二氧化硅诱导巨噬细胞中核转录因子Sp1的表达活化

Expression and role of nuclear transcription factor Sp1 in macrophages stimulated by silicon dioxide

目的研究二氧化硅(SiO_2)刺激的巨噬细胞中核转录因子(Sp1)表达和定位的动态变化。方法将40只健康SD大鼠随机分为实验组和对照组,每组各20只。采用非暴露式气管插管,实验组每只大鼠一次性注入1ml无菌生理盐水与SiO_2混悬液(0．2g/kg)；对照组大鼠注入等量的无菌生理盐水；分别于灌注后第1、7、14、21和28天各处死4只。免疫组织化学检测体内SiO_2刺激的肺巨噬细胞中Sp1蛋白表达的定位及其动态变化。体外培养小鼠腹腔巨噬细胞系(RAW264．7细胞),逆转录-聚合酶链反应(RT-PCR)检测体外SiO_2刺激的巨噬细胞中Sp1 mRNA的动态变化；免疫细胞化学、Western blot法检测体外SiO_2刺激的巨噬细胞中Sp1蛋白表达的定位及其动态变化。结果实验组与对照组比较,肺巨噬细胞中Sp1蛋白表达上调,于第14天达高峰；体外SiO_2作用RAW264．7细胞30～960min,Sp1 mRNA表达明显升高,呈现先升高而后下降的趋势,峰值位于240min；Sp1总蛋白和核蛋白表达亦呈现先升高而后下降的趋势,总蛋白表达峰值位于240min,核蛋白峰值位于480min；Sp1蛋白于120min开始发生明显的核移位,480min时核移位最明显。结论SiO_2能激活巨噬细胞中Sp1,Sp1可能在矽肺的发生发展过程中起重要的调控作用。

Objective To study the expression and localization of nuclear transcription factor Spl in macrophages after stimulated by silicon dioxide in vivo and in vitro. Methods Forty Sprague Dawley rats were randomly divided into the control group and the silica exposure group, 20 in each group. The rat silicosis models were established by direct tracheal instillation of silica into rat hing（0.2 g/kg） only once while the control group was instilled with equal amount of saline. Animals were killed at 1st,7th, 14th,21st and 28th day after instillation. Dynamic changes of Spl protein expression and its cellular localization were detected by immunohistochemistry in pulmonary macrophages. In vitro,Spl mRNA and protein expression and their dynamic changes were monitored by RT-PCR and western blotting after stimulated by silicon dioxide in cultured RAW264.7 macrophages respectively. Cellular localization of Spl protein was characterized by immtmocytochemistry. Results Compared to the control group, the Spl protein expression was increased in ptdrnonary macrophages and reached the peak at the 14th day in the silica exposure group. In vitro, the Spl mRNA level began to rise at 30 minutes after the administration of sili- con dioxide and reached the peak at 240 minutes and then decreased to the minimal level at 960 minutes. The Spl total protein and nuclear protein also exhibited the similar trend. The former reached the peak at 240 minutes and the latter at 480 minutes. The significant nuclear translocation of Spl protein was observed at 120 minutes after the administration of silicon dioxide and became most significant at 480 minutes. Conclusion Silicon dioxide can activate nuclear transcription factor Spl in macrophages in vivo and in vitro. Spl might play an important pathogenic role in the development of silicosis.