靶向沉默Mcl-1基因对感染不同毒力结核杆菌小鼠腹腔巨噬细胞凋亡的影响

发布时间：2018-04-19 05:26

本文选题：结核分枝杆菌 + Mcl-基因　；参考：《中国病理生理杂志》2015年12期

【摘要】：目的:通过RNA干扰技术特异性沉默Mcl-1基因,探讨下调Mcl-1基因对感染不同毒力结核杆菌小鼠腹腔巨噬细胞凋亡的影响。方法:分别用制备好的新疆地区流行的优势强毒结核分枝杆菌临床分离株(简称强毒株)、结核分枝杆菌国际标准强毒株H37Rv(简称H37Rv)、结核分枝杆菌国际标准无毒株H37Ra(简称H37Ra)和卡介苗(BCG)菌悬液感染BALB/c小鼠,再以筛选并构建好的Mcl-1-shRNA作用于感染的小鼠模型,同时设立相应对照组,于作用后1 d、3 d、5 d和7 d提取小鼠腹腔巨噬细胞,应用实时荧光定量PCR和Western blot检测各组小鼠腹腔巨噬细胞中Mcl-1 mRNA和蛋白的表达;应用流式细胞术检测各组巨噬细胞的凋亡水平。结果:小鼠被不同毒力的结核杆菌感染后其腹腔巨噬细胞中Mcl-1 mRNA和蛋白的表达水平均有不同程度的升高,其中以感染了强毒株和H37Rv的腹腔巨噬细胞升高最为明显(P0.05);应用RNA干扰技术沉默Mcl-1基因后,Mcl-1mRNA和蛋白的表达水平明显低于对照组(P0.05);流式细胞术分析显示,下调Mcl-1基因的表达可诱导小鼠腹腔巨噬细胞凋亡。结论:应用Mcl-1-shRNA可有效沉默Mcl-1在感染了不同毒力结核杆菌小鼠腹腔巨噬细胞中的表达,并能上调巨噬细胞的凋亡水平。
[Abstract]:Aim: to study the effect of down-regulating Mcl-1 gene on peritoneal macrophage apoptosis in mice infected with different virulence Mycobacterium tuberculosis by RNA interference technique.Methods: the clinical isolates of dominant virulent Mycobacterium tuberculosis (virulent strain), virulent strain H37Rv1 (International Standard strain H37Rvv) and International Standard strain H37Ra( no strain of Mycobacterium tuberculosis) were prepared in Xinjiang, respectively.H37Raand Bacillus Calmette-Guerin Bacillus Calmette-Guerin (BCG) suspension infected BALB/c mice,Then Mcl-1-shRNA was used to screen and construct the infected mouse model, and the corresponding control group was set up. The peritoneal macrophages were extracted from mice on the 1st day, 3rd day, 5th day, and 7th day, respectively.The expression of Mcl-1 mRNA and protein in peritoneal macrophages was detected by real-time fluorescence quantitative PCR and Western blot, and the apoptotic level of macrophages in each group was detected by flow cytometry.Results: the expression levels of Mcl-1 mRNA and protein in peritoneal macrophages of mice infected with different virulence Mycobacterium tuberculosis were increased in varying degrees.Among them, the peritoneal macrophages infected with virulent strain and H37Rv increased the most significantly (P 0.05), the expression level of mcl-1 mRNA and protein in Mcl-1 gene silenced by RNA interference technique was significantly lower than that in control group (P 0.05). Flow cytometry analysis showed that the expression level of Mcl-1 mRNA and protein was significantly lower than that of control group.Down-regulating the expression of Mcl-1 gene can induce mouse peritoneal macrophage apoptosis.Conclusion: Mcl-1-shRNA can effectively silence the expression of Mcl-1 in the peritoneal macrophages of mice infected with different virulent Mycobacterium tuberculosis and can up-regulate the apoptosis level of macrophages.
【作者单位】：石河子大学医学院病理生理学教研室;石河子大学医学院新疆地方与民族高发病教育部重点实验室;石河子大学医学院第一附属医院泌尿外科;石河子大学医学院病原生物学与免疫学教研室;
【基金】：国家自然科学基金资助项目(No.81260241)
【分类号】：R378.911

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