Endotoxin impairs agonist-induced calcium mobilization in rat mesangial cells.
We hypothesized that endotoxin would impair agonist-induced calcium (Ca2+) mobilization in rat mesangial cells, owing to the induction of nitric oxide synthase (NOS) and augmented nitric oxide (NO) synthesis. We measured basal and bradykinin-induced peak free cytosolic Ca2+ concentrations through microspectrofluorimetry with fura-2 in confluent mesangial cells, and assayed conditioned medium for nitrite accumulation. Prior to measurement, cells were incubated overnight in serum-supplemented medium, with or without endotoxin, 1-arginine, indomethacin, meclofenamate, or N omega-nitro-L-arginine methyl ester (L-NAME). Endotoxin (1 mg/ml) decreased bradykinin-induced peak Ca2+ responses by 35 to 60% (p < 0.0001) and increased nitrite accumulation > 6-fold (p < 0.01). Arginine supplementation further (> 9-fold, p < 0.0001) increased nitrite accumulation without changing the effect on Ca2+. Inhibition of NOS abolished increments in nitrite concentration but had no effect on impaired Ca2+ responses. Cyclooxygenase (COX) inhibitors, present during incubation with endotoxin, but not afterward, normalized bradykinin-stimulated calcium responses. Thrombin-stimulated Ca2+ responses were similarly affected. We conclude that neither NO nor prostaglandins act directly to impair agonist-induced Ca2+ mobilization following endotoxin exposure; however, this effect may be an indirect effect of COX products, including reactive oxygen intermediates.
Murray
P T
PT
Department of Anesthesia, University of Chicago, Illinois, USA.
Wylam
M E
ME
Umans
J G
JG
D000818:Animals; D001120:Arginine / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D002470:Cell Survival; D002478:Cells, Cultured; D017077:Culture Media, Conditioned; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004731:Endotoxins / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D007213:Indomethacin / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008469:Meclofenamic Acid / Q000494:pharmacology; D015591:Microspectrophotometry; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000502:physiology; D009573:Nitrites / Q000378:metabolism; D051381:Rats; D013917:Thrombin / Q000494:pharmacology
D017077:Culture Media, Conditioned; D016861:Cyclooxygenase Inhibitors; D004731:Endotoxins; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D009573:Nitrites; D009569:Nitric Oxide; D008469:Meclofenamic Acid; D001120:Arginine; D019001:Nitric Oxide Synthase; D013917:Thrombin; D001920:Bradykinin; D019331:NG-Nitroarginine Methyl Ester; D007213:Indomethacin