Cumulative clinical and experimental evidence has revealed a cardinal role for mitochondrial integrity in cardiac aging. Parkin-mediated mitophagy is essential to ensure mitochondrial quality control in myocardium. This study was designed to examine the impact of Parkin overexpression on aging-induced myocardial anomalies and the underlying mechanisms with a focus on Parkin-regulated mitophagy. Cardiac function, myocardial apoptosis, mitochondrial ultrastructure and mitophagy were examined in young (3 mo) and old (24–26 mo) wild-type (WT) and Parkin transgenic mice. Our data revealed compromised myocardial function and mitochondrial morphology along with overtly apoptosis with advanced aging, the effects of which were attenuated by Parkin overexpression. Advanced aging dampened mitophagy as evidenced by decreased levels of Parkin, LC3II, phosphorylation of p62 and TBK1 in isolated mitochondria as well as reduced mitochondria autophagosomes, the effects of which were mitigated by restoration of mitophagy via Parkin overexpression. Using the low-dose doxorubicin (DOX) in vitro model of cell senescence, we noted that Parkin-offered beneficial effect against senescence was abolished by the TBK1 kinase inhibitor BX795. With TBK1 overexpression in cardiomyocytes, we uncovered the interaction of Parkin with TBK1 using a Co-immunoprecipitation (Co-IP) assay. The interaction of Parkin with TBK1 contributed to K63-linked polyubiquitination of TBK1. Our study also noted that DOX disturbed K63-linked polyubiquitination of TBK1 with downregulation of Parkin. Parkin overexpression promoted K63-linked polyubiquitination of TBK1 through Lys30 and Lys401 residues to foster TBK1 phosphorylation to facilitate efficient mitophagy. In summary, these findings suggested that Parkin effectively rescued cardiac aging through promoting K63-linked polyubiquitination of TBK1 to facilitate mitophagy.

累积的临床和实验证据表明，心脏衰老中线粒体完整性的重要作用。帕金介导的线粒体吞噬对于确保心肌线粒体的质量控制至关重要。本研究旨在检查帕金过表达对衰老引起的心肌异常的影响及其潜在机制，重点是帕金调节的线粒体。在年轻（3 mo）和老（24-26 mo）野生型（WT）和Parkin转基因小鼠中检查了心脏功能，心肌细胞凋亡，线粒体超微结构和线粒体。我们的数据显示，受损的心肌功能和线粒体形态以及明显的细胞凋亡伴随着衰老，其作用被帕金过表达所减弱。帕金森，LC3II，分离的线粒体中p62和TBK1的磷酸化以及线粒体自噬体的减少，通过帕金过表达恢复线粒体，减轻了其影响。使用低剂量的阿霉素（DOX）体外细胞衰老体外模型，我们注意到TBK1激酶抑制剂BX795取消了帕金提供的抗衰老有益作用。借助TBK1在心肌细胞中的过表达，我们使用共免疫沉淀（Co-IP）分析揭示了Parkin与TBK1的相互作用。帕金与TBK1的相互作用促进了TBK1的K63连接多泛素化。我们的研究还指出，DOX会干扰Parkin的下调，从而干扰TBK1的K63连锁多聚泛素化。Parkin过表达通过Lys30和Lys401残基促进TBK1的K63连接多泛素化，从而促进TBK1磷酸化，从而促进有效的线粒体吞噬。总之，这些发现表明，帕金通过促进TBK1的K63连锁多聚泛素化以促进线粒体吞噬，有效地挽救了心脏衰老。