心脏细胞衰老与Uvrag基因的缺失

Ultraviolet resistance-associated gene(Uvrag)deficiency promotes cellular senescence in the heart

背景:前期工作发现自噬相关基因Uvrag缺失导致年龄相关心肌病,但细胞衰老在此过程中的作用尚不清楚。目的:研究Uvrag基因缺失对心脏细胞衰老的影响。方法:从Uvrag基因缺失纯合子小鼠和野生型小鼠中各取14只雄性小鼠分两批分别饲养到3月龄和8月龄。采用real time RT-PCR检测小鼠心脏组织衰老相关分泌表型相关因子mRNA表达变化,苏木精-伊红染色、天狼猩红染色、衰老相关β-半乳糖苷酶染色进行心肌组织学观察,透射电镜观察小鼠心肌细胞超微结构的变化,蛋白免疫印迹检测小鼠心脏组织p53蛋白表达量。实验方案经上海交通大学动物实验伦理委员会批准。结果与结论:①苏木精-伊红染色和天狼猩红染色结果显示,Uvrag基因缺失导致单个心肌细胞显著增大及心脏纤维化;②透射电镜显示,Uvrag基因缺失小鼠心肌细胞线粒体形态异常、排列紊乱,肌浆网肿胀;③衰老相关β-半乳糖苷酶染色发现,Uvrag基因缺失小鼠心脏衰老细胞显著增多;④real time RT-PCR结果表明,Uvrag基因缺失小鼠心脏衰老相关分泌表型因子表达显著上调;⑤Western blot结果显示Uvrag基因缺失导致心脏组织衰老调控关键蛋白p53表达显著升高;⑥结果说明,自噬相关基因Uvrag缺失促进小鼠心脏细胞衰老;Uvrag基因是潜在的延缓心脏衰老及抗衰老相关心脏疾病的靶分子。

BACKGROUND:Previously we have shown that ultraviolet resistance-associated gene(Uvrag)deficient mice develop age-related cardiomyopathy.However,whether cellular senescence contributes to the pathogenesis remains unknown.OBJECTIVE:To determine cellular senescence in Uvrag-deficient hearts.METHODS:Wild-type and Uvrag-deficient male mice at 3 and 8 months of age were utilized.Real-time RT-PCR was used to detect the changes in the mRNA expression of senescence-associated secretory phenotype-related factors in mouse heart tissue,and myocardial histological observation was performed using hematoxylin-eosin staining,Sirius scarlet staining,and aging-relatedβ-galactosidase staining.Transmission electron microscopy was used to observe the ultrastructure changes of mouse cardiomyocytes.Western blot was used to detect the expression of p53 protein in mouse heart tissue.An approval was obtained from the Experimental Animal Ethics Committee of Shanghai Jiaotong University.RESULTS AND CONCLUSION:Hematoxylin-eosin staining and Sirius red staining showed that cross-sectional area of individual cardiomyocytes was enlarged and cardiac fibrosis was enhanced in the Uvrag-deficient mice.In addition,cardiomyocytes in the Uvrag-deficient mice exhibited abnormal mitochondrial morphology,disordered arrangement,and swollen sarcoplasmic reticulum as revealed by transmission electron microscopy.Senescence-associated?-galactosidase staining suggested that Uvrag deficiency significantly increased senescent cells in the hearts from Uvrag-deficient mice.Furthermore,real-time RT-PCR demonstrated a marked increase in the expression of the senescence-associated secretory phenotype genes in Uvrag-deficient hearts.Finally,western blot showed that the expression of p53,a key regulator of cellular senescence,was upregulated in Uvrag-deficient hearts.To conclude,Uvrag deficiency promotes cellular senescence in the mouse heart.Uvrag is a potential target for delaying senescence of the heart and anti-aging-related heart disease.