The Role Of PKCs In Propofol-induced Excessive Vasodilation In Spontaneously Hypertensive Rats

Objective: To verify the role of PKC?2 and PKC? in propofol-induced vasodilation in aortic smooth muscle and mesenteric artery smooth muscle in SHR.To further investigate the downstream mechanisms of PKC?2 and PKC? in propofol-induced vasodilation in arteries of SHR using primary cultured aortic VSMCs.Functional studies were designed to determine the role of PKC?2 and PKC? in etomidate,midazolam and dexmedetomidine-induced vasodilation of aortic rings and mesenteric artery rings in SHR.Methods: We isolated aortic smooth musle and cultured primary aortic VSMCs and used western blot to detect the protein expression of PKCs in WKY and SHR aortic smooth muscle and VSMCs.ELISA assay was used to compare the kinase activity of PKC?2 and PKC? in WKY and SHR aortic smooth muscle and VSMCs.The isolated endothelium-denuded aortic and mesenteric rings were mounted in the organ bath system to investigate the effect of LY333531 and PKC? pseudo-substrate inhibitor(PSS)on propofol-induced vasodilation of artery rings in WKY and SHR.Western blot was used to measure the effect of propofol on the phosphorylation level and translocation of PKC?2 and PKC? in WKY and SHR aortic smooth muscle and VSMCs.Besides,western blot was also used to determine the effects of propofol,LY333531 and PSS on calcium-sensitization signaling pathway(CPI-17,MYPT,MLC),F-actin content and F-actin/G-actin ratio in WKY and SHR VSMCs.Furthermore,we used lentivirus-mediated shRNA targeting PKC?2 and PKC? to detect the effect of propofol,LY333531 and PSS on calcium-sensitivity and F-actin/G-actin ratio in WKY and SHR VSMCs.The endothelium-denuded aortic andmesenteric artery rings were used to investigate the effect of LY333531 and PSS on etomidate,midazolam and dexmedetomidine-induced vasodilation of artery rings in WKY and SHR.Results: The protein expression levels,as well as the kinase activity of PKC?2 and PKC? in aortic smooth muscle and VSMCs in SHR VSMCs were significantly higher than those in WKY.LY333531 and PSS inhibited propofol-induced vasodilation of aortic and mesenteric artery rings in SHR but not in WKY.NE significantly increased the phophorylation level and translocation from cytoplasm to membrane of PKC?2and PKC? in WKY and SHR VSMCs.Propofol significantly inhibited the activation process of PKC?2 and PKC? induced by NE in SHR VSMCs but not in WKY VSMCs.NE significantly increased the phophorylation level of CPI-17,MYPT and MLC.Propofol and LY333531 both inhibited the phophorylation level of CPI-17,MYPT and MLC in SHR VSMCs remarkably.However,PSS did not influence calcium-sensitization.NE significantly increased F-actin content and F-actin/G-actin ratio in WKY and SHR VSMCs.Propofol,LY333531 and PSS significantly inhibited actin polymerization in SHR VSMCs.Propofol and transfection with shRNA(PKC?2)both significantly inhibited CPI-17,MYPT and MLC phophorylation in SHR VSMCs,while transfection with shRNA(PKC?)did not have any effect on calcium-sensitization.Propofol,transfection with shRNA(PKC?2)and transfection with shRNA(PKC?)all significantly inhibited F-actin/G-actin ratio in SHR VSMCs.LY333531 significantly inhibited etomidate,midazolam and dexmedetomidine-induced vasodilation in SHR mesenteric artery rings,but only significantly inhibited etomidate and midazolam-induced relaxation in SHR aortic rings.PSS significantly inhibited etomidate and midazolam-induced relaxation in SHR mesenteric artery rings,while PSS did not exert any influence on etomidate,midazolam and dexmedetomidine-induced vasodilation in SHR aorta.Conclusions: Propofol has a vasodilation effect on NE pre-contracted WKY and SHR aorta and mesenteric artery smooth muscle.The inhibitory effect of propofol on PKC?2-mediated calcium-sensitization pathway and actin polymerization is at least partially involved in the effect of the drug-induced vasodilation of SHR arterial smooth muscle.In addition,the vasodilatory effect of propofol is also related to its inhibition of PKC?-mediated actin polymerization.Intravenous anesthetics(etomidate,midazolam and dexmedetomidine)are able to cause relaxation of NE pre-contracted WKY and SHR aorta and mesenteric arteries.PKC?2 mediates the diastolic effect of etomidate,midazolam and dexmedetomidine on SHR mesenteric artery smooth muscle,and also mediates the diastolic effect of etomidate and midazolam on SHR aortic smooth muscle.However,PKC? only mediates the diastolic effect of etomidate and midazolam on SHR mesenteric artery smooth muscle.