Impairments in macroautophagy/autophagy, which degrades dysfunctional organelles as well as long-lived and aggregate proteins, are associated with several cardiomyopathies; however, the regulation of cardiac autophagy remains insufficiently understood. In this regard, ULK1 and ULK2 are thought to play primarily redundant roles in autophagy initiation, but whether their function is developmentally determined, potentially having an impact on cardiac integrity and function remains unknown. Here, we demonstrate that perinatal loss of ULK1 or ULK2 in cardiomyocytes (cU1-KO and cU2-KO mice, respectively) enhances basal autophagy without altering autophagy machinery content while preserving cardiac function. This increased basal autophagy is dependent on the remaining ULK protein given that perinatal loss of both ULK1 and ULK2 in cU1/2-DKO mice impaired autophagy causing age-related cardiomyopathy and reduced survival. Conversely, adult loss of cardiac ULK1, but not of ULK2 (i.e., icU1-KO and icU2-KO mice, respectively), led to a rapidly developing cardiomyopathy, heart failure and early death. icU1-KO mice had impaired autophagy with robust deficits in mitochondrial respiration and ATP synthesis. Trehalose ameliorated autophagy impairments in icU1-KO hearts but did not delay cardiac dysfunction suggesting that ULK1 plays other critical, autophagy-independent, functions in the adult heart. Collectively, these results indicate that cardiac ULK1 and ULK2 are functionally redundant in the developing heart, while ULK1 assumes a more unique, prominent role in the adult heart. Abbreviations: ATG4: autophagy related 4, cysteine peptidase; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG9: autophagy related 9; ATG13: autophagy related 13; CYCS: Cytochrome C; DNM1L, dynamin 1-like; MAP1LC3A: microtubule-associated protein 1 light chain 3 alpha; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MFN1: mitofusin 1; MFN2: mitofusin 2; MT-CO1: mitochondrially encoded cytochrome c oxidase I; MYH: myosin, heavy polypeptide; NBR1: NBR1 autophagy cargo receptor; NDUFA9: NADH:ubiquinone oxidoreductase subunit A9; OPA1: OPA1, mitochondrial dynamin like GTPase; PPARGC1A, peroxisome proliferator activated receptor, gamma, coactivator 1 alpha; SDHA: succinate dehydrogenase complex, subunit A, flavoprotein (Fp); SQSTM1: sequestosome 1; ULK1: unc-51 like kinase 1; ULK2: unc-51 like kinase 2; UQCRC1: ubiquinol-cytochrome c reductase core protein 1

Nitric oxide (NO) stimulates mitochondrial biogenesis in skeletal muscle. However, NO metabolism is disrupted in individuals with type 2 diabetes mellitus (T2DM) potentially contributing to their decreased cardiorespiratory fitness (i.e., VO₂max) and skeletal muscle oxidative capacity. We used a randomized, double-blind, placebo-controlled, 8-week trial with beetroot juice containing nitrate (NO₃⁻) and nitrite (NO₂⁻) (250 mg and 20 mg/day) to test potential benefits on VO₂max and skeletal muscle oxidative capacity in T2DM. T2DM (N = 36, Age = 59 ± 9 years; BMI = 31.9 ± 5.0 kg/m²) and age- and BMI-matched non-diabetic controls (N = 15, Age = 60 ± 9 years; BMI = 29.5 ± 4.6 kg/m²) were studied. Mitochondrial respiratory capacity was assessed in muscle biopsies from a subgroup of T2DM and controls (N = 19 and N = 10, respectively). At baseline, T2DM had higher plasma NO₃⁻ (100%; p < 0.001) and lower plasma NO₂⁻ levels (−46.8%; p < 0.0001) than controls. VO₂max was lower in T2DM (−26.4%; p < 0.001), as was maximal carbohydrate- and fatty acid-supported oxygen consumption in permeabilized muscle fibers (−26.1% and −25.5%, respectively; p < 0.05). NO₃⁻/NO₂⁻ supplementation increased VO₂max (5.3%; p < 0.01). Further, circulating NO₂⁻, but not NO₃⁻, positively correlated with VO₂max after supplementation (R²= 0.40; p < 0.05). Within the NO₃⁻/NO₂⁻ group, 42% of subjects presented improvements in both carbohydrate- and fatty acid-supported oxygen consumption in skeletal muscle (vs. 0% in placebo; p < 0.05). VO₂max improvements in these individuals tended to be larger than in the rest of the NO₃⁻/NO₂⁻ group (1.21 ± 0.51 mL/(kg*min) vs. 0.31 ± 0.10 mL/(kg*min); p = 0.09). NO₃⁻/NO₂⁻ supplementation increases VO₂max in T2DM individuals and improvements in skeletal muscle oxidative capacity appear to occur in those with more pronounced increases in VO₂max.

Non-alcoholic fatty liver disease (NAFLD) has recently become a public health concern concurrent with the obesity crisis. Previous work has shown aberrant mitochondrial content/quality and autophagy in models of NAFLD, whereas exercise is known to improve these derangements. The purpose of this study was to examine the effect of different weight-loss modalities on hepatic mitochondrial content, autophagy and mitophagy in NAFLD. Forty-eight male C57BL/6J mice were divided into 1 of 4 groups: low fat diet (LFD, 10% fat, 18 weeks), high fat diet (HFD, 60% fat diet, 18 weeks), weight-loss by diet (D, 60% fat diet for 10 weeks then 10% fat diet for 8 weeks) or weight-loss by diet and physical activity (D/PA, 60% fat diet for 10 weeks, then 10% fat diet plus a running wheel for 8 weeks). Immunoblot data were analyzed by one-way analysis of variance (ANOVA) with significance denoted at p ​< ​0.05. COX-IV protein contents were approximately 50% less in HFD compared to LFD. D/PA had 50% more BNIP3 compared to HFD. PINK1 content was 40% higher in D and D/PA compared to LFD. P-PARKIN/PARKIN levels were 40% lower in HFD, D, and D/PA compared to LFD. Whereas p-Ub^Ser65 was 3-fold higher in HFD. LC3II/I ratio was 50% greater in HFD and D/PA, yet p62 protein content was 2.5 fold higher in HFD. High-fat diet causes disruptions in markers of mitochondrial quality control. Physical activity combined with diet were able to ameliorate these derangements and seemingly improve hepatic mitochondrial quality above control values.