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【目的】探讨小鼠巨噬细胞系RAW264.7体外感染卡介苗的应答。【方法】体外感染RAW264.7细胞23h后,分析细胞形态和细胞表面共刺激分子的表达。然后去除培养上清中的卡介苗,继续培养不同时间,通过CFSE、annexin V/PI和Rh123标记,分析宿主细胞的应答。【结果】卡介苗感染23h后,细胞生长状态良好,细胞内能明显观察到吞噬泡中的BCG。细胞表面共刺激分子CD40、CD54、CD80、CD86、CD11b的表达明显升高,CD11c、I-Ad以及H-2Kd的表达变化不明显。CFSE标记卡介苗后,随着培养时间的延长,荧光强度逐渐减弱,但是4天后仍然明显地高于对照组。除去培养上清中的卡介苗后继续培养,含有卡介苗的细胞逐渐减少,继续培养60h后基本检测不到。另外,卡介苗感染不能诱导细胞凋亡,线粒体膜电位先升高后降低,5d后,基本上与对照组一致。【结论】通过以上分析,为卡介苗免疫机理的研究提供了重要数据。
【Objective】 To investigate the response of murine macrophage cell line RAW264.7 to BCG infection in vitro. 【Method】 In vitro infection of RAW264.7 cells 23 h after the analysis of cell morphology and cell surface costimulatory molecules. The BCG in the culture supernatant was then removed and cultured for different times. The host cell response was analyzed by CFSE, annexin V / PI and Rh123 labeling. 【Results】 After BCG infection for 23h, the cell growth was good, and BCG in the phagocytic vacuole was clearly observed in the cells. The expressions of costimulatory molecules CD40, CD54, CD80, CD86 and CD11b on the cell surface were significantly increased, while the expressions of CD11c, I-Ad and H-2Kd did not change significantly. After CFSE labeled BCG, the fluorescence intensity gradually decreased with the extension of culture time, but still significantly higher than that of the control group after 4 days. Remove the culture supernatant in the BCG continue to culture, cells containing BCG gradually decreased, continue to culture after 60h basically undetectable. In addition, BCG infection can not induce apoptosis, mitochondrial membrane potential first increased and then decreased, after 5d, basically consistent with the control group. 【Conclusion】 The above analysis provides important data for the study of BCG immune mechanism.