首页  期刊介绍 征稿简则 编委会 期刊征订 广告服务 留言板 联系我们 English

中国应用生理学杂志 ›› 2020, Vol. 36 ›› Issue (3): 211-215.doi: 10.12047/j.cjap.5973.2020.047

• 研究论文 • 上一篇    下一篇

过表达miR-31对结肠炎模型小鼠TLR4/NF-κB信号通路及凋亡蛋白的调控

刘凯丽, 都新新, 张文琴, 吴亚俐, 王茜, 王春芳, 崔香丽   

  1. 山西医科大学生理学系, 太原 030001
  • 收稿日期:2019-10-15 修回日期:2020-03-21 发布日期:2020-09-25
  • 通讯作者: Tel: 15035164438; E-mail: cuixlcxl@sina.com
  • 基金资助:
    国家自然科学基金(81272695);山西省自然科学基金(201801D121311);山西省“1331工程”重点学科建设计划经费;山西省国际合作计划项目(2015081038)

Overexpression of miR-31 regulates TLR4/NF-κB signaling pathway and apoptotic protein in colitis model mice

LIU Kai-li, DU Xin-xin, ZHANG Wen-qin, WU Ya-li, WANG Xi, WANG Chun-fang, CUI Xiang-li   

  1. Department of Cell Physiology, Shanxi Medical University, Taiyuan 030001, China
  • Received:2019-10-15 Revised:2020-03-21 Published:2020-09-25

摘要: 目的: 探讨miR-31对DSS诱发结肠炎小鼠TLR4/NF-κB信号通路和凋亡相关蛋白的影响。方法: ①小鼠结肠炎实验:用1%葡聚糖硫酸钠(DSS)诱发小鼠溃疡性结肠炎(UC)。14只FVB非转基因小鼠随机分为control组(n=6),DSS组(n=8),16只FVB miR-31转基因小鼠随机分为miR-31过表达组(n=8),miR-31过表达+DSS 组(n=8),DSS溶于水后通过饮水给予小鼠。DSS组和miR-31+DSS组第一周饮用1%DSS水,第二周饮用正常无菌水,第三周饮用1%DSS水,如此5周后造模完成,之后留取小鼠的结肠组织,通过Western blot和IHC检测小鼠结肠组织NF-κB p65、TLR4、Bax、Bcl-2蛋白的表达;TUNEL检测小鼠结肠组织细胞凋亡。②细胞培养实验:在人结肠上皮细胞系HCT 116细胞中通过脂质体转染的方法转染miR-31 mimic和inhibitor,使miR-31过表达或敲低,每组均进行三次重复,48 h后收取细胞,通过Western blot检测NF-κB p65、TLR4蛋白的表达。结果: ①动物实验中,与control组相比,小鼠结肠组织中DSS组和miR-31过表达组NF-κB p65、TLR4蛋白表达水平和凋亡细胞指数均显著升高(P<0.05或P<0.01),Bcl-2/Bax比值显著降低(P<0.05或P<0.01);且与DSS组相比,miR-31+DSS组NF-κB p65、TLR4蛋白表达水平和凋亡细胞指数也显著升高(P<0.01),Bcl-2/Bax比值显著降低(P<0.01)。②细胞实验中,与control组相比, HCT 116细胞过表达miR-31组的NF-κB p65、TLR4蛋白表达水平均显著升高(P<0.05或P<0.01),敲低miR-31组的NF-κB p65、TLR4蛋白表达水平下降(P<0.05)。结论: miR-31通过促进TLR4/NF-κB信号通路和介导肠上皮细胞凋亡促进结肠炎的发展。

关键词: miR-31, 溃疡性结肠炎, 葡聚糖硫酸钠, 小鼠, 人结肠上皮细胞系HCT 116, TLR4/NF-κB信号通路, 细胞凋亡

Abstract: Objective: To investigate the effects of miR-31 on TLR4/NF-κB signaling pathway and apoptosis-related proteins in dextran sulfate sodium (DSS) induced mouse colon colitis. Methods: ① Mouse model of colon colitis: 1% DSS was used to induce mouse ulcerative colitis (UC). Fourteen FVB non-transgenic mice were randomly divided into control group (n= 6), DSS group (n= 8), and 16 FVB miR-31 transgenic mice were randomly divided into miR-31 overexpression group (n= 8), miR-31 overexpression +DSS group (n= 8). DSS was dissolved in water and administered to mice by drinking water. The DSS group and miR-31+DSS group drank 1% DSS water in the first week, normal sterilized water in the second week, and 1% DSS water in the third week, after 5 weeks, the modeling was completed, then the colon tissues of the mice were collected. Western blot and IHC were used to detect the expressions of NF-κB p65, TLR4, Bax and Bcl-2 proteins in mouse colon tissue, TUNEL was used to detect apoptosis of mouse colon tissues. ② Cell culture experiments: Transfection of miR-31mimic and inhibitor by lipofectamine resulted in overexpression or knockdown of miR-31 in human colon epithelial cell line HCT 116 cells, each group was repeated three times and cells were collected 48 h later, Western blot was used to detect the expressions of NF-κB p65 and TLR4 protein. Results: ① In animal experiments, compared with the control group, the expression levels of NF-κB p65, TLR4 protein and apoptotic cell index in the DSS group and miR-31 overexpression group in mouse colon tissue were significantly increased (P<0.05 or P<0.01), and the Bcl-2 / Bax ratio was significantly reduced (P<0.05 or P<0.01); and compared with the DSS group, the expression levels of NF-κB p65, TLR4 protein and apoptotic cell index in the miR-31+DSS group were significantly increased (P<0.01), while the Bcl-2/Bax ratio was significantly decreased (P<0.01). ② In cell experiments, compared with the control group, the expression levels of NF-κB p65 and TLR4 protein in the over-expressed miR-31 group of HCT 116 cells were significantly increased (P<0.05 or P<0.01), the expressions of NF-κB p65 and TLR4 protein in miR-31 knockdown group were decreased (P<0.05). Conclusion: miR-31 promotes the development of colitis by promoting TLR4/NF-κB signaling pathway and mediating apoptosis of intestinal epithelial cells.

Key words: miR-31, ulcerative colitis, dextran sulfate sodium (DSS), mouse, human colon epithelial cell line HCT 116, TLR4/NF-κB signaling pathway, apoptosis

中图分类号: 

版权所有 © 2015 《中国应用生理学杂志》编辑部
京ICP备16058274号-1
地址:天津市和平区大理道1号,邮编:300050  电话:022-84655184  E-mail:tjzgyish@163.com
本系统由北京玛格泰克科技发展有限公司设计开发 技术支持:support@magtech.com.cn