CYTOSOLIC 85-KDA PHOSPHOLIPASE A(2)-MEDIATED RELEASE OF ARACHIDONIC-ACID IS CRITICAL FOR PROLIFERATION OF VASCULAR SMOOTH-MUSCLE CELLS

Recent evidence suggests that arachidonic acid (AA) may be involved in regulating cellular proliferation, The predominant mechanism of AA re lease from cellular phospholipids is via phospholipase A(2) (PLA(2)) h ydrolysis, The purpose of this study was to examine the roles of the d istinct 14-kDa and 85-kDa PLA(2) enzymes in human coronary artery vasc ular smooth muscle cell (hCAVSMC) proliferation, Cultured hCAVSMCs pro liferate in the presence of growth medium with a typical doubling time of 30-40 h, grow at a slower proliferative rate upon reaching conflue ncy (day 8), and eventually undergo contact inhibition of growth (day 10), Neither Type II 14-kDa PLA(2) activity nor mass changed over a 10 -day culture period, In contrast, 85-kDa PLA(2) protein activity and m RNA decreased as time in culture progressed. This reduction in 85-kDa PLA(2) correlated with reductions in DNA synthesis and suggested a pos sible association between 85-kDa PLA(2) and proliferation, To directly evaluate the role of the 85-kDa PLA(2) in proliferation we examined t he effects of an 85-kDa PLA(2) inhibitor (AACOCF(3)) and 85-kDa PLA(2) antisense oligonucleotides on proliferation. Both reagents dose depen dently inhibited proliferation, whereas a 14-kDa PLA(2) inhibitor (SB2 03347), a calcium-independent PLA(2) inhibitor (HELSS), all 85-kDa sen se oligonucleotide, and a nonrelevant scrambled control oligonucleotid e had no effect, The mechanism by which 85-kDa PLA(2) influences cellu lar proliferation remains unclear. Inhibition of 85-kDa PLA(2) activit y produced neither phase-specific cell cycle arrest nor apoptosis (flu orescence-activated cell sorter analysis). Addition of AA (20 mu M) at tenuated the effects of both AACOCF(3) and 85-kDa antisense oligonucle otides implicating AA as a key mediator in cellular proliferation Howe ver, although prostaglandin E-2 (PGE(2)) was present in the culture me dium, it peaked early (day 3) in culture, and indomethacin had no effe ct on cellular proliferation indicating that hCAVSMC proliferation was not mediated through PGE(2). These data provide the first direct evid ence that PLA(2) is involved in control of VSMC proliferation and indi cate that 85-kDa PLA(2)-mediated liberation of AA is critical for cell ular proliferation.