Very low density lipoprotein-mediated signal transduction and plasminogen activator inhibitor type 1 in cultured HepG2 cells

In normal subjects and in patients with cardiovascular disease, plasma trig lycerides are positively correlated with plasminogen activator inhibitor ty pe 1 (PAI-1) levels, Moreover, in vitro studies indicate that VLDLs induce PAI-1 synthesis in cultured cells, ie, endothelial and HepG2 cells. However , the signaling pathways involved in the effect of VLDL on PAI-1 synthesis have not yet been investigated. We report that VLDLs induce a signaling cas cade that leads to an enhanced secretion of PAI-1 by HepG2 cells. In myo-[H -3]inositol-labeled HepG2 cells, VLDL (100 mu g/mL) caused a time-dependent increase in [H-3]inositol phosphates, the temporal sequence being tris>bis >monophosphate. VLDL brought about a time-dependent stimulation of membrane -associated protein kinase C (PKC) activity and arachidonate release. Final ly, VLDL stimulated mitogen-activated protein (MAP) kinase, and this effect was reduced by 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H7), which suggests that PKC plays a pivotal role in MAP kinase phosphorylation. VLDL- induced PAI-1 secretion was completely prevented by U73122, a specific inhi bitor of phosphatidylinositol-specific phospholipase C, by H7 or by PKC dow nregulation, and by mepacrine (all P<0.01 versus VLDL-treated cells), 3,4,5 -Trimethoxybenzoic acid 8-(diethylamino)-octyl ester, which prevents Ca2+ r elease from intracellular stores, inhibited VLDL-induced PAI-1 secretion by 60% (P<0.05), and the MAP kinase/extracellular signal-regulated kinase kin ase (MEK) inhibitor PD98059 completely suppressed both basal and VLDL-induc ed PAI-1 secretion. These data demonstrate that VLDL-induced PAI-1 biosynth esis results from a principal signaling pathway involving PKC-mediated MAP kinase activation.