Novel and Differential Roles of Ca 2+ ‐Independent Phospholipase A 2 Isoforms in Renal Cell Growth and Phospholipid Metabolism

Abstract

Recent studies demonstrate that human renal cells express both cytosolic (iPLA2β) and microsomal (iPLA2γ) Ca2+-independent phospholipase A2 (iPLA2). However, the roles of these iPLA2 in renal cell physiology are not fully understood. The effect of R-bromonenol lactone (R-BEL, iPLA2γ inhibitor) and S-BEL (iPLA2β inhibitor) on cell growth was assessed in HEK293 and Caki-1 (human renal epithelial) cells using 3-(4, dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide (MTT) staining. Treatment of cells with S-BEL (0-5.0 μM) decreased MTT staining 35% after 48 hr. In contrast, R-BEL (0–5.0 μM) only decreased MTT staining 15%, compared to control cells. Transfection of cells with iPLA2β siRNA reduced MTT staining 35%, while iPLA2γ siRNA only decreased MTT staining 10–15%. siRNA against iPLA2β also resulted in 25% decreases in cell number and protein, while siRNA against iPLA2γ only slightly decreased cell number, and did not alter cellular protein. To determine the roles of iPLA2β and γ in phospholipid metabolism the effect of R- and S-BEL on phospholipid profiles was determined using electrospray ionization-mass spectrometry. Treatment of cells with R-BEL alone decreased the expression of 14:0–16:0 phosphatidylcholine (PtdCho), 16:0–20:4 PtdCho and increased the expression of 16:0–20:1 PtdCho. In comparison, treatment of cells with S-BEL increased the expression of 16:1–20:4 PtdCho and 20:1–22:4 plasmenylcholine. These data demonstrate a novel role for iPLA2β and γ in renal cell growth and suggest that these isoforms have differential roles in phospholipid metabolism. Supported by a Georgia Cancer Coalition Distinguished Scholar Award.