重组脂联素对氧化损伤下内皮细胞内质网应激相关信号分子的影响

发布时间：2018-01-07 16:00

本文关键词：重组脂联素对氧化损伤下内皮细胞内质网应激相关信号分子的影响　出处：《福建医科大学》2010年硕士论文　论文类型：学位论文

【摘要】： 目的:探讨重组脂联素对第三丁基过氧化氢(t-BHP)诱导的人脐静脉内皮细胞(HUVECs)凋亡是否存在影响,同时观察重组脂联素对内质网应激相关信号通路IRE1α-JNK的影响。方法:应用不同浓度t-BHP(25、50、100、200、400μmol/L)诱导HUVECs 1～24 h,MTT法观察t-BHP作用后细胞的存活率;Hochest/PI染色荧光显微镜观测细胞凋亡形态;Western blot检测t-BHP诱导HUVECs凋亡过程中细胞内内质网应激相关分子IRE1α、p-JNK及Caspase-3蛋白表达水平的变化。在此基础上,用携带人脂联素基因的重组腺病毒感染HUVECs处理,观察重组脂联素对t-BHP诱导的HUVECs凋亡是否有保护作用,以及细胞内内质网应激相关信号分子IRE1α、p-JNK及Caspase-3蛋白表达水平的变化。结果:MTT显示,不同浓度t-BHP(25、50、100、200、400μmol/L)诱导HUVECs 1～24 h后,发现随着t-BHP作用浓度的增加和时间的延长,可明显抑制HUVECs的生长,并呈现一定的量效和时效关系。不同浓度t-BHP处理8 h后, Hochest/PI染色荧光显微镜下显示,随着t-BHP浓度的增加,HUVECs的损伤更明显,细胞凋亡率随之升高;ER应激相关信号蛋白IRE1α、p-JNK及凋亡蛋白Caspase-3表达增加。采用携带人脂联素基因的重组腺病毒感染HUVECs预处理,可以明显减轻t-BHP诱导的HUVECs凋亡,IRE1α蛋白表达无明显改变,p-JNK、Caspase-3蛋白表达均明显降低。结论:(1)t-BHP可明显诱导HUVECs发生凋亡性损伤;(2)内质网应激相关信号分子IRE1α、JNK参与了t-BHP诱导HUVECs产生凋亡效应的过程;(3)重组脂联素可以有效抑制t-BHP诱导HUVECs产生的氧化损伤;(4)重组脂联素能够减轻t-BHP诱导的HUVECs凋亡,但不是通过IRE1α发挥作用的。
[Abstract]:Objective: to investigate the effect of recombinant adiponectin on apoptosis of human umbilical vein endothelial cells (HUVECs) induced by third Ding Ji. The effects of recombinant adiponectin on endoplasmic reticulum stress-related signaling pathway (IRE1 伪 -JNK) were also observed. Methods: the HUVECs was induced for 24 h with different concentrations of t-BHP2550U (200渭 mol / L). MTT assay was used to observe the survival rate of the cells treated with t-BHP. The morphology of apoptosis was observed by fluorescence microscope with Hochest/PI staining. Western blot was used to detect the intracellular endoplasmic reticulum stress-related molecule IRE1 伪 during t-BHP induced HUVECs apoptosis. On the basis of p-JNK and Caspase-3 protein expression, HUVECs was infected with recombinant adenovirus carrying human adiponectin gene. To observe the protective effect of recombinant adiponectin on HUVECs apoptosis induced by t-BHP and the intracellular endoplasmic reticulum stress-related signaling molecule IRE1 伪. Changes of p-JNK and Caspase-3 protein expression levels. Results HUVECs was induced by different concentrations of t-BHP1 2550m ~ (100 渭 mol / L) at 400 渭 mol 路L ~ (-1) for 24 h. It was found that with the increase of t-BHP concentration and the prolongation of time, the growth of HUVECs could be inhibited obviously, and there was a dose-effect and time-dependent relationship. Hochest/PI staining fluorescence microscope showed that with the increase of t-BHP concentration, the damage of HUVECs was more obvious, and the apoptosis rate increased with the increase of t-BHP concentration. The expression of ER stress-related signal protein IRE1 伪 p-JNK and apoptotic protein Caspase-3 was increased. HUVECs was pretreated with recombinant adenovirus carrying human adiponectin gene. The expression of IRE1 伪 protein in HUVECs apoptosis induced by t-BHP was significantly alleviated, and the expression of p-JNKK Caspase-3 protein was significantly decreased. Conclusion T- BHP can induce apoptotic injury in HUVECs. 2) endoplasmic reticulum stress-related signaling molecule IRE1 伪 -JNK is involved in the process of HUVECs apoptosis induced by t-BHP. 3) Recombinant adiponectin can effectively inhibit the oxidative damage induced by t-BHP induced HUVECs. 4) Recombinant adiponectin could attenuate the apoptosis of HUVECs induced by t-BHP, but not through IRE1 伪.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R363

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