辐射诱导对小鼠巨噬细胞钙结合蛋白S100A8表达及调控影响的初步研究  

Preliminary study on γ-ray induced expression and regulation of calcium-binding protein S100A8 in mouse macrophage cell line RAW264.7

作　　者：丛悦[1] 饶亚岚[1] 陈肖华[1] 董波[1] 李峰生[1] 王治东[1] 高荣莲[1] 高玲[1] 罗庆良[1] 

机构地区：[1]军事医学科学院放射与辐射医学研究所,北京100850

出　　处：《解放军医学杂志》2009年第5期580-582,共3页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金资助课题(30400119)

摘　　要：目的探讨辐射诱导后小鼠巨噬细胞中钙结合蛋白S100A8 mRNA的表达情况及其调控机制。方法体外培养小鼠巨噬细胞RAW264.7,按处置方法不同分为4组:A组正常培养48h;B组经10Gy射线照射后培养48h;C组用5g/ml LPS培养24h后,更换新鲜培养基继续培养24h;D组经10Gy射线照射后培养24h,然后添加5g/ml LPS继续培养24h。采用RT-PCR及定量PCR方法检测各组细胞S100A8 mRNA表达的变化。分别采用H2O2和喜树碱处理细胞,定量PCR检测S100A8 mRNA表达的变化。构建上游5′端系列报告基因表达载体并转染细胞,采用双荧光素酶法检测报告基因的表达水平。结果经γ射线或LPS刺激后,RAW264.7细胞S100A mRNA表达明显增加,二者联合作用后增加更为明显(P<0.05);H2O2处理后S100A8 mRNA表达增加,而喜树碱处理后其表达没有明显变化。单独经γ射线或LPS刺激后,报告基因的表达无明显变化,而二者联合作用后报告基因表达明显增加(P<0.05)。结论辐射诱导可促进RAW264.7细胞中钙结合蛋白S100A8 mRNA的表达。Objective To study the expression and regulation of S100A8, a calcium-binding protein, in mouse rnacrophage cell line RAW264. 7, which was used as cellular model of radiation injury. Methods RAW264.7 cells were exposed to γ-ray irradiation and/or lipopolysaccharide （LPS）, then the mRNA level of S100A8 was detected by RTPCR and real-time quantitative PCR methods. Real-time quantitative PCR was used to investigate the mRNA level of S100A8 in H2O2 or camptothecin treated cells. The second messenger in signal-transduction pathway was explored. To find γ-ray irradiation response element in the region 5＇ to the transcription start site, a series of 5＇ deletion fragments of this sequence linking luciferase reporter gene were constructed, and relative luciferase activity was measured after transfection and irradiation. Results The expression level of S100A8 mRNA was increased by irradiation. The expression of S100A8 in H2O2 treated cells was increased. However, no change was found in the expression of camptothecin treated cdls. The expression level of luciferase reporter gene didn＇t change either after irradiation. Conclusions It has been revealed in present study that γ-ray irradiation or LPS challenge can induce the mRNA expression of S100A8, and that the cooperative effect between γ-rays and LPS can further increase the mRNA expression of S100A8 in RAW264. 7. The second messenger of γ-ray induced S100A8 expression appears to be ROS rather than DNA damage. The responsive element to γ-ray radiation was not at position --877 to 0. It is likely that γ-ray radiation may regulate the expression of S100A8 by increasing its mRNA stability rather than influencing transcription frequency.

关 键 词：辐射效应 RAW264.7细胞 基因 S100A8 

分 类 号：R349.64[医药卫生—基础医学]