Date Published: July 05, 2017
Publisher: John Wiley and Sons Inc.
Author(s): Jun Ren, Lifang Yang, Li Zhu, Xihui Xu, Asli F. Ceylan, Wei Guo, Jian Yang, Yingmei Zhang.
Aging is accompanied with unfavorable geometric and functional changes in the heart involving dysregulation of Akt and autophagy. This study examined the impact of Akt2 ablation on life span and cardiac aging as well as the mechanisms involved with a focus on autophagy and mitochondrial integrity. Cardiac geometry, contractile, and intracellular Ca2+ properties were evaluated using echocardiography, IonOptix® edge‐detection and fura‐2 techniques. Levels of Sirt1, mitochondrial integrity, autophagy, and mitophagy markers were evaluated using Western blot. Our results revealed that Akt2 ablation prolonged life span (by 9.1%) and alleviated aging (24 months)‐induced unfavorable changes in myocardial function and intracellular Ca2+ handling (SERCA2a oxidation) albeit with more pronounced cardiac hypertrophy (58.1%, 47.8%, and 14.5% rises in heart weight, wall thickness, and cardiomyocyte cross‐sectional area). Aging downregulated levels of Sirt1, increased phosphorylation of Akt, and the nuclear transcriptional factor Foxo1, as well as facilitated acetylation of Foxo1, the effects of which (except Sirt1 and Foxo1 acetylation) were significantly attenuated or negated by Akt2 ablation. Advanced aging disturbed autophagy, mitophagy, and mitochondrial integrity as evidenced by increased p62, decreased levels of beclin‐1, Atg7, LC3B, BNIP3, PTEN‐induced putative kinase 1 (PINK1), Parkin, UCP‐2, PGC‐1α, and aconitase activity, the effects of which were reversed by Akt2 ablation. Aging‐induced cardiomyocyte contractile dysfunction and loss of mitophagy were improved by rapamycin and the Sirt1 activator SRT1720. Activation of Akt using insulin or Parkin deficiency prevented SRT1720‐induced beneficial effects against aging. In conclusion, our data indicate that Akt2 ablation protects against cardiac aging through restored Foxo1‐related autophagy and mitochondrial integrity.
Biological aging is an irreversible process featured by progressive myocardial remodeling, reduced cardiac reserve, diastolic dysfunction, and a higher cardiac morbidity and mortality in the elderly (Lakatta, 1999; Sussman & Anversa, 2004; Yang et al., 2005; Shimizu & Minamino, 2016). A number of hypotheses have been postulated for cardiac aging including oxidative stress, depletion of cellular fuel, myosin heavy chain isozyme switch, apoptosis, mitochondrial injury, autophagy dysregulation, and intracellular Ca2+ mishandling (Lakatta, 1999; Yang et al., 2005, 2006; Boengler et al., 2007; Feridooni et al., 2015; Horn, 2015). Nonetheless, the precise machineries behind cardiac aging still remain somewhat elusive. Recent evidence from our laboratory and others has depicted a unique role for phosphoinositide 3‐kinase (PI3K) and its downstream‐signaling target protein kinase B (Akt) in aging‐induced pathological changes in the heart (Inuzuka et al., 2009; Hua et al., 2011; Pillai et al., 2014). It was shown that the on‐and‐off switching of the PI3K/Akt pathway, particularly by insulin and insulin‐like growth factor‐1 (IGF‐1), serves as a powerful physiological integrator rudimentary to life span and aging (Nair & Ren, 2012; O’Neill, 2013). Our data have revealed an essential role for diminished autophagy, an evolutionarily conserved lysosome‐dependent process for turnover of proteins and organelles, in Akt overactivation‐induced accentuation of cardiac aging process (Hua et al., 2011). Autophagy plays a key role for biological aging process and cardiac homeostasis. Diminished autophagy has been shown to accelerate mammalian aging (Hars et al., 2007; Simonsen et al., 2008; Toth et al., 2008; Alvers et al., 2009; Taneike et al., 2010), in association with accumulation of damaged intracellular components including protein aggregate (Cuervo et al., 2005; Zhang & Cuervo, 2008; Gottlieb et al., 2009; Taneike et al., 2010). Moreover, defective autophagy facilitates ventricular remodeling, contractile defects, and heart failure (Nakai et al., 2007; Yitzhaki et al., 2009). Given the critical role for Akt in the regulation of cardiac survival and life span (Hay & Sonenberg, 2004; Hahn‐Windgassen et al., 2005), this study was designed to examine the role of Akt2 ablation on aging‐induced geometric, functional, and intracellular Ca2+ homeostatic changes in the heart, with a focus on autophagy and mitochondrial integrity. The Akt downstream‐signaling molecules including Forkhead transcriptional factors of Forkhead box O class (Foxo) were examined in wild‐type and Akt2 knockout mice. FOXOs in particular Foxo1 are associated with life span and serve as a paradigm for the understanding of aging‐related chronic diseases such as cancer and diabetes (Charitou & Burgering, 2013). Levels of autophagy and mitophagy protein markers including phosphatase and tensin homolog (PTEN)‐induced kinase 1 (PINK1), Parkin, and BCL2/adenovirus E1B 19‐kDa‐interacting protein 3 (BNIP3) were examined in young or aged mouse hearts. It has been depicted that mitochondrial priming is mediated either by the PINK1‐Parkin signaling pathway or the mitophagic receptors Nix and BNIP3 (Ding & Yin, 2012), in an effort to eliminate damaged mitochondria and to preserve mitochondrial integrity. Our findings indicated that Akt2 ablation prolongs life span and improves myocardial contractile function with a possible adaptive cardiac remodeling through the Sirt1‐mediated autophagy regulation.
Our salient findings revealed that ablation of Akt2 prolonged life span and rescued against aging‐associated contractile and intracellular Ca2+ defects, despite more pronounced ‘adaptive’ cardiac hypertrophy. In addition, Akt2 ablation alleviated aging‐associated mitochondrial injury. Our data further depicted an essential role for the Akt‐Foxo1 phosphorylation and Sirt1‐Foxo1 deacetylation cascades in regulation of autophagy in cardiac aging (as depicted in Fig. S3, Supporting information). The permissive roles of autophagy and Sirt1 in Akt2 knockout‐offered beneficial action received consolidation from the observations that autophagy inhibition using 3‐MA negated Akt2 ablation‐offered beneficial response against cardiac aging, whereas autophagy induction using rapamycin or Sirt1 activation using SRT1720 alleviated aging‐induced cardiomyocyte mitophagy and mechanical derangements. The fact that SRT1720‐offered beneficial response in aging was nullified or attenuated by insulin suggests possible offsetting effects between Akt and Sirt1 in cardiac homeostasis in aging. Our finding that Parkin knockout obliterated SRT1720‐offered benefit against aging depicted a mitophagy‐mediated mechanism in Sirt1‐offered benefit in aging. These results collectively suggest a possible role for Akt2 (the major Akt isozyme in the heart) and autophagy in cardiac aging and more importantly, the therapeutic potential for autophagy or mitophagy in cardiac aging.
This work was supported in part by grants from the National Institute of Health/National Institute of Aging (R03 AG21324), and the National Natural Science Foundation of China (81522004, 81570225, 81370195, 81521001).
JR, LY, and YZ designed the experiments, conducted the study, and prepared the manuscript; JR, LY, XX, AFC, WG conducted the study; LZ, WG, and JY reviewed the manuscript and contributed to the discussion.