抑制Mcl-1表达调控结核感染的小鼠腹腔巨噬细胞凋亡的机制初探  

Preliminary mechanism of Mcl-1 silencing in regulating apoptosis of mouse peritoneal macrophages infected with MTB

作　　者：王新敏[1] 王小芳[2] 卢洋 杨菩 韩玲 王英姿[2] 张万江[2] 章乐[2] WANG Xin-min;WANG Xiao-fang;LU Yang;YANG Pu;HAN Ling;WANG Ying-zi;ZHANG Wan-jiang;ZHANG Le(Department of Urinary Surgery,The First Affiliated Hospital of Shihezi University,Shihezi 832002,China;Department of Pathophysiology,Medical College of Shihezi University,Shihezi 832002,China.)

机构地区：[1]石河子大学医学院第一附属医院泌尿外科,新疆石河子832002 [2]石河子大学医学院病理生理学教研室,新疆石河子832002

出　　处：《中国病理生理杂志》2018年第10期1861-1868,共8页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81660330)

摘　　要：目的:应用Mcl-1-shRNA质粒抑制不同毒力结核分枝杆菌(MTB)菌株感染的小鼠腹腔巨噬细胞中Mcl-1的表达,通过观察Bcl-2和Bax表达的变化探讨其调控机制。方法:制备不同毒力MTB菌株悬液,分别感染BALB/c小鼠,再用Mcl-1-shRNA质粒处理感染小鼠模型,并同时设立对应的对照组,于处理后1 d、3 d、5 d和7d处死小鼠并收集腹腔巨噬细胞。应用流式细胞术检测不同处理时间、不同毒力菌株感染时小鼠腹腔巨噬细胞的凋亡率,real-time PCR和Western blot检测Bcl-2和Bax的表达。结果:Mcl-1-shRNA质粒处理后,不同毒力MTB菌株感染的小鼠巨噬细胞凋亡率均比对照组有不同程度的增高,其中以BCG和H37Ra组最明显(P <0. 05); Bcl-2的mRNA和蛋白水平显著减少,而Bax的mRNA和蛋白的表达均显著增加,以BCG感染组较为显著,且二者mRNA的比值与菌株毒力呈负相关(P <0. 05)。结论:抑制Mcl-1的表达可显著促进不同毒力MTB菌株感染的小鼠腹腔巨噬细胞凋亡,其调控机制可能与Bcl-2和Bax蛋白的表达及MTB菌株毒力密切相关。AIM:To investigate the mechanism of myeloid cell leukemia-1(Mcl-1)silencing in regulating the apoptosis of mouse peritoneal macrophages infected with Mycobacterium tuberculosis(MTB)by observing the changes of Bcl-2 and Bax expression.METHODS:The suspensions of MTB strains with different virulence,BCG,H37Ra,H37Rv and XJ-MTB,were prepared to infect BALB/c mice.The transfection of Mcl-1-shRNA plasmid was used to establish a mouse model,and a corresponding control group at the same time was set up.The mice were executed and their peritoneal macrophages were collected 1 d,3 d,5 d and 7 d after the treatment.The apoptosis of the macrophages treated with diffe-rent virulence of MTB strains at different time points was analyzed by flow cytometry.The expression of Bcl-2 and Bax at mRNA and protein levels was determined by real-time PCR and Western blot.RESULTS:The apoptotic rate of mouse peritoneal macrophages increased to some extent after transfection with Mcl-1-shRNA plasmid compared with control group.The order of apoptotic rates was BCG>H37Ra>H37Rv≈XJ-MTB(P<0.05).The expression of Bcl-2 at mRNA and protein levels was significantly decreased,while the expression of Bax at mRNA and protein levels was significantly increased.The changes in BCG infection group were the most significant,and the negative correlation between the Bcl-2/Bax ratio at mRNA level and the virulence of the MTB strains was observed(P<0.05).CONCLUSION:Inhibition of Mcl-1 expression significantly promotes the apoptosis of peritoneal macrophages in mice infected with different virulence of MTB strains.The regulatory mechanism may be closely related to the protein expression of Bcl-2 and Bax and the virulence of MTB strains.

关 键 词：结核分枝杆菌 BAX 巨噬细胞 细胞凋亡 MCL-1基因 

分 类 号：R542.1[医药卫生—心血管疾病] R363[医药卫生—内科学]