肺泡Ⅱ型上皮细胞自噬体与结核分枝杆菌相互作用的研究  

Interaction Between Mycobacterium Tuberculosis and Autophagosome in Alveolar Epithelial Cell TypeⅡ

作　　者：郭旭光[1] 马越云[1] 周珊[1] 刘家云[1] 苏明权[1] 郝晓柯[1] 

机构地区：[1]第四军医大学西京医院全军临床检验医学中心,陕西西安710032

出　　处：《现代生物医学进展》2010年第5期810-813,共4页Progress in Modern Biomedicine

基　　金：国家自然科学基金(面上项目)(30872358)

摘　　要：目的:检测LC3在肺泡Ⅱ型上皮细胞A549上的表达情况,及结核分枝杆菌刺激后对其表达的影响,探讨自噬在结核分枝杆菌感染上皮细胞中所起的作用。方法:体外培养肺泡Ⅱ型上皮细胞A549,在结核分枝杆菌感染A549细胞0h,24h分别提取RNA,采用RT-PCR的方法检测LC3mRNA的表达情况。采用凋亡坏死染色试剂盒在结核分枝杆菌感染24h后检测对照组,3-MA组,MTB组和3-MA+MTB组的细胞坏死情况。在结核分枝杆菌感染A549细胞4h,8h,16,24h采用Non-Radioactive Cytocity Assay的方法检测对照组,3-MA组,MTB组和3-MA+MTB组上清液LDH的OD值。结果:LC3在肺泡Ⅱ型上皮细胞显著表达,结核分枝杆菌感染后LC3表达降低。细胞凋亡和坏死染色结果显示空白组和3-MA组没有明显差异(P>0.05),MTB组和3-MA+MTB组有明显差异(P<0.05)。LDH检测显示MTB组和3-MA+MTB组上清液LDH的OD值数据两两之间有明显差异(P<0.05)并且有时间依赖性。结论:肺泡II型上皮细胞自噬体在抵抗结核分枝杆菌的感染过程中起一定的作用。Objective：To detect the expression of LC3 in human alveolar type Ⅱ epithelial cells A549 and the effect of Mycobacterium tuberculosis on it, and to lay the foundation for studying autophagosome resistance in the process of Mycobacterium tuberculosis infection. Methods：Human pulmonary type II epithelial cells were cultured in vitro and stimulated with Myeobaeterinm tuberculosis, Extract the RNA ofA549 cells at Oh and 24h and detect LC3 mRNA expression by RT-PCR. Test the necrosis of control group ,3-MA group, MTB group and 3-MA ＋ MTB group with the necrosis and apoptosis staining kit after 24h. Detect the OD value of LDH of the control group,3-MA group,MTB group and 3-MA＋MTB group at 4h, 8h, 16h and 24h by Non-Radioactive Cytotoxieity Assay respectively. Results： The expression of LC3 mRNA deteced by RT-PCR was significantly different.The apoptosis and necrosis staining showed the blank group and 3-MA group was not significantly different （P〉0.05）, MTB group and 3-MA ＋ MTB group significantly different （P〈0.05）. The OD value of LDH test showed MTB group and 3-MA ＋ MTB Group was significantly different （P〈0.05） and time dependent. Conclusions：Autophagy is a defense mechanism inhibiting Mycobacterium tuberculosis survival in infected Human pulmonary type Ⅱ epithelial cells.

关 键 词：LC3 结核分枝杆菌 肺泡Ⅱ型上皮细胞 3-MA 自噬体 

分 类 号：R378.911[医药卫生—病原生物学]