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PubMed1995

Eosinophil cationic protein and eosinophil-derived neurotoxin. Evolution of novel function in a primate ribonuclease gene family.

Human eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP) are members of a unique subfamily of rapidly evolving primate ribonuclease genes that emerged via a gene duplication event occurring after the divergence of Old World from New World monkeys (Rosenberg, H. F., Dyer, K. D., Tiffany, H. L., and Gonzalez, M. (1995) Nature Genet. 10, 219-223). In this work, we studied the activity of the protein encoded by the EDN/ECP homolog of the New World monkey, Saguinus oedipus (marmoset), a representative of the "ancestral" single sequences. Although the nucleotide sequence of the single marmoset gene (mEDN) was equally homologous (82%) to both human genes, the encoded amino acid sequence, calculated isoelectric point, and immunoreactivity all suggested a closer relationship with EDN. Furthermore, mEDN (at 0.3-1.0 microM concentrations) had no measurable anti-staphylococcal activity, suggesting functional as well as structural similarity to EDN. However, with yeast tRNA as substrate, mEDN had significantly less ribonuclease activity than EDN; Michaelis constants were nearly identical (Km (mEDN) = 0.67 microM; Km (EDN) = 0.70 microM), while turnover numbers differed by a factor of 100 (kcat (mEDN) = 0.91 s-1; kcat (EDN) = 0.64 x 10(-2) s-1). Thus, evolutionary constraints appear to have promoted two novel functions: increased cationicity/toxicity (ECP) and enhanced ribonuclease activity (EDN). The latter result is particularly intriguing, as it suggests a crucial role for ribonuclease activity in the (as yet to be determined) physiologic function of EDN.

Amino Acid Sequence↗
PubMed1999

Eosinophil peroxidase nitrates protein tyrosyl residues. Implications for oxidative damage by nitrating intermediates in eosinophilic inflammatory disorders.

Eosinophil peroxidase (EPO) has been implicated in promoting oxidative tissue injury in conditions ranging from asthma and other allergic inflammatory disorders to cancer and parasitic/helminthic infections. Studies thus far on this unique peroxidase have primarily focused on its unusual substrate preference for bromide (Br(-)) and the pseudohalide thiocyanate (SCN(-)) forming potent hypohalous acids as cytotoxic oxidants. However, the ability of EPO to generate reactive nitrogen species has not yet been reported. We now demonstrate that EPO readily uses nitrite (NO(2)(-)), a major end-product of nitric oxide ((.)NO) metabolism, as substrate to generate a reactive intermediate that nitrates protein tyrosyl residues in high yield. EPO-catalyzed nitration of tyrosine occurred more readily than bromination at neutral pH, plasma levels of halides, and pathophysiologically relevant concentrations of NO(2)(-). Furthermore, EPO was significantly more effective than MPO at promoting tyrosine nitration in the presence of plasma levels of halides. Whereas recent studies suggest that MPO can also promote protein nitration through indirect oxidation of NO(2)(-) with HOCl, we found no evidence that EPO can indirectly mediate protein nitration by a similar reaction between HOBr and NO(2)(-). EPO-dependent nitration of tyrosine was modulated over a physiologically relevant range of SCN(-) concentrations and was accompanied by formation of tyrosyl radical addition products (e.g. o,o'-dityrosine, pulcherosine, trityrosine). The potential role of specific antioxidants and nucleophilic scavengers on yields of tyrosine nitration and bromination by EPO are examined. Thus, EPO may contribute to nitrotyrosine formation in inflammatory conditions characterized by recruitment and activation of eosinophils.

Animals↗
PubMed1988

Purification and characterization of eosinophil cationic protein from normal human eosinophils.

ECP (eosinophil cationic protein) was purified in high yield from the granules of human buffy coat eosinophils obtained from healthy individuals. The separation procedure included gel filtration on Sephadex G-75, ion-exchange chromatography on Bio Rex 70, and chelating chromatography on zinc-chelate Sepharose 6B. The normal ECP is a single-chain, highly cationic glycoprotein which separates on SDS-polyacrylamide gel electrophoresis into at least 3 molecular weight forms, with molecular weights of 18.5, 20 and 22 kDa. A heterogeneity in charge was also observed, with the 18.5 kDa form being the most cationic one. The various molecular species of ECP exhibited antigenic identity, identical amino acid compositions, and identical amino-terminal amino acid sequences. The molecular heterogeneity was shown to be caused by differences in glycosylation of the protein.

Amino Acid Sequence↗
PubMed1993

Effect of butyric acid on induction of differentiation into eosinophil-like cells in human eosinophilic leukemia cells, EoL-1 cell line: possible role of granulocyte-macrophage colony-stimulating factor as an autocrine differentiating factor.

EoL-1 cells differentiated into mature eosinophil-like cells when incubated with butyric acid (BA). The differentiated cells possessed many granules that stained with Luxol fast blue and acquired the ability to produce leukotriene C4 and eosinophil cationic protein (ECP), and migrated toward some eosinophillotactic attractants, including PAF, IL-5, IL-3 and GM-CSF. Monoclonal antibody to GM-CSF and IL-5, especially GM-CSF, strongly decreased the ability of BA to induce differentiation, whereas antibody to IL-3 did not. Recombinant human (rh) IL-3, rhGM-CSF and recombinant mouse (rm) IL-5, by themselves, did not have the ability to induce differentiation. On the other hand, the presence of these cytokines significantly (p < 0.01) augmented BA-induced cell differentiation. Other cytokines including rhIL-1 alpha/beta, rhIL-2, rhIL-4, rhIL-6, rhIL-8, rhTNF-alpha and rhIFN-gamma had no effect on differentiation. Expression of GM-CSF and GM-CSF receptor mRNA was detected in cellular RNA obtained from BA-stimulated EoL-1 cells. Moreover, about 5-10 pg/ml of GM-CSF was detected in the culture supernatant of BA-stimulated EoL-1 cells. These findings suggested that BA-induced EoL-1 cell differentiation was elicited under the influence of GM-CSF endogenously produced in these cells. GM-CSF also seemed to be an autocrine differentiating factor in EoL-1 cells.

Antibodies↗
PubMed1994

Mechanism of eosinophil infiltration in the patient with subcutaneous angioblastic lymphoid hyperplasia with eosinophilia (Kimura's disease). Mechanism of eosinophil chemotaxis mediated by candida antigen and IL-5.

Kimura's disease is a chronic granulomatous disease of unknown etiology. Although eosinophilia is one of the characteristic features in this disease, little is known about the mechanism of eosinophilia. In the present study it was demonstrated that interleukin-5 (IL-5) was produced and released from the site of a granuloma and lymph nodes after stimulation with candida antigen. It was also shown that peripheral blood eosinophils from patients with Kimura's disease contained a large proportion of hypodense eosinophils and that their viability was prolonged. These results strongly suggest that locally produced IL-5 induced by candida antigen contributes to the eosinophilia in this disease.

Adult↗
PubMed2005

The principal eosinophil peroxidase product, HOSCN, is a uniquely potent phagocyte oxidant inducer of endothelial cell tissue factor activity: a potential mechanism for thrombosis in eosinophilic inflammatory states.

In vivo, bromide (Br(-)), nitrite (NO(2)(-)), and thiocyanate (SCN(-)) compete for oxidation by eosinophil peroxidase (EPO) and H(2)O(2), yielding, respectively, HOBr, NO(2)., and HOSCN. We have recently shown that SCN(-) is the strongly preferred substrate for EPO in vivo and that HOSCN, in contrast with other EPO-generated oxidants and HOCl, is a relatively weak, cell-permeant, sulfhydryl (SH)-reactive oxidant. We here show that HOSCN is a uniquely potent (up to 100-fold) phagocyte oxidant inducer of tissue factor (TF) activity in human umbilical vein endothelial cells (HUVECs). This induction is attributable to transcriptional up-regulation of TF gene expression dependent upon both activation of the p65/c-Rel TF-kappaB transcription factor and activity of the ERK1/2 kinase pathway upstream of Egr-1 and was markedly further enhanced in the presence of wortmannin, an inhibitor of the PI3 kinase/Akt pathway. HOSCN also markedly activates the proinflammatory p65/p50 NF-kappaB pathway. Based on these findings we hypothesize that HOSCN generated by adherent and infiltrating eosinophils may provoke the development of a prothrombotic and proinflammatory endothelial/endocardial phenotype that promotes the pronounced thrombotic diathesis characteristic of the hypereosinophilic syndrome.

Early Growth Response Protein 1↗
PubMed1983

Eosinophil chemotactic factor in schistosome eggs: a comparative study of eosinophil chemotactic factors in the eggs of Schistosoma japonicum and S. mansoni in vitro.

Significant chemotactic activity for eosinophils was detected in soluble egg antigen (SEA) preparations of both Schistosoma japonicum and S. mansoni in dose-dependent fashion. The activity of S. japonicum SEA was higher than that of S. mansoni SEA. Gel filtration on Sephadex G-150 showed that S. japonicum SEA was composed of two groups of eosinophil chemotactic factors (ECFs), one of high molecular weight (JEE-H) and the other of low molecular weight (JEE-L). S. mansoni SEA showed ECF composition similar to that of S. japonicum SEA. JEE-H was stable on heating (100 degrees C, 60 min) and resistant to pronase digestion, but was sensitive to periodate oxidation. JEE-L was also stable on heating and resistant to pronase and carboxypeptidase A digestions. These properties of the ECFs were also held in common with those of S. mansoni SEA. JEE-L was extractable by toluene, indicating a hydrophobic nature. These results suggest that schistosome eggs themselves contain ECFs, and that the composition of S. mansoni and S. japonicum SEA-derived ECFs is essentially the same. However, they differ from the other ECFs which have already been described in schistosome infections.

Animals↗
PubMed2005

Genetic polymorphisms of eosinophil-derived neurotoxin and eosinophil cationic protein in tropical pulmonary eosinophilia.

Because eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP) are critical in the pathogenesis of tropical pulmonary eosinophilia (TPE), we analyzed genetic polymorphisms of both in 181 individuals from southern India with varying clinical manifestations of Wuchereria bancrofti infection (including 26 with TPE). Using haplotype frequency analysis, we identified four known (of nine) and two novel haplotypes for EDN (1, 2, 7, 8, 10, and 11). For ECP, five (of seven known) haplotypes (1-5) were identified. Although we found no significant association between frequencies of EDN and ECP polymorphisms and TPE development, we observed a unique pattern of EDN and ECP polymorphism distribution among this population. Genotype TT at locus 1088 of ECP in one TPE patient was not observed in any other clinical group. Although the EDN and ECP polymorphisms appear unlikely to be associated with the development of TPE, further analyses will be more definitive.

Animals↗
PubMed2006

[Eosinophil cationic protein as a marker of eosinophil activity].

Eosinophil cationic protein (ECP) is one of the markers released by eosinophils. It suggests that ECP can be a good laboratory measurement for late allergic phase assessment. ECP can be measured in serum, BAL, sputum or nasal lavage. There are many studies that document the use of ECP in such diseases like: bronchial asthma, pollinosis or food allergy, but their results are not uniform.

Asthma↗
PubMed1998

[Peripheral eosinophil counts relates the improvement of the skin lesions of atopic dermatitis patients more sensitive than serum eosinophil cationic protein levels].

We described the severity of skin lesions and detected peripheral eosinophil counts (Eos) and serum eosinophil cationic protein levels (s-ECP) in 81 patients with atopic dermatitis (male 33, female 48, age 20.7 +/- 8.8) at the first consultation. Eos were 583.3 +/- 560.2/mm3 and s-ECP were 18.0 +/- 21.7 micrograms/l (normal 6.88 +/- 3.46). After approximately 30 days treatment, the improvement of dermatitis were estimated and those lavatory findings were rechecked. Eos were 446.7 +/- 367.6/mm3 and s-ECP were 13.2 +/- 11.7 micrograms/l. At 5 percent level, positive correlations were observed between Eos and s-ECP (R = 0.75), Eos and the severity of dermatitis (R = 0.52) and s-ECP and the severity of dermatitis (R = 0.51) at the first consultation. We found a positive correlation between Eos and the improvement of dermatitis (R = 0.40) but no correlation between s-ECP and the improvement of dermatitis (R = 0.10). Eos related the improvement of the skin lesions of atopic dermatitis more sensitive than s-ECP.

Adolescent↗
PubMed1985

Human lymphocyte-eosinophil interactions. I. Modulation of phytohemagglutinin-induced lymphocyte proliferation by eosinophils.

Do eosinophils modulate lymphocyte function? This question was studied by examining the effect of purified eosinophils (eos) on lectin-induced human lymphocyte proliferation. Intact resting or zymosan-stimulated eos or their extracts were cocultured with phytohemagglutinin-stimulated mononuclear cells in vitro and [3H]thymidine uptake was measured at 72 hr. Zymosan-stimulated eos consistently suppressed (up to 90%) the lectin-induced proliferative response by a noncytotoxic mechanism. Freeze-thaw extracts from zymosan-stimulated eos also significantly suppressed lymphocyte proliferation to a similar degree. The amount of suppression was directly proportional to the number of eos or the amount of extract added to the lymphocyte cultures. Intact resting eos and their extracts occasionally exhibited suppressive effects (up to 40%) on lymphocyte proliferation; this suppression, however, was always less than that of activated eos or their extracts. Eos pretreated with the protein synthesis inhibitor, pactamycin, exhibited significantly less suppressive activity, suggesting that a protein was responsible in part for the reduction in proliferation. The addition of superoxide dismutase or catalase to the eos-mononuclear cell cocultures did not reduce the amount of suppression observed, thus making it unlikely that active oxygen products were involved in the mechanism of suppression. Heating extracts from stimulated eos to 80 degrees C for 30 min resulted in partial loss of suppressive activity while extensive dialysis of the extracts had no effect. The studies reported here provide evidence that a nondialyzable and heat sensitive factor(s) produced by stimulated eos may exert feedback inhibition of lymphocyte function.

Catalase↗
PubMed1985

Activation of human eosinophils by monokines and lymphokines: source and biochemical characteristics of the eosinophil cytotoxicity-enhancing activity produced by blood mononuclear cells.

We and others have previously reported that human blood mononuclear cells release in culture certain substances that enhance the capacity of purified human blood eosinophils to kill the antibody-coated larvae of Schistosoma mansoni. The present study shows that this eosinophil cytotoxicity-enhancing activity (ECEA) is released by monocytes and T lymphocytes. Monocytes produce ECEA in resting and in LPS-stimulated cultures; T lymphocytes release such activity when stimulated by mitogens such as concanavalin A. Furthermore, the human monocytic line U-937 also releases ECEA-like activity when stimulated by LPS. The enhancing activity produced by monocytes has been partially characterized: it is sensitive to proteolysis by trypsin, relatively heat stable, and associated with molecules that have an apparent molecular weight of 14,000 to 65,000 daltons and isoelectric points of 3.8-3.9, 4.2, 4.5, 4.8-4.9. This shows that while ECEA produced by monocytes is heterogeneous in size and charge, it is probably different from interleukin 1.

Chromatography, Gel↗
PubMed1986

Human eosinophils and parasitic diseases--III. Beta-interferon increases eosinophil IgG-Fc receptor expression and capacity.

Eosinophilic granulocytes from the blood of patients with parasitic diseases were preincubated in the presence or absence of beta-interferon (beta-IFN). The number of cells bearing IgG-Fc receptors was determined by rosette formation with chicken erythrocytes coated with IgG antibodies. beta-IFN augmented the expression of IgG-Fc receptors on the cells within 1 h incubation. Treatment of beta-IFN with heat or acid did not abolish the activity. The doses of beta-IFN needed to induce the modulations of IgG-Fc receptors could also augment the eosinophil-mediated antibody-dependent cytotoxicity (ADCC) against chicken red blood cells which resulted most pronounced when the IgG antibodies were present in suboptimal amount. These data suggest the role of beta-IFN in the host resistance to parasitic diseases, mainly in the initial phases of parasitic infestation when the specific IgG antibody levels are very low.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1988

Periarteritis of coronary arteries with severe eosinophilic infiltration. A new pathologic entity (eosinophilic periarteritis)?

A 40 year-old male presented symptoms of angina pectoris for about nine years and expired with symptoms of unstable angina, changing pattern at the terminal stage. At autopsy, both right and left coronary arteries of the subepicardial region were grayish white and elastic hard. Histologically, inflammatory infiltration was localized in adventitia of coronary arteries located in the subepicardial region. Inflammatory cells infiltrated into the adventitia were mostly eosinophiles. The medial smooth muscle cells were well preserved and the intima showed irregular thickening with fibrosis. Vascular obstruction or recanalization could not be observed. As a result of these findings, it was considered that these inflammatory changes of the coronary arteries could be termed eosinophilic periarteritis. These inflammatory changes could not be found in the intramural coronary arteries. Rather extensive fibrosis could be seen in the muscle layer centering about the posterior wall of the left ventricle. No findings of angiitis could be detected in the blood vessels except subepicardial coronary arteries.

Adult↗
PubMed1997

Macrophage-mediated candidacidal activity is augmented by exposure to eosinophil peroxidase: a paradigm for eosinophil-macrophage interaction.

Various disease states are associated with eosinophilia and the release of eosinophil peroxidase (EPO) into the microenvironment. The present study targets the effects of low levels of EPO on macrophage (M phi) phagocytosis and intracellular killing of Candida albicans as well as M phi oxidative activity measured as the luminescence product of luminol dioxygenation. Resident murine peritoneal M phi were exposed to various concentrations of EPO. Chemiluminescence data indicate that nanomolar concentrations of EPO markedly enhanced the dioxygenation activity (respiratory burst) of M phi. In other studies, the exposure of M phi to 0.17 microM EPO for 10 min. enhanced M phi-mediated candidacidal activity 10 fold. The above data indicate that EPO enhances certain M phi functions. Also the results illustrate a previously un-recognized interaction between eosinophils and M phi and implicate yet another possible role for EPO in host defenses against disease.

Animals↗
PubMed1975

Studies of the responses of basophil and eosinophil leucocytes and mast cells to the nematode Trichostrongylus colubriformis. Ultrastructural changes in basophils and eosinophils at the site of infection.

Basophil and eosinophil leucocytes infiltrate the small intestinal lamina propria of guinea-pigs infected with the nematode Trichostrongylus colubriformis. By comparing the morphology of both cell types at the site of infection with bone marrow and buffy coat cells, it was found that, after entering the lamina propria, basophils developed an electron lucent halo beneath the granule-limiting membrane while the characteristic orderly periodicity of the granules changed to a fibrillar or amorphous appearance. The granules also tended to coalesce but remained within the cell. Approximately half the eosinophils at the site of infection developed deficiencies in the amorphous outer matrix of their granules but showed no obvious change in the central electron-dense cores.

Animals↗