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Eosinophils are classically known as proinflammatory cells, as they are equipped with a variety of preformed cytotoxic mediators and have been shown to definitively contribute to asthma. The connection between eosinophils and asthma development has led to a new class of asthma therapeutics based on blocking eosinophils with humanized antibodies that neutralize IL-5, a potent eosinophil growth, activation, and survival factor. Yet, recent studies have led to an increasing appreciation that eosinophils have a variety of homeostatic functions, including immunomodulation. In this issue of the JCI, Mesnil et al. identify a notable population of lung-resident eosinophils and demonstrate that, compared with traditional eosinophils, these cells have distinct characteristics, including nuclear structure, surface markers, IL-5 independence, and immunoregulatory function that is capable of polarizing adaptive immune responses, at least in vitro. Thus, these results reinforce a key homeostatic role for this enigmatic cell population, particularly in residing and regulating immunity in the lung., Eosinophils: traditionally proinflammatory Eosinophils are traditionally viewed as proinflammatory cells, as they are equipped with a variety of preformed cytotoxic mediators, including the granule proteins major basic protein (MBP) and [...]

Eosinophilic inflammation is a cornerstone of chronic asthma that often culminates in subepithelial fibrosis with variable airway obstruction. Pulmonary eosinophils (Eos) are a predominant source of TGF-[beta]1, which drives fibroblast proliferation and extracellular matrix deposition. We investigated the regulation of TGF-[beta]1 and show here that the peptidyl-prolyl isomerase (PPIase) Pin1 promoted the stability of TGF-[beta]1 mRNA in human Eos. In addition, Pin1 regulated cytokine production by both in vitro and in vivo activated human Eos. We found that Pin1 interacted with both PKC-[alpha] and protein phosphatase 2A, which together control Pin1 isomerase activity. Pharmacologic blockade of Pin1 in a rat asthma model selectively reduced eosinophilic pulmonary inflammation, TGF-[beta]1 and collagen expression, and airway remodeling. Furthermore, chronically challenged Pin1-/- mice showed reduced peribronchiolar collagen deposition compared with wild-type controls. These data suggest that pharmacologic suppression of Pin1 may be a novel therapeutic option to prevent airway fibrosis in individuals with chronic asthma., Introduction Persistent asthma is characterized by chronic pulmonary inflammation, which often culminates in airway fibrosis. The development of airway disease is referred to as remodeling and includes the deposition of [...]

INTRODUCTION: We investigated whether markers of airway and systemic inflammation, as well as heart rate variability (HRV) in asthmatics, change in response to fluctuations in ambient particulate matter (PM) in [...]

A PCR-based method was developed to detect Angiostrongylus cantonensis directly from molluscan tissue to control the outbreak of human eosinophilic meningitis.

Lymphopenia and restricted T cell repertoires in humans are often associated with severe eosinophilic disease and a T cell Th2 bias. To examine the pathogenesis of this phenomenon, C57BL/6 Rag2-/-mice received limited (3 x [10.sup.4]) or large (2 x [10.sup.6]) numbers of CD4 T cells. Three to 5 months after transfer, mice that had received 3 x [10.sup.4] T cells, but not those that received 2 x [10.sup.6], developed fulminant macrophage pneumonia with eosinophilia, Ym1 deposition, and methacholine-induced airway hyperresponsiveness, as well as eosinophilic gastritis; esophagitis and other organ damage occurred in some cases. Donor cells were enriched for IL-4, IL-5, and IL-13 producers. When 3 x [10.sup.4] cells were transferred into CD3[epsilon]-/-hosts, the mice developed strikingly elevated serum IgE. Prior transfer of 3 x [10.sup.5] CD25+ CD4 T cells into Rag2-/- recipients prevented disease upon subsequent transfer of CD25- CD4 T cells, whereas 3 x [10.sup.4] regulatory T cells (Tregs) did not, despite the fact that there were equal total numbers of Tregs in the host at the time of transfer of CD25- CD4 T cells. Limited repertoire complexity of Tregs may lead to a failure to control induction of immunopathologic responses, and limitation in repertoire complexity of conventional cells may be responsible for the Th2 phenotype. eosinophils | IgE ILL-4 | macrophages | pneumonia

Although leukocytes of the innate immune system, including eosinophils, contain within their granules preformed stores of cytokines available for selective and rapid release, little is known about the mechanisms governing the mobilization and secretion of these cytokines. Here we show that a cytokine receptor, the IL-4 receptor [alpha] chain, mediates eotaxin-stimulated mobilization of preformed IL-4 from eosinophil granules into secretory vesicles. Eosinophils contain substantial intracellular quantities of several granule- and vesicle-associated cytokine receptors, including IL-4, IL-6, and IL-13 receptors as well as CCR3. Both IL-4 and IL-4 receptor [alpha] chain colocalized in eosinophil granules; and after eotaxin-stimulation, IL-4 receptor [alpha] chain, bearing bound IL-4, was mobilized into secretory vesicles. These findings indicate that intracellular cytokine receptors within secretory vesicles transport their cognate cytokines requisite for the secretion of cytokines preformed in innate immune leukocytes. intracellular cytokine receptor | intracellular cytokine trafficking piecemeal degranulation | vesicular transport

The visualization, by confocal Raman microscopy, of the intracellular distribution of two enzymes important in the immune eosinophil peroxidase (EPO) in eosinophils is reported. The effect of photobleaching of the Raman signal at 413.1 nm excitation on the reconstructed Raman images is discussed.

The NADPH--oxidase is a plasma membrane enzyme complex that enables phagocytes to generate superoxide in order to kill invading pathogens, a critical step in the host defense against infections. The oxidase transfers electrons from cytosolic NADPH to extracellular oxygen, a process that requires concomitant [H.sup.+] extrusion through depolarization-activated [H.sup.+] channels. Whether [H.sup.+] fluxes are mediated by the oxidase itself is controversial, but there is a general agreement that the oxidase and [H.sup.+] channel are intimately connected. Oxidase activation evokes profound changes in whole-cell [H.sup.+] current ([I.sub.H]), causing an approximately--40-mV shift in the activation threshold that leads to the appearance of inward [I.sub.H]. To further explore the relationship between the oxidase and proton channel, we performed voltage-clamp experiments on inside-out patches from both resting and phorbol-12-myristate-13-acetate (PMA)-activated human eosinophils. Proton currents from resting cells displayed slow voltage-dependent activation, long-term stability, and were blocked by micromolar internal [[Zn.sup.2+]]. [I.sub.H] from PMA-treated cells activated faster and at lower voltages, enabling sustained [H.sup.+] influx, but ran down within minutes, regaining the current properties of nonactivated cells. Bath application of NADPH to patches excised from PMA-treated cells evoked electron currents ([I.sub.e]), which also ran down within minutes and were blocked by diphenylene iodonium (DPI). Run-down of both [I.sub.H] and [I.sub.e], was delayed, and sometimes prevented, by cytosolic ATP and GTP-[gamma]-S. A good correlation was observed between the amplitude of [I.sub.e] and both inward and outward [I.sub.H] when a stable driving force for [e.sup.-] was imposed. Combined application of NADPH and DPI reduced the inward [I.sub.H] amplitude, even in the absence of concomitant oxidase activity. The strict correlation between [I.sub.e] and [I.sub.H] amplitudes and the sensitivity of [I.sub.H] to oxidase-specific agents suggest that the proton channel is either part of the oxidase complex or linked by a membrane-limited mediator. KEY WORDS: physiology * NADPH-oxidase * zinc * phagocyte * patch-clamp

Eosinophils are implicated prominently in allergic diseases and the host response to parasitic infections. Eosinophils may be activated in vitro by diverse classes of agonists such as immunoglobulins, lipid mediators, and cytokines. The leukocyte Ig-like receptors (LIRs) comprise a family of inhibitory and activating cell-surface receptors. Inhibitory MRs down-regulate cellular responses through cytoplasmic immunoreceptor tyrosine-based inhibitory motifs. There are limited data on the action of the activating LIRs, which are thought to signal through the Fc receptor [gamma] chain, which contains an immunoreceptor tyrosine-based activation motif. We now demonstrate the expression of LIR1 (inhibitory), LIR2 (inhibitory), LIR3 (inhibitory), and LIR7 (activating) on eosinophiis from 4, 4, 12, and 11, respectively, of 12 healthy donors. Cross-linking of LIR7 with plate-bound antibody elicited the dose- and time-dependent release of eosinophil-derived neurotoxin and leukotriene [C.sub.4]. Eosinophils activated with antibodies to LIR7 embedded in gelphase EliCell preparations showed leukotriene [C.sub.4] generation at the nuclear envelope and the release of IL-12 but not IL-4 by vesicular transport. Thus, LIR7 is an activating receptor for eosinophils that elicited the release of cytotoxic granule proteins, de novo lipid mediator generation, and cytokine release through vesicular transport.

Insights into structure-function relations of many proteins opens the possibility of engineering peptides to selectively interfere with a protein's activity. To facilitate the use of peptides as probes of cellular processes, we have developed caged peptides whose influence on specific proteins can be suddenly and uniformly changed by near-UV light. Two peptides are described which, on photolysis of a caging moiety, block the action of calcium-calmodulin or myosin light chain kinase (MLCK). The efficacy of theses peptides is demonstrated in vitro and in vivo by determining their effect before and after photolysis on activities of isolated enzymes and cellular functions known to depend on calcium-calmodulin and MLCK. These caged peptides each were injected into motile, polarized eosinophils, and when exposed to light promptly blocked cell locomotion in a similar manner. The results indicate that the action of calcium-calmodulin and MLCK, and by inference myosin II, are required for the ameboid locomotion of these cells. This methodology provides a powerful means for assessing the role of these and other proteins in a wide range of spatio-temporally complex functions in intact living cells.

Eosinophil migration in vivo is markedly attenuated in rats treated chronically with the NO synthase (NOS) inhibitor [N.sup.[Omega]]-nitro-L-arginine methyl ester (L-NAME). In this study, we investigated the existence of a NOS system in eosinophils. Our results demonstrated that rat peritoneal eosinophils strongly express both type II (30.2 [+ or -] 11.6% of counted cells) and type III (24.7 [+ or -] 7.4% of counted cells) NOS, as detected by immunohistochemistry using affinity purified mouse mAbs. Eosinophil migration in vitro was evaluated by using 48-well microchemotaxis chambers and the chemotactic agents used were N-formyl-methionyl-leucyl-phenylalanine (fMLP, 5 x [10.sup.-8] M) and leukotriene [B.sub.4] ([LTB.sub.4], [10.sup.-8] M). L-NAME (but not D-NAME) significantly inhibited the eosinophil migration induced by both fMLP (54% reduction for 1.0 mM; P < 0.05) and [LTB.sub.4] (61% reduction for 1.0 mM; P < 0.05). In addition, the type II NOS inhibitor 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine and the type I/II NOS inhibitor 1-(2-trifluoromethylphenyl) imidazole also markedly (P < 0.05) attenuated fMLP- (52% and 38% reduction for 1.0 mM, respectively) and [LTB.sub.4]- (52% and 51% reduction for 1.0 mM, respectively) induced migration. The inhibition of eosinophil migration by L-NAME was mimicked by the soluble guanylate cyclase inhibitor 1H-[1,2,4] oxadiazolo [4,3,-a] quinoxalin-1-one (0.01 and 0.1 mM) and reversed by either sodium nitroprusside (0.1 mM) or dibutyryl cyclic GMP (1 mM). We conclude that eosinophils do express NO synthase(s) and that nitric oxide plays an essential role in eosinophil locomotion by acting through a cyclic GMP transduction mechanism.

The CC chemokines regulated on activation normal T expressed and secreted (RANTES) and monocyte chemotactic protein 3 (MCP-3), and the anaphylatoxin C5a, induce activation, degranulation, chemotaxis, and transendothelial migration of eosinophils. Adhesion assays on purified ligands showed differential regulation of [Beta]1 and [Beta]2 integrin avidity in eosinophils. Adhesiveness of VLA-4 ([Alpha]4[Beta]1, CD29/CD49d) for vascular cell adhesion molecule 1 or fibronectin was rapidly increased but subsequently reduced by RANTES, MCP-3, or C5a. The deactivation of VLA-4 lead to cell detachment, whereas phorbol 12-myristate 13-acetate induced sustained activation of VLA-4. In contrast, chemoattractants stimulated a prolonged increase in the adhesiveness of Mac-1 ([Alpha]M[Beta]2, CD11b/CD18) for intercellular adhesion molecule 1. Inhibition by pertussis toxin confirmed signaling via G protein-coupled receptors. Chemoattractants induced transient, while phorbol 12-myristate 13-acetate induced sustained actin polymerization. Disruption of actin filaments by cytochalasins inhibited increases in avidity of VLA-4 but not of Mac-1. Chemoattractants did not upregulate a [Mn.sup.2+]-inducible [Beta]1 neoepitope defined by the mAb 9EG7, but induced prolonged expression of a Mac-1 activation epitope recognized by the mAb CBRM1/5. This mAb inhibited chemoattractant-stimulated adhesion of eosinophils to intercellular adhesion molecule 1. Thus, regulation of VLA-4 was dependent on the actin cytoskeleton, whereas conformational changes appeared to be crucial for activation of Mac-1. To our knowledge, this is the first demonstration that physiological agonists, such as chemoattractants, can differentially regulate the avidity of a [Beta]1 and a [Beta]2 integrin expressed on the same leukocyte.

Normal human blood cells exposed for 14 days to recombinant human interleukin develop into granulocytes that have the characteristics of basophils and eosinophils. The hybrid granulocytes developed into hypodense mononuclear eosinophils when they were removed from the matrigel and cytokine growth medium. When they were transferred into a growth medium that did not contain cytokines, the eosinophils produced granulocyte-macrophage colony-stimulating factor, pointing to the role of undifferentiated eosinophils in autocrine processes.

Immunochemical and spectroscopic analyses of eosinophils deficient in hereditary eosinophil peroxidase (EPO) reveal that these eosinophils do not exhibit the characteristic EPO absorption spectrum, but have peptides that are antigenically related to EPO. This deficiency has a compound heterozygosity with mutations that lead to substitution of an arginine residues with histidine. Eosinophil precursors in these eosinophils generate a spectroscopically different EPO that however, has normal cytochemical reaction for peroxidase and immunoreactivity with monoclonal and polyclonal anti-EPO antibodies.

The high affinity immunoglobulin E (IgE) receptor (Fc-epsilon-RI) mediates eosinophil-dependent cytotoxicity against Schistosoma and participates in physiological protective immune response against metazoan parasites. Fe-epsilon-RI is a tetrameric structure that comprises an IgE-binding alpha-chain, a beta-chain and two disulphide-linked gamma-chains. Fe-epsilon-RI is also involved in eosinophil degranulation.

The concentration of intracellular free calcium ([[Ca.sup.2+]]i) in polarized eosinophils was imaged during chemotaxis by monitoring fluorescence of the calcium-sensitive dye Fura-2 with a modified digital imaging microscope. Chemotactic stimuli caused [[Ca.sup.2+]]i to increase in a nonuniform manner that was related to cell activity. In cells moving persistently in one direction, [[Ca.sup.2+]]i was highest at the rear and lowest at the front of the cell. Before cells turned, [[Ca.sup.2+]]i transiently increased. The region of the cell that became the new leading edge had the lowest [[Ca.sup.2+]]i. These changes in [[Ca.sup.2+]]i provide a basis for understanding the organization and local activity of cytoskeletal proteins thought to underlie the directed migration of many cells., THE ABILITY OF CELLS TO MIGRATE toward a directional stimulus is a basic property of virtually all cells at some stage in their development [1] Directed migration results from the [...]

A relatively obscure immune cell, the eosinophil, has a dramatic way of defending against pathogens. It rapidly ejects mitochondrial DNA, ensnaring bacteria and hastening their demise (pages 949-953). Eosinophils, a [...]

The ongoing epidemic of allergic asthma in the developed world has lent urgency to the quest to understand the pathogenesis of this chronic, often debilitating, inflammatory disease. There is a [...]

The mammalian RNase A superfamily comprises a diverse array of ribonucleolytic proteins that have a variety of biochemical activities and physiological functions. Two rapidly evolving RNases of higher primates are of particular interest as they are major secretory proteins of eosinophilic leukocytes and have been found to possess anti-pathogen activities in vitro. To understand how these RNases acquired this function during evolution and to develop animal models for the study of their functions in vivo, it is necessary to investigate these genes in many species. Here, we report the sequences of 38 functional genes and 23 pseudogenes of the eosinophil-associated RNase (EAR) family from 5 rodent species. Our phylogenetic analysis of these genes showed a clear pattern of evolution by a rapid birth-and-death process and gene sorting, a process characterized by rapid gene duplication and deactivation occurring differentially among lineages. This process ultimately generates distinct or only partially overlapping inventories of the genes, even in closely related species. Positive Darwinian selection also contributed to the diversification of these EAR genes. The striking similarity between the evolutionary patterns of the EAR genes and those of the major histocompatibility complex, immunoglobulin, and T cell receptor genes stands in strong support of the hypothesis that host-defense and generation of diversity are among the primary physiological function of the rodent EARs. The discovery of a large number of divergent EARs suggests the intriguing possibility that these proteins have been specifically tailored to fight against distinct rodent pathogens.