水通道蛋白1在切应力调节血管内皮细胞迁移和血管生成中的作用及机制  

Role of aquaporin 1 in shear stress-regulated endothelial cell migration and angiogenesis

作　　者：张敏 孙玉 杜大鹏 王小波 于建新 王汉琴 ZHANG Min;SUN Yu;Du Da-peng;WANG Xiao-bo;YU Jian-xin;WANG Hanqin(Center for Translational Medicine,Suizhou Hospital,Hubei University of Medicine,Suizhou 441300,China;Department of Anatomy,Basic Medical College,Hubei University of Medicine,Shiyan 442000,China)

机构地区：[1]湖北医药学院附属随州医院转化医学研究中心,湖北随州441300 [2]湖北医药学院基础医学院解剖学教研室,湖北十堰442000

出　　处：《中国病理生理杂志》2022年第3期403-411,共9页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.31670961);湖北医药学院研究生科技创新项目(No.YC2019038)。

摘　　要：目的:探讨流体切应力作用下水通道蛋白1(AQP1)的表达对血管内皮细胞迁移和血管生成的影响及可能的机制。方法:取雄性C57BL/6小鼠主动脉弓和胸主动脉血管壁组织,用qPCR和Western blot检测体内不同切应力作用部位血管壁AQP1表达的差异。原代培养人脐静脉内皮细胞(HUVECs),体外采用平行平板流动腔系统给HUVECs分别加载层流(LF,15 dyn/cm^(2)单向层切应力)和扰流[DF,(0.5±4)dyn/cm^(2)振荡切应力],用特异性小干扰RNA(siRNA)转染技术沉默AQP1基因,采用Transwell实验和Matrigel小管形成实验检测HUVECs迁移和血管生成能力;用Western blot检测内皮型一氧化氮合酶(eNOS)Ser1177和Ser633磷酸化水平。结果:在体胸主动脉中AQP1的mRNA和蛋白表达显著高于主动脉弓(P<0.05)。HUVECs静态下敲减AQP1可以显著抑制细胞迁移和血管生成(P<0.01)。与DF组相比,LF显著上调HUVECs中AQP1的mRNA和蛋白表达(P<0.05),促进HUVECs迁移(P<0.01)和血管生成能力(P<0.05),同时显著增加eNOS Ser1177(P<0.01)和Ser633(P<0.05)磷酸化水平。转染siAQP1后,LF诱导的AQP1表达增强被抑制(P<0.01),HUVECs的迁移和血管生成能力也随之降低(P<0.01),同时eNOS Ser1177和Ser633的磷酸化水平降低(P<0.05或P<0.01)。eNOS抑制剂N^(G)-硝基-L-精氨酸甲酯(L-NAME)预处理HUVECs后,LF诱导的细胞迁移和血管生成能力被抑制(P<0.01)。结论:AQP1在流体切应力调节血管内皮细胞迁移和血管生成中发挥作用,其机制可能和eNOS信号有关。AIM:To investigate the role of aquaporin 1(AQP1)in shear stress-regulated migration and angiogenesis of endothelial cells,and to explore the underlying mechanisms.METHODS:The aortic arches and straight segments of thoracic aortas were obtained from C57BL/6 mice.Human umbilical vein endothelial cells(HUVECs)were isolated from fresh human umbilical cord.The parallel plate flow chamber system was used to load unidirectional laminar flow(LF,15 dyn/cm^(2) laminar shear stress)and disturbed flow[DF,(0.5±4)dyn/cm^(2) oscillatory shear stress]on HUVECs.qPCR and Western blot were used to detect the expression of AQP1 at mRNA and protein levels,respectively.Silencing of AQP1 was performed by AQP1-specific siRNA.Assays of Transwell and tube formation in Matrigel were performed to detect cell migration and angiogenesis.The protein levels of endothelial nitric oxide synthase(eNOS)and phosphorylated eNOS at Ser633 and Ser1177 were measured by Western blot.RESULTS:The mRNA and protein expression of AQP1was up-regulated in the straight thoracic aorta compared with the inner of the aortic arch(P<0.05).Knockdown of AQP1in HUVECs at static condition markedly reduced cell migration and angiogenesis(P<0.01).Exposure of HUVECs to LF for8 h obviously induced AQP1 expression and promoted HUVEC migration and angiogenesis(P<0.05 or P<0.01).LF also significantly increased eNOS phosphorylation both at Ser1177(P<0.05)and Ser633(P<0.01).As expected,AQP1 silencing abolished the above effects of LF-stimulated HUVECs(P<0.05 or P<0.01).N^(G)-nitro-L-arginine methyl ester(L-NAME),an eNOS inhibitor,also prevented LF-induced HUVEC migration and angiogenesis(P<0.01).CONCLU-SION:AQP1 plays a crucial role in the regulation of migration and angiogenesis in shear stress-stimulated endothelial cells,and its mechanism may be related to the eNOS signaling pathway.

关 键 词：水通道蛋白1 切应力 血管生成 血管内皮细胞 内皮型一氧化氮合酶 

分 类 号：R363.2[医药卫生—病理学] R329.25[医药卫生—基础医学]