Oxidized low-density lipoprotein downregulates endothelial basic fibroblast growth factor through a pertussis toxin-sensitive G-protein pathway - Mediator role of platelet-activating factor-like phospholipids

Background-Oxidized LDL (oxLDL) inhibits angiogenesis in part by downregula ting endothelial basic fibroblast growth factor (bFGF). To determine the me chanism of the downregulation, we investigated the signal transduction path way involving potential phospholipid mediators. Methods and Results-Cultured bovine aortic endothelial cells were incubated with PBS (lipoprotein-free control), LDL, or copper oxLDL under serum-free conditions. At 24 hours, oxLDL (50 mug/mL) decreased bFGF mRNA (Northern b lot), bFGF protein (Western blot and ELISA), and concomitant DNA synthesis, all by 40% to 50% compared with PBS. LDL had no effect. Pretreating the ce lls with 100 ng/mL pertussis toxin (PTX) for 18 hours before oxLDL exposure almost completely blocked the inhibitory effects of oxLDL. In contrast, in hibiting other major cellular signal transduction pathways with PD-98059 (m itogen-activated protein kinase kinase inhibitor), HA-1004 (inhibitor of cG MP- and cAMP-dependent protein kinase), or Ro-31-8220 (protein kinase C inh ibitor) or chelating intracellular Ca2+ with BAPTA-AM failed to attenuate a ny of the oxLDL effects assayed. Addition to the cultures of WEB 2086, a sp ecific antagonist of the PTX-sensitive. G protein-coupled platelet-activati ng factor (PA-F) receptor, blocked the action of oxLDL. Whereas PAY dispers ed in the culture medium failed to produce oxLDL-like effects, degradation of PAF and PAP-like phospholipids accumulated in oxLDL with a recombinant h uman PAY acetylhydrolase eliminated the inhibitory effects of oxLDL on bFGF expression and DNA synthesis. Conclusions-OxLDL suppresses endothelial bFGF expression and DNA synthesis through a PTX-sensitive heterotrimeric G-protein pathway involving mediator phospholipids similar, but not identical, to PAF.