Studies of the role of group VI phospholipase A(2) in fatty acid incorporation, phospholipid remodeling, lysophosphatidylcholine generation, and secretagogue-inducecd arachidonic acid release in pancreatic islets and insulinoma cells

An 84-kDa group VI phospholipase A(2) (iPLA(2)) that does not require Ca2for catalysis has been cloned from Chinese hamster ovary cells, murine P388 D1 cells, and pancreatic islet beta-cells, A housekeeping role for iPLA(2) in generating lysophosphatidylcholine (LPC) accepters for arachidonic acid incorporation into phosphatidylcholine (PC) has been proposed because iPLA( 2) inhibition reduces LPC levels and suppresses arachidonate incorporation and phospholipid remodeling in P388D1 cells. Because islet beta-cell phosph olipids are enriched in arachidonate, we have examined the role of iPLA(2) in arachidonate incorporation into islets and INS-1 insulinoma cells. Inhib ition of iPLA(2) with a bromoenol lactone (BEL) suicide substrate did not s uppress and generally enhanced [H-3]arachidonate incorporation into these c ells in the presence or absence of extracellular calcium at varied time poi nts and EEL concentrations. Arachidonate incorporation into islet phospholi pids involved deacylation-reacylation and not de novo synthesis, as indicat ed by experiments with varied extracellular glucose concentrations and by e xamining [C-14]glucose incorporation into phospholipids. EEL also inhibited islet cytosolic phosphatidate phosphohydrolase (PAPH), but the PAPH inhibi tor propranolol did not affect arachidonate incorporation into islet or INS -1 cell phospholipids. Inhibition of islet iPLA(2) did not alter the phosph olipid head-group classes into which [H-3]arachidonate was initially incorp orated or its subsequent transfer from PC to other lipids. Electrospray ion ization mass spectrometric measurements indicated that inhibition of INS-1 cell iPLA(2) accelerated arachidonate incorporation into PC and that inhibi tion of islet iPLA(2) reduced LPC levels by 25%, suggesting that LPC mass d oes not limit arachidonate incorporation into islet PC. Gas chromatography/ mass spectrometry measurements indicated that EEL but not propranolol suppr essed insulin secretagogue-induced hydrolysis of arachidonate from islet ph ospholipids. In islets and INS-1 cells, iPLA(2) is thus not required for ar achidonate incorporation or phospholipid remodeling and may play other role s in these cells.