桑葚提取物对脂多糖诱导的心肌细胞炎症反应和细胞凋亡的影响

Effect of mulberry extract on cardiomyocyte inflammatory response and apoptosis induced by lipopolysaccharide

目的探讨桑葚提取物(ME)对脂多糖(LPS)致心肌细胞炎症反应和细胞凋亡的影响及分子机制。方法以正常培养的H9c2细胞作为对照组;用10 mg/L LPS处理的H9c2细胞作为LPS组;分别用50、100、200 mg/L的ME处理H9c2细胞,再用10 mg/L LPS处理,作为ME低、中、高浓度组。将si-NC、si-S100B转染至H9c2细胞中,再用10 mg/L LPS处理,作为LPS+si-NC组、LPS+si-S100B组;将pcDNA、pcDNA-S100B转染至H9c2细胞中,用200 mg/L ME处理,再用10 mg/L LPS处理,作为LPS+ME+pcDNA组、LPS+ME+pcDNA-S100B组。酶联免疫吸附法检测肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)水平;流式细胞术检测心肌细胞凋亡;Western blot检测Bcl-2、Bax、S100钙结合蛋白B(S100B)的蛋白表达;实时荧光定量PCR检测S100B mRNA表达水平。结果不同浓度ME处理的心肌细胞中TNF-α、IL-6的分泌水平降低,细胞凋亡率显著降低,Bcl-2表达水平显著升高,Bax和S100B表达水平均显著降低,且呈浓度依赖性。干扰S100B表达能抑制LPS诱导的心肌细胞炎症反应和细胞凋亡,S100B过表达则逆转了ME对LPS诱导的心肌细胞炎症反应和细胞凋亡的抑制作用。结论ME可抑制LPS诱导的心肌细胞炎症反应和细胞凋亡,其机制可能与下调S100B有关。

Aim To investigate the effect of mulberry extract(ME)on inflammatory response and apoptosis of cardiomyocytes induced by lipopolysaccharide(LPS)and its molecular mechanism.Methods H9c2 cells in normal culture were cultured as control group;H9c2 cells were treated with 10 mg/L LPS as LPS group;H9c2 cells were treated with 50,100 and 200 mg/L ME,and then treated with 10 mg/L LPS,as low,medium and high concentration ME groups.After transfection of si-NC and si-S100B into H9c2 cells,the cells were treated with 10 mg/L LPS,as LPS+si-NC group and LPS+si-S100B group.After transfection of pcDNA and pcDNA-S100B into H9c2 cells,the cells were treated with 200 mg/L ME followed by 10 mg/L LPS,as LPS+ME+pcDNA group and LPS+ME+pcDNA-S100B group.The levels of tumor necrosis factorα(TNF-α)and interleukin-6(IL-6)were detected by enzyme-linked immunosorbent assay.Flow cytometry was used to detect apoptosis;Western blot was used to detect the expressions of Bcl-2,Bax and S100 calcium binding protein B(S100B);Real-time quantitative PCR was used to detect the expression of S100B mRNA.Results The levels of TNF-αand IL-6 in cardiomyocytes treated with different concentrations of ME were significantly decreased,the apoptosis rate was significantly decreased,the expression of Bcl-2 was significantly increased,the expression of Bax was significantly decreased,and the expression of S100B was significantly decreased,with concentration-dependent manner(P<0.05).Interference of S100B expression inhibited LPS-induced inflammatory response and apoptosis of cardiomyocytes.Overexpression of S100B reversed the inhibitory effect of ME on LPS-induced inflammatory response and apoptosis of cardiomyocytes.Conclusion ME can inhibit LPS-induced inflammatory response and apoptosis of cardiomyocytes,and its mechanism may be related to the down-regulation of S100B.