Neutralization of TNF by the Antibody cA2 Reveals Differential Regulation of Adhesion Molecule Expression on TNF-Activated Endothelial Cells

Upregulation of adhesion proteins plays an important role in mediating inflammation. The induction of adhesive molecules has been well studied, but the reversibility of their expression has not been well characterized. A neutralizing anti-TNF monoclonal antibody (cA2) was used to study the down regulation of TNF-induced E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) on cultured human umbilical vein endothelial cells (HUVECs). Addition of cA2 following TNF stimulation of HUVECs enhanced the rate of E-selectin and VCAM-1 down-regulation from the cell surface and also reduced steady state E-selectin and VCAM-1 mRNA levels. The cA2-mediated disappearance of E-selectin, but not VCAM-1 protein was microtubule and not microfilament dependent. Neutralization of TNF only slightly reduced ICAM-1 cell surface levels following initial TNF stimulation, suggesting a slower turnover of ICAM-1 compared to E-selectin and VCAM-1. Microtubule inhibition during TNF stimulation partially inhibited E-selectin, VCAM-1 and ICAM-1 mRNA upregulation. VCAM-1 and ICAM-1 cell surface expression were similarly partially inhibited, however, E-selectin levels were unaffected, presumably due to the dual, opposing effect of inhibiting protein expression and inhibiting internalization. Microfilament inhibition during protein induction specifically inhibited the maximal expression of VCAM-1 protein and mRNA, without affecting E-selectin or ICAM-1. These data support the notion that E-selectin, VCAM-1, and ICAM-1 expression are differentially regulated on HUVECs and suggest that TNF neutralizing therapies may be effective because of their ability to reduce the levels of pre-existing adhesion proteins.     

Different patterns of IL-1β secretion, adhesion molecule expression and apoptosis induction in human endothelial cells treated with 7α-, 7β-hydroxycholesterol, or 7-ketocholesterol

Among oxysterols oxidized at C7 (7α-, 7β-hydroxycholesterol, and 7-ketocholesterol), 7β-hydroxycholesterol and 7-ketocholesterol involved in the cytotoxicity of oxidized low density lipoproteins (LDL) are potent inducers of apoptosis. Here, we asked whether all oxysterols oxidized at C7 were able to trigger apoptosis, to stimulate interleukin (IL)-1β and/or tumor necrosis factor (TNF)-α secretion, and to enhance adhesion molecule expression (intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin) on human umbilical venous endothelial cells (HUVECs). Only 7β-hydroxycholesterol and 7-ketocholesterol were potent inducers of apoptosis and of IL-1β secretion. TNF-α secretion was never detected. Depending on the oxysterol considered, various levels of ICAM-1, VCAM-1 and E-selectin expression were observed. So, oxysterols oxidized at C7 differently injure and activate HUVECs, and the α- or β-hydroxyl radical position plays a key role in apoptosis and IL-1β secretion.     

Cryptotanshinone inhibits oxidized LDL-induced adhesion molecule expression via ROS dependent NF-κB pathways

Adhesion molecules, such as intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin, play important roles in the initial stage of atherosclerosis. Cryptotanshinone (CPT), a natural compound isolated from Salvia miltiorrhiza Bunge, exhibits anti-atherosclerotic activity although the underlying mechanisms remain elusive. In this study, the protective effect of CPT against oxidized low-density lipoprotein (ox-LDL)-induced adhesion molecule expression was investigated in human umbilical vein endothelial cells. Ox-LDL significantly induced ICAM-1, VCAM-1, and E-selectin expression at the mRNA and protein levels but reduced eNOS phosphorylation and NO generation, which were reversed by CPT pretreatment. Sodium nitroprusside, a NO donor, N-acetyl-L-cysteine (NAC), a reactive oxygen species (ROS) scavenger, and BAY117082, a NF-κB inhibitor, inhibited ox-LDL-induced ICAM-1, VCAM-1, and E-selectin expression. Ox-LDL-induced ROS production was significantly inhibited by CPT and NAC. Furthermore, ox-LDL activated the NF-κB signaling pathway by inducing phosphorylation of IKKβ and IκBα, promoting the interaction of IKKβ and IκBα, and increasing p65 nuclear translocation, which were significantly inhibited by CPT. In addition, CPT, NAC, and BAY117082 inhibited ox-LDL-induced membrane expression of ICAM-1, VCAM-1, E-selectin, and endothelial–monocyte adhesion and restored eNOS phosphorylation and NO generation. Results suggested that CPT inhibited ox-LDL-induced adhesion molecule expression by decreasing ROS and inhibiting the NF-κB pathways, which provides new insight into the anti-atherosclerotic mechanism of CPT.     

Oxidized LDL further enhances expression of adhesion molecules in Chlamydophila pneumoniae-infected endothelial cells

Chlamydophila pneumoniae is a common respiratory pathogen that has been shown to be associated with coronary artery disease. Recent studies have shown that one of the possible mechanisms of the atherogenicity of C. pneumoniae is overexpression of cell adhesion molecules (CAMs) in infected endothelial cells. We investigated whether exposure of C. pneumoniae-infected endothelial cells to oxidized LDL (oxLDL) leads to further upregulation of CAMs. Flow cytometry and immunoblot analysis of human aortic endothelial cells (HAECs) was performed for intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin. ICAM-1 was expressed in 78.7% of C. pneumoniae-infected HAECs. The addition of oxLDL (100 microg/ml) to infected HAECs increased the proportion of ICAM-1-positive cells to 92%. VCAM-1 was only observed in 9.3% of infected HAECs, and the addition of oxLDL had no further effect on the surface expression of VCAM-1. C. pneumoniae also upregulated the surface expression of E-selectin on 52.2% of the cells, and incubation with oxLDL further increased the proportion of positive cells to 63.64%. In conclusion, C. pneumoniae upregulated the expression of the adhesion molecules ICAM-1, VCAM-1, and E-selectin on HAECs. The addition of oxLDL to the infected cells further enhanced the surface expression of ICAM-1 and E-selectin.     

Attenuation of leukocyte-endothelium interaction by antioxidant enzymes

This report assessed the effect of overexpressing Cu,Zn superoxide dismutase (SOD) and/or catalase on the interaction of mononuclear cells (MNCs) and endothelial cells (ECs). ECs were obtained from the aorta of wild-type mice and transgenic mice overexpressing Cu,ZnSOD and/or catalase. MNCs were obtained from wild-type mice. Treatment of wild-type ECs with CuSO4-oxidized low-density lipoprotein (oxLDL) significantly elevated the expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) and increased the adherence of MNCs. Overexpression of Cu,ZnSOD and/or catalase in ECs attenuated the adherence of MNCs and the expression of cell adhesion molecules induced by oxLDL. For example, ECs overexpressing Cu,ZnSOD and/or catalase showed significantly less expression of VCAM-1 and ICAM-1 and less number of adherent MNCs than wild-type ECs. Moreover, ECs overexpressing Cu,ZnSOD and catalase in combination showed significantly less expression of VCAM-1 and ICAM-1 and less number of adherent MNCs than those overexpressing either Cu,ZnSOD or catalase alone. These results suggest that combinational overexpression of Cu,ZnSOD and catalase can reduce the expression of cell adhesion molecules and inhibit the adherence of leukocyte to ECs more efficiently than overexpression of Cu,ZnSOD or catalase alone.     

Inhibitory effects of vitamin E on endothelial-dependent adhesive interactions with leukocytes induced by oxidized low density lipoprotein

Leukocyte-endothelial cell interactions, which are mediated by various adhesion molecules, are a crucial event in inflammatory reactions including atherosclerosis. Alpha-tocopherol (alpha-Toc) has been used for protection and therapy of vascular diseases because of its antioxidant activity. The objective of the present study was to determine effect of alpha-Toc on endothelial-dependent adhesive interactions with leukocytes elicited by oxidized low density lipoprotein (oxLDL). Incubation of HUVEC with oxLDL (100 microg/mL) increased expression of proteins and messenger RNA of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) on enzyme immunoassay and northern blotting assay; pretreatment with alpha-Toc reduced in a dose dependent manner. Adherence of polymorphonuclear leukocytes (PMN) or mononuclear leukocytes (MNC) to oxLDL-activated HUVEC was much increased compared with that to unstimulated HUVEC. Treatment of HUVEC with alpha-Toc, monoclonal antibody to ICAM-1 or VCAM-1 inhibited adherence of PMN or MNC in a dose dependent manner. These results suggest that alpha-Toc works as anti-atherogenic agent through inhibiting endothelial-dependent adhesive interactions with leukocytes induced by oxLDL.     

7-Ketocholesterol enhances the expression of adhesion molecules on human aortic endothelial cells by increasing the production of reactive oxygen species

The aim of the present study was to assess the expression of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), monocytic adhesion of human aortic endothelial cells (HAECs), and the production of intracellular reactive oxygen species (ROS), when HAECs were stimulated by 7-ketocholesterol. 7-ketocholesterol enhances surface expression of ICAM-1 and VCAM-1 as determined by EIA, induces their mRNA expression by RT-PCR, and stimulates adhesiveness of HAECs to U937 monocytic cells. We confirmed up-regulation of ROS production of HAECs treated with 7-ketocholesterol. Although the surface expression of ICAM-1 and VCAM-1 on HAECs treated with 7-ketocholesterol increased in a time-dependent manner, alpha-tocopherol inhibited this increase of the surface expression of ICAM-1 and VCAM-1. In the monocytic adhesion assay, adhesion of U937 to HAECs treated with 7-ketocholesterol was enhanced, but monoclonal anti-ICAM-1 and VCAM-1 antibodies reduced the endothelial adhesiveness. In conclusion, this study suggests that the endothelial adhesiveness to monocytic cells that was increased by 7-ketocholesterol was associated with enhanced expression of ICAM-1 and VCAM-1 mediated by ROS production.     

Artichoke juice improves endothelial function in hyperlipemia

Artichoke extracts have been shown to produce various pharmacological effects, such as the inhibition of cholesterol biosynthesis and of LDL oxidation. Endothelial dysfunction represents the first stage of atherosclerotic disease; it is usually evaluated in humans by a noninvasive ultrasound method as brachial flow-mediated vasodilation (FMV) and by the determination of several humoral markers such as vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin. Aim of the study was to investigate the effects of dietary supplementation with artichoke juice on brachial FMV of hyperlipemics. We studied 18 moderately hyperlipemic patients (LDL cholesterol > 130 <200 mg/dl and/or triglycerides >150 <250 mg/dl) of both genders and 10 hyperlipemic patients, matched for age, sex and lipid parameters. All subjects were under isocaloric hypolipidic diet. A basal determination of serum lipids, soluble VCAM-1, ICAM-1, E-selectin and brachial FMV was performed. Thereafter patients were given 20 ml/die of frozen artichoke juice. The same parameters were repeated after 6 weeks. After artichoke treatment there was an increase of triglycerides (156 +/- 54 vs 165 +/- 76 mg/dL, p <0.05) and a reduction of total cholesterol (261 +/- 37 vs 244 +/- 38 mg/dL, p <0.05) and LDL cholesterol (174 +/- 31 vs 160 +/- 34 mg/dL, p <0.05). Controls showed a significant decrease in total and LDL cholesterol (respectively: 267 +/- 22 vs 249 +/- 20 mg/dL and 180 +/- 24 vs 164 +/- 23 mg/dL, both p <0.001). After artichoke there was a decrease in VCAM-1(1633 +/- 1293 vs 1139 +/- 883 ng/mL, p <0.05) and ICAM-1(477 +/- 123 vs 397 +/- 102 ng/mL, p <0.05), brachial FMV increased (3.3 +/- 2.7 vs 4.5 +/- 2.4%, p <0.01), while controls did not exhibit significant changes in VCAM-1, ICAM-1, E-selectin and brachial FMV. Univariate analysis showed that, in artichoke patients, changes of VCAM-1 and ICAM-1 were significantly related to changes in brachial FMV (respectively: r=-0.66 and r=-0.62; both p <0.05). In conclusion, artichoke dietary supplementation seems to positively modulate endothelial function in hypercholesterolemia.     

秦皮甲素对AGEs致损的ECV304细胞增殖及氧化应激的影响

为研究秦皮甲素对血管内皮细胞的保护作用,采用CCK-8法观察秦皮甲素对体外AGEs培养的人脐静脉内皮细胞增殖的影响。检测不同浓度AGEs以及秦皮甲素作用后对内皮细胞一氧化氮(NO)、不对称二甲基精氨酸(ADMA)水平的影响以及内皮细胞氧化应激有关指标:活性氧簇(reactive oxygen species,ROS)、丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxide dismutase,SOD);脂肪代谢相关指标:乳酸脱氢酶(lactic dehydrogenase,LDH)、总胆固醇(total cholesterol,CHO)、甘油三酯(triglyceride,TG)和低密度脂蛋白(low density lipoprotein,LDL),同时分别检测粘附相关因子:血管细胞粘附分子-1(VCAM-1)和细胞间粘附分子-1(ICAM-1)的表达水平。结果显示200 mg/L AGEs对人内皮细胞ECV304增殖有显著抑制作用,秦皮甲素可对抗AGEs导致的内皮细胞增殖抑制,并呈浓度依赖性。在25 mg/L时,保护效应达到最高。秦皮甲素可抵抗ROS生成。同时可改善细胞的脂类代谢:胆固醇、LDL以及TG含量在秦皮甲素作用后改善明显。秦皮甲素可显著抑制内皮粘附因子VCAM-1的表达。秦皮甲素还可上调NO水平,下调ADMA水平。总之,秦皮甲素可有效促进人血管内皮细胞增殖并在改善氧化应激,脂代谢,粘附因子和NO释放等方面发挥作用。     

Effects of TNF-alpha on leukocyte adhesion molecule expressions in cultured human lymphatic endothelium.

TNF-alpha alters leukocyte adhesion molecule expression of cultured endothelial cells like human umbilical vein endothelial cells (HUVEC). This study was designed to investigate the changes in vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and platelet endothelial cell adhesion molecule-1 (PECAM-1) expression with TNF-alpha stimulation in cultured human neonatal dermal lymphatic endothelial cells (HNDLEC). The real-time quantitative PCR analysis on HNDLEC showed that TNF-alpha treatment leads to increases of VCAM-1 and ICAM-1 mRNAs to the 10.8- and 48.2-fold levels of untreated cells and leads to a reduction of PECAM-1 mRNA to the 0.42-fold level of untreated cells. Western blot and immunohistochemical analysis showed that TNF-alpha leads to VCAM-1 and ICAM-1 expressions that were inhibited by antiserum to human TNF receptor or by AP-1 inhibitor nobiletin. In flow cytometry analysis, the number of VCAM-1- and ICAM-1-positive cells increased, and PECAM-1-positive cells decreased with TNF-alpha treatment. Regarding protein amounts produced in cells and amounts expressed on the cell surface, VCAM-1 and ICAM-1 increased in HNDLEC and HUVEC, and PECAM-1 decreased in HNDLEC in a TNF-alpha concentration-dependent manner. VCAM-1, ICAM-1, and PECAM-1 protein amounts in TNF-alpha-stimulated cells were lower in HNDLEC than in HUVEC. This suggests that the lymphatic endothelium has the TNF-alpha-induced signaling pathway, resulting in increased VCAM-1 and ICAM-1 expression to a weaker extent than blood endothelium and PECAM-1 reduction to a stronger extent than blood endothelium.     

Simvastatin modulates TNFalpha-induced adhesion molecules expression in human endothelial cells

Adhesion and transendothelial migration of leukocytes into the vascular wall is a crucial step in atherogenesis. Expression of cell adhesion molecules by endothelial cells plays a leading role in this process. We investigated the effect of simvastatin, an inhibitor of HMG-CoA reductase administered to reduce plasma levels of LDL-cholesterol, on the expression of vascular cell adhesion molecule-1 (VCAM-1) and intracellular cell adhesion molecule-1 (ICAM-1) by human umbilical vein endothelial cells (HUVEC) stimulated with tumor necrosis factor alpha (TNFalpha). We found the expression to be significantly inhibited by the drug in a time and concentration-dependent manner and to a greater extent in the case of VCAM-1 as compared with ICAM-1. In TNFalpha-stimulated HUVEC, simvastatin decreased VCAM-1 and ICAM-1 mRNA levels, inhibited TNFalpha-induced activation of nuclear factor kappaB (NF-kappaB) and enhanced expression of peroxisome proliferator-activated receptor alpha (PPARalpha). These effects were associated with reduction of adherence of monocytes and lymphocytes to HUVEC. The present findings suggest that the benefits of statins in vascular disease may include the inhibition of expression of VCAM-1 and ICAM-1 through effects on NF-kappaB.     

7-Ketocholesterol enhances the expression of adhesion molecules on human aortic endothelial cells by increasing the production of reactive oxygen species

Abstract

The aim of the present study was to assess the expression of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), monocytic adhesion of human aortic endothelial cells (HAECs), and the production of intracellular reactive oxygen species (ROS), when HAECs were stimulated by 7-ketocholesterol. 7-ketocholesterol enhances surface expression of ICAM-1 and VCAM-1 as determined by EIA, induces their mRNA expression by RT-PCR, and stimulates adhesiveness of HAECs to U937 monocytic cells. We confirmed up-regulation of ROS production of HAECs treated with 7-ketocholesterol. Although the surface expression of ICAM-1 and VCAM-1 on HAECs treated with 7-ketocholesterol increased in a time-dependent manner, α-tocopherol inhibited this increase of the surface expression of ICAM-1 and VCAM-1. In the monocytic adhesion assay, adhesion of U937 to HAECs treated with 7-ketocholesterol was enhanced, but monoclonal anti-ICAM-1 and VCAM-1 antibodies reduced the endothelial adhesiveness. In conclusion, this study suggests that the endothelial adhesiveness to monocytic cells that was increased by 7-ketocholesterol was associated with enhanced expression of ICAM-1 and VCAM-1 mediated by ROS production.     

Butyrate inhibits cytokine-induced VCAM-1 and ICAM-1 expression in cultured endothelial cells: the role of NF-kappaB and PPARalpha

Adhesion and migration of leukocytes into the surrounding tissues is a crucial step in inflammation, immunity, and atherogenesis. Expression of cell adhesion molecules by endothelial cells plays a leading role in this process. Butyrate, a natural short-chain fatty acid produced by bacterial fermentation of dietary fiber, has been attributed with anti-inflammatory activity in inflammatory bowel disease. Butyrate in vitro is active in colonocytes and several other cell types. We have studied the effect of butyrate on expression of endothelial leukocyte adhesion molecules by cytokine-stimulated human umbilical vein endothelial cells (HUVEC). Pretreatment of HUVEC with butyrate-inhibited tumor necrosis factor-alpha (TNFalpha)-induced expression of vascular cell adhesion molecule-1 (VCAM-1) and intracellular cell adhesion molecule-1 (ICAM-1) in a time and concentration-dependent manner. Butyrate at 10 mM/L inhibited interleukin-1 (IL-1)-stimulated VCAM-1 and ICAM-1 expression. The effect of butyrate on cytokine-stimulated VCAM-1 expression was more pronounced than in the case of ICAM-1. Butyrate decreased TNFalpha-induced expression of mRNA for VCAM-1 and ICAM-1. Suppressed expression of VCAM-1 and ICAM-1 was associated with reduced adherence of monocytes and lymphocytes to cytokine-stimulated HUVEC. Butyrate inhibited TNFalpha-induced activation of nuclear factor-kappaB (NF-kappaB) in HUVEC. Finally, butyrate enhanced peroxisome proliferator-activated receptor-alpha (PPARalpha) expression in HUVEC. These results demonstrate that butyrate may have anti-inflammatory properties not only in colonocytes but also in endothelial cells. The anti-inflammatory and (perhaps) antiatherogenic properties of butyrate may partly be attributed to an effect on activation of NF-kappaB and PPARalpha and to the associated expression of VCAM-1 and ICAM-1. The present findings support further investigations on the therapeutic benefits of butyrate in several pathological events involving leukocyte recruitment.     

Involvement of NAD(P)H oxidase in the enhanced expression of cell adhesion molecules in the aorta of diabetic mice

The increased levels of cell adhesion molecules (CAM) have been identified in diabetic vasculatures, but the underlying mechanisms remain unclear. To determine the relationship among vascular production of superoxide, expression of CAM and diabetes, superoxide generation and expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), E- and P-selectin in the aorta from control (C57BL/6J) and diabetic mice (ob/ob) were measured. In situ staining for superoxide using dihydroethidium showed an increased superoxide production in diabetic aorta in association with an enhanced NAD(P)H oxidase activity. Immunohistochemical analysis revealed that the endothelial expression of ICAM-1 (3.5 +/- 0.4) and VCAM-1 (3.8 +/- 0.3) in diabetic aorta was significantly higher than that in control aorta (0.9 +/- 0.5 and 1.6 +/- 0.3, respectively). Furthermore, there was a strong positive correlation (r = 0.89, p < 0.01 in ICAM-1 and r = 0.88, p < 0.01 in VCAM-1) between ICAM-1/VCAM-1 expression and vascular production of superoxide. The present data indicate that the increased production of superoxide via NAD(P)H oxidase may explain the enhanced expression of CAM in diabetic vasculatures.     

Murine Endothelial Cell Line Cells,F-2: Interaction with Leukocytes and Cytokines Production

Flowcytometry demonstrated that murine endothelial cell line F-2 expresses MHC class I antigen, FcR II, Mac-1 and vascular cell adhesion molecule-1 (VCAM-1), but not intercellular adhesion molecule-1 (ICAM-1) and class II antigen. However, co-culturing with TNF-α for 24 hr resulted in the increased expression of ICAM-1, and the decreased expression of VCAM-1. IL-1α and IFN-γ exerted this regulatory effect on VCAM-1 but not on ICAM-1. T (Con A blast) and B (LPS blast) cells adhered to F-2 cells at almost equal levels, and the adhesion was enhanced 20 to 50% when the cells were precultured with TNF-α for 24 hr. The inhibition assay using either (anti-ICAM-1 + anti-LFA-1, lymphocyte function-associated antigen-1) or (anti-VCAM-1 + anti-VLA-4, very late antigen-4) mAbs demonstrated that the ICAM-1 system was utilized more preferentially by T than B blasts when F-2 cells were stimulated with TNF-α, and the VCAM-1 system was vice versa under the unstimulated and stimulated conditions. Granulocytes also adhered to F-2 cells, but no mAbs could inhibit the adhesion. Although F-2 cells produced a considerable amount of IL-6, GM-CSF and neutrophil chemotactic activity, a 24 hr incubation with TNF-α resulted in an increase of 12 fold in IL-6 and 3 fold in neutrophil chemotactic activity production.     

Eosinophil adhesion to cholinergic nerves via ICAM-1 and VCAM-1 and associated eosinophil degranulation

In vivo, eosinophils localize to airway cholinergic nerves in antigen-challenged animals, and inhibition of this localization prevents antigen-induced hyperreactivity. In this study, the mechanism of eosinophil localization to nerves was investigated by examining adhesion molecule expression by cholinergic nerves. Immunohistochemical and functional studies demonstrated that primary cultures of parasympathetic nerves express vascular cell adhesion molecule-1 (VCAM-1) and after cytokine pretreatment with tumor necrosis factor-alpha and interferon-gamma intercellular adhesion molecule-1 (ICAM-1). Eosinophils adhere to these parasympathetic neurones after cytokine pretreatment via a CD11/18-dependent pathway. Immunohistochemistry and Western blotting showed that a human cholinergic nerve cell line (IMR-32) expressed VCAM-1 and ICAM-1. Inhibitory experiments using monoclonal blocking antibodies to ICAM-1, VCAM-1, or CD11/18 and with the very late antigen-4 peptide inhibitor ZD-7349 showed that eosinophils adhered to IMR-32 cells via these adhesion molecules. The protein kinase C signaling pathway is involved in this process as a specific inhibitor-attenuated adhesion. Eosinophil adhesion to IMR-32 cells was associated with the release of eosinophil peroxidase and leukotriene C(4). Thus eosinophils adhere to cholinergic nerves via specific adhesion molecules, and this leads to eosinophil activation and degranulation; this may be part of the mechanism of eosinophil-induced vagal hyperreactivity.