低氧损伤下调大鼠脑血管内皮细胞中硫化氢及其介导的Rho A-ROCK通路激活  

Anoxic injury down-regulates hydrogen sulfide in rat cerebrovascular endothelial cells and H_2S-mediated activation of RhoA-ROCK pathway

作　　者：王良芳 陈志武[1] Wang Liangfang;Chen Zhiwu(Dept of Pharmacology,Anhui Medical University,Hefei 230032)

机构地区：[1]安徽医科大学药理学教研室,合肥230032

出　　处：《安徽医科大学学报》2019年第1期50-55,共6页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81374002)

摘　　要：目的观察大鼠脑血管内皮细胞中内皮源性硫化氢(H_2S)、一氧化氮(NO)及Ras同源基因家族成员A-Rho相关的卷曲螺旋激酶(RhoA-ROCK)信号通路在低氧性损伤中的动态变化,并探讨内皮源性H_2S对RhoA-ROCK通路的影响。方法胶原酶消化法原代培养大鼠脑血管内皮细胞,内皮细胞分别低氧培养1、2、4、8、24 h后,测定H_2S和NO含量,G-LISA检测RhoA活性,细胞裂解后对相关蛋白进行免疫印迹分析。结果低氧1 h后H_2S含量显著下降; NO含量在低氧4 h后开始显著下降; RhoA活性低氧8 h后才有显著增加。低氧4 h时内源性H_2S合成酶胱硫醚γ-裂解酶(CSE)表达就有了显著下降;低氧8 h后内皮型一氧化氮合成酶(e NOS)才开始明显降低;低氧8 h时Rho激酶ROCK1、ROCK2表达显著增加。内源性和外源性H_2S均可抑制RhoA的激活。结论在大鼠脑血管内皮细胞低氧损伤过程中,内皮源性H_2S降低发生最早,其次为NO,而RhoA-ROCK通路激活发生在后,可能是继发于H_2S的降低。Objective To observe the effect of different hypoxic time on hydrogen sulfide( H2S),nitric oxide( NO)and Ras homolog gene family,member A/Rho associated coiled coil-forming kinase( RhoA-ROCK) pathway in rat cerebrovascular endothelial cells( EC),and investigate the effect of dermatogenous H2S on the RhoA-ROCK pathway. Methods Rat brain vascular EC was cultured by collagenase digestion. The EC was measured for H2S and NO after hypoxia for 1,2,4,8 and 24 h respectively. G-LISA was used to detect RhoA activity. Proteins expression changes were detected by Western blot. Results After 1 hour of hypoxia,the content of H2S decreased significantly,the NO content decreased significantly after hypoxia of 4 hours,the activity of RhoA increased significantly after hypoxia of 8 h. The expression of CSE protein decreased significantly after 4 h of hypoxia,the expression level of eNOS protein decreased significantly after 8 h of hypoxia,and the expression of ROCK1 and ROCK2 increased significantly at 8 h of hypoxia. Both endogenous and exogenous H2S inhibited RhoA activity. Conclusion During the hypoxic injury of rat cerebrovascular endothelial cells. The decrease of endogenous H2S occurred first,followed by NO,and the activation of RhoA-ROCK pathway occurred later,which may be secondary to the decrease of H2S.

关 键 词：内皮细胞 低氧 硫化氢 RhoA-ROCK通路 

分 类 号：R965[医药卫生—药理学]