[1]潘永露,何淑芳,韩正怡,等.Fas/caspase-8/3和ERK信号通路在Fas siRNA减轻心肌细胞缺氧/复氧损伤中的作用[J].中国药理学通报,2018,(08):1150-1157.[doi:10.3969/j.issn.1001-1978.2018.08.023]
 PAN Yong-lu,HE Shu-fang,HAN Zheng-yi,et al.Roles of Fas/caspase-8/3 and ERK signaling pathways in Fas siRNA induced reduction of hypoxia/reoxygenation injury to cardiomyocytes[J].Chinese Pharmacological Bulletin,2018,(08):1150-1157.[doi:10.3969/j.issn.1001-1978.2018.08.023]
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Fas/caspase-8/3和ERK信号通路在Fas siRNA减轻心肌细胞缺氧/复氧损伤中的作用()
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《中国药理学通报》[ISSN:/CN:]

卷:
期数:
2018年08期
页码:
1150-1157
栏目:
论著
出版日期:
2018-08-26

文章信息/Info

Title:
Roles of Fas/caspase-8/3 and ERK signaling pathways in Fas siRNA induced reduction of hypoxia/reoxygenation injury to cardiomyocytes
文章编号:
1001-1978(2018)08-1150-08
作者:
潘永露1何淑芳1韩正怡1窦梦云1杨 婉1程 洁1陈志武2张 野1
1.安徽医科大学第二附属医院麻醉科,安徽 合肥 230601; 2.安徽医科大学基础医学院药理学教研室,安徽 合肥 230032
Author(s):
PAN Yong-lu1 HE Shu-fang1 HAN Zheng-yi1 DOU Meng-yun1 YANG Wan1 CHENG Jie1 CHEN Zhi-wu2 ZHANG Ye1
1.Dept of Anesthesiology, the Second Hospital of Anhui Medical University, Hefei 230601, China; 2.Dept of Pharmacology, Anhui Medical University, Hefei 230032, China
关键词:
Fas caspase 细胞外信号调节激酶 心肌细胞 缺氧/复氧损伤 凋亡
Keywords:
Fas caspase extracellular signaling-regulated kinase cardiomyocytes hypoxia/reoxygenation injury apoptosis
分类号:
R-332; R322.11; R329.25; R329.411; R342.2; R345.57; R845.22
DOI:
10.3969/j.issn.1001-1978.2018.08.023
文献标志码:
A
摘要:
目的 评价Fas/caspase-8/3和细胞外信号调节激酶(extracellular signaling-regulated kinase,ERK)信号通路在Fas siRNA减轻心肌细胞缺氧/复氧(hypoxia/reoxygenation,H/R)损伤中的作用。方法 培养H9c2心肌细胞,随机分为4组(n=3):①正常对照组:H9c2细胞置于DMEM/F12细胞培养液中培养; ②H/R组:将H9c2细胞进行5 h缺氧和1 h复氧处理; ③Fas siRNA组(H/R+Fas-siRNA):H9c2细胞转染Fas siRNA 24 h后进行H/R损伤; ④siRNA阴性对照组(H/R+siRNA-NC):H9c2细胞转染Fas siRNA-NC 24 h后进行H/R损伤。处理结束后,采用CCK-8法检测细胞活力; 微量酶标法测定培养液中乳酸脱氢酶(LDH)的活性; Annexin-V-FITC/PI双染法检测细胞凋亡; 荧光定量RT-PCR法检测细胞 Fas mRNA表达水平; Western blot法检测细胞内Fas、caspase-8/3、ERK和GSK-3β蛋白表达。结果 与对照组相比,H/R组细胞活力降低,LDH活性和细胞凋亡增加,Fas mRNA及蛋白水平升高,cleaved caspase-8/3、p-ERK、p-GSK-3β蛋白表达升高(P<0.05); 与H/R组相比,H/R+Fas-siRNA组细胞活力明显提高,LDH活性和细胞凋亡率降低,Fas mRNA及蛋白表达水平降低,cleaved caspase-8/3蛋白表达降低,p-ERK、p-GSK-3β蛋白表达增高(P<0.05),而H/R+siRNA-NC组上述指标差异无统计学意义。结论 Fas/caspase-8/3和ERK信号通路在Fas siRNA减轻心肌细胞H/R损伤中发挥着重要作用。
Abstract:
Aim To evaluate the roles of Fas/caspase-8/3 and extracellular signaling-regulated kinase(ERK)signaling pathways in Fas siRNA induced reduction of hypoxia/reoxygenation(H/R)injury to cardiomyocytes.Methods H9c2 cardiomyocytes were cultured in DME/F-12 culture medium supplemented with 10% fetal bovine serum and randomly divided into four groups(n=3 each)using a random number table: control group, H/R group, Fas siRNA group(H/R+Fas-siRNA)and siRNA control group(H/R+siRNA-NC).H9c2 cardiomyocytes were cultured in normal culture atmosphere in control group; treatment with 5 h hypoxia and 1 h reoxygenation was carried out to H9c2 cells to establish H/R injury in group H/R.In addition, the H9c2 cells in groups H/R+Fas siRNA and H/R+siRNA-NC were transfected with Fas siRNA and Fas siRNA-NC for 24 h before H/R injury, respectively.Cell viability was determined using CCK-8 method, lactate dehydrogenase(LDH)activity in the culture medium was detected using chemical colorimetry, and cell apoptosis was assessed using Annexin-V-FITC/PI flow cytometry in each group at the end of treatment.Relative expression of Fas mRNA was detected by fluorescent quantitative RT-PCR, while the levels of Fas, caspase-8/3, ERK, GSK-3β protein were observed by Western blot.Results Compared with control group, cell viability was significantly repressed, and LDH activity and apoptotic rate as well as the expression of Fas mRNA,protein, and cleaved caspase-8/3 protein and p-ERK, p-GSK-3β protein all increased in group H/R(P<0.05), while Fas siRNA alleviated H/R injury when compared with group H/R, as evidenced by the significantly increased cell viability, and further proved by the suppressed LDH activity and apoptotic rate as well as the expression of Fas mRNA,protein and cleaved caspase-8/3 protein.In addition, p-ERK, p-GSK-3β protein expression increased in group H/R+Fas siRNA(P<0.05).There was no difference in the parameters mentioned above in group H/R+siRNA-NC.Conclusion Fas/caspase-8/3 and ERK signaling pathways play an important role in the alleviation of H/R injury by Fas siRNA in cardiomyocytes.

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备注/Memo

备注/Memo:
收稿日期:2018-03-14,修回日期:2018-04-18
基金项目:安徽省科技厅重点项目(No 1301043030); 安徽省高校省级自然科学研究重大项目(No KJ2014ZD16)
作者简介:潘永露(1990-),男,硕士生,研究方向:麻醉药理学,E-mail: 18255116393@163.com;
张 野(1968-),男,博士,教授,主任医师,博士生导师,研究方向:麻醉药理学,通讯作者,E-mail: zhangye_hassan@sina.com
更新日期/Last Update: 2018-07-26