摘要
研究木犀草素(luteolin,Lut)抗H9c2心肌细胞损伤的作用及其机制。采用大鼠H9c2心肌细胞株,以1μM阿霉素(doxorubicin,Dox)建立心肌细胞损伤模型,不同浓度Lut(5、10、20μM)进行干预。细胞增殖法(MTT)检测细胞活力。通过高通量转录组测序筛选Lut(20μM)改善H9c2心肌细胞损伤的靶点基因。利用分子克隆技术,将心肌细胞中线粒体延长因子1(mitochondrial elongation factor 1,MIEF1)表达抑制,蛋白免疫印迹法(Western blot)检测Lut对低表达MIEF1心肌细胞中MIEF1,Ser637位点磷酸化-Drp1(p-Drp1,Ser637),Caspase-3蛋白表达水平。与模型Dox组比较,Lut显著改善H9c2细胞活力(P<0.05)。Dox组与正常组进行比较得到3582个差异基因;Lut加Dox组与Dox组进行比较得到1981个差异基因。利用小干扰RNA(siRNA)技术成功构建低表达MIEF1心肌细胞,Western blot结果显示,Lut能够增加低表达MIEF1心肌细胞中MIEF1、p-Drp1(Ser637)蛋白表达水平(P<0.05),降低Caspase-3蛋白表达水平(P<0.05)。总之,Lut对H9c2心肌细胞的保护作用可能与促进MIEF1表达有关,本实验结果为Lut治疗阿霉素导致的心肌细胞损伤提供理论依据。
To study the effect and mechanism of luteolin(Lut)on H9c2 myocardial cell injury.The rat H9c2 myocardial cell line was used to establish a myocardial cell injury model with 1μM doxorubicin(Dox).Different concentrations of Lut(5,10,20μM)were used for intervention.Cell viability was measured by MTT assay.High-throughput transcriptome sequencing was used to screen the target genes of Lut(20μM)for improving H9c2 myocardial cell injury.The extend myocardial cell mitochondria Factor 1(mitochondrial elongation factor 1,MIEF1)expression was suppressed by use molecular cloning technology.Western blot was performed to detect the expression levels of MIEF1,phosphorylated-Drp1(p-Drp1,Ser637),and Caspase-3 in myocytes with low MIEF1 expression by Lut.Compared with Dox group,Lut significantly improved the cell viability of H9c2(P<0.05).3582 differentially expressed genes were found in Dox group compared with the normal group.1981 differentially expressed genes were obtained in Lut plus Dox group compared with Dox group.Small interfering RNA(siRNA)was used to interfere with the construction of MIEF1 low expression cardiomyocytes.Western blot results showed that,Lut increased the expression level of MIEF1 and p-Drp1(Ser637)protein in myocytes with low expression of MIEF1(P<0.05)and decreased the expression level of Caspase-3(P<0.05).In conclusion,the protective effect of Lut on H9c2 cardiomyocytes may be related to the promotion of MIEF1 expression,the results of this experiment provide a theoretical basis for the treatment of doxorubicin induced cardiomyocyte injury.
作者
史有阳
杨瑞
张洋
孙霃平
刘胜
SHI You-yang;YANG Rui;ZHANG Yang;SUN Chen-ping;LIU Sheng(Department of Breast Surgery (Integrated Traditional and Western Medicine),Longhua hospital affiliated to Shanghai university of traditional Chinese medicine,Shanghai 200032,China)
出处
《天然产物研究与开发》
CAS
CSCD
北大核心
2020年第5期820-825,共6页
Natural Product Research and Development
基金
国家自然科学基金(81603629)
上海中医药大学研究生创新能力培养专项(201812017053)。
