title: Possible involvement of a chloride-bicarbonate exchanger in apoptosis of endothelial cells and cardiomyocytes.
abstract: We examined the role of ion movement in staurosporine-induced apoptosis of vascular endothelial cells. Cultured vascular endothelial cells from bovine carotid arteries were used. Apoptosis was determined by propidium iodide assay. Treatment of the endothelial cells with staurosporine (10 nmol/l-1 micromol/l) for 6 h induced nuclear fragmentation in a dose-dependent manner. Staurosporine (1 micromol/l) elicited apoptosis in 70.5+/-1.5% of cells. Concomitant treatment of endothelial cells with 1 mmol/l of 4, 4-diisothiocyanatostilbene-2,2-disulfonic acid (DIDS), a chloride-bicarbonate exchange blocker, completely inhibited staurosporine-induced apoptosis. Other ion transporter inhibitors such as dimethyl amiloride and anthracene-9 carboxylic acid were less effective inhibitors of staurosporine-induced apoptosis of endothelial cells. DIDS prevented staurosporine-induced apoptosis of endothelial cells as well as cardiomyocytes. Next, we determined whether chloride ions or bicarbonate are involved in apoptosis. Incubation with a chloride ion removal buffer did not inhibit staurosporine-induced apoptosis of endothelial cells. However, endothelial cell apoptosis was completely suppressed by an inhibitor of caspase, benzyloxycarbonyl-Asp-CH(2)-O(C)O-dichlorobenzene (zD-dcb, 50 micromol/l). Staurosporine (1 micromol/l) increased the intracellular pH of endothelial cells, and DIDS (1 mmol/l), but not a caspase inhibitor, inhibited this increase in pH caused by staurosporine. Our findings suggest that endothelial cell apoptosis induced by staurosporine may be associated with the Cl(-)and bicarbonate (HCO-3) ions. Thus, Cl(-)efflux from cells or HCO-3 influx to cells (which increases pH) may play an important role in signal transduction leading events such as activation of caspase in staurosporine-induced apoptosis.
authors.affiliation: Second Department of Internal Medicine, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka, Tokyo, 181, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Fujita
authors.forename: Y
authors.initials: Y
authors.lastname: Ishizaki
authors.forename: A
authors.initials: A
authors.lastname: Yanagisawa
authors.forename: I
authors.initials: I
authors.lastname: Morita
authors.forename: S I
authors.initials: SI
authors.lastname: Murota
authors.forename: K
authors.initials: K
authors.lastname: Ishikawa
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D017920:Antiporters / Q000037:antagonists & inhibitors / Q000502:physiology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001639:Bicarbonates / Q000378:metabolism; D002339:Carotid Arteries / Q000166:cytology; D020169:Caspases / Q000502:physiology; D002417:Cattle; D002478:Cells, Cultured; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D004791:Enzyme Inhibitors; D005260:Female; D005333:Fetus; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D009206:Myocardium / Q000166:cytology*; D011247:Pregnancy; D047428:Protein Kinase Inhibitors; D051381:Rats; D017208:Rats, Wistar; D019311:Staurosporine / Q000378:metabolism / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors
chemical_list: D017920:Antiporters; D001639:Bicarbonates; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D004791:Enzyme Inhibitors; D047428:Protein Kinase Inhibitors; D014409:Tumor Necrosis Factor-alpha; D006861:Hydrogen Peroxide; D020169:Caspases; D019311:Staurosporine; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid