黏着斑激酶抑制剂Y15对中心静脉导管联合5-氟脲嘧啶诱导的细胞氧化损伤的作用机制研究  

Effect and mechanism of focal adhesion kinase inhibitor Y15 on the oxidative damage of EA.hy926 cell induced by central venous catheter combined with 5-fluorouracil

作　　者：王彦茹[1,2] 赵艳杰[1] 林素兰[1] 郑玉建[2] WANG Yan-ru;ZHAO Yan-jie;LIN Su-lan;ZHENG Yu-jian(Nursing School,Xinjiang Medical University,Urumqi,830011,Xinjiang Uyghur Autonomous Region,China;Public Health School,Xinjiang Medical University,Urumqi,830011,Xinjiang Uyghur Autonomous Region,China)

机构地区：[1]新疆医科大学护理学院,新疆乌鲁木齐830011 [2]新疆医科大学公共卫生学院,新疆乌鲁木齐830011

出　　处：《中国临床药理学杂志》2020年第17期2699-2702,共4页The Chinese Journal of Clinical Pharmacology

基　　金：新疆维吾尔自治区自然科学基金资助项目(2016D01C175);新疆教育厅研究生科研创新基金资助项目(XJGRI2016083);新疆乌鲁木齐市科学技术计划基金资助项目(H161318002);新疆医科大学研究生创新创业基金资助项目(CXCY050)。

摘　　要：目的研究抑制黏着斑激酶(FAK)-蛋白激酶B(Akt)信号通路活化在减轻中心静脉导管(CVC)联合5-氟脲嘧啶(5-FU)诱导的血管内皮细胞氧化性损伤中的作用及其机制。方法将EA.hy926细胞随机分为3组:正常组、模型组和实验组,每组3个时间点:24,48,72 h,每个时间点8个复孔。模型组(CVCs+划痕+5-FU),细胞划痕后,放入截好的CVC节段3个,再加入5-FU(40μg·mL^-1),建立EA.hy926细胞氧化损伤模型;实验组在模型组基础上加入FAK抑制剂(Y15,50μmol·L^-1)。以噻唑蓝比色法检测各时间点(24,48,72 h)细胞活性(OD值),以2,7-二乙酸二氯荧光素检测细胞活性氧(ROS)水平,硝酸还原酶法测定细胞培养液中一氧化氮(NO)含量,以蛋白质印迹法检测细胞中p-FAK Tyr 397和p-Akt Ser 473的表达水平(灰度值)。结果干预72 h后,正常组、模型组和实验组的细胞存活率分别为(94.80±3.27)%,(50.20±3.96)%和(71.00±2.92)%;这3组的ROS相对水平分别为(98.82±6.63)%,(469.83±19.60)%和(178.01±11.49)%;这3组的NO含量分别为(59.33±2.44),(22.10±2.57)和(43.39±2.58)μmol·L^-1;这3组的p-FAK Tyr 397相对表达水平为0.19±0.02,1.27±0.04和0.88±0.01;这3组的p-Akt Ser 473相对表达水平为0.14±0.02,1.17±0.04和1.05±0.09。上述指标:正常组与模型组比较、或实验组与模型组相比,差异均有统计学意义(P<0.01或P<0.001)。结论抑制黏着斑激酶-蛋白激酶B信号通路活化能够减轻CVC联合5-FU诱导的EA.hy926细胞氧化损伤。Objective To explore the role and mechanism of inhibition of focal adhesion kinase(FAK)-protein kinase B(Akt)signaling pathway activation in reducing oxidative injury of vascular endothelial cells induced by central venous catheter(CVC)combined with 5-fluorouracil(5-FU).Methods EA.hy926 cells were randomly divided into three groups:normal group,model group and experimental group,with 3 time points in each group:24,48,72 h,and 8 multiple wells at each time point.In the model group(CVCs+scratch+5-FU),after the cells were scratched,placed 3 CVC segments,and then added 5-FU(final concentration 40μg·mL^-1)to simulate the injury of vascular endothelial cells induced by administration of CVC after insertion and indwelling into veins,and establish the EA.hy 926 cell oxidative damage model;in the experimental group,basis on the model group,the FAK inhibitor(Y15)was added 50μmol·L^-1.MTT colorimetric method was used to detect cell activity(OD value)at each time point(24,48,72 h).The level of reactive oxygen species(ROS)was detected by 2,7-dichlorofluorescein diacetate.The content of nitric oxide(NO)in cell culture medium was determined by nitrate reductase method.The expression levels(gray value)of p-FAK Tyr 397 and p-Akt Ser 473 were detected by Western blot.Results After72 h of intervention,the cell survival rates in normal,model and experimental groups were(94.80±3.27)%,(50.20±3.96)%and(71.00±2.92)%,respectively;the relative levels of cellular ROS in the 3 groups were(98.82±6.63)%,(469.83±19.60)%and(178.01±11.49)%,respectively;the NO contents in the 3 groups were(59.33±2.44),(22.10±2.57)and(43.39±2.58)μmol·L^-1,respectively;the relative expression levels of p-FAK Tyr 397 in the 3 groups were 0.19±0.02,1.27±0.04 and 0.88±0.01,respectively;the relative expression levels of p-Akt Ser 473 in the 3 groups were 0.14±0.02,1.17±0.04 and 1.05±0.09,respectively.Comparison between model group and normal group,or between experimental group and model group,the difference of the factors were significant(P<0.01 or

关 键 词：黏着斑激酶抑制剂 中心静脉导管 5-氟脲嘧啶 细胞氧化损伤 黏着斑激酶-蛋白激酶B信号通路 

分 类 号：R97[医药卫生—药品]