28Aging is associated with an increased risk of cardiovascular disease and death. Here we 29show that oral supplementation of the natural polyamine spermidine extends the lifespan of 30 mice and exerts cardioprotective effects, reducing cardiac hypertrophy and preserving 31 diastolic function in old mice. Spermidine feeding enhanced cardiac autophagy, mitophagy 32 and mitochondrial respiration, and it also improved the mechano-elastical properties of 33 cardiomyocytes in vivo, coinciding with increased titin phosphorylation and suppressed 34 subclinical inflammation. Spermidine feeding failed to provide cardioprotection in mice that 35 lack the autophagy-related protein Atg5 in cardiomyocytes. In Dahl salt-sensitive rats that 36 were fed a high-salt diet, a model for hypertension-induced congestive heart failure, 37 spermidine feeding reduced systemic blood pressure, increased titin phosphorylation and 38 prevented cardiac hypertrophy and a decline in diastolic function, thus delaying the 39 progression to heart failure. In humans, high levels of dietary spermidine, as assessed from 40 food questionnaires, correlated with reduced blood pressure and a lower incidence of 41 cardiovascular disease. Our results suggest a new and feasible strategy for the protection 42 from cardiovascular disease. 43Author's manuscript to Eisenberg et al.

Diabetic patients develop cardiomyopathy characterized by hypertrophy, diastolic dysfunction, lipotoxicity, and mitochondrial dysfunction. How mitochondrial function is regulated in diabetic cardiomyopathy remains poorly understood. Mice were fed either a normal diet (ND) or a high fat diet (HFD, 60 kcal % fat). Mitophagy, evaluated with Mito‐Keima, was increased after 3 weeks of HFD feeding (mitophagy area: 8.3% per cell with ND and 12.4% with HFD) and continued to increase after 20 weeks (p<0.05). Although we have shown recently that mitophagy during the early phase of HFD feeding is mediated by Atg7‐dependent mechanisms, the mechanisms mediating mitophagy in the heart during the chronic phase of HFD feeding remain poorly understood. Phosphorylation of ULK1 was activated and Rab9 protein level was increased in the mitochondrial fraction within 20 weeks of HFD consumption (p<0.05). By isolating adult cardiomyocytes from GFP‐Rab9 transgenic mice fed HFD, we discovered that mitochondria were sequestrated by Rab9‐positive ring‐like structures. Since ULK1 regulates Rab9‐dependent mitophagy, we fed ULK1 cKO mice with HFD for 20 weeks. In wild type (WT) mice, cardiac hypertrophy and diastolic dysfunction (EDPVR = 0.051±0.009 in ND and 0.115±0.006 in HFD) were induced after 20 weeks of HFD feeding (p<0.05). By crossing Tg‐Mito‐Keima mice with ULK1 cKO mice, we found that downregulation of ULK1 impaired mitophagy in response to ND or 20 weeks of HFD consumption (p<0.05). Deletion of ULK1 exacerbated diastolic dysfunction (EDPVR=0.115±0.006 in WT and 0.162±0.021 in ULK1 cKO, p<0.05) and even induced systolic dysfunction (ESPVR=22.74±2.13 in WT and 16.78±2.12 in ULK1 cKO, p<0.05) during HFD feeding. Electron microscopic analyses indicated that the mitochondrial cristae structure was disrupted more severely in ULK1 cKO mice with HFD feeding than control mice (p<0.05). In summary, genetic disruption of ULK1‐Rab9‐dependent mitophagy during the chronic phase of HFD feeding exacerbates mitochondrial dysfunction, thereby facilitating the development of diabetic cardiomyopathy. ULK1‐Rab9‐dependent mitophagy serves as an essential quality control mechanism for cardiac mitochondria during HFD feeding. Support or Funding Information The project was supported by AHA and NIH (5R01HL138720‐02). This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

TRE reduced MI-induced cardiac remodeling and dysfunction through activation of autophagy.

Rationale: Obesity-associated cardiomyopathy characterized by hypertrophy and mitochondrial dysfunction. Mitochondrial quality control mechanisms, including mitophagy, are essential for the maintenance of cardiac function in obesity-associated cardiomyopathy. However, autophagic flux peaks at around 6 weeks of high-fat diet (HFD) consumption and declines thereafter. Objective: We investigated whether mitophagy is activated during the chronic phase of cardiomyopathy associated with obesity (obesity cardiomyopathy) after general autophagy is downregulated and, if so, what the underlying mechanism and the functional significance are. Methods and Results: Mice were fed either a normal diet or a HFD (60 kcal% fat). Mitophagy, evaluated using Mito-Keima, was increased after 3 weeks of HFD consumption and continued to increase after conventional mechanisms of autophagy were inactivated, at least until 24 weeks. HFD consumption time-dependently upregulated both Ser555-phosphorylated Ulk1 (unc-51 like kinase 1) and Rab9 (Ras-related protein Rab-9) in the mitochondrial fraction. Mitochondria were sequestrated by Rab9-positive ring-like structures in cardiomyocytes isolated from mice after 20 weeks of HFD consumption, consistent with the activation of alternative mitophagy. Increases in mitophagy induced by HFD consumption for 20 weeks were abolished in cardiac-specific ulk1 knockout mouse hearts, in which both diastolic and systolic dysfunction were exacerbated. Rab9 S179A knock-in mice, in which alternative mitophagy is selectively suppressed, exhibited impaired mitophagy and more severe cardiac dysfunction than control mice following HFD consumption for 20 weeks. Overexpression of Rab9 in the heart increased mitophagy and protected against cardiac dysfunction during HFD consumption. HFD-induced activation of Rab9-dependent mitophagy was accompanied by upregulation of TFE3 (transcription factor binding to IGHM enhancer 3), which plays an essential role in transcriptional activation of mitophagy. Conclusions: Ulk1-Rab9-dependent alternative mitophagy is activated during the chronic phase of HFD consumption and serves as an essential mitochondrial quality control mechanism, thereby protecting the heart against obesity cardiomyopathy.

SummaryThe age-associated deterioration in cellular and organismal functions associates with dysregulation of nutrient-sensing pathways and disabled autophagy. The reactivation of autophagic flux may prevent or ameliorate age-related metabolic dysfunctions. Non-toxic compounds endowed with the capacity to reduce the overall levels of protein acetylation and to induce autophagy have been categorized as caloric restriction mimetics (CRMs). Here, we show that aspirin or its active metabolite salicylate induce autophagy by virtue of their capacity to inhibit the acetyltransferase activity of EP300. While salicylate readily stimulates autophagic flux in control cells, it fails to further increase autophagy levels in EP300-deficient cells, as well as in cells in which endogenous EP300 has been replaced by salicylate-resistant EP300 mutants. Accordingly, the pro-autophagic activity of aspirin and salicylate on the nematode Caenorhabditis elegans is lost when the expression of the EP300 ortholog cpb-1 is reduced. Altogether, these findings identify aspirin as an evolutionary conserved CRM.