Progesterone Promotes Endothelial NO Synthase Expression And Vascular Relaxation Through Enhancing Profesterone Receptor-SP1 Formation

Objective: The present study aimed to investigate the effect of progesterone on the expression of endothelial nitric oxide synthase(eNOS)and vasodilatation function and further explore the related genetic mechanism in Human umbilical vein endothelial cells(HUVECs).Methods: In vivo,progesterone receptor antagonist RU486 was intraperitoneally injected into SD rats to detect eNOS expression and vasodilation in thoracic aorta and mesenteric artery.In vitro,the eNOS?transcription factor SP1(Specificity protein 1)and PR(Progesterone receptor)expression was detected after HUVECs(Human umbilical vein endothelial cell)treated with progesterone.The transcription mechanism was further explored by CHIP,IP and Luciferase assay.Results: RU486 decreased the protein and mRNA levels of eNOS and impaired vasodilation in thoracic aorta and mesenteric artery in female rats.In male rats,RU486 reduced the protein and mRNA expression of eNOS in mesenteric artery and damaged its diastolic function,but had nosignificant effect on the expression of eNOS and diastolic function in thoracic aorta.In vitro,progesterone increased eNOS protein and mRNA expression in HUVECs in a concentration-and time-dependent manner.In vitro,treatment with progesterone increased the expression of eNOS,and the transcription factors SP1,but no affected the expression of nuclear progesterone receptor(PR-A/B)in HUVECs.Interestingly,immunoprecipitation assay(IP)illustrated that progesterone increased the formation of PR-A-SP1 complex.In contrast,blockade of PR activity abolished this effect.Chromatin immunoprecipitation assay(CHIP)showed that progesterone increased PR-A/B and SP1 binding to eNOS promoter.Mutation of the SP1 or PR binding motif on the eNOS promoter completely abolished progesterone-induced increasing of eNOS promoter activity.Conclusion: Progesterone activated eNOS transcription and up-regulated eNOS protein expression by increasing the formation of PR-A-SP1 complex,thereby improving endothelial relaxation.