The Role Of PKCs In Propofol-induced ENOS Activation In Umbilical Vein Vascular Endothelial Cells

Obiective：The purpose of this study is to investigate the role of PKCs in medatingpropofol-induced eNOS phosphorylation and subsequent NO production in humanumbilical venous endothelial cells (HUVECs).Methods：Cell within passages2to5were used in all experiments. To determinewether propofol has an effect on eNOS activation, HUVECs were treated with propofol(1-100μmol/L) for up to24hrs, and phosphorylation of eNOS at serine1177was analyzedby immunoblot assay. To investigate the possible role of phosphatidylinositol3-kinase(PI3K) and protein kinase C (PKC) in propofol-induced eNOS phosphorylation, cultureswere preincubated with LY294002(10μmol/L), an inhibitor of The PI3K, orbisindolylmaleimide I(1μmol/L), an inhibitor of PKCs, and the phosphorylation of eNOSto propofol (1-hour and24-hour incubation at5and50μmol/L, respectively) was assessed.To distinguish the involvement of different PKC isoforms (PKC-а, PKC-δ, PKC-ε, PKC-θand PKC-ξ) in propofol activation of eNOS, the individual PKC isoform-specific inhibitors,siRNA or Adv-caPKC was used, and the phosphorylation of eNOS following treatmentwith propofol was also assessed as above.Results: Propofol increased the expression of P-Ser1177-eNOS in bothconcentrations-and time-dependent manners. The PI3K inhibitor, LY294002, did notaffect eNOS phosphorylation by propofol. However, the PKC inhibitor,bisindolylmaleimide I, significantly blocked propofol-induced eNOS activation.Furthermore, selective inhibition or overexpression of these PKC isoforms by theirinhibitors, siRNA or Adv-caPKC differentially regulated propofol-induced eNOSactivation, which was enhanced by PKC-а, PKC-εand PKC-θ, and inhibited by PKC-δ, respectively.Conclusion：Propofol induces the activation of eNOS in HUVECs in concentration-and time-dependent manners. This activation is independent of the PI3K/Akt signalingpathway. PKC isoforms defferentially modulate propofol-induced eNOS phosphorylation,with an enhanceing effect by PKC-а, PKC-ε and PKC-θ, while an inhibiting action byPKC-δ...