Objective To explore the role of RAGE in the progress of high mobility protein Bl activating hepatic stellate cell. Methods The recovery HSC-T6 cells were cultured until the cells on logarithmic phase were prepared for the experiment. The trials are divided into five groups. Control group: DMEM culture medium 1 ml; HMGB1 group: 0.1 μg/ml HMGB1 1 ml; anti-RAGE group: after cultivated with 20 μg/ml RAGE antibody for 2 hours, plus 0.1 μg/ml HMGB1; sRAGE group: after cultivated with 20 μg/ml sRAGE for 2 hours, plus 0.1 μg/ml HMGB1; AGE group: after cultivated with 160 μg/ml AGE for 2 hours, plus 0.1 μg/ml HMGB1. Each group cells were stimulated for 24 h, then the levels of HA, PⅢP and CⅣ in the supernatant were measured by ELISA and the expression of α-SMA and TGF-β1 in HSC-T6 were tested by immunohistochemistry. Results Compare with HMGB1 group, the levels of α-SMA, TGF- β1 in HSC was significantly decreased, and the expression of HA,PⅢP and CⅣ in supernatant was greatly lower in anti-RAGE group and sRAGE group. Compare with control group, the levels of α-SMA, TGF- β1 in HSC was significantly increased, and the expression of HA, PⅢP and CⅣ in supernatant was greatly higher in AGE group. Conclusions RAGE bound to HMGB1 contributes to HSC activation and extracellular matrix releasion.%目的 探讨晚期糖基化终末产物受体(RAGE)在高迁移率族蛋白B1(HMGB1)激活肝星状细胞(HSC)中的作用.方法 大鼠HSC-T6细胞系培养至对数生长期,分为5组,对照组加DMEM培养液1 ml;HMGB1组加0.1 μg/ml 的HMGB1 1 ml;抗RAGE组将20 μg/ml RAGE抗体加入培养板2 h后,再加0.1 μg/ml HMGB1;sRAGE组将20 μg/ml sRAGE加入培养板2 h后,再加0.1 μg/ml HMGB1;AGE组:将160 μg/ml AGE 加入培养板2 h后,再加0.1 μg/ml HMGB1.刺激 24 h 后,收集各组上清培养液酶联免疫吸附试验(ELISA)方法检测透明质酸(HA)、Ⅲ型胶原(PⅢP)和Ⅳ型胶原(CⅣ)的含量;小心取出六孔板内载玻片,免疫组织化学法检测α-SMA、转化生长因子(TGF)-β1的表达.结果 抗RAGE组、sRAGE组与HMGB1组相比,HSC细胞质中的α-SMA和TGF-β1表达水平和培养液中HA、PⅢP和CⅣ的含量均显著下降(P＜0.05);AGE组与对照组相比,HSC细胞质中的α-SMA和TGF-β1表达水平和培养液中的HA、PIIIP和CⅣ含量均明显增高(P＜0.05).结论 HMGB1与RAGE结合,使HSC活化,合成释放细胞外基质增加.
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