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目的:探究大黄酸对脂多糖(LPS)诱导RAW264.7细胞高迁移率族蛋白B1(HMGB1)表达的影响。方法:将培养的RAW264.7细胞分为空白对照组、模型组(LPS)、正丁酸钠阳性对照组及大黄酸低(140μmol/L)、高(210μmol/L)浓度组共5组,给药24 h后收集培养液上清并提取细胞总RNA、总蛋白。采用ELISA法检测培养液上清中HMGB1的含量;RT-PCR法检测HMGB1 mRNA表达;Western Blot检测HMGB1、HDAC3蛋白表达;免疫细胞化学共聚焦显微镜观察HMGB1定位。结果:与LPS组比较,大黄酸低、高浓度组细胞培养液上清HMGB1含量和细胞HMGB1总蛋白及HMGB1 mRNA表达均显著降低(P<0.05或P<0.01);激光共聚焦显微镜结果表明,LPS组红色标记的HMGB1出现在细胞质内,大黄酸组胞质内的HMGB1显著减少,胞核增多。与LPS组比较,大黄酸低、高浓度组细胞HDAC3总蛋白表达显著升高(P<0.05或P<0.01)。结论:大黄酸能有效抑制LPS刺激下RAW264.7细胞HMGB1的表达和释放。
Objective: To investigate the effect of rhein on the expression of HMGB1 in RAW264.7 cells induced by lipopolysaccharide (LPS). Methods: The cultured RAW264.7 cells were divided into blank control group, model group (LPS), normal sodium butyrate control group and low (140μmol / L) and high (210μmol / L) After 24 h, the supernatant of the culture supernatant was collected and the total RNA and total protein of the cells were extracted. The content of HMGB1 in culture supernatant was detected by ELISA. The expression of HMGB1 mRNA was detected by RT-PCR. The expression of HMGB1 and HDAC3 protein was detected by Western Blot. The localization of HMGB1 was detected by immunocytochemistry confocal microscopy. Results: Compared with LPS group, HMGB1 and HMGB1 mRNA and HMGB1 mRNA in the cell culture medium of low and high concentrations of rhein were significantly decreased (P <0.05 or P <0.01). Confocal laser scanning microscopy The red labeled HMGB1 in LPS group appeared in the cytoplasm, HMGB1 in the cytoplasm of rhein group decreased significantly and the nucleus increased. Compared with LPS group, the total HDAC3 protein expression of rhein low and high concentration groups was significantly increased (P <0.05 or P <0.01). Conclusion: Rhein can effectively inhibit the expression and release of HMGB1 in LPS-stimulated RAW264.7 cells.