Endothelin-1 stimulates cytosolic phospholipase A2 activity and gene expression in rat glomerular mesangial cells

Endothelin-1 (ET-1) stimulates vascular smooth muscle and mesangial cells to release prostaglandin E2 (PGE2), which can attenuate the vasoconstrictor and mitogenic effects of this peptide. Phospholipase A2 (PLA2)-mediated release of arachidonic acid from the sn-2 position of membrane phospholipids is thought to be one of the rate-limiting steps in prostaglandin (PG) synthesis. We evaluated the role of ET-1 to regulate gene expression, protein synthesis and enzymatic activity of cytosolic PLA2 (cPLA2), an intracellular form of the PLA2 enzyme family, in cultured rat mesangial cells using both acute and chronic incubation protocols. Acute ET-1-induced stimulation of cPLA2 activity was maximal after 10 minutes (181.1 +/- 6.84% of control), persisted for 40 minutes and did not require new protein synthesis. Heparin, a potent inhibitor of intracellular Ca2+ increase as well as mitogen-activated protein (MAP) kinase activation and cell proliferation, did not affect the rapid cPLA2 stimulation by ET-1. Chronic incubation of glomerular mesangial cells with ET-1 (1 to 24 hr) led to time- and dose-dependent increases in cPLA2 mRNA expression which was maximal after six hours, persisted up to 24 hours and which was accompanied by both cPLA2 protein formation, as assessed by Western analysis, as well as by stimulation of enzymatic activity. Inhibition of protein synthesis by cycloheximide increased basal cPLA2 mRNA accumulation in quiescent mesangial cells, and the combination of ET-1 and cycloheximide resulted in a greater induction of cPLA2 gene expression when compared to ET-1 alone. Actinomycin D treatment blocked the effect of ET-1 on cPLA2 mRNA accumulation.(ABSTRACT TRUNCATED AT 250 WORDS)