Cilostazol suppresses adhesion of human neutrophils to HUVECs stimulated by FMLP and its mechanisms.
The interaction between neutrophils and endothelial cells (ECs) is of great importance in many physiological and pathological progresses. Although cilostazol (CLZ), a novel selective phosphodiesterase (PDE) type 3 inhibitor, has been proved to be useful in vasodilatation and inhibition of platelet aggregation, its effect on adhesion is not clearly known. In this study, we examined the effects and investigated the mechanisms of cilostazol on neutrophil adhesion to human umbilical endothelial cells (HUVECs) triggered by N-formyl-methionyl-leucyl-phenylal-anine (FMLP), a chemotactic peptide. The soluble vascular cell adhesive molecule-1 (sVCAM-1) release from FMLP (10 microM)-stimulated HUVECs was determined by ELISA kits. Fluo-2, a fluorescent indicator, was used to investigate intracellular free calcium concentration ([Ca2+]i) in HUVECs. HL-60 cells were induced to be neutrophilic by DMSO and loaded with Fluo-3, another fluorescent indicator, to detect [Ca2+]i, and CLA was used as a chemiluminescent indicator to determine superoxide production in neutrophilic cells. The result showed that CLZ (1-100 microM) significantly inhibited neutrophil adhesion to FMLP-stimulated HUVECs. In HUVECs, CLZ obviously downregulated sVCAM-1 level, while it had no meaningful influence [Ca2)]i. But in neutrophils, FMLP-activated superoxide generation and [Ca2+]i increase were found being inhibited by exposure to CLZ . Furthermore, we also demonstrated that Ca2+ increase was preceded to the superoxide generation in neutrophils. The results suggest that CLZ involves in adhesion reactions between neutrophil and ECs, partly via VCAM-1 expression in ECs, and decreasing [Ca2+]i induced activation of neutrophils, which means a lot to prevent atherosclerosis and other cardiovascular diseases.
Yang
Yingbao
Y
Department of Clinical Pharmacology and Therapeutics, Hamamatsu University School of Medicine, Hamamatsu 431-3192, Japan.
Luo
Jinghui
J
Kazumura
Kimiko
K
Takeuchi
Kazuhiko
K
Inui
Naoki
N
Hayashi
Hideharu
H
Ohashi
Kyoichi
K
Watanabe
Hiroshi
H
D000328:Adult; D002118:Calcium / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects; D002454:Cell Differentiation; D002469:Cell Separation; D002478:Cells, Cultured; D000077407:Cilostazol; D042783:Endothelial Cells / Q000187:drug effects*; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D018922:HL-60 Cells; D006801:Humans; D066298:In Vitro Techniques; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects*; D010727:Phosphoric Diester Hydrolases / Q000494:pharmacology*; D013268:Stimulation, Chemical; D013481:Superoxides / Q000378:metabolism; D013777:Tetrazoles / Q000494:pharmacology*; D014470:Umbilical Cord / Q000166:cytology; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
D013777:Tetrazoles; D019010:Vascular Cell Adhesion Molecule-1; D013481:Superoxides; D009240:N-Formylmethionine Leucyl-Phenylalanine; D010727:Phosphoric Diester Hydrolases; D000077407:Cilostazol; D002118:Calcium