SUPEROXIDE RESPONSES OF ENDOTHELIAL-CELLS TO C5A AND TNF-ALPHA - DIVERGENT SIGNAL TRANSDUCTION PATHWAYS

There is increasing evidence that endothelial cells respond to a variety of mediators. In the current studies rat pulmonary artery endothelial cells (RPAEC) responded to human recombinant C5a and tumor necrosis factor-alpha (TNF-alpha) with the generation of superoxide (O2(-)). RPAEC responsiveness was dependent on whether cells had been obtained from confluent or subconfluent cell monolayers. RPAEC responded to C5a and TNF-alpha in a dose-dependent manner, with increases in intracellular Ca2+ (Ca(i)2+), formation of D-myo-inositol 1,4,5-trisphosphate (Ins(1,4,5)P3], and generation of O2(-). Optimal O2(-) responses occurred in cells that had been pretreated with the inhibitor of superoxide dismutase (SOD), diethyldithiocarbamate, and O2(-) responses were allopurinol insensitive. Pertussis toxin pretreatment abolished the ability of C5a to cause increases in Ins(1,4,5)P3 and Ca(i)2+ and formation of O2(-) but did not inhibit the changes in Ca(i)2+ and formation of O2(-) after addition of TNF-alpha. The O2(-) response to C5a but not to TNF-alpha was abolished by pretreatment with the inhibitor of protein kinase C, staurosporine. These data indicate that signal transduction events in response to C5a and TNF-alpha were fundamentally different.