异氟烷预处理激活Nrf2-ARE通路保护H9c2心肌细胞缺氧-复氧损伤  

Protective effect of isoflurane on H9c2 cells from hypoxia-reoxygenation injury via Nrf2-ARE signaling pathway

作　　者：梁冰[1] 方洁[1] 

机构地区：[1]河南中医药大学第一附属医院麻醉科,郑州450000

出　　处：《第三军医大学学报》2016年第14期1615-1621,共7页Journal of Third Military Medical University

摘　　要：目的研究异氟烷预处理对大鼠胚胎心肌细胞(H9c2)缺氧-复氧损伤及Nrf2-ARE信号通路的影响。方法用大鼠H9c2细胞低氧模拟缺血性损伤,并将细胞分为空白对照组、缺氧-复氧组、异氟烷预处理组、转染Nrf2 siRNA组、转染非特异性siRNA组(Scramble组)、异氟烷预处理的Scramble组和异氟烷预处理的siRNA组。采用MTT法检测H9c2细胞存活率,TUNEL染色检测细胞凋亡,分别使用硫代巴比妥酸法(TBA)和分光光度法测定MDA和GSH水平,qRT-PCR及Western blot检测Nrf2、HO-1和NQO1基因的表达。结果与空白对照组比较,缺氧-复氧组细胞活力降低,凋亡细胞数上升,MDA水平升高,GSH水平降低,同时H9c2细胞中Nrf2、HO-1和NQO1的mRNA和蛋白表达降低,差异均具有统计学意义(P<0.01)。与缺氧-复氧组比较,异氟烷可提高H9c2细胞活力,降低凋亡细胞数,并降低MDA水平,升高GSH水平,上调Nrf2、HO-1和NQO1的表达(P<0.05)。siRNA转染组Nrf2的mRNA和蛋白表达与缺氧-复氧组和Scramble组相比均降低(P<0.05),2%异氟烷对siRNA转染组Nrf2 mRNA和蛋白表达均无影响(P>0.05);经2%异氟烷处理的siRNA转染组H9c2细胞存活率与空白对照组、2%异氟烷处理组和2%异氟烷处理的Scramble组相比降低,凋亡细胞数增多,MDA水平升高,GSH水平降低(P<0.05)。结论异氟烷预处理对H9c2细胞缺氧-复氧损伤具有保护作用,这种保护作用与激活Nrf2-ARE信号通路有关。Objective To investigate the protective effect of isoflurane on H9c2 cell injury induced by hypoxia-reoxygenation via Nrf2-ARE signaling pathway. Methods H9e2 cells were cultured and divided into control group, hypoxia-reoxygenation group, isoflurane pre-treatment group, Nrf2 siRNA transfection group, nonspecific transfection group （scramble group）, nonspecific transfection with isoflurane pretreatment group, and Nrf2 siRNA transfection with isoflurane pretreatment group. Cell viability and apoptosis were evaluated by MTT assay and TUNEL assay, respectively. The levels of MDA and GSH were tested with TBA method and spectrophotometric method, qRT-PCR and Western blotting were performed to detect the expression of Nrf2, HO-1 and NQO1 at mRNA and protein levels. Results Compared with the control group, the decreased cell viability, increased cell apoptosis, raised MDA level, and descended GSH level were observed in the hypoxia-reoxygenation group. Meanwhile, both of the mRNA and protein levels of Nrf2, HO-1 and NQO1 were remarkably down-regulated in the hypoxia-reoxygenation group compared with the control group （P 〈0. 05 ）. Isoflurane pre-treatment significantly improved H9c2 cell viability, reduced the number of apoptotic cells, decreased MDA level and increased GSH level compared with the hypoxiareoxygenation group （P 〈0.05）. Furthermore, the mRNA and protein levels ot Nrf2, HO-1 and NQO1 m H9c2 cells were also up-regulated after isoflurane treatment （P 〈 0. 05 ）. Compared with the hypoxia- reoxygenation group and the scramble group, the expression of Nrf2 in the siRNA transfection group was significantly decreased （ P 〈 0. 05 ）, and 2% isoflurane had no significant effect on the expression of Nrf2. The cell viability was significantly decreased, and apoptotic cells were notably increased in the Nrf2 siRNA transfection with isoflurane pretreatment group compared with the control group, isoflurane pretreatment group and nonspecific transfection with isoflurane pretreatment group （P �

关 键 词：异氟烷 缺氧-复氧 Nrf2-ARE信号通路 H9C2细胞 

分 类 号：R322.11[医药卫生—人体解剖和组织胚胎学] R364.4[医药卫生—基础医学]