白藜芦醇通过上调miR-149-5p抑制铁死亡改善LPS诱导的心肌细胞损伤

Resveratrol Improves LPS-induced Cardiomyocyte Injury by Upregulating miR-149-5p to Inhibit Ferroptosis

目的探究白藜芦醇(Rsv)对于抑制miR-149-5p介导的铁死亡改善脂多糖(lipopolysaccharide,LPS)诱导的心肌细胞(H9C2)损伤的作用。方法采用不同浓度Rsv处理H9C2细胞,然后再用LPS刺激,观察Rsv对LPS诱导的H9C2细胞损伤的影响。将H9C2细胞分为不同组别,包括Control组、LPS组、LPS+Rsv组、LPS+miRNA-inhibitor-NC组、LPS+miR149-5p-inhibitor组、LPS+miR149-5p-inhibitor+Rsv组。通过CCK-8检测细胞活力,使用试剂盒检测H9C2细胞乳酸脱氢酶(lactate dehydrogenase,LDH)、谷胱甘肽(glutathione,GSH)和丙二醛(malondialdehyde,MDA)释放变化,采用DCFH-DA荧光探针测定H9C2细胞活性氧(reactive oxygen species,ROS)水平,采用铁离子比色法观察心肌细胞中Fe^(2+)的变化并通过RT-qPCR检测细胞miR-149-5p的表达情况。结果Rsv可以显著降低LPS对H9C2细胞的损伤。与LPS组比较,Rsv+LPS组的细胞活力提高、LDH分泌量降低、GSH释放增加、脂质ROS生成减少、Fe^(2+)含量降低、脂质过氧化物MDA的释放减少并且miR-149-5p的表达增加。与LPS+miR-149-5p-inhibitor组比较,LPS+miR149-5p-inhibitor+Rsv组细胞活力增加,LDH含量减少,GSH增加,ROS产生减少,脂质过氧化和铁积累减少并且miR-149-5p的表达增加。结论Rsv可能通过上调miR-149-5p的表达抑制铁死亡,从而减轻LPS诱导的H9C2细胞损伤。

Objective To investigate the effect of resveratrol(Rsv)on inhibiting miR-149-5p-mediated ferroptosis and improving lipopolysaccharide(LPS)-induced cardiomyocyte(H9C2)injury.Methods Different concentrations of Rsv were used to treat H9C2 cells,followed by LPS stimulation to observe the effect of Rsv on LPS-induced injury in H9C2 cells.The H9C2 cells were divided into different groups,including the Control group,LPS group,LPS+Rsv group,LPS+miRNA-inhibitor-NC group,LPS+miR149-5p-inhibitor group,and LPS+miR149-5p-inhibitor+Rsv group.CCK-8 assay was used to detect cell viability,and a kit was used to measure changes in lactate dehydrogenase(LDH),glutathione(GSH),and malondialdehyde(MDA)release in H9C2 cells.DCFH-DA fluorescent probe was used to determine the level of reactive oxygen species(ROS)in H9C2 cells.Iron ion colorimetry was used to observe changes in Fe^(2+)content in cardiomyocytes,and RT-qPCR was used to detect the expression of miR-149-5p in cells.Results Rsv significantly reduced LPS-induced injury in H9C2 cells.Compared with the LPS group,the Rsv+LPS group showed increased cell viability,reduced LDH secretion,increased GSH release,reduced lipid ROS generation,decreased Fe^(2+)content,reduced MDA release,and increased miR-149-5p expression.Compared with the LPS+miR-149-5p-inhibitor group,the LPS+miR-149-5p-inhibitor+Rsv group showed increased cell viability,reduced LDH content,increased GSH,reduced ROS production,reduced lipid peroxidation and iron accumulation,and increased miR-149-5p expression.Conclusion Rsv may inhibit ferroptosis by upregulating miR-149-5p expression and thus alleviate LPS-induced injury in H9C2 cells.