HMGB1对LPS诱导A549细胞自噬表达的调控作用

Regulatory effect of HMGB1 on LPS-induced autophagy expression of A549 cells

目的明确脂多糖(LPS)对A549细胞自噬的影响,初步探讨高迁移率族蛋白B1(HMGB1)对LPS诱导的A549细胞自噬表达的影响。方法CCK-8实验检测不同剂量的LPS对A549细胞存活率的影响,取最低毒性剂量的LPS诱导A549细胞构建急性呼吸窘迫综合征(ARDS)肺上皮细胞损伤模型;采用流式细胞术和透射电镜检测细胞死亡形式及细胞自噬情况;联合使用siRNA干扰技术干预、LPS处理A549细胞,随机分为空白组、LPS组、Si-HMGB1组、Si-HMGB1+LPS组。通过Western blot实验检测各组自噬表达水平。结果①LPS处理A549细胞24 h的最低毒性剂量为100μg/ml,且细胞的死亡形式为自噬性细胞死亡;②siRNA干扰技术抑制HMGB1后,与LPS组相比,Si-HMGB1+LPS组的HMGB1和LC3B-Ⅱ/LC3B-Ⅰ的蛋白表达量明显下降,表达量介于空白组和LPS组之间,差异有统计学意义(P<0.05)。结论LPS可引起A549细胞自噬及自噬性细胞死亡,Si-HMGB1能抑制LPS诱导A549细胞自噬。

Objective To investigate the effect of lipopolysaccharide(LPS)on the autophagy of A549 cells and the effect of high mobility group box-1 protein(HMGB1)on the autophagy expression induced by LPS.Methods CCK-8 test was used to detect the effect of different doses of LPS on the survival rate of A549 cells.The lung epithelial cell injury model of acute respiratory distress syndrome(ARDS)was established by inducing A549 cells with the lowest toxic dose of LPS.Death and autophagy of cells were detected by flow cytometry and transmission electron microscopy.A549 cells were treated with siRNA interference technology as well as LPS.These cells were randomly divided into the blank group,the LPS group,the Si-HMGB1 group,and the Si-HMGB1+LPS group.The expression of autophagy in each group was detected by Western blot.Results①The lowest toxic dose of LPS for treatment of A549 cells for 24 h was 100μg/ml,and the cells died of autophagy.②In comparison with the LPS group,the Si-HMGB1+LPS group had significantly decreased protein expressions of HMGB1 and LC3B-Ⅱ/LC3B-Ⅰafter HMGB1 was inhibited by siRNA interference technique.The above mentioned expressions in the Si-HMGB1+LPS group were between those of the blank group and those of the LPS group,and the differences were statistically significant(P<0.05).Conclusion LPS can induce autophagy and autophagic death of A549 cells,and Si-HMGB1 can inhibit autophagy induced by LPS in A549 cells.