血管紧张素Ⅱ对H9C2细胞中NLRP3炎性体的影响
发布时间：2018-10-23 07:36
【摘要】：研究背景:慢性心力衰竭是多种心血管疾病的终末阶段。国内外研究者普遍认为心室重构是慢性心力衰竭的主要发病机制。血管紧张素II(AngII)可通过激活神经内分泌系统而导致心室重塑的发生。但越来越多的证据表明,AngII还可通过激活白介素-6(IL-6)、转化生长因子-β(TNF-β)等炎症细胞因子导致心室重塑的发生。NLRP3炎性小体及其下游产物参与了各种病因所导致的心肌炎症,进而可导致心室重塑。但是目前关于AngII能否直接通过NLRP3炎性小体诱导心肌细胞发生炎症的报道较少,本研究拟对此作一些探讨。目的:探讨血管紧张素II对H9C2细胞(大鼠心肌细胞)中NLRP3的作用。方法:培养H9C2细胞株,选用对数生长期的H9C2细胞进行试验,用10-6mol/l的AngII予以刺激,于不同的时间点(0小时(0h)、1小时(1h)、3小时(3h)、6小时(6h)、12小时(12h))收集细胞,通过实时荧光定量PCR(RT-qPCR)及蛋白质印迹法(Western blot)分别检测细胞中NLRP3、ASC、Caspase-1、IL-1β的基因和蛋白水平;通过酶联免疫吸附法(ELISA)检测细胞培养液上清液中IL-1β的含量。结果:1.AngII分别刺激H9C2细胞0h、1h、3h、6h及12h后,NLRP3、ASC、Caspase-1及IL-1β的基因相对表达量在各处理组与0h组相比,除1h组的Caspase-1差异无统计学意义外(P0.05),余各处理组差异均有统计学意义(P0.05)。1h组、3h组、6h组、12h组的基因相对表达量进行两两比较,除1h组与3h组之间NLRP3的基因相对表达量差异无统计学意义(P0.05)外,余各组间差异均有统计学意义(P0.05)。2.AngII分别刺激H9C2细胞0h、1h、3h、6h及12h后,NLRP3、ASC、Caspase-1及IL-1β的蛋白相对表达量在各处理组与0h组相比,差异有统计学意义(P0.05),且1h组、3h组、6h组及12h组间差异有统计学意义(P0.05)。3.AngII分别刺激H9C2细胞0h、1h、3h、6h及12h后,细胞培养液上清液中IL-1β的含量在各处理组与0h组相比,差异有统计学意义(P0.05),且1h组、3h组、6h组及12h组间差异有统计学意义(P0.05)。结论:1.经浓度为10-6mol/l的血管紧张素II刺激后,H9C2细胞生长状态良好,细胞形态未见明显异常。2.浓度为10-6mol/l的血管紧张素II可促使H9C2细胞中NLRP3、ASC、Caspase-1和IL-1β的基因及蛋白表达水平增加,同时也可导致细胞培养液上清液中IL-1β的含量增加。3.AngII可通过直接激活NLRP3炎性小体而导致心肌细胞炎症的发生,这一发病机制可能与AngII激活心力衰竭相关,为临床上药物治疗心肌炎症所致心功能不全及心肌重构提供理论依据。
[Abstract]:Background: chronic heart failure is the terminal stage of many cardiovascular diseases. Researchers at home and abroad generally believe that ventricular remodeling is the main pathogenesis of chronic heart failure. Angiotensin II (AngII) can induce ventricular remodeling by activating the neuroendocrine system. But there is growing evidence that AngII can also cause ventricular remodeling by activating inflammatory cytokines such as interleukin-6 (IL-6) and transforming growth factor- 尾 (TNF- 尾). NLRP3 inflammatory bodies and their downstream products are involved in various etiological causes of myocarditis. In turn, it can lead to ventricular remodeling. However, there are few reports on whether AngII can induce cardiomyocyte inflammation directly through inflammatory corpuscles of NLRP3. Aim: to investigate the effect of angiotensin II on NLRP3 in H9C2 cells (rat cardiomyocytes). Methods: H9C2 cell lines were cultured. H9C2 cells in logarithmic growth phase were tested and stimulated with AngII of 10-6mol/l. The cells were collected at different time points (0 h (0 h), 1 hour (1 h), 3 h (3 h), 6 h (6 h), 12 h (12 h). The gene and protein levels of NLRP3,ASC,Caspase-1,IL-1 尾 were detected by real-time fluorescence quantitative PCR (RT-qPCR) and Western blot (Western blot), and the content of IL-1 尾 in the supernatant of cell culture medium was detected by enzyme-linked immunosorbent assay (ELISA). Results: the relative expression of NLRP3,ASC,Caspase-1 and IL-1 尾 genes in H9C2 cells stimulated by 1.AngII for 6 h and 12 h were compared with those in 0 h group. There was no significant difference in Caspase-1 between 1h group and 6h group (P0.05), but there were significant differences among the other treatment groups (P0.05). The relative gene expression levels of 1h group, 3h group, 6h group and 12h group were compared. There was no significant difference in the relative expression of NLRP3 gene between 1h group and 3h group (P0.05), but there was significant difference between the other groups (P0.05). The relative expression of NLRP3,ASC,Caspase-1 and IL-1 尾 protein in the treatment group was higher than that in the 0h group after 2.AngII stimulation for 6 h and 12 h, respectively. The difference was statistically significant (P0.05), and there was significant difference between 1h group, 3h group, 6h group and 12h group (P0.05). After 3.AngII stimulated H9C2 cells for 6 h and 12 h, the content of IL-1 尾 in supernatant of cell culture medium was higher than that of 0 h group. The difference was statistically significant (P0.05), and there was significant difference between 1h group, 3h group, 6h group and 12h group (P0.05). Conclusion: 1. After stimulation of angiotensin II with concentration of 10-6mol/l, H9C2 cells grew well and the morphology of H9C2 cells was not abnormal. 2. Angiotensin II at concentration of 10-6mol/l increased the gene and protein expression of NLRP3,ASC,Caspase-1 and IL-1 尾 in H9C2 cells. It can also increase the content of IL-1 尾 in the supernatant of cell culture medium. 3.AngII can directly activate the inflammatory body of NLRP3 and induce the inflammation of cardiomyocytes, which may be related to the activation of heart failure by AngII. To provide a theoretical basis for the clinical treatment of cardiac insufficiency and myocardial remodeling caused by myocardial inflammation.
【学位授予单位】：川北医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R541.6
本文编号：2288523
[Abstract]:Background: chronic heart failure is the terminal stage of many cardiovascular diseases. Researchers at home and abroad generally believe that ventricular remodeling is the main pathogenesis of chronic heart failure. Angiotensin II (AngII) can induce ventricular remodeling by activating the neuroendocrine system. But there is growing evidence that AngII can also cause ventricular remodeling by activating inflammatory cytokines such as interleukin-6 (IL-6) and transforming growth factor- 尾 (TNF- 尾). NLRP3 inflammatory bodies and their downstream products are involved in various etiological causes of myocarditis. In turn, it can lead to ventricular remodeling. However, there are few reports on whether AngII can induce cardiomyocyte inflammation directly through inflammatory corpuscles of NLRP3. Aim: to investigate the effect of angiotensin II on NLRP3 in H9C2 cells (rat cardiomyocytes). Methods: H9C2 cell lines were cultured. H9C2 cells in logarithmic growth phase were tested and stimulated with AngII of 10-6mol/l. The cells were collected at different time points (0 h (0 h), 1 hour (1 h), 3 h (3 h), 6 h (6 h), 12 h (12 h). The gene and protein levels of NLRP3,ASC,Caspase-1,IL-1 尾 were detected by real-time fluorescence quantitative PCR (RT-qPCR) and Western blot (Western blot), and the content of IL-1 尾 in the supernatant of cell culture medium was detected by enzyme-linked immunosorbent assay (ELISA). Results: the relative expression of NLRP3,ASC,Caspase-1 and IL-1 尾 genes in H9C2 cells stimulated by 1.AngII for 6 h and 12 h were compared with those in 0 h group. There was no significant difference in Caspase-1 between 1h group and 6h group (P0.05), but there were significant differences among the other treatment groups (P0.05). The relative gene expression levels of 1h group, 3h group, 6h group and 12h group were compared. There was no significant difference in the relative expression of NLRP3 gene between 1h group and 3h group (P0.05), but there was significant difference between the other groups (P0.05). The relative expression of NLRP3,ASC,Caspase-1 and IL-1 尾 protein in the treatment group was higher than that in the 0h group after 2.AngII stimulation for 6 h and 12 h, respectively. The difference was statistically significant (P0.05), and there was significant difference between 1h group, 3h group, 6h group and 12h group (P0.05). After 3.AngII stimulated H9C2 cells for 6 h and 12 h, the content of IL-1 尾 in supernatant of cell culture medium was higher than that of 0 h group. The difference was statistically significant (P0.05), and there was significant difference between 1h group, 3h group, 6h group and 12h group (P0.05). Conclusion: 1. After stimulation of angiotensin II with concentration of 10-6mol/l, H9C2 cells grew well and the morphology of H9C2 cells was not abnormal. 2. Angiotensin II at concentration of 10-6mol/l increased the gene and protein expression of NLRP3,ASC,Caspase-1 and IL-1 尾 in H9C2 cells. It can also increase the content of IL-1 尾 in the supernatant of cell culture medium. 3.AngII can directly activate the inflammatory body of NLRP3 and induce the inflammation of cardiomyocytes, which may be related to the activation of heart failure by AngII. To provide a theoretical basis for the clinical treatment of cardiac insufficiency and myocardial remodeling caused by myocardial inflammation.
【学位授予单位】：川北医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R541.6
【参考文献】
相关期刊论文 前10条
1 杨仁强;黄玲;马晓欣;金思一;王丹;李旭;;NLRP3炎症小体介导血管紧张素Ⅱ诱导的人脐静脉血管内皮细胞炎症因子IL-1β的表达[J];南方医科大学学报;2016年06期
2 赵欢;傅豪;解营利;杨涛;张明;张海燕;徐晖;;NLRP3炎症小体在大鼠严重烫伤早期心肌内的表达及意义[J];心脏杂志;2016年05期
3 张金生;张宝霞;杜梅梅;朱慧芳;张阳阳;;体外心肌微环境下益气活血化瘀方药诱导骨髓间充质干细胞向心肌样细胞分化的研究[J];北京中医药大学学报;2016年02期
4 李志园;杨刚;黄华;肖正华;任小梅;;双相脉冲电刺激可促进脂肪干细胞分化为心肌细胞样细胞[J];细胞与分子免疫学杂志;2015年09期
5 阿希;李玉琳;王绿娅;吴依娜;刘燕;杜杰;;炎症小体Nlrp3在高血压小鼠心肌纤维化中的作用[J];心肺血管病杂志;2015年06期
6 况薇;唐敏;何学令;吴文超;刘小菁;李良;;周期性拉伸应变对大鼠骨髓间充质干细胞向心肌样细胞分化的影响[J];生物医学工程学杂志;2014年03期
7 李慧;陈晓虎;;中医药治疗冠心病心绞痛概述[J];四川中医;2014年02期
8 王艳;王海萍;吕洋;;中药制剂诱导骨髓间充质干细胞向心肌细胞的分化[J];中国组织工程研究;2014年01期
9 严中琴;杨刚;崔燎;何学令;况薇;吴文超;刘小菁;李良;;电刺激促骨髓间充质干细胞向心肌样细胞分化[J];生物医学工程学杂志;2013年03期
10 王秀力;李春梅;吕茜;王雷;王洁;宫海滨;;胰岛素样生长因子1诱导骨髓间充质干细胞分化为心肌样细胞的实验研究[J];中华心血管病杂志;2013年02期
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