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目的本研究利用A族链球菌(GAS)感染小鼠巨噬细胞,来分析SOCS-1的表达情况,以此探索GAS进行免疫逃逸的可能机制。方法同时培养RAW264.7巨噬细胞和C57小鼠骨髓巨噬细胞(BMDMs),以感染复数(MOI)100∶1加入对数生长期GAS及经热灭活处理的GAS(70℃加热处理60 min)刺激RAW264.7及BMDMs巨噬细胞1 h,弃去细菌后,继续培养2,4,6,8,10 h,分别收集细胞,通过RT-PCR及Western blot方法检测JAK/STAT通路的活化及SOCS-1的表达情况。结果 GAS感染RAW264.7及BMDMs细胞4 h后p-STAT1的水平明显高于热灭活组(P<0.05),6 h达到高峰,8 h后开始降低;同时,两组细胞SOCS-1基因的表达,GAS组4 h开始明显升高,6 h达到高峰,8 h开始降低;而热灭活GAS组SOCS-1基因在感染后6 h后开始升高,且升高的幅度明显小于GAS组(P<0.05)。SOCS-1蛋白在GAS感染后6 h可以检测到,而热灭活GAS刺激细胞10h检测不到SOCS-1的表达。结论GAS感染小鼠初期通过快速活化JAK/STAT通路引起SOCS-1的表达,以此来逃避机体的免疫攻击。
Objective To investigate the possible mechanism of GAS immune escape by analyzing the expression of SOCS-1 in murine macrophages infected with Group A Streptococcus (GAS). Methods RAW264.7 macrophages and C57 mouse bone marrow macrophages (BMDMs) were cultured at the same time. The logarithmic growth phase (GAS) and heat-inactivated GAS (70 ℃ heat treatment 60 min) to stimulate macrophages of RAW264.7 and BMDMs for 1 h. After removing the bacteria, the cells were cultured for further 2, 4, 6, 8 and 10 h. The cells were collected and the expression of JAK / STAT pathway was detected by RT-PCR and Western blot Activation and SOCS-1 expression. Results The levels of p-STAT1 in GAS-infected RAW264.7 and BMDMs cells after 4 h were significantly higher than those in heat-inactivated groups (P <0.05), peaked at 6 h, and then decreased at 8 h. Meanwhile, the SOCS-1 gene GAS group began to rise significantly at 4 h, peaked at 6 h, and began to decrease at 8 h; however, the SOCS-1 gene in heat-inactivated GAS group began to increase 6 h after infection, and the amplitude of increase was significantly lower than that of GAS Group (P <0.05). SOCS-1 protein was detectable at 6 h after GAS infection, whereas SOCS-1 expression was undetectable by heat-inactivated GAS for 10 h. CONCLUSIONS: GAS-infected mice evolve SOCS-1 expression through rapid activation of JAK / STAT pathway in order to evade immune attack.