蛇床子素调控circ_0084043/miR-338-3p对高糖诱导的心肌细胞损伤的影响

Effects of osthole on cardiomyocyte damage induced by high glucose and its regulatory effects on circ_0084043/miR-338-3p molecular axis

目的探讨蛇床子素对高糖诱导的心肌细胞损伤的影响及其对circ_0084043/miR-338-3p分子轴的调控作用。方法采用高糖诱导心肌细胞H9C2建立细胞损伤模型(HG组),加入不同浓度的蛇床子素处理细胞(HG+蛇床子素低剂量组、HG+蛇床子素中剂量组、HG+蛇床子素高剂量组),将正常培养的H9C2细胞记为NC组;si-NC、si-circ_0084043分别转染入H9C2细胞后加入30 mmol/L葡萄糖处理细胞(HG+si-NC组、HG+si-circ_0084043组),pcDNA、pcDNA-circ_0084043分别转染入H9C2细胞后加入100 nmol/L蛇床子素与30 mmol/L葡萄糖处理细胞(HG+蛇床子素高剂量+pcDNA组、HG+蛇床子素高剂量+pcDNA-circ_0084043组);采用qRT-PCR法检测circ_0084043、miR-338-3p的表达量;采用MTT实验检测细胞活力;采用流式细胞术检测细胞凋亡率;根据试剂盒检测MDA、SOD的水平;双荧光素酶报告基因实验检测circ_0084043与miR-338-3p的靶向关系;采用Western blot法检测Bax、Bcl-2蛋白表达量。结果与NC组比较,HG组circ_0084043的表达水平升高(P<0.05),miR-338-3p的表达水平降低(P<0.05),细胞活力与Bcl-2蛋白水平和SOD的活性降低(P<0.05),凋亡率与Bax蛋白水平和MDA的水平升高(P<0.05);与HG组比较,蛇床子素不同剂量组circ_0084043的表达水平降低(P<0.05),miR-338-3p的表达水平升高(P<0.05),细胞活力与Bcl-2蛋白水平和SOD的活性升高(P<0.05),凋亡率与Bax蛋白水平和MDA的水平降低(P<0.05);circ_0084043可靶向调控miR-338-3p;与HG+si-NC组比较,HG+si-circ_0084043组细胞活力与Bcl-2蛋白水平和SOD的活性升高(P<0.05),凋亡率与Bax蛋白水平和MDA的水平降低(P<0.05);与HG+蛇床子素高剂量+pcDNA组比较,HG+蛇床子素高剂量+pcDNA-circ_0084043组细胞活力与Bcl-2蛋白水平和SOD的活性降低(P<0.05),凋亡率与Bax蛋白水平和MDA的水平升高(P<0.05)。结论蛇床子素可抑制circ_0084043的表达而促进miR-338-3p的表达从而促进细胞增殖及抑制细胞凋亡、氧化应激进而减轻高糖诱导的心肌细胞损伤。

Objective To investigate the efects of osthole on cardiomyocyte damage induced by high glucose and its regulatory effects on circ_0084043/miR-338-3p molecular axis.Methods High glucose-induced cardiomyocyte H9C2 was used to establish a cell injury model(HG group),and the cells were treated with different concentrations of osthol(low,medium,high dose groups),The normal cultured H9C2 cells were taken as NC group.After si-NC and si-circ_0084043 were transfected into H9C2 cells respectively,30mmol/L glucose-treated cells(si-NC group,si-circ_0084043 group)were added,and pcDNA and pcDNA-circ_0084043 were respectively transfected into H9C2 cells,which were treated by 100nmol/L osthole and 30mmol/L glucose(osthole high dose+pcDNA group,osthole high dose+pcDNA-circ_0084043 group).The qRT-PCR was used to detect the expression levels of circ_0084043 and miR-338-3p,and MTT detected the cell viability,and flow cytometry detected the apoptosis rate,and the levels of MDA and SOD were detected.Moreover dual luciferase reporter gene experiment was used to detect the targeting correlation between circ_0084043 and miR-338-3p,and Western Blot was used to detect the expression levels of Bax and Bcl-2 protein.Results Compared with those in NC group,the expression levels of circ_0084043 and Bax protein,and the MDA le3vels and the apoptosis rate in HG group were significantly increased(P<0.05),however,the expression levels of miR-338-3p and Bcl-2 protein as well as the activity of SOD were significantly decreased(P<0.05).Compared with those in HG group,the expression levels of circ_0084043 and Bax protein as well as MDA levels in different doses of osthole groups were significantly decreased(P<0.05),however,the expression levels of miR-338-3p and Bcl-2 as well as the cell viability and the activity of SOD were significantly increased(P<0.05).Compared with those in si-NC group,the cell viability,the protein level of Bcl-2 and the activity of SOD in si-circ_0084043 group were significantly increased(P<0.05),while the apoptosis rate,the protein level of Bax and MDA levels were significantly decreased(P<0.05).Compared with those in osthole high dose+pcDNA group,the cell viability,the protein level of Bcl-2 and the activity of SOD in osthole high dose+pcDNA-circ_0084043 group were significantly decreased(P<0.05),while the apoptosis rate and the protein level of Bax and MDA levels were significantly increased(P<0.05).Conclusion Osthole can inhibit the expression of circ_0084043,so as to promote the expression of miR-338-3p to promote the cell proliferation and inhibit cell apoptosis,oxidative stress,thereby relieving the cardiomyocyte damage induced by high glucose.