Mitophagy In Cardiomyocytes Research Articles

Prolylcarboxypeptidase Mitigates Myocardial Ischemia/Reperfusion Injury by Stabilizing Mitophagy.

The role of prolylcarboxypeptidase (PRCP) in myocardial ischemia/reperfusion (I/R) injury is unclear. Herein, we aimed to evaluate the protective effect of the PRCP–angiotensin-(1–7) [Ang-(1–7)]/bradykinin-(1–9) [BK-(1–9)] axis on myocardial I/R injury and identify the mechanisms involved. Plasma PRCP level and activity, as well as Ang-(1–7) and BK-(1–9) levels, were compared in healthy subjects, patients with unstable angina, and those with ST-segment–elevated acute myocardial infarction (AMI). Thereafter, the effects of PRCP overexpression and knockdown on left ventricular function, mitophagy, and levels of Ang-(1–7) and BK-(1–9) were examined in rats during myocardial I/R. Finally, the effects of Ang-(1–7) and BK-(1–9) on I/R-induced mitophagy and the signaling pathways involved were investigated in vitro in rat cardiomyocytes. AMI patients showed increased plasma level and activity of PRCP and levels of Ang-(1–7) and BK-(1–9) as compared with healthy subjects and those with unstable angina. PRCP protected against myocardial I/R injury in rats by paradoxical regulation of cardiomyocyte mitophagy during the ischemia and reperfusion phases, which was mediated by downstream Ang-(1–7) and BK-(1–9). We further depicted a possible role of activation of AMPK in mitophagy induction during ischemia and activation of Akt in mitophagy inhibition during reperfusion in the beneficial effects of Ang-(1–7) and BK-(1–9). Thus, the PRCP–Ang-(1–7)/BK-(1–9) axis may protect against myocardial I/R injury by paradoxical regulation of cardiomyocyte mitophagy during ischemia and reperfusion phases.

Role of Parkin-mediated mitophagy in glucocorticoid-induced cardiomyocyte maturation

Glucocorticoids can promote cardiomyocyte maturation. However, the mechanism underlying glucocorticoid-mediated cardiomyocyte maturation is still unclear. Mitophagy plays a key role in cardiomyocyte maturation. Based on current knowledge, our study evaluated the effects of the glucocorticoid dexamethasone (100 nM) on the maturation of mouse embryonic stem cell-derived cardiomyocytes and the role of mitophagy in this maturation. The results showed that dexamethasone can promote embryonic stem cell-derived cardiomyocyte maturation, inhibit cardiomyocyte proliferation, and promote myocardial fiber arrangement. However, dexamethasone did not affect mitochondrial morphology in cardiomyocytes. Glucocorticoid receptor inhibitors (RU486, 1 nM) can inhibit dexamethasone-mediated cardiomyocyte maturation. Additionally, dexamethasone can promote mitophagy in embryonic stem cell-derived cardiomyocytes and induce LC3 and lysosomal aggregation in mitochondria. The inhibition of mitophagy can inhibit the cardiomyocyte maturation effect of dexamethasone. Furthermore, our research found that dexamethasone may mediate the occurrence of mitophagy in cardiomyocytes through Parkin. The siRNA-mediated inhibition of Parkin expression can inhibit mitochondrial autophagy caused by dexamethasone, thus inhibiting cardiomyocyte maturation. Overall, our study found that dexamethasone can promote embryonic stem cell-derived cardiomyocyte maturation through Parkin-mediated mitophagy.

Aldehyde dehydrogenase 2 preserves mitochondrial morphology and attenuates hypoxia/reoxygenation-induced cardiomyocyte injury.

Disturbance of mitochondrial fission and fusion (termed mitochondrial dynamics) is one of the leading causes of ischemia/reperfusion (I/R)-induced myocardial injury. Previous studies showed that mitochondrial aldehyde dehydrogenase 2 (ALDH2) conferred cardioprotective effect against myocardial I/R injury and suppressed I/R-induced excessive mitophagy in cardiomyocytes. However, whether ALDH2 participates in the regulation of mitochondrial dynamics during myocardial I/R injury remains unknown. In the present study, we investigated the effect of ALDH2 on mitochondrial dynamics and the underlying mechanisms using the H9c2 cells exposed to hypoxia/reoxygenation (H/R) as an in vitro model of myocardial I/R injury. Cardiomyocyte apoptosis was significantly increased after oxygen-glucose deprivation and reoxygenation (OGD/R), and ALDH2 activation largely decreased the cardiomyocyte apoptosis. Additionally, we found that both ALDH2 activation and overexpression significantly inhibited the increased mitochondrial fission after OGD/R. Furthermore, we found that ALDH2 dominantly suppressed dynamin-related protein 1 (Drp1) phosphorylation (Ser616) and adenosine monophosphate-activated protein kinase (AMPK) phosphorylation (Thr172) but not interfered with the expression levels of mitochondrial shaping proteins. We demonstrate the protective effect of ALDH2 against cardiomyocyte H/R injury with a novel mechanism on mitochondrial fission/fusion.

Poly (ADP‐ribose) polymerase inhibition protects against myocardial ischaemia/reperfusion injury via suppressing mitophagy

Myocardial ischaemia/reperfusion (I/R) injury attenuates the beneficial effects of reperfusion therapy. Poly(ADP‐ribose) polymerase (PARP) is overactivated during myocardial I/R injury. Mitophagy plays a critical role in the development of myocardial I/R injury. However, the effect of PARP activation on mitophagy in cardiomyocytes is unknown. In this study, we found that I/R induced PARP activation and mitophagy in mouse hearts. Poly(ADP‐ribose) polymerase inhibition reduced the infarct size and suppressed mitophagy after myocardial I/R injury. In vitro, hypoxia/reoxygenation (H/R) activated PARP, promoted mitophagy and induced cell apoptosis in cardiomyocytes. Poly(ADP‐ribose) polymerase inhibition suppressed H/R‐induced mitophagy and cell apoptosis. Parkin knockdown with lentivirus vectors inhibited mitophagy and prevented cell apoptosis in H/R‐treated cells. Poly(ADP‐ribose) polymerase inhibition prevented the loss of the mitochondrial membrane potential (ΔΨm). Cyclosporin A maintained ΔΨm and suppressed mitophagy but FCCP reduced the effect of PARP inhibition on ΔΨm and promoted mitophagy, indicating the critical role of ΔΨm in H/R‐induced mitophagy. Furthermore, reactive oxygen species (ROS) and poly(ADP‐ribosylation) of CypD and TSPO might contribute to the regulation of ΔΨm by PARP. Our findings thus suggest that PARP inhibition protects against I/R‐induced cell apoptosis by suppressing excessive mitophagy via the ΔΨm/Parkin pathway.

Abstract 160: Atg7-Dependent Activation of Mitochondrial Autophagy in Cardiomyocytes

Introduction: Autophagy-related protein 7 (Atg7) has been studied for its role in cellular macroautophagy induction and regulation in different cell systems. We have previously reported that Atg7 regulates cardiac macroautophagy. However, whether Atg7 plays any role in the initiation and regulation of mitochondrial autophagy (mitophagy) in cardiomyocytes remains elusive. Objective: We investigated subcellular localization and molecular function of Atg7 in the regulation of mitophagy through a series of in vitro and in vivo experiments. Methods and Results: We isolated mitochondria from mouse hearts by subcellular fractionation and conducted biochemical experiments which revealed localization of Atg7 on the mitochondrial membrane. Next, we co-stained for Atg7 in isolated heart mitochondria stained with Mitotracker dye and found Atg7 localizes to mitochondrial membrane. To elucidate molecular function of Atg7 on mitophagy, adenovirus-mediated Atg7 overexpressed cultured neonatal rat cardiomyocytes (NRCs) were treated with mitophagy inducer carbonyl cyanide 3-chlorophenylhydrazone (CCCP) in vitro and stained with organelle-specific mitochondria and lysosome dyes to count mitophagy fluorescent dots by confocal microscopy. Atg7 overexpressed NRCs exhibited an increase in mitophagy dots than β-gal treated NRCs following CCCP induction. Further, we have treated cardiac specific Atg7 overexpressed (Atg7 Tg) mice and littermate non-transgenic (Ntg) mice with CCCP to assess mitophagy induction in vivo . Immunoblotting confirmed enhanced accumulation of mature autophagolysosomal marker LC3-II protein in mitochondrial fraction from Atg7 Tg hearts. Conclusions: Our in vitro and in vivo findings indicate that Atg7 localizes to the mitochondrial membrane, regulates recruitment and initiation of mitophagy machinery, and facilitates mitophagy in cardiomyocytes.

Abstract 438: GRAF1 is a Novel Mediator of Parkin-dependent Mitophagy

Selective autophagic removal of damaged mitochondria by Parkin-mediated mitophagy is essential for metabolic homeostasis of long-lived cells and defects in this clearance pathway leads to the accumulation of toxic intermediates and accelerates the pathogenesis of numerous cardiometabolic diseases. This process is facilitated by the recruitment of the autophagosomal protein, LC3, by LC3-binding receptors that accumulate on damaged mitochondria. However, little is known about the precise LC3-receptors that mediate Parkin-dependent mitophagy in cardiomyocytes. Having previously identified a cardioprotective role for GRAF1, our current data indicate that GRAF1 plays critical role in promoting Parkin-dependent mitochondrial clearance. GRAF1 is expressed at high levels in the heart from E17 onwards and is poised to co-regulate actin- and lipid-dynamics by virtue of its multi-domain structure that includes a lipid binding/bending BAR domain, a phospholipid binding PH domain, a Rho-GAP domain, and a protein-interaction SH3 domain. Importantly, GRAF1 depletion in primary cardiomyocytes led to impaired mitochondrial OXPHOX-mediated ATP generation, mitochondrial membrane depolarization, increased mitochondrial-associated ROS, and increased ischemia/reperfusion-dependent myocyte death. These outcomes were accompanied by reduced LC3 mediated autophagic flux and accumulation of damaged mitochondria (and Parkin) in GRAF1 deficient cardiomyocytes and hearts. Mechanistically, we showed that GRAF1 facilitates Parkin-dependent mitophagy by serving as a novel LC3 receptor. GRAF1 is recruited to damaged mitochondria, is phosphorylated in a PINK1/Parkin-dependent fashion, and physically interacts with Parkin and LC3. While multiple LC3 receptors are recruited to depolarized mitochondria and act in a redundant function to promote mitochondrial engulfment, our studies reveal a unique and indispensable role for GRAF1 in this process. Our studies reveal that GRAF1 is distinctive because it facilitates mitochondrial clearance by further facilitating recruitment of Parkin and additional down-stream receptors to damaged mitochondria in a feed-forward fashion.

Abstract 164: Hypoxia-Induced Cardiomyocyte Mitophagy and Mitochondrial Permeability Transition are Inhibited by Bnip3 Phosphorylation

Bnip3 is a hypoxia-inducible initiator of cardiomyocyte cell death and has also been implicated as a mitochondrial receptor for the cellular mitophagy machinery. Previous work from our group demonstrates that the prostaglandin E1 analogue, misoprostol, prevents hypoxia-induced mitochondrial dysfunction and is a potent activator of PKA. We hypothesize that misoprostol alters the phosphorylation status of Bnip3, inhibiting its ability to induce cardiomyocyte mitophagy, mitochondrial dysfunction and cell death. Using a rodent model of neonatal hypoxia, in combination with rat primary ventricular neonatal cardiomyocytes (PVNC’s) and H9c2 cells, we assessed the effect of hypoxia and misoprostol drug treatments on mitochondrial function, mitophagy, and cell viability. In postnatal day 5 rats, hypoxia caused a 30% increase in the concentration of serum cardiac troponin-I, a clinically relevant marker for cardiomyocyte cell death, which was absent with the addition of misoprostol drug treatments (n=3). Using PVNC’s we further demonstrated that hypoxia reduced measures of mitochondrial function including membrane potential (47%) and maximal respiration (46%), which were restored back to control levels with the addition of misoprostol (p<0.01). Furthermore, hypoxia induced a 36% increase in mitophagy, concurrent with a 210% increase in mitochondrial permeability transition, both of which were reversed with misoprostol drug treatment (p<0.01). Using a combination of mass spectrometry and mutagenesis, we also show that PKA directly phosphorylates the transmembrane domain of Bnip3 to inhibit its function. Mechanistically, when the PKA phosphorylation site on Bnip3 was neutralized, the protective effect of misoprostol on mitochondrial membrane potential, mitophagy and permeability transition was lost. Taken together, these results demonstrate a foundational role for Bnip3 phosphorylation in the molecular regulation of cardiomyocyte mitochondrial dysfunction. These findings further identify a pharmacological mechanism, through PKA, which may ultimately be able to prevent hypoxia-induced myocardial injury.

Melatonin activates Parkin translocation and rescues the impaired mitophagy activity of diabetic cardiomyopathy through Mst1 inhibition.

Mitophagy eliminates dysfunctional mitochondria and thus plays a cardinal role in diabetic cardiomyopathy (DCM). We observed the favourable effects of melatonin on cardiomyocyte mitophagy in mice with DCM and elucidated their underlying mechanisms. Electron microscopy and flow cytometric analysis revealed that melatonin reduced the number of impaired mitochondria in the diabetic heart. Other than decreasing mitochondrial biogenesis, melatonin increased the clearance of dysfunctional mitochondria in mice with DCM. Melatonin increased LC3 II expression as well as the colocalization of mitochondria and lysosomes in HG‐treated cardiomyocytes and the number of typical autophagosomes engulfing mitochondria in the DCM heart. These results indicated that melatonin promoted mitophagy. When probing the mechanism, increased Parkin translocation to the mitochondria may be responsible for the up‐regulated mitophagy exerted by melatonin. Parkin knockout counteracted the beneficial effects of melatonin on the cardiac mitochondrial morphology and bioenergetic disorders, thus abolishing the substantial effects of melatonin on cardiac remodelling with DCM. Furthermore, melatonin inhibited Mammalian sterile 20‐like kinase 1 (Mst1) phosphorylation, thus enhancing Parkin‐mediated mitophagy, which contributed to mitochondrial quality control. In summary, this study confirms that melatonin rescues the impaired mitophagy activity of DCM. The underlying mechanism may be attributed to activation of Parkin translocation via inhibition of Mst1.

Mst1 inhibits Sirt3 expression and contributes to diabetic cardiomyopathy through inhibiting Parkin-dependent mitophagy

Mitochondrial dysfunction contributes to heart failure induced mortality in approximately 80% of diabetic patients. Mitophagy degrades defective mitochondria and maintains a healthy mitochondrial population, which is essential for cardiomyocyte survival in diabetic stress. Herein, we determined whether Mst1 regulated mitophagy and investigated the downstream signaling pathway in the development of diabetic cardiomyopathy (DCM). Mst1 deficiency promoted elimination of dysfunctional mitochondria in diabetic cardiomyopathy without affecting mitochondrial biogenesis. Enhanced mitophagy was observed in Mst1 interfering cardiomyocytes subjected to high glucose treatment using 3-Methyladenine and Chloroquine. Consistent with these results, in vivo and in vitro loss of function experiments indicated that Mst1 participated in the development of DCM by inhibiting Parkin-dependent mitophagy. Mst1 deficiency alleviated the detrimental phenotype of DCM. Interestingly, the protective effects of Mst1 knockout on DCM were compromised in diabetic Parkin−/− mice. Mechanistically, Mst1 knockdown significantly enhanced Parkin expression and translocation to the mitochondria, as evidenced by immunofluorescence study and Western blot analysis. Furthermore, Sirt3 deletion abolished the detrimental effects of Mst1 on DCM. Collectively, Mst1 inhibits Sirt3 expression thus participates in the development of DCM by inhibiting cardiomyocyte mitophagy. The mechanism is associated with Parkin inhibition.

P21‐Activated Kinase 1 is Necessary for Autophagy and Mitophagy in Cardiomyocytes

P21‐Activated Kinase 1 (PAK1) is a serine‐threonine kinase that regulates multiple signal transduction pathways in cardiomyocytes. Despite the large body of evidence indicating a key role for PAK1 in cardiac protective signalling, the mechanisms underlying the cardioprotective effects of PAK1 remain elusive. Autophagy is a cellular process in which unwanted organelles and proteins are degraded and recycled to maintain cellular homeostasis. Mitophagy is a form of selective autophagy that eliminates damaged mitochondria from the cell via the lysosome system to control mitochondrial quality. Autophagy and mitophagy are essential for cell survival in response to starvation, but may become detrimental under other conditions. How autophagy and mitophagy are regulated for adaptation in different contexts is under intense investigation. In this study, we explored whether PAK1 regulates autophagy and mitophagy in cardiomyocytes. PAK1 was knocked down by using siRNA in H9c2 cardiac myoblasts and then the expression levels of the autophagosome marker LC3‐II were analysed by Western blot. Interestingly, downregulation of PAK1 reduced LC3‐II levels in the total cell lysates, which was not significantly increased by the lysosomal protease inhibitors pepstatin A (pepA) and E64d, suggesting that autophagy flux is reduced by downregulation of PAK1. Similarly, PAK1 knockdown reduced LC3‐II expression levels in mitochondria either in the presence or absence of pepA and E64d, indicating limited mitochondrial degradation or mitophagy in the absence of PAK1. These results were reproduced in mouse hearts lacking PAK1. Together, our results suggest that PAK1 is essential to maintain the activities of autophagy and mitophagy in cardiomyocytes in vitro and in vivo. Current studies in our lab are investigating the functional significance of autophagy and mitophagy in PAK1‐induced myocardial protection.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Metformin inhibits autophagy and mitophagy in cardiomyocytes

Metformin, a first‐line medication for the treatment of type 2 diabetes mellitus, has been suggested by several clinical studies to be cardioprotective. However, the mechanisms underlying metformin's cardioprotection remain debated. One current hypothesis proposes that metformin protects the heart by increasing autophagy in cardiomyocytes, leading to increased removal of protein aggregates and damaged organelles including mitochondria. In this study, we tested this hypothesis using both in vitro and in vivo models. Rat H9C2 cardiac myoblasts were treated with 3 mM metformin for 24 hours and autophagy/mitophagy were determined by Western blot analysis of LC3‐II, a well‐established marker of autophagic vesicles. Surprisingly, metformin significantly reduced LC3‐II levels in either total cell lysates or mitochondrial fractions; and lysosomal inhibitors increased LC3‐II in metformin‐treated cells to a less degree as compared with vehicle‐treated cells. These results suggest that metformin inhibited rather than stimulated autophagy and mitophagy, contrary to the current belief. This was confirmed by a novel dual fluorescent mitophagy reporter showing reduced mitochondrial fragments being degraded in the lysosomes in the presence of metformin. The reduced mitophagy was also found in the heart from the mitophagy reporter mice that received 200 mg/kg of metformin through gavage. Collectively, these results indicate that the cardioprotective benefit of metformin observed in many different models may be through mechanisms other than increased autophagy and mitophagy.Support or Funding InformationThis work was supported by the National Institutes of Health National Center for Research Resources.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

PEDF protects cardiomyocytes by promoting FUNDC1‑mediated mitophagy via PEDF-R under hypoxic condition.

Pigment epithelial-derived factor (PEDF) is known to exert diverse physiological activities. Previous studies suggest that hypoxia could induce mitophagy. Astoundingly, under hypoxic condition, we found that PEDF decreased the mitochondrial density of cardiomyocytes. In this study, we evaluated whether PEDF could decrease the mitochondrial density and play a protective role in hypoxic cardiomyocytes via promoting mitophagy. Immunostaining and western blotting were used to analyze mitochondrial density and mitophagy of hypoxic cardiomyocytes. Gas chromatography-mass spectrometry and ELISA were used to analyze levels of palmitic acid and diacylglycerol. Transmission Electron Microscopy was used to detect mitophagy and the mitochondrial density in adult male Sprague-Dawley rat model of acute myocardial infarction. Compared to the control group, we observed that PEDF decreased mitochondrial density through promoting hypoxic cardiomyocyte mitophagy. PEDF increased the levels of palmitic acid and diacylglycerol, and then upregulated the levels of protein kinase Cα (PKC-α) and its activation. Furthermore, inhibition of PKC-α by Go6976 could effectively suppress PEDF-induced mitophagy. Besides, we found that PEDF promoted FUNDC1-mediated cardiomyocyte mitophagy via ULK1, which depended on the activation of PKC-α. Finally, we discovered that mitophagy was increased and mitochondrial density was reduced in adult male Sprague-Dawley rat model of acute myocardial infarction. We concluded that PEDF promotes mitophagy to protect hypoxic cardiomyocytes, through PEDF/PEDF-R/PA/DAG/PKC-α/ULK1/FUNDC1 pathway.

Mitophagy in Cardiomyocytes and in Platelets: A Major Mechanism of Cardioprotection Against Ischemia/Reperfusion Injury.

Mitophagy, a process that selectively removes damaged organelles by autolysosomal degradation, is an early cellular response to ischemia. Mitophagy is activated in both cardiomyocytes and platelets during ischemia/reperfusion (I/R) and heart disease conditions. We focus on the molecular regulation of mitophagy and highlight the role of mitophagy in cardioprotection.

AMPK activation serves a critical role in mitochondria quality control via modulating mitophagy in the heart under chronic hypoxia.

Mitochondrial biogenesis is one of the generally accepted regulatory mechanisms in the heart under chronic hypoxia. The precise quantity and quality control of mitochondria is critical for the survival and function of cardiomyocytes. Mitochondrial autophagy, also known as mitophagy, which selectively eliminates dysfunctional and unwanted mitochondria, is the most important type of mitochondrial quality control. However, the detailed molecular mechanisms of mitophagy in cardiomyocytes have been largely undefined. The present study investigated the role of adenosine 5′-monophosphate-activated protein kinase (AMPK) in mitophagy regulation in cardiomyocytes under chronic hypoxia. H9c2 cells were cultured under hypoxic conditions (1% O2) for different time periods. Mitochondrial biogenesis was confirmed and hypoxia was found to induce the collapse of mitochondrial membrane potential (ΛΨm) and increase the number of dysfunctional mitochondria. As expected, mitochondrial autophagy was increased significantly in cardiomyocytes exposed to hypoxic conditions for 48 h. AMPK was activated under hypoxia. Notably, when the activation of AMPK was enhanced by the AMPK agonist AICAR, mitochondrial autophagy was increased accordingly. By contrast, when AMPK activation was blocked, mitochondrial autophagy was decreased and cardiomyocyte apoptosis was increased. In conclusion, in the present study, mitophagy was activated and played a crucial role in cardioprotection under chronic hypoxia. AMPK was involved in mitophagy regulation, thereby providing a potential therapeutic target for heart diseases associated with chronic hypoxia.

BRG1 and BRM SWI/SNF ATPases redundantly maintain cardiomyocyte homeostasis by regulating cardiomyocyte mitophagy and mitochondrial dynamics in vivo.

There has been an increasing recognition that mitochondrial perturbations play a central role in human heart failure. Mitochondrial networks, whose function is to maintain the regulation of mitochondrial biogenesis, autophagy ('mitophagy') and mitochondrial fusion/fission, are new potential therapeutic targets. Yet our understanding of the molecular underpinning of these processes is just emerging. We recently identified a role of the SWI/SNF ATP-dependent chromatin remodeling complexes in the metabolic homeostasis of the adult cardiomyocyte using cardiomyocyte-specific and inducible deletion of the SWI/SNF ATPases BRG1 and BRM in adult mice (Brg1/Brm double mutant mice). To build upon these observations in early altered metabolism, the present study looks at the subsequent alterations in mitochondrial quality control mechanisms in the impaired adult cardiomyocyte. We identified that Brg1/Brm double-mutant mice exhibited increased mitochondrial biogenesis, increases in 'mitophagy', and alterations in mitochondrial fission and fusion that led to small, fragmented mitochondria. Mechanistically, increases in the autophagy and mitophagy-regulated proteins Beclin1 and Bnip3 were identified, paralleling changes seen in human heart failure. Evidence for perturbed cardiac mitochondrial dynamics included decreased mitochondria size, reduced numbers of mitochondria, and an altered expression of genes regulating fusion (Mfn1, Opa1) and fission (Drp1). We also identified cardiac protein amyloid accumulation (aggregated fibrils) during disease progression along with an increase in pre-amyloid oligomers and an upregulated unfolded protein response including increased GRP78, CHOP, and IRE-1 signaling. Together, these findings described a role for BRG1 and BRM in mitochondrial quality control, by regulating mitochondrial number, mitophagy, and mitochondrial dynamics not previously recognized in the adult cardiomyocyte. As critical to the pathogenesis of heart failure, epigenetic mechanisms like SWI/SNF chromatin remodeling seem more intimately linked to cardiac function and mitochondrial quality control mechanisms than previously realized.

BRG1 and BRM SWI/SNF ATPases redundantly maintain cardiomyocyte homeostasis by regulating cardiomyocyte mitophagy and mitochondrial dynamics in vivo

BRG1 and BRM SWI/SNF ATPases redundantly maintain cardiomyocyte homeostasis by regulating cardiomyocyte mitophagy and mitochondrial dynamics in vivo

A change of heart (mitochondria)

Heart Mitochondria Mitochondria provide an essential source of energy to drive cellular processes and are particularly important in heart muscle cells (see the Perspective by Gottlieb and Bernstein). After birth, the availability of oxygen and nutrients to organs and tissues changes. This invokes changes in metabolism. Gong et al. studied the developmental transitions in mouse heart mitochondria soon after birth. Mitochondria were replaced wholesale via mitophagy in cardiomyocytes over the first 3 weeks after birth. Preventing this turnover by interfering with parkin-mediated mitophagy specifically in cardiomyocytes prevented the normal metabolic transition and caused heart failure. Thus, the heart has coopted a quality-control pathway to facilitate a major developmental transition after birth. Wai et al. examined the role of mitochondrial fission and fusion in mouse cardiomyocytes. Disruption of these processes led to “middle-aged” death from a form of dilated cardiomyopathy. Mice destined to develop cardiomyopathy were protected by feeding with a high-fat diet, which altered cardiac metabolism. Science , this issue p. [10.1126/science.aad2459][1], p. [10.1126/science.aad0116][2]; see also p. [1162][3] [1]: /lookup/doi/10.1126/science.aad2459 [2]: /lookup/doi/10.1126/science.aad0116 [3]: /lookup/volpage/350/1162?iss=6265

Imbalanced OPA1 processing and mitochondrial fragmentation cause heart failure in mice.

Mitochondrial morphology is shaped by fusion and division of their membranes. Here, we found that adult myocardial function depends on balanced mitochondrial fusion and fission, maintained by processing of the dynamin-like guanosine triphosphatase OPA1 by the mitochondrial peptidases YME1L and OMA1. Cardiac-specific ablation of Yme1l in mice activated OMA1 and accelerated OPA1 proteolysis, which triggered mitochondrial fragmentation and altered cardiac metabolism. This caused dilated cardiomyopathy and heart failure. Cardiac function and mitochondrial morphology were rescued by Oma1 deletion, which prevented OPA1 cleavage. Feeding mice a high-fat diet or ablating Yme1l in skeletal muscle restored cardiac metabolism and preserved heart function without suppressing mitochondrial fragmentation. Thus, unprocessed OPA1 is sufficient to maintain heart function, OMA1 is a critical regulator of cardiomyocyte survival, and mitochondrial morphology and cardiac metabolism are intimately linked.

Parkin-mediated mitophagy directs perinatal cardiac metabolic maturation in mice.

In developing hearts, changes in the cardiac metabolic milieu during the perinatal period redirect mitochondrial substrate preference from carbohydrates to fatty acids. Mechanisms responsible for this mitochondrial plasticity are unknown. Here, we found that PINK1-Mfn2-Parkin-mediated mitophagy directs this metabolic transformation in mouse hearts. A mitofusin (Mfn) 2 mutant lacking PINK1 phosphorylation sites necessary for Parkin binding (Mfn2 AA) inhibited mitochondrial Parkin translocation, suppressing mitophagy without impairing mitochondrial fusion. Cardiac Parkin deletion or expression of Mfn2 AA from birth, but not after weaning, prevented postnatal mitochondrial maturation essential to survival. Five-week-old Mfn2 AA hearts retained a fetal mitochondrial transcriptional signature without normal increases in fatty acid metabolism and mitochondrial biogenesis genes. Myocardial fatty acylcarnitine levels and cardiomyocyte respiration induced by palmitoylcarnitine were concordantly depressed. Thus, instead of transcriptional reprogramming, fetal cardiomyocyte mitochondria undergo perinatal Parkin-mediated mitophagy and replacement by mature adult mitochondria. Mitophagic mitochondrial removal underlies developmental cardiomyocyte mitochondrial plasticity and metabolic transitioning of perinatal hearts.

Abstract 19536: Lipid Oversupply to Cardiomyocytes Induces Sphingolipid-dependent Oxidative Stress and Induction of Mitophagy Through Ceramide Synthase 2

Introduction: Diabetic cardiomyopathy (DbCM) contributes to the high risk of heart failure (HF) in diabetics, but mechanisms underlying DbCM remain unclear. We previously showed that highly saturated fat feeding in mice altered cardiomyocyte sphingolipid profiles leading to DbCM and that some of these maladaptations were dependent on autophagy and ceramide synthase 5 (CerS5). Hypothesis: In the present study, we sought to further understand the cellular processes in which lipid overload leads to DbCM in in the context of sphingolipids. Methods: Mice fed an obesogenic diet and in vitro studies using H9c2 cardiomyocytes were utilized in this study. Results: Increased oxidative stress and enhanced apoptosis were identified in the hearts of animals subjected to lipid overload, while animals treated with the sphingolipid synthesis inhibitor myriocin were protected. Similarly, cardiomyocytes treated with palmitate showed a sphingolipid-dependent increase of reactive oxygen species (ROS) and mitophagy. Treatment with mitochondria-targeted ceramide analogs induced cardiomyocyte cell death in an increasing chain length manner, which was exacerbated by inhibiting mitophagy. This suggested that lipotoxicity to cardiomyocytes occurs in part through oxidative stress in a sphingolipid-dependent manner, and mitophagy may occur to prevent further damage. Our observation that only longer chain ceramides led to these outcomes suggested involvement of CerS2 which specifically produces very long chain ceramides (C20 and higher). Overexpression of CerS2 showed increased mitophagy in cardiomyocytes and knockdown of CerS2 by CRISPR-CAS9 technology decreased mitophagy. Conclusions: Taken together, our data suggest that lipid overload induces mitophagy potentially as a protective measure in defense from CerS2-induced mitochondrial damage, oxidative stress and cell death in DbCM.