不同毒力结核分枝杆菌对THP-1细胞凋亡及细胞因子表达的影响

Effect of Different Virulence Mycobacterium tuberculosis on Cytokine Expression and Apoptosis of THP-1 Cells

为探讨不同毒力结核分枝杆菌(MTB)感染巨噬细胞后不同时间点细胞凋亡情况与其分泌的几种主要细胞因子转录与表达之间的关系,建立THP-1体外巨噬细胞模型,用不同毒力MTB(强毒株H37Rv、弱毒株BCG)感染细胞,应用流式细胞术检测巨噬细胞在六个时间点的凋亡情况,应用荧光定量PCR检测细胞因子TNF-α、IL-10、IL-6的转录水平,应用ELISA方法检测细胞上清液中三种细胞因子的分泌量。结果显示:BCG组和H37Rv组的早期凋亡率均显著高于对照组(P<0.05)。巨噬细胞细胞因子TNF-α、IL-6的转录水平与分泌水平相符,弱毒菌株巨噬细胞凋亡情况随时间延长成上升趋势,其分泌的细胞因子TNF-α表现为相同趋势,IL-6表现为相反趋势。强毒株巨噬细胞凋亡情况低于弱毒株,细胞因子分泌情况复杂,一般表现为一种先增强后抑制的情况。本试验通过对不同时间点细胞因子与凋亡情况关系的研究为结核分枝杆菌对巨噬细胞凋亡机制提供新思路。

In order to investigate the relationship between apoptosis and transcription and expression of several major cytokines during Mycobacterium tuberculosis(MTB)infection,THP-1derived macrophage was selected to establish in vitro model.Infected cells with different virulence MTB(H37Rv,BCG),flow cytometry was used to detect macrophage apoptosis at six time points,quantitative PCR was used to detect transcription level of cytokines TNF-α,IL-10,IL-6,ELISA was used to detect the secretion of cell supernatants of three cytokines.The results showed:the early apoptotic rate of BCG and H37 Rv group were both higher than control group and the difference was statistically significant(P<0.05).Transcription levels of cytokines,TNF-α,IL-6wereconsistent with the level of secretion.There was an increase with time on apoptosis of low virulent strain,and there is a same trend on secretion of TNF-α,an opposite tendency on secretion of IL-6.Apoptosis of virulent strains was less than attenuated vaccine strain.Secretion of virulent strains was complex,like increase first and then suppression.The relationship between cytokines and apoptosis of cells at different time points may provide new ideas for study of macrophage apoptosis mechanisms.