studied examples of this are CMV and OR IMMUNOPATHOLOGY
EBV. T-cell responses to these viruses are prominent and readily
detectable in humans, and yet the immune system is unable to Typically, individual humans respond to a virus infection in
clear either pathogen completely. However, these viruses generally different ways. When the common cold or even pandemic
remain undetectable in immunocompetent individuals. Other influenza infection occurs, only a small percentage of exposed
viral infections, such as those caused by the herpes viruses HSV persons may develop overt clinical disease. In the prevaccine
and VZV, are marked by periods of latency when no virus can days, poliomyelitis was a much-feared consequence of poliovirus
be detected. Yet, periods of viral reactivation, often triggered by infection, but only a very small percentage of infected persons
stress, can lead to episodes of disease. These are controlled by developed the paralyzing complications. Similarly, only an
the immune response, which plays a central role in controlling unfortunate few develop life-threatening meningoencephalitis
herpes virus latency. 29 following infection with the insect-transmitted West Nile virus.
Many of the most medically important human viruses are It is particularly characteristic of chronic viral infections that
associated with persistent viremia. These include those causing clinical expression is highly variable. With HCV, for example,
chronic infections, such as HIV, HCV, HBV, and human in 70–80% of patients, some form of chronic liver disease develops,
T-lymphotropic virus (HTLV), among others. Such chronic viral and the virus is not cleared. However, in up to 30%, the infection
infections are marked by high levels of persisting antigen and is controlled, the virus is cleared, and immunity to reinfection
can result in skewed T-cell immunodominance hierarchies, altered develops. The latter group of individuals make a type of immune
tissue localization of immune cells, and severely impaired T-cell response that includes protective antibodies along with an
30
function. This altered T-cell function is hierarchical and results appropriate pattern of T-cell responsiveness. 32
in functional T-cell defects ranging from reduced cytokine We do not fully understand the reasons for the varying
production and altered proliferative capacity (exhaustion) to outcomes of virus infections in different persons, and almost
death (deletion) of the responding T cells (Fig. 25.4). certainly multiple factors are involved. Many of these factors
Sustained viral antigen levels and inflammation are responsible impact the response pattern made by the innate immune system,
for this immune dysfunction. This is in stark contrast to normal which, in turn, affects the magnitude and type of adaptive immune
memory T-cell development, which occurs in the absence of response that occurs. Some of the circumstances that do influence
persisting antigen (see previous section). Studies have demon- the outcome of infection include genetic susceptibility of the
strated that signaling through multiple inhibitory receptors host, the age of the host when infected, the dose and route of
expressed on the cell surface contributes to exhaustion during infection, the variable induction in the host of antiinflammatory
30
chronic infections. This includes the receptor programmed cells and proteins, and the presence of concurrent infections
death (PD)-1, expression of which may be essential for preventing and past exposure to cross-reactive antigens. 32
excessive immunopathology by effector T cells and yet appears
to contribute directly to failed immunity to HIV infection and IMMUNOPATHOLOGY AND AUTOIMMUNITY
other chronic human viral infections. Although the molecular
mechanisms of exhaustion remain unclear, differential involve- Immune responses against virus-infected cells often result in
ment of transcription factors and altered gene expression define tissue damage, especially if cell killing is involved or if there is
extensive recruitment and activation of inflammatory cell types,
such as macrophages and sometimes neutrophils. If the response
Cytokines/ Proliferative Antigen PD-I is brief and is quickly repaired, it is usually deemed an immu-
killing potential load expression
noprotective event. A prolonged tissue-damaging effect resulting
Functional T cells from an immune reaction against viruses is considered immu-
nopathology. Such situations most commonly involve persistent
Partial exhaustion
viruses, which are themselves often mildly cytodestructive in the
Full exhaustion absence of an immune reaction. Chronic tissue damage initiated
by viruses can also result in development of an autoreactive and
Deletion (death)
an occasionally oncogenic response. For example, some auto-
FIG 25.4 Hierarchical Model of T-Cell Exhaustion During immune diseases may be initiated or exacerbated by viral
Persistent Viral Infection. T-cell function (cytokine production, infections, but no named virus has been regularly incriminated
33
killing, and proliferative potential) is negatively influenced by as a cause of human autoimmune disease. Circumstantial
increasing levels of antigen. Low levels of persistent antigen evidence exists for a virus link in multiple sclerosis (MS), insulin-
may lead to partial loss of function and intermediate levels of dependent diabetes, and possibly systemic lupus erythematosus
programmed death (PD)-1 expression. High, sustained levels of (SLE). In MS, many viruses have been isolated from patients,
antigen over time can lead to full loss of function, high levels although no specific one has been tied to the disease etiology.
of PD-1, and eventually cell death (deletion). The current hypothesis is that viral infections set up an

372 Part tHrEE Host Defenses to Infectious Agents



TABLE 25.4 Lesions resulting
From Immunopathology
Primarily involving CD8 T Murine lymphocytic choriomeningitis
+
cells acting as cytotoxic T virus;
lymphocytes or sources of Hepatitis B virus (HBV)–induced
proinflammatory cytokines chronic hepatitis
Coxsackie B virus–induced diabetes
Coxsackie B virus–induced
myocarditis
Demyelination caused by some
strains of mouse coronavirus and
Theiler virus
+
Primarily involving CD4 T Demyelination caused by some
cells that produce Th1 strains of mouse coronavirus and
cytokines Theiler’s virus;
Herpes simplex virus (HSV)–induced
stromal keratitis FIG 25.5 Example of Herpetic Stromal Keratitis (Hsk) in the
Involvement of CD4 T cells Respiratory syncytial virus (RSV)– Human Eye After Herpes Simplex Virus-1 (HSV-1) Infection.
+
that produce Th2 cytokines induced pulmonary lesions
Involvement of antibody Glomerulonephritis in chronic Inflammation of the eye and eyelid can be observed, as well as
hepatitis B neovascularization and substantial necrosis, ulceration, and opacity
Dengue hemorrhagic fever of the cornea.



inflammatory environment that may exacerbate or tip the balance metalloproteinases, and components of the oxygen burst. Although
toward disease in genetically susceptible individuals. coxsackie virus can be a cause of diabetes in the mouse, attempts
Immunopathological reactions involving viruses have several to relate viral infection directly to the etiology of human diabetes
mechanisms, but T cells are usually involved as orchestrators of have so far failed.
inflammatory events (Table 25.4). The clearest example of
immunopathology involving a virus is lymphocytic choriomen- CLINICaL rELEVaNCE
ingitis virus (LCMV) in the mouse. This model has dominated
ideas and has set several paradigms in viral immunology in Hypothesized Role of Viruses in Autoimmunity
general. The first virus-induced immunopathological lesions Molecular mimicry: similar epitopes shared by virus and host
recognized were glomerulonephritis and arteritis, noted in mice Bystander activation: chronic release of cytokines and host antigens
persistently infected with LCMV. The lesions were assumed to activates local autoreactive lymphocytes
represent inflammatory reactions to tissue-entrapped immune Viral persistence: chronic viral antigen presentation on host cells leads
complexes that activate complement. Similar immune complex– to prolonged immunopathology
mediated lesions occur in other infections, including lung lesions
found in severe influenza, respiratory syncytial virus infection, Immunopathological reactions against viruses can also involve
+
viral hepatitis, and arthritis. However, only rarely have viral subsets of CD4 T cells, which can be either Th1 or Th17 or
antigens been shown to contribute to the antigen component both. One well-studied example involves persistent infection with
34
of the complex. An example where the inclusion of viral antigen Theiler virus in mice. This infection causes a demyelinating
in immune complexes has been demonstrated is chronic HBV syndrome that resembles the autoimmune disease experimental
+
infection of humans. Autoimmune diseases, such as SLE, also allergic encephalomyelitis. In both situations, CD4 T cells that
result from immune complex–mediated tissue damage. However, produce Th1 cytokines appear to serve as pathological mediators.
evidence linking viruses to the etiology or pathogenesis of SLE Furthermore, in both models an increase in the involvement of
is scarce, since the immune complexes in SLE do not appear to myelin-derived autoantigens occurs as the disease progresses.
include viral antigens at any stage. Once again, such observations indicate the possible role of a
+
Thanks largely to the LCMV model, it is clear that CD8 virus in an autoimmune disease. With the Theiler virus model,
T-cell recognition of viral antigens can result in tissue damage. the virus persists in the nervous system and chronically stimulates
+
In LCMV infection, damage occurs in the leptomeninges of CD4 T cells to secrete an array of cytokines. The demyelinating
immunocompetent mice infected intracerebrally. Hepatitis can events appear to result from cytokine action on oligodendrocytes.
also occur in mice infected intravenously. Neither lesion becomes Myelin components, such as myelin basic protein, proteolipid
+
+
evident if the CD8 T-cell response is suppressed. CD8 T protein, and myelin oligodendroglial glycoprotein, may be released
cell–mediated immunopathology can be a causative mechanism and can participate as additional antigen in immunoinflammatory
of chronic hepatitis associated with HCV and HBV infection, events. This scenario is referred to as epitope spreading.
+
although the tissue damage also involves inflammatory CD4 T Another model of virus-induced immunopathology that
+
cells. Additional viral immunopathology models where lesions mainly involves the Th1 subset of CD4 T cells is stromal keratitis
+
35
result primarily from CD8 T-cell involvement include myocarditis caused by HSV infection (Fig. 25.5). The pathogenesis of this
and insulin-dependent diabetes associated with coxsackie B virus immunopathological lesion is unusual in that it occurs and
+
infection. In both instances, CD8 T cells mainly orchestrate progresses when viral antigens can no longer be demonstrated.
events, but tissue damage may result from the bystander effects The chronic immunoinflammatory lesions are mainly orchestrated
+
of cytokines and other molecules, such as lipid mediators, by CD4 T cells, but multiple early events induce the subsequent

CHaPtEr 25 Host Defenses to Viruses 373


pathology. Viral replication, the production of certain cytokines biology approaches are expected to result in novel screening for
and chemokines (IL-1, IL-6, IL-12, and CXCL8), recruitment of immune protection parameters after vaccination. In the near
inflammatory cells (e.g., neutrophils), and neovascularization future, this should also assist in the formulation of new vaccines
of the avascular cornea all precede immunopathology. Recently, containing key immune activators, such as those that stimulate
it has become evident that Th17 T cells participate in stromal certain subsets of T cells or induce appropriate homing molecule
keratitis lesions. The role of Th17 T cells as orchestrators of expression on these cells to direct them to tissues where they
inflammatory reactions has been a major research focus, especially are required to mediate protection (e.g., mucosal sites, or skin).
36
in lesions of autoimmune diseases. When Th17 T cells are the
principal mediators of tissue damage, abundant neutrophils are ON tHE HOrIZON
recruited to inflammatory sites, with such cells being mainly Pressing Issues in Need of Solutions
responsible for tissue damage.
A further mechanism of viral-induced immunopathology Design of new vaccines that induce broadly neutralizing antibodies
33
and autoimmunity is molecular mimicry. Molecular mimicry Design of new vaccines that induce tissue-resident and circulating memory
represents shared antigenic epitopes, either B- or T-cell antigen, T-cell subsets
between the host and virus. This concept originated with Overcoming immune dysfunction during chronic viral infections for
successful viral clearance
streptococci and their association with rheumatic fever. With Improving the efficacy of vaccines to viruses using systems biology
human autoimmune disease, there is little direct support for approaches
viral molecular mimicry; however, some animal models have Therapies for reducing immunopathology during viral infections
been used to prove the theoretical case, where a viral antigen is
expressed as a self-protein in the islet cells of the pancreas. In In some individuals, viral infections cause mild, or sometimes
this model, subsequent infection with the virus induces diabetes. debilitating, tissue damage. Factors that influence whether a viral
However, this is not true mimicry and may be more closely infection results in immunopathology varies from individual to
related to viral antigen persistence in a model such as Theiler individual. These factors include age, the route of infection,
disease. preexisting immunity, host genetics, and the host’s viral burden
As discussed previously, the outcome of a T-cell response to or virome. Our knowledge of the influence of these factors on
a virus may be critically dependent on the balance of the T-cell- the outcome of viral infection is expected to improve rapidly in
type response thus, tissue damage is likely to be more severe the coming decades. Recent advances have shed considerable
+
+
and prolonged if CD8 or Th1 and Th17 CD4 T cells are light on the various proinflammatory and antiinflammatory
predominant. Lesions become milder and may resolve when the mediators produced during viral infections. These represent key
balance favors Tregs. Accordingly, therapeutic approaches that targets for novel therapies in the near future via the use of
can shift the balance of T cells are under trial. small-molecule inhibitors.
KEY CONCEPtS CONCLUSIONS
Phases of Immunity Affected by Regulatory
T Cells (Tregs) Humans are infected by many pathogenic viruses. In most cases,
these infections are controlled by the immune system with limited
Interference with antigen presentation by dendritic cells damage to the host. However, certain viruses, particularly in
Inhibition of T-cell proliferation cases where the host’s immune system is impaired, can cause
Inhibition of molecules involved in tissue-specific migration of effector significant damage to the host’s tissues. As our understanding
cells of the mechanisms underlying innate immune defenses, antigen
Inhibition of T-cell effector functions in lymphoid and nonlymphoid tissues
presentation, T- and B-cell responses, and Tregs continues to
improve, so too does the ability to design better vaccines and
TRANSLATIONAL RESEARCH OPPORTUNITIES therapies to boost the immune control of viral infections.
Although this remains a challenging goal, particularly for many
Reversing T-cell exhaustion in patients suffering from chronic human viruses, such as HIV, HCV, and HSV, these rapid advances
infections or cancer will be a key clinical target in the near future. continue to provide many avenues for further investigation.
The discovery of multiple inhibitory receptors on exhausted T
cells (e.g., PD-1, LAG-3, 2B4, TIM-3) has provided the opportunity ACKNOWLEDGMENTS
to selectively improve T-cell function through blockade of these
inhibitory receptors. This may be combined with blockade of Barry T. Rouse is supported by grants from the National Institutes
immunosuppressive cytokines (e.g., IL-10) or enhancement of of Health and Scott N. Mueller by the Australian Research Council
signals stimulatory to the response (e.g., IL-7 therapy), as well and the Australian National Health and Medical Research Council.
as with more traditional antiviral therapies and vaccination. The
challenge that lies ahead will be in determining which combination Please check your eBook at https://expertconsult.inkling.com/
of inhibitory and stimulatory signals will need to be manipulated for self-assessment questions. See inside cover for registration
in different diseases and in different groups of patients. details.
The design of a new generation of vaccines to target diseases,
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52–64. 36. Korn T, Bettelli E, Oukka M, et al. IL-17 and Th17 Cells. Annu Rev
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19. Ahmed R, Akondy RS. Insights into human CD8 T-cell memory using Immunol 2009;27:485–517.
the yellow fever and smallpox vaccines. Immunol Cell Biol 2011;89: 37. Pulendran B. Systems vaccinology: probing humanity’s diverse immune
340–5. systems with vaccines. Proc Natl Acad Sci USA 2014;111:12300–6.

CHaPtEr 25 Host Defenses to Viruses 374.e1


MUL t IPLE-CHOICE QUES t IONS

1. Many of the most medically important human viruses are 3. Immunological memory is a cardinal feature of adaptive
associated with persistent viremia, including human immu- immunity to virus infection. Memory T cells can be divided
nodeficiency virus (HIV) and hepatitis C virus (HCV). What into multiple subsets. Which of the following statements
are the effects of high levels of persisting antigen on the accurately describe the major subsets of memory T cells?
immune response? A. Effector memory T cells migrate through lymphoid tissues,
A. Increased killing capacity by cytotoxic T cells central memory T cells migrate through nonlymphoid
B. Altered tissue distribution and impaired functions of T tissues, and tissue-resident memory T cells circulate in
cells blood.
C. Upregulation of T-cell proliferation B. Effector memory T cells migrate through nonlymphoid
D. Enhanced cytokine production by virus-specific T cells tissues, central memory T cells persist in nonlymphoid
E. Reduced expression of coinhibitory molecules by the tissues, and tissue-resident memory T cells persist pre-
responding virus-specific T cells dominantly in lymphoid tissues.
C. Effector memory T cells persist in nonlymphoid tissues
2. Many viral infections can cause immunopathological reactions,
such as herpetic stromal keratitis induced by herpes simplex and do not enter blood, central memory T cells persist in
virus. What is a major parameter in the pathogenesis of this lymphoid tissues, and tissue-resident memory T cells
disease? migrate predominantly through nonlymphoid tissues.
A. Inhibition of inflammatory cell recruitment into the infected D. Effector memory T cells migrate exclusively in blood, central
tissues memory T cells migrate through lymphoid tissues, and
B. Recruitment of cytotoxic CD4 T cells that cause ocular tissue-resident memory T cells are retained in nonlymphoid
pathology tissues.
C. Recruitment and retention of pathogenic CD8 T cells in E. Effector memory T cells migrate through nonlymphoid
the late stage of the response tissues, central memory T cells migrate through lymphoid
D. Recruitment and retention of pathogenic Th1 CD4 T cells tissues, and tissue-resident memory T cells are retained
into the ocular tissues in tissues.
E. Reduction in Th17 CD4 T cells in the ocular tissues

26









Host Defenses to Intracellular Bacteria



Stephen T. Reece, Stefan H.E. Kaufmann







The evolutionary relationship between humans and bacteria is CLINICAL PEARLS
so intimate that it is impossible to imagine the development of
1
one without the other. Although this coexistence is generally Distinguishing Clinical Characteristics of
mutually beneficial, clear boundaries do exist between the two Infections With Intracellular Bacteria
and are intensely defended. We tend to think of the human host Nonsterilizing immunity
as the defender and bacteria as transgressors of these boundaries. Persistent bacteria, sometimes latent infection
Evolution of human immunity has been accompanied by evolu- Formation of long-lasting tissue granulomas containing low numbers of
tion of ingenious bacterial mechanisms to not only survive its viable bacteria
onslaught but also to manipulate it to enhance survival. This idea Critical role of T cells in protection, role of antibodies less well established
is instructively mirrored in the lifestyle of intracellular bacteria. but likely to play an as-yet unappreciated role
These bacteria actively seek out an environment inside human Critical role of immune response in pathology
Lack of effective vaccines
cells where they can flourish; this is not an easy environment Host-directed therapies toward enhancing antimicrobial mechanisms
in which to survive. Human cells have developed an ability to while limiting host pathology
differentiate bacterial from host components and direct host cells
to clear the invader. The most successful intracellular pathogens
have adapted to the intracellular environment of a particular without causing clinical signs of illness, but bacterial growth
cell target, proliferate only slowly, and can live for long periods can be reactivated to cause disease if the immune response
completely undetected by the immune system, as we see in the case becomes compromised. This occurs in M. tuberculosis infection,
of the tuberculosis (TB) bacterium Mycobacterium tuberculosis. In resulting in disease years or decades after primary infection. In
other instances, for example, in listeriosis, intracellular infection is fact, disease need not arise from infection at all. In many regions,
more explosive, with the rich intracellular environment harnessed for example, the majority of adults harbor M. tuberculosis without
to rapidly amplify bacterial growth. In some cases, intracellular suffering from clinical disease. However, disease can develop
bacteria live for a very long time in the human body, sometimes directly after primary infection, during maturation of the immune
for a person’s entire lifetime. A wide spectrum of pathologies response, or with regression once the immune response is suf-
ensues from intracellular infection, making most intracellular ficiently strong. Yet, sterile eradication of the pathogen is rarely
bacteria highly clinically relevant. Moreover, new concepts on achieved: bacteria persist latently, and illness may reemerge at
the influence of intracellular bacteria on host cell differentiation a later time. For example, Rickettsia prowazekii may persist for
point to their ability to change infected cell phenotype to enhance decades after convalescence from typhus to cause Brill-Zinsser
survival. disease later.
This chapter evaluates the current interpretation of this Several intracellular bacteria possess components that can
fascinating interplay between human and microbe, sheds light profoundly influence the course of disease, for example, the
on how the human immune system functions, and how cellular lipopolysaccharides (LPSs) of brucellae and salmonellae. Chronic
phenotype can be molded in cells whose fates were previously persistence inside host cells, however, depends on the target cell
believed to be strictly predetermined. Finally, such insights can remaining intact and physiologically active. Accordingly, many
inform new therapeutic and prophylactic approaches to keep intracellular bacteria are of low toxicity and do not have dramatic
intracellular bacterial infections under control. direct effects on their host. Instead, pathogenesis is largely
determined by the immune response. Classic examples of this
concept include granuloma liquefaction in acute TB, which
BALANCE OF PROTECTION AND PATHOLOGY severely affects lung function, and eye scarring as a consequence
DEFINES THE CHRONIC NATURE OF of chronic or recurring Chlamydia trachomatis infection that
INTRACELLULAR BACTERIAL INFECTION ultimately leads to trachoma.
The survival of intracellular bacteria has major consequences
Some bacteria, such as Listeria monocytogenes, are fully eradicated for pathology. Although many intracellular bacteria show some
once the host immune response has reached its peak activity. organ tropism, dissemination to other organs frequently occurs,
Most often, the intracellular habitat provides a protective niche resulting in different disease forms. For example, TB is generally
that promotes persistent infection in the face of an ongoing manifested in the lung in 80% of cases, yet many other organs
immune response. Here, the bacteria can persist for long periods can be affected. In contrast to other Salmonella enterica serovars,

375

376 PARt tHREE Host Defenses to Infectious Agents



TABLE 26.1 Major Infectious Diseases Caused by Intracellular Bacteria
Disease Pathogen Prevalence Incubation time Route of Infection target Cell
Granulomatous Intracellular Bacteria
Tuberculosis Mycobacterium tuberculosis Worldwide Years (latency after Inhalation of Macrophage
primary infection and bacteria-containing
disease reactivation) microdroplets
Weeks (miliary TB)
Leprosy Mycobacterium leprae South America Years Smear infection Macrophage
Africa through mucosa/ Schwann cell
India inhalation
Southeast Asia
Typhoid fever Salmonella enterica serovars Worldwide 7–10 days Fecal–oral Macrophage
Typhi and Paratyphi
Brucellosis Brucella spp. Worldwide Weeks to months Zoonosis; cows, Macrophage
goats, pigs;
inhalation, gut, skin
abrasion
Listeriosis Listeria monocytogenes Worldwide Days to months Fecal–oral Macrophage
Hepatocyte
Nongranulomatous Intracellular Bacteria
Legionnaires’ disease Legionella pneumophila Worldwide 2–10 days Inhalation Macrophage
Rocky Mountain Rickettsia rickettsiae Western 1 week Tick bite Vascular endothelial cell
spotted fever hemisphere Smooth muscle cell
Urogenital infection Chlamydia trachomatis Worldwide 1–3 weeks Sexual intercourse Epithelial cell
serovars D-K
Conjunctivitis, Chlamydia trachomatis Africa Conjunctivitis: 1–3 weeks Eye Epithelial cell
trachoma serovars A-C Trachoma: years
Cat scratch disease Bartonella henselae Worldwide Bacillary angiomatosis Flea, sandfly, Erythrocyte
B. quintana Peliosis hepatitis or mosquito bite; Endothelial cell
B. bacilliformis Endocarditis animal scratch or
Bacteremia with fever bite
Neuroretinitis: 1–3 weeks




the serovars Typhi and Paratyphi are not restricted to the carrier state or the so-called latent TB infection (LTBI). Infection
gastrointestinal (GI) tract but are disseminated to internal organs, with M. tuberculosis starts with the so-called Ghon complex,
primarily the liver and spleen. In these cases, the type of clinical characterized by a caseous lesion in the midlung as well as in
disease depends markedly on the infected tissue type. the draining lymph nodes. These primary lesions can progress,
2
but their development rarely causes disease directly. Moreover,
KEY CONCEPtS bacteria from these sites can disseminate to other regions of
Characteristic Features of Intracellular the lung and systemically, causing disease of the kidneys, liver,
Bacterial Infections and central nervous system (CNS). Containment of the primary
lesions, which leads to LTBI, is a function of an effective, pre-
Persistence of bacteria inside mononuclear phagocytes (i.e., dominately cellular antitubercular immune response. Infection
macrophages) of immunocompromised patients, notably those with acquired
Low to absent bacterial-mediated toxicity to the host immunodeficiency syndrome (AIDS), or newborns frequently
Protection requires cytokine-mediated activation of infected results in systemic disease (miliary TB). TB represents a major
phagocytes health problem worldwide, including an increasing incidence
Interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) produced by
antigen-specific T cells are key cytokines for protection in many industrialized countries. In 2015, the World Health
Organization (WHO) estimated that 10.4 million active TB
cases were diagnosed worldwide and close to 1.8 million people
3
died of the disease. The much larger estimated number of 2
INTRACELLULAR BACTERIAL INFECTIONS OF billion individuals infected with M. tuberculosis well illustrates
CLINICAL RELEVANCE (Table 26.1) the dissociation of infection from disease. The emergence of
multidrug-resistant strains and extremely drug-resistant strains
Granulomatous Infections has complicated treatment with currently available antibiotic
Tuberculosis therapy, and even when treatment is successful, recurrence
The major entry of tubercle bacilli into the human body is via of disease can occur. The currently available live whole cell
inhalation into the lung. These inhaled bacteria are then engulfed vaccine BCG (Bacille Calmette–Guérin), an attenuated strain
by alveolar macrophages, which transport the pathogens to the derived from the etiological agent of bovine TB Mycobacte-
lung interstitia. Their exact fate after these events is enigmatic. rium bovis, shows only low and variable protection against
Moreover, most infections in humans result in an asymptomatic pulmonary TB.

CHAPtER 26 Host Defenses to Intracellular Bacteria 377


Leprosy Gastroenteritis
Mycobacterium leprae is most likely transmitted by contact S. enterica serovars Typhimurium and Enteritidis, often referred
with patients who shed microorganisms in nasal secretions and to as nontyphoidal salmonella (NTS), are the major causes of
lesion exudates. It primarily affects the nerves and the skin, salmonella gastroenteritis in humans, which occurs mainly as a
frequently leading to stigmatizing deformities. In skin, bacilli result of the ingestion of contaminated food or water. The bacteria
target keratinocytes, histiocytes, and macrophages, whereas rapidly cross the intestinal epithelia and replicate in the lamina
in peripheral nerves, Schwann cells are the major target for propria, inducing an influx of polymorphonuclear neutrophils
entry. Leprosy is a spectral disease. The tuberculoid pole is (PMNs), which is generally sufficient to resolve the infection
characterized by rigorous T-cell responses, which succeed in within 7 days. In rare cases, bacteria enter the bloodstream and
restricting microbial growth in well-defined lesions contain- cause systemic bacteremia, most notably in patients with AIDS,
ing few bacilli. In contrast, at the lepromatous pole, bacterial where death can occur as a result of septic shock. 8
growth is unrestricted and lesions contain abundant bacilli
within macrophages lacking signs of activation. Several types Listeriosis
of immunosuppression have been implicated in this latter type Listeriosis monocytogenes is increasingly recognized to cause
of disease. Infection of Schwann cells promotes nerve damage foodborne gastroenteritis. Clinical listeriosis affects mainly
and anesthesia. This results in injuries and secondary infections pregnant women, older adults, fetuses, and neonates. Disease
that significantly exaggerate the disease. Despite the success of manifestations are most severe in patients with a compromised
multidrug therapy in reducing the number of registered leprosy immune system, in whom the CNS becomes involved and fatal
cases worldwide, some 216 000 new cases were reported in bacteremia can result. Additionally, as these bacteria are able to
2013. This suggests that active transmission of M. leprae is still cross the placenta, listeriosis is a major cause of perinatal and
occurring and that more effective interventions are required to neonatal disease, typically resulting in abortion. Listeria outbreaks
prevent it. 4 are sporadic with low incidence but high fatality and affect
high-income countries, such as the United States.
Atypical Mycobacterial Infections
Mycobacterial species present in the environment are typi- Brucellosis
cally unable to persist within activated macrophages and thus Brucellosis is the most common global zoonosis of humans with
9
rarely cause disease in individuals with competent immune approximately 500 000 cases per year. It is caused by Brucella
status. 5 abortus, B. melitensis, or B. suis, which primarily infect cows,
As a consequence of human immunodeficiency virus (HIV) goats, and pigs, respectively. The bacteria are transmitted to
infection, however, nontuberculous mycobacteria (NTM), primar- humans via inhalation, through abraded skin or the GI tract.
ily Mycobacterium avium/Mycobacterium intracellulare, have Lesions are primarily found within macrophage-rich tissues,
gained clinical importance, and these infections are recognized especially the spleen and bone marrow. Human brucellosis is
as one of the most common complications of AIDS in industrial- characterized by systemic symptoms, particularly undulant fever.
ized nations. Although the disease often remains subclinical, it becomes chronic
Mycobacterium scrofulaceum occasionally causes lymphadenitis in some patients, and relapses and remissions may occur. Interest
in children, and Mycobacterium kansasii primarily causes infec- in brucellosis has increased in the last 5 years because of elevated
tions in older men with preexisting lung disease. levels of detection resulting from better surveillance.
Mycobacterium ulcerans causes a severe subcutaneous infection
characterized by chronic skin ulcerations, known as Buruli ulcer. Lymphogranuloma Venereum
This pathology is caused—at least in part—by elaboration of a Lymphogranuloma venereum (LGV), a sexually transmitted
mycolactone toxin by the bacillus that exhibits highly cytopathic disease, is highly prevalent in Africa, Southeast Asia, and Latin
6
effects. Buruli ulcer is most predominant in West African America. LGV has recently emerged as infection of sexually active
countries that accounted for most of the 2251 cases reported homosexual men in Europe and the United States. It is caused
globally in 2014. 7 by the L1, L2, and L3 serotypes of Chlamydia trachomatis, which
are disseminated from the urogenital tract to local lymph nodes
Typhoid or Enteric Fever and then to skin. Accordingly, LGV is characterized by lymph
Salmonella enterica serovars Typhi and Paratyphi A, Paratyphi node swelling and skin lesions, which are accompanied by systemic
B, and Paratyphi C are leading causes of community-acquired complications. 10
bloodstream infections in low- and middle-income countries.
The route of transmission is fecal–oral and largely occurs via Melioidosis
contaminated water sources. Bacteria are disseminated within Burkholderia pseudomallei is a gram-negative bacillus and the
mononuclear phagocytes (MPs) from the GI tract to macrophage- causative agent of melioidosis, endemic in Southeast Asia and
rich organs, particularly the liver, spleen, and lymph nodes. Northern Australia. The disease can be acquired through inhala-
Accordingly, typhoid is characterized by systemic symptoms, tion and ingestion or through cuts in the skin. Susceptible hosts
such as prolonged fever and malaise, with sustained bacteremia, can suffer abscess formation in multiple organs and, in some cases,
although diarrhea or constipation may also be present. In some disseminated infection, resulting in septic shock accompanied by
cases, an asymptomatic carrier state can persist as a result of pneumonia. There are an estimated 165 000 cases of melioidosis
chronic infection of the gallbladder, which maintains the envi- per year globally, resulting in approximately 89 000 deaths. 11
ronmental reservoir of infection in endemic areas. Typhoid fever
remains a major cause of morbidity and mortality, with approxi- Tularemia
mately 21 million new cases and over 190 000 deaths per year This rare zoonosis in humans caused by Francisella tularensis is
worldwide. 8 mainly found in rabbits and has recently gained wider recognition

378 PARt tHREE Host Defenses to Infectious Agents


12
because of its potential for dual use. Infection can be spread Bartonella
to humans via contaminated animals or tick bites. This gram- Bartonella spp. represent gram-negative facultative intracellular
negative bacterium survives in macrophages and primarily causes pathogens transmitted by insect vectors, such as fleas, sandflies,
18
acute pneumonia as well as skin sores, with subsequent involve- and mosquitoes. The most clinically relevant species are B.
ment of lymph nodes. henselae, B. quintana, and B. bacilliformis. B. henselae causes cat
scratch disease (CSD) resulting in local lymphadenopathy in the
Nongranulomatous Infections lymph node draining the scratch site accompanied by fever,
Legionnaires’ Disease or Legionellosis headache, and splenomegaly. Oculoglandular involvement
Legionnaires’ disease is caused by Legionella pneumophila, an (Parinaud syndrome), encephalopathy, neuroretinitis, or osteo-
environmental bacterium that persists within amoeba living in myelitis can occur, albeit in rare cases. In immunosuppressed
water reservoirs (e.g., air-cooling systems), from where it is spread patients, bacillary angiomatosis and peliosis can occur, character-
aerogenically. Infection is exacerbated by a compromised immune ized by pseudotumoral proliferation of endothelial cells. Bacteria
status. Characteristically, Legionnaires’ disease presents as atypical persist within erythrocytes with the intracellular location provid-
pneumonia associated with general symptoms and is complicated ing a protective niche.
by extrapulmonary infection, renal failure, and lung abscesses.
Cases of Legionnaires’ disease in the United States increased GRANULOMA PATHOLOGY AS HALLMARK OF
from 0.39 to 1.36 per 100 000 people from 2000 to 2011. 13
INTRACELLULAR BACTERIAL INFECTION
Chlamydial Urethritis, Cervicitis, and Conjunctivitis
C. trachomatis serovars D–K enter and persist in epithelial cells KEY CONCEPtS
of the urogenital tract, causing cervicitis and urethritis. In women, Balance of Protection and Host Pathology
infertility can develop as a result of chronic or recurrent infection. in Granulomas
In neonates, congenital infection during birth may result in
conjunctivitis and pneumonia. Urogenital infections by chla- Macrophage activation results in bacterial death (protective)
mydiae occur worldwide and are now considered the most Intracellular bacterial killing by “killer molecules” from T cells
(protective)
common sexually transmitted bacterial disease, with an estimated Lysis of infected macrophages by T cells results in release of bacteria
100 million new infections occurring annually. 14 and killing by more effective effector cells (protective) or bacterial
dissemination (pathogenic)
Trachoma Development of central necrosis in granulomas results in death of tissue
Smear infections of the eye with C. trachomatis serovars A, and bacteria (protective/pathogenic)
B, and C cause inclusion conjunctivitis. As a consequence of Fibrotic encapsulation of granuloma results in containment of infection
(protective)
multiple chronic infections and of the resulting immune response, Overexuberant tissue fibrosis and necrosis (pathogenic)
scars develop that eventually injure the cornea, leading to tra- Liquefaction of central necrotic tissue in granulomas results in bacterial
choma. Approximately 84 million people are infected with C. replication, cavity formation, and transmission of bacteria (pathogenic
trachomatis worldwide, 7.6 million of whom suffer from visual and contagious)
impairment. 15
A characteristic feature of many infections caused by intracellular
Chlamydia Pneumoniae bacteria is the eventual need for tissue remodeling by the host at
C. pneumoniae (formerly known as C. trachomatis TWAR strain) the site of infection. Granulomas are the result of an inability to
is the cause of mild respiratory disease in young adults and may rapidly clear host tissue of intracellular bacteria and represent a
cause serious infections in older, more debilitated patients. fascinating site of the host–pathogen interface (Fig. 26.1). The
Atypical pneumonia may also be caused by Chlamydia psittaci, longevity of the granuloma depends directly on the continuous
although this zoonosis, transmitted by birds, is relatively rare. presence of the microbial pathogen, and the lesion generally
disappears after its sterile eradication. Granulomas form the focus
Typhus of the coordinated cross-talk between different types of T cells,
Rickettsia prowazekii, R. typhi, and R. tsutsugamushi cause diseases B cells, and infected and uninfected mononuclear phagocytes
16
of varying severity. They are transmitted by arthropods and (MPs) and dendritic cells (DCs). Even if the immune system fails
infect vascular endothelial cells at the site of an insect bite or to completely eliminate bacteria inside the granuloma, the latter
scratch, causing skin reactions. Subsequently, pathogens are performs a protective function by containing microbes within
disseminated to the central organs, and more general symptoms distinct foci and preventing their dissemination. At the same
develop. Globally, typhus is of minor importance. time, the granuloma can be detrimental to the host because it
19
can interfere with physiological organ functions. More detailed
Rocky Mountain Spotted Fever, Ehrlichiosis study of cellular phenotype within granulomas is starting to
Rocky Mountain spotted fever is caused by Rickettsia rickettsii. establish how cellular differentiation is orchestrated and how
Infection of the vascular endothelium leads to systemic symptoms the granuloma develops.
and skin manifestations that may be followed by shock and Granulomatous lesions are generally initiated by nonspecific
16
neurological complications. Worldwide, this disease, as well as inflammatory signals mediated by bacterial products, chemokines,
Mediterranean spotted fever caused by Rickettsia conorii, is of and proinflammatory cytokines that are produced by endothelial
minor importance; as is probably Ehrlichiosis, a newly emerging cells and MPs at the site of infection. Inflammatory phagocytes
zoonosis transmitted by ticks and caused by various Ehrlichia (of both monocytic and granulocytic origin) are attracted to
17
spp., mainly E. chaffeensis. Disease manifestations include the site of microbial replication, and an infiltrative, sometimes
generalized symptoms, such as fever and muscle pain. exudative, lesion develops. Following the accumulation and

CHAPtER 26 Host Defenses to Intracellular Bacteria 379


FIG 26.1 Development of Granuloma Pathology and Implica-
tions for Tuberculosis (TB). This figure depicts three distinct
yet continuous stages of granuloma pathology in the lung due
to Mycobacterium tuberculosis infection. (A) Solid granuloma:
Composed largely of T cells and infected and uninfected MPs.
These granulomas are defined by a lack of central necrosis and
likely are representative of an ability to control M. tuberculosis
replication. (B) Caseous/necrotic granulomas: These structures
contain a central region of demarcated necrotic cell death. Bacteria
are often detected within the caseous necrotic region and in
proximal cells, notably mononuclear phagocytes (MPs). Since
calcified caseous granulomas containing few bacteria have been
observed, development of central necrosis may be a consequence
of antibacterial mechanisms resulting in sacrifice of host cells
to contain infection. (C) Cavity formation: These structures result
from inability of caseous granulomas to contain bacterial replica-
A Solid granuloma tion. The acellular necrotic region, containing a large number of
extracellular bacteria, increases in size and can liquefy and empty
into the lung airways, resulting in transmission of viable bacteria
via cough. Therefore granuloma formation is central to human-
to-human spread of TB. Dissemination of bacteria through the
bloodstream results in disease manifestation in other organs,
such as the meninges and the urinary bladder.



activation of increasing numbers of MPs and DCs, this lesion
takes an increasingly structured granulomatous form. A significant
number of B cells is also found, which seem to influence granu-
loma morphology. Once specific T cells have been attracted to
the lesion, it transforms into a productive granuloma that provides
the most appropriate tissue site for antibacterial protection. Here,
activation of MPs by interferon-γ (IFN-γ) and tumor necrosis
factor-α (TNF-α) inhibits microbial growth. However, unbridled
B Necrotic granuloma macrophage activation can have tissue-damaging effects, and
mechanisms within the granuloma tightly regulate these effects.
Eventually, the granuloma is encapsulated by a fibrotic wall, and
its center becomes necrotic. Both tissue reactions are primarily
protective, the former by promoting bacterial containment and
the latter by reducing the nutrient and oxygen supply to the
pathogen. The combined effects of chronic macrophage activation,
persistence of intracellular bacteria, and hypoxia likely lead to
enhanced cell death in the center of granulomas, resulting in
the formation of a caseum. Caseation may favor the local replica-
tion of normally facultative intracellular bacteria in the cellular
detritus, as well as microbial dissemination to distant tissue sites
and to the environment to transmit infection. Hypoxia also has
pronounced effects on enzyme functions that can dictate mac-
rophage phenotype.


THE INTERDEPENDENCE OF INNATE AND
C Caseous granuloma ADAPTIVE IMMUNITY IN PROTECTION AGAINST
INTRACELLULAR BACTERIA
Innate Immune Mechanisms as First-Line Defense
The interaction between host cell and pathogen that defines the
intracellular lifestyle consists of a number of different layers.
The first layer that differentiates intracellular bacteria from other
bacteria, notably commensal bacteria that colonize the host but
do not cause infection, is that of host cell entry. Extracellular
bacteria are typically engulfed by professional phagocytes, which

380 PARt tHREE Host Defenses to Infectious Agents


include tissue macrophages, DCs, and PMNs. This uptake is TABLE 26.2 Major Pattern Recognition
enhanced by host components of the complement system and Receptors Involved in Sensing of
antibodies, which bind to complement receptors (CRs) and Fc Intracellular Bacteria
20
receptors, respectively, on professional phagocytes. M. tuberculosis
actively targets macrophages, where it must counteract numerous Pattern Recognition
antimicrobial mechanisms operative in these cells (see below). Receptor Location Ligand
Intracellular bacteria also use elaborate mechanisms to enter toll-Like Receptors
nonprofessional phagocytes, by which they must subvert host TLR1 Plasma membrane Triacyl lipoprotein
endocytic processes that are normally engaged in traffic of cellular TLR2 Plasma membrane PGA, porins, LAM
cargoes. In some cases, this provides a less hostile environment TLR4 Plasma membrane LPS
because of their inability to efficiently mobilize antibacterial TLR5 Plasma membrane Flagellin
effector mechanisms. Bartonella spp., unique among intracellular TLR6 Plasma membrane Diacyl lipoprotein
bacteria, can enter red blood cells, thus allowing transmission TLR7 (human TLR8) Endosome ssRNA
CpG DNA
Endosome
TLR9
via blood-sucking insect vectors. This represents a particularly
advantageous niche as red blood cells lack the machinery to Scavenger Receptors
drive the adaptive immune responses required for protection. SR-A Plasma membrane LPS, LTA, CpG DNA,
Entry into nonphagocytic host cells requires bacteria to induce proteins
their own internalization. Bacteria that colonize the GI tract MARCO Plasma membrane LPS, proteins
(i.e., L. monocytogenes or salmonellae) or mucosal membranes CD36 Plasma membrane Diacyl lipoprotein
of the urogenital tract (i.e., C. trachomatis) must mediate tight LOX-1 Plasma membrane Protein
adhesion to the host cell membrane and be capable of mediating SREC Plasma membrane Protein
the uptake process. Broadly, two processes are utilized by bacteria C-type Lectins
to induce uptake into a nonphagocytic cell. The “zipper” mecha- DC-SIGN Plasma membrane LPS, ManLAM,
nism is mediated by binding of a bacterial cell surface protein capsular
binding a cognate receptor on the host cell membrane. L. polysaccharide
monocytogenes entry into intestinal epithelial cells depends on MINCLE Plasma membrane Mycobacterial cord
engagement of InIA to E-cadherin to mediate uptake. factor: TDM
Salmonellae and C. trachomatis use a “trigger” mechanism
to induce internalization and inject multiple factors into the NOD-Like Receptors
host cell cytoplasm to mediate uptake. These proteins are delivered NOD1 Cytoplasm D-glutamyl-meso-
diaminopimelic acid
by the needle-like structures that form part of bacterial type III NOD2 Cytoplasm MDP
secretion systems (T3SSs). These injected proteins then target NLRP1 Cytoplasm MDP
host proteins involved in host cell signaling and actin remodeling NLRP3 Cytoplasm RNA, LPS, LTA, MDP
to induce bacterial entry. The C. trachomatis–secreted proteins NLRC4 Cytoplasm Flagellin
Tarp, CT166, and CT694 reversibly stimulate the Rho-family Naip5 Cytoplasm Flagellin
guanosine triphosphatase (GTPase) Rac1 to trigger internalization.
Similarly, salmonellae inject T3SS factors to stimulate the Rho- AIM2-Like Receptors
family GTPases Cdc42 and Rac1. The success of these mechanisms AIM2 Cytoplasm dsDNA
of induced uptake enables intracellular bacteria to persist inside IFI16 Cytoplasm dsDNA
diverse cell types. Rickettsia spp., C. trachomatis, M. leprae, and StING/cGAS Pathway
L. monocytogenes ultimately target vascular endothelial cells, cGAS Cytoplasm dsDNA
epithelial cells, Schwann cells, and hepatocytes, respectively, as
their preferred intracellular habitats. Note: We omit PRRs (e.g., TLR3, which binds viral produced double-stranded RNA)
To prevent intracellular infection, the host depends on its not classically associated with intracellular bacteria. AIM2, absent in melanoma-2;
ability to discriminate between host and bacterial molecules. CD36, cluster of differentiation 36; CpGDNA, cytosine-phosphatidyl-guanine DNA;
DC-SIGN, dendritic cell–specific intercellular adhesion molecule-3-grabbing
As already mentioned, bacteria targeting the intracellular envi- nonintegrin; dsDNA, double-stranded DNA; LAM, lipoarabinomannan;
ronment often do so via mucosal surfaces already populated LOX-1, lipoxygenase-1; LPS, lipopolysaccharide; LTA, lipoteichoic acid;
ManLAM, mannose lipoarabinomannan; MARCO, macrophage receptor with
by commensal organisms (the microbiome) that do not alert collagenous structure; MDP, muramyl dipeptide; MINCLE, macrophage-inducible
host defenses. The host must, therefore, discriminate between C-type lectin; NLR, NOD-like receptor; NOD, nucleotide-binding domain;
commensal and pathogenic bacteria via recognition of conserved PGA, peptidoglycans; PRR, pattern recognition receptor; SR, scavenger receptor;
ssRNA, single-stranded RNA; SREC, scavenger receptor expressed by endothelial
molecular motifs of bacteria, named pathogen-associated molecular cell-I; TDM, trehalose dimycolate; TLR, Toll-like receptor.
patterns (PAMPs). This occurs via host receptors broadly defined
as pattern recognition receptors (PRRs, Table 26.2). The best-
characterized group of PRRs is that of the so-called Toll-like lipids, such as lipoarabinomannan (LAM), trehalose dimycolate
receptors (TLRs). The TLR system constitutes an innate scan- (TDM), and phosphatidylinositol mannosides (PIMs) bind either
ning mechanism of microbial pattern recognition to distinguish TLR-2 or TLR-4. Lipoteichoic acid (LTA) of gram-positive bacteria
between a wide spectrum of bacteria and viruses. TLRs are present is recognized by TLR-2. TLR-9 binds low-methylated bacterial
as homo- or heterodimers on the plasma membrane or within DNA containing CpG motifs within endosomes.
21
intracellular endosome/phagosome compartments. PAMPs of Scavenger receptors and C-type lectins are also PRRs and
22
bacterial origin comprise di- and triacyl lipoproteins, LPSs, and function at the cell membrane. Scavenger receptors were first
flagellin, which are recognized by TLR-2/6, TLR-2/1, TLR-4/4, or defined by their ability to transport modified forms of low-
TLR-5/5, respectively. The vast array of mycobacterial cell wall density lipoproteins inside cells, indicating their ability to also

CHAPtER 26 Host Defenses to Intracellular Bacteria 381


interact with host molecules. However, receptors, such as SR-A, that induce innate immune mechanisms and, subsequently, the
MARCO, CD36, LOX-1, and SREC, can bind a wide array of mobilization of the adaptive immune response. These include
bacterial molecules, such as lipids, CpG DNA, and proteins cytokines that act both locally and systemically and are important
(see Table 26.2 for binding specificities). SR-A is important for mediators of protection against intracellular bacteria via specific
clearance of extracellular bacteria from the spleen and liver. signaling through engagement of host cell surface receptors. Such
MARCO expressed on alveolar macrophages is implicated in engagement mobilizes both critical mechanisms of host protection
clearance of pneumococcal bacteria preventing pneumonia. and orchestration of adaptive immune responses.
C-type lectins are similarly membrane-expressed and include
DC-specific intercellular adhesion molecule–grabbing nonintegrin Macrophage Training by Epigenetic Mechanisms
(DC-SIGN); mannose receptor; dectin-1; dectin-2, which chiefly Following infection with, for example, M. tuberculosis, macro-
recognize fungal components; and MINCLE, which recognizes phages express elevated effector mechanisms over long periods.
trehalose dimycolate (TDM), the cord factor of M. tuberculosis. More recent findings have revealed that this is caused, at least
It has been suggested that scavenger receptors and C-type in part, by epigenetic changes that are induced in MPs during
lectins are required to bind and internalize the bacillus, whereas infection. Similar training can also occur after vaccination with
it is primarily the TLRs that discriminate between the pathogens live vaccines. Moreover, BCG vaccination was shown to induce
and initiate the necessary intracellular signaling events. It should, changes in methylation patterns of the NOD2 gene in humans.
however, be noted that intracellular signaling events can also be These epigenetic changes led to increased production of proin-
triggered by other interactions, such as ligand binding to mac- flammatory cytokines by MPs, namely, TNF and IL-1β, a tendency
rophage mannose receptor (MMR), dectin 1, or DC-SIGN. Far that persisted for 3 months after vaccination. Furthermore, this
from a one-ligand, one-receptor binary mechanism of sensing enhanced innate immunity led to increased resistance not only
and signaling, PRRs often collaborate to produce multiprotein against M. tuberculosis but also against other bacterial pathogens.
complexes. CD14, MD2, and TLR4 collaborate for LPS sensing Indeed, it has been speculated that epigenetic alterations in MPs
and signaling. Similarly, MARCO and TLR2 synergize to recognize following vaccination are responsible for the nonspecific reduction
TDM. To allow signaling, these complexes interact with adaptor of mortality in BCG-vaccinated infants in resource-poor regions
proteins containing immunoreceptor tyrosine-based activation of the world. 23
motif (ITAM)-like or Toll/interleukin-1 receptor (TIR) domain
motifs. TLR signaling occurs via the adaptor proteins MyD88,
TIRAP/Mal, and Trif. These molecules then orchestrate a down- Cytokines as Mediators of Defense Against
stream signaling cascade, which culminates in induced patterns Intracellular Bacteria
of gene transcription that mediate innate and, ultimately, adaptive We have already mentioned that a range of cytokines are induced
immune mechanisms that aim at combating the intracellular by the signaling mechanisms that result from engagement of
bacteria. PRRs. These serve by both enhancing intracellular mechanisms
The cellular cytoplasm is monitored for presence of molecules of bacterial killing and mobilizing adaptive immune responses,
of bacterial origin by a further group of PRRs, the nucleotide representing the next layer of host defense. Because these responses
oligomerization domain protein-like receptors (NLRs). These allow an amplification of the initial innate immune responses,
molecules are characterized by presence of a nucleotide-binding they must be carefully regulated by the host to prevent extensive
domain and leucine-rich repeat motifs. Molecules from this tissue pathology. In fact, we might view the development of a
group recognizing bacterial components are nucleotide-binding granuloma as the sequela of a balance between bacterial killing
oligomerization domain (NOD)–containing proteins NOD1 and mechanisms and the need to restrict tissue pathology orchestrated
NOD2, NOD-like receptor P1 (NLRP1), NLRP3, and Naip5. Other by adaptive immunity. At the onset of infection, initial cytokine
cytosolic PRRs include the absent in melanoma-2-like receptor secretion occurs in the cell type that initially encounters intracel-
(ALR) family, cyclic guanosine monophosphate–adenosine lular bacteria and on initiation of signaling cascades by PRRs.
monophosphate (cGMP-AMP) synthase (cGAS), and stimulator These small molecules can act locally and systemically to directly
of interferon (IFN) genes (STING), all of which can be activated instruct cells, to produce antibacterial molecules, to combat
by bacterial DNA (see Table 26.2). intracellular infection, and to both increase numbers of immune
Engagement of NLRs and ALRs leads to activation of the cells and direct the composition of the cellular infiltrate that
multiprotein complex called the inflammasome, leading to cleavage will ultimately attempt to resolve intracellular bacterial infection.
of pro-IL-1β and pro-IL-18 to produce their active forms. In Cytokines are ultimately produced by multiple cell types, including
addition, activation of the NLRs, NOD1, and NOD2 results in adaptive T cells, B cells, unconventional T cells, MPs, DCs, PMNs,
inflammatory cytokine secretion. Certain PRRs are also receptive and even epithelial and endothelial cells. We will first consider
to certain endogenous “danger” signals produced by tissues the hierarchy by which these cytokines act in the control of
undergoing stress, damage, or cell death. These signals are trig- intracellular bacterial infection and the antibacterial mechanisms
gered by self-proteins, named danger-associated molecular patterns they regulate. We will then return to the generation and regulation
(DAMPs), include endogenous heat shock proteins, host of the cells that produce them.
nucleotides, and the chromatin component HMGB1. Therefore
PRRs mediate signals not only emanating from intracellular IFN-γ, TNF-α, IL-12, and IL-18
bacteria but also from host cells damaged by the infection process. By far, the cytokine with the clearest demonstrable potency
Understanding how PAMP and DAMP PRR signaling meshes against intracellular bacteria is IFN-γ. Extensive studies on the
to produce a coherent disease-specific output remains an exciting activation of antibacterial effector functions in macrophages have
challenge for future research. revealed a central role for IFN-γ. Accordingly, IFN-γ neutralization
As already alluded to, the culmination of PRR collaborative with antibodies, or deletion of the IFN-γ gene by homologous
sensing and signaling is the induction of inflammatory genes recombination, markedly exacerbates infectious diseases, such as