摘要
目的探讨含YTH域家族蛋白1(YTHDF1)对缺氧复氧(H/R)条件下H9c2细胞凋亡和氧化应激的影响。方法用H9c2细胞构建H/R模型,并用YTHDF1小干扰RNA(siRNA)和过表达质粒转染细胞,采用MTT比色法测定细胞活力,并用赫斯特染色及流式细胞术测定细胞凋亡及细胞中超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量和细胞培养基上清中乳酸脱氢酶(LDH)活性,Western blot法测定细胞中YTHDF1、cleaved caspase-3、Bax和FasL的蛋白水平,比较对照组(control组)、H/R组、YTHDF1基因干扰和过表达组H9c2细胞凋亡和氧化损伤情况。结果H/R损伤后LDH活性和MDA含量明显升高(P<0.05),SOD活性和细胞活力显著降低(P<0.05),H/R处理后H9c2细胞中YTHDF1的蛋白水平均显著升高(P<0.05);YTHDF1干扰减轻了H/R损伤引起的形态学变化,YTHDF1过表达增加了H/R损伤引起的细胞形态变化;H/R损伤处理后cleaved caspase-3、Bax和FasL蛋白表达显著高于control组(P<0.05);H/R+YTHDF1-siRNA组cleaved caspase-3、Bax和FasL的蛋白水平显著低于H/R组(P<0.05),H/R+pcDNA3.1-YTHDF1组cleaved caspase-3、Bax和FasL蛋白水平显著高于H/R组(P<0.05)。结论下调YTHDF1可以抑制H/R处理的H9c2细胞cleaved caspase-3、Bax和FasL蛋白水平的上调并降低H/R处理的心肌细胞氧化损伤。
Objective To investigate the effect of YTH domain family protein 1(YTHDF1)on apoptosis and oxidative damage in H9c2 cells under hypoxic reoxygenation(H/R)conditions.Methods The H/R model was constructed with H9c2 cells.Cells were transfected with YTHDF1 small interfering RNA(siRNA)and overexpression plasmid,and cell viability was measured by MTT colorimetric assay.Apoptosis,superoxide dismutase(SOD)activity,malondialdehyde(MDA)content in cells and lactate dehydrogenase(LDH)activity in cell culture medium supernatant were measured by Hirst staining and flow cytometry.The protein levels of YTHDF1,cleaved caspase-3,Bax and FasL in the cells were determined by Western blot,and the apoptosis and oxidative damage of H9c2 cells in the control,H/R,YTHDF1 gene interference and overexpression groups were compared.Results LDH activity and MDA content were significantly increased,SOD activity and cell viability were significantly decreased after H/R injury(P<0.05),and the protein levels of YTHDF1 in H9c2 cells were significantly increased after H/R treatment(P<0.05),YTHDF1 interference alleviated the morphological changes caused by H/R injury,and YTHDF1 overexpression increased the morphological changes caused by H/R injury;cleaved caspase-3,Bax and FasL protein expression was significantly higher after H/R injury treatment than in the control group(P<0.05),cleaved caspase-3,Bax and FasL protein levels were significantly lower in the H/R+pcDNA3.1-YTHDF1 group than in the H/R group(P<0.05)and cleaved caspase-3,Bax and FasL protein levels were significantly higher in the H/R+pcDNA3.1-YTHDF1 group than in the H/R group(P<0.05).Conclusion Downregulation of YTH-DF1 inhibited the up-regulation of cleaved caspase-3,Bax and FasL protein levels and decreased oxidative damage in H/R-treated H9c2 cells.
作者
涅鲁排尔·热依木江
金颖
罗荔
张雅玲
刘刚
李雪
罗梅
Nielupaier Reyimujiang;Jin Ying;Luo Li;Zhang Yaling;Liu Gang;Li Xue;Luo Mei(The Fifth Affiliated Hospital of Xinjiang Medical University,Urumqi 830000)
出处
《国际老年医学杂志》
2024年第1期46-52,共7页
International Journal of Geriatrics
基金
新疆维吾尔自治区自然科学基金项目(2022D01C323)。
