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952 Part VII: Neutrophils, Eosinophils, Basophils, and Mast Cells Chapter 62: Eosinophils and Related Disorders 953

ECP is an arginine-rich protein. The complementary DNA (cDNA) effective in generating reactive oxygen species with about 1.7-fold
encodes for a 27-amino-acid leader sequence and a 133-amino-acid greater generation than neutrophils after stimulation with PMA (phor-
mature polypeptide with a molecular mass of 15.6 kDa. ECP has bol myristate acetate), possibly as a result of a 10-fold greater expression
66 percent amino-acid-sequence homology with EDN and 31 percent of the voltage-gated channel Hv1. 119
homology with human pancreatic ribonuclease, but it has low ribonu- The killing of schistosomula opsonized with nonimmune serum
clease activity compared to EDN. ECP is toxic for helminthic parasites, is presumed to be mediated via the complement receptors, CR1 and
isolated myocardial cells, and guinea pig tracheal epithelium. ECP also CR3. Incubation of eosinophils with serum-coated beads results in
inhibits lymphocyte proliferation in vitro. Both ECP and EDN produce the release of 15 percent of ECP. Similarly, opsonized zymosan inter-
neurotoxicity (the Gordon phenomenon) when injected into the cere- acts with eosinophils, causing generation of hydrogen peroxide and the
brospinal fluid of experimental animals. ECP may damage cells by a col- phagocytosis of the zymosan. Soluble mediators, such as PAF, LTB , and
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loid osmotic process, as it can induce non–ion-selective pores in both 5-oxoeicosatetraenoate, can elicit the direct secretion of both granule
cellular and synthetic membranes. proteins and lipid mediators, although only with highly activated eosin-
EDN, also called EPX, is a 16-kDa, glycosylated protein possess- ophils or when used in conjunction with cytochalasin B, which inhibits
ing marked ribonuclease activity. The cDNA predicts a 134-amino-acid, microtubule assembly. Eosinophils release their granule components by
mature polypeptide that is identical to human urinary ribonuclease. exocytosis, with individual granules fusing with the plasma membrane.
Like ECP, it is a member of a ribonuclease multigene family. EDN This process involves a guanosine triphosphate (GTP)-binding protein
expression is not restricted to eosinophils, as it is found in mononuclear and is modulated by the intracellular calcium concentration. In com-
cells and possibly neutrophils. It is also probably secreted by the liver. mon with other secretory cells degranulation is controlled by soluble
It does not appear to be toxic to parasites or mammalian cells, and its N-ethylmaleimide–sensitive factor attachment protein (SNAP) recep-
only known effect, other than its ribonuclease activity, is neurotoxicity. tor complex (SNARE) proteins that guide the granules to the cell sur-
A major constituent of eosinophil is CLC protein, which is a lyso- face. Vesicular SNAREs expressed on granules bind to target SNAREs
phospholipase. It constitutes up to 10 percent of eosinophil protein and expressed on the plasma membrane. The SNARE proteins VAMP 2 and
is also found in large quantities in basophils. It was thought to possess VAMP 7 are important in eosinophil secretion from granules and their
lysophospholipase activity, but this is not the case. It is a member of the function in eosinophils is regulated by cyclin-dependent kinase 5. 120
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galactin family (galectin 10). Its function is unknown.
EOSINOPHILS IN DISEASE
EOSINOPHIL SECRETION AND ACTIVATION
A striking feature of eosinophil-rich inflammatory reactions is the high MEASUREMENT OF EOSINOPHILS
concentration of granule proteins, often in the presence of relatively IN THE BLOOD
small numbers of intact eosinophils. Sometimes free granules can be Eosinophils can be enumerated in the blood either by “wet counts” in
seen independent of eosinophils. This may occur as part of a cyto- modified Neubauer chambers, differential counts on dried films, or by
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lytic nonapoptotic process involving nicotinamide adenine dinucleo- automated cell counting by flow cytometry. Automated counting that
tide phosphate oxidase (NADPH) and the formation of extracellular uses detection of eosinophil peroxidase is the most accurate method,
DNA traps. 114,115 Mediator secretion can be triggered physiologically followed by counting in a cell chamber. Counting on films is least accu-
by engagement of immunoglobulin Fc receptors, especially after eos- rate because of the tendency for eosinophils to congregate at the mar-
inophil activation has been primed with soluble mediators such as PAF, gins of the slide. Common wet stains for eosinophils include eosin in
IL-5, and GM-CSF. Priming by GM-CSF involves phosphorylation of acetone, phloxine, and Kimura stain, which was originally developed
L-plastin and PKCβII. The eosinophil expresses receptors for IgG, to stain basophils. Many stains, including May-Grunewald-Giemsa,
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IgA, and IgD. The eosinophil also binds IgE, and eosinophils can under- Romanowsky, chromotrope 2R, and Biebrich scarlet, will identify eos-
take a number of IgE-dependent functions, including killing of schisto- inophils in blood films, cytospin preparations, or tissues.
somes opsonized with specific IgE. The receptor involved is unclear as The eosinophil count should preferentially be evaluated in abso-
the accumulated evidence now suggests that eosinophils do not express lute numbers rather than as a percentage of white cells, as the latter
either the low-affinity (FcεRII) or high-affinity (FcεRI) IgE receptor to will depend on the total white cell count. The normal eosinophil count
any degree although they do express high intracellular levels of the α is generally taken as less than 0.4 × 10 /L, although healthy medical
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chain of FcεRI. 117 students in the United States had a range of 0.015 to 0.65 × 10 /L.
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Three receptors for IgG have been described: the high-affinity Eosinophil counts are higher in neonates (Chap. 7). Clinical trials of
receptor FcγR1 (CD64), and two low-affinity receptors FcγRII (CDw32) anti-Th2 therapies are using a lower cut off in the region of 0.3 × 10 /L
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and FcγRIII (CD16). CD16 is expressed both as a transmembrane form to identify patients with eosinophilic asthma suggesting that the
and a form with a phosphatidylinositol anchor, transcribed from two “normal” eosinophil count in people without allergic disease is below
distinct genes. Only FcγRII is constitutively expressed by eosinophils this level. The eosinophil count varies with age, time of day, exercise
to any significant degree. A number of eosinophil functions are medi- status, and environmental stimuli, particularly allergen exposure. Blood
ated via this receptor, including schistosomula killing, phagocytosis, eosinophil counts undergo diurnal variation, being lowest in the morn-
secretion of granule proteins, and generation of newly formed, mem- ing and highest at night. This effect results in a greater than 40 per-
brane-derived lipid mediators such as PAF and LTC . After stimulation cent variation and may be related to the reciprocal diurnal variation
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for 2 days in vitro with interferon (IFN)-γ, eosinophils express CD16 in cortisol levels, which are highest in the morning. The factors that
and CD64 as well as CD32. Perhaps the most potent stimulus for eosin- control blood eosinophil counts in health are imperfectly understood.
ophil degranulation is crosslinking of IgA receptors, especially when the Concentrations of eosinophil growth factors are likely to be important,
cells have been primed with growth factors. Consistent with the prefer- but other factors may be involved, including an element of genetic con-
ence of eosinophils to secrete their mediators onto a large surface, Fc- trol. Normal counts vary by up to 40-fold, and, in populations where
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mediated degranulation is enhanced if the eosinophils are adherent to a eosinophilia is common, such as endemically parasitized areas, there
protein-coated surface via integrin α β . Eosinophils are particularly are marked variations in the blood eosinophil level, independent of
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