[1]张泽英,张辉,赵昕,等.shRNA沉默C5aR对LPS诱导的大鼠肾小管上皮细胞损伤的影响[J].江苏农业学报,2017,(06):1327-1332.[doi:doi:10.3969/j.issn.1000-4440.2017.06.019]
　ZHANG Ze-ying,ZHANG Hui,ZHAO Xin,et al.Effect of silencing C5a receptor expression with shRNA on rat kidney epithelial cells with acute injury induced by lipopolysaccharide[J].,2017,(06):1327-1332.[doi:doi:10.3969/j.issn.1000-4440.2017.06.019]
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shRNA沉默C5aR对LPS诱导的大鼠肾小管上皮细胞损伤的影响()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2017年06期
页码:
1327-1332
栏目:
畜牧兽医·水产养殖
出版日期:
2017-12-30

文章信息/Info

Title:
Effect of silencing C5a receptor expression with shRNA on rat kidney epithelial cells with acute injury induced by lipopolysaccharide
作者:
张泽英1; 2; 张辉1; 赵昕1; 黄爱妮1; 蒋思婧2
（1.武昌工学院食品与环境工程学院，湖北武汉430065；2.湖北大学生命科学学院，湖北武汉430062）
Author(s):
ZHANG Ze-ying1; 2; ZHANG Hui1; ZHAO Xin1; HUANG Ai-ni1; JIANG Si-jing2
(1.College of Food and Environmental Engineering, Wuchang Institute of Technology, Wuhan 430065, China;2.School of Life Sciences, Hubei University, Wuhan 430062, China)
关键词:
C5a受体; 脂多糖; 基因沉默; 脓毒症
Keywords:
C5a receptor; lipopolysaccharide; gene silence; sepsis
分类号:
S852.2
DOI:
doi:10.3969/j.issn.1000-4440.2017.06.019
文献标志码:
A
摘要:
为观察短发夹状RNA（Short hairpin RNA , shRNA）阻断大鼠C5a受体（C5a receptor, C5aR）基因表达对抑制脂多糖（Lipopolysaccharide, LPS）诱导的大鼠肾小管上皮细胞（NRK-52E细胞）损伤的影响，将构建好的针对大鼠C5aR基因的shRNA表达质粒转染NRK-52E细胞系，经G418筛选后，形成稳定的表达C5aR shRNA的细胞系。设置空白对照组（NC组）、正常对照组（LPS组）、阴性对照组（LPS+shRNAc组）和处理组（LPS+C5aR-shRNA组）。采用荧光定量反转录PCR(RT-PCR)检测细胞中白介素-6（Interleukin-6, IL-6）、肿瘤坏死因子α（Tumor necrosis factor-alpha, TNF-α）mRNA的表达，酶联免疫吸附方法(Enzyme-linked immunosorbent assay, ELISA)检测细胞上清液中TNF-α、IL-6和髓过氧化物酶(MPO)活性，流式细胞仪检测各组细胞凋亡率。结果显示,与正常对照组和阴性对照组相比，试验组中C5aR shRNA能有效抑制炎症因子TNF-α、IL-6的释放，降低细胞凋亡率和细胞中MPO活性（P<0.05）。可见，C5aR shRNA能显著抑制由LPS诱导的TNF-α、IL-6等炎症因子的释放，改善大鼠肾脏功能，对LPS诱导的肾脓毒症损伤有明显保护作用。
Abstract:
To investigate the effect of silencing C5a receptor (C5aR) expression with shRNA on rat kidney epithelial cells with acute injury induced by lipopolysaccharide (LPS), eukaryotic expression vector of small hairpin RNA targeting rat C5aR gene was constructed and transfected into rat renal tubular epithelial cell line NRK-52E, after G418 selection, stable cell line expressing C5aR shRNA was got. The experiment was designed as follows: blank control group (NC group), normal control group (LPS group), negative control group (LPS+shRNAc group), experimental group (LPS+C5aR-shRNA group). After incubation with C5aR shRNA, the expression of interleukin-6(IL-6) and tumor necrosis factor-alpha(TNF-α) mRNA were detected by fluorescence quantitative reverse transcription PCR, the content of TNF-α, IL-6 and myeloperoxidase(MPO) in the supernatant of cells were tested by enzyme-linked immunosorbent assay(ELISA), and the apoptosis ratio of each group was detected by flow cytometry. The results showed that C5aR shRNA in the experimental group could effectively inhibit the release of inflammatory factors TNF-α and IL-6, reduce the apoptosis rate and MPO content in cells (P<0.05) compared with that in the normal control group and negative control group. In conclusion, specific short hairpin RNA targeting C5aR can significantly inhibit the release of inflammatory factors such as TNF-α and IL-6 induced by LPS, improve the renal function in rats and has obvious protective effect on renal sepsis injury induced by LPS.

参考文献/References:

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备注/Memo

备注/Memo:
收稿日期：2017-03-24 基金项目：国家自然科学基金项目（31070776） 作者简介：张泽英（1980-），女，四川泸州人，博士，副教授，研究方向为食品科学与免疫学。(E-mail)qianyezzy@163.com 通讯作者：蒋思婧，(E-mail)jiangsijing@hubei.edu.cn
更新日期/Last Update: 2018-01-03