Introduction

Research reveals cardiac regeneration exists at levels previously deemed unattainable. Clinical trials using stem cells demonstrate promising cardiomyogenic and regenerative potential but insufficient contractile recovery. Incomplete understanding of the biology of administered cells likely contributes to inconsistent patient outcomes. Metabolism is a core component of many well-characterized stem cell types, and metabolic changes fundamentally alter stem cell fate from self-renewal to lineage commitment, and vice versa. However, the metabolism of stem cells currently studied for cardiac regeneration remains incompletely understood. Areas covered: Key metabolic features of stem cells are reviewed and unique stem cell metabolic characteristics are discussed. Metabolic changes altering stem cell fate are considered from quiescence and self-renewal to lineage commitment. Key metabolic concepts are applied toward examining cardiac regeneration through stem cell-based approaches, and clinical implications of current cell therapies are evaluated to identify potential areas of improvement. Expert commentary: The metabolism and biology of stem cells used for cardiac therapy remain poorly characterized. A growing appreciation for the fundamental relationship between stem cell functionality and metabolic phenotype is developing. Future studies unraveling links between cardiac stem cell metabolism and regenerative potential may considerably improve treatment strategies and therapeutic outcomes.

The lysosomal degradation of misfolded proteins and dysfunctional organelles by the process of autophagy is critical for the maintenance of heart function. Parkin is an E3 ubiquitin ligase that participates in the removal of dysfunctional mitochondria via mitochondrial autophagy (mitophagy). Mutations in the PARK2 gene encoding Parkin are linked to juvenile recessive forms of Parkinson's disease. As a result, Parkin has mainly been studied in the context of neurodegenerative disease. However, Parkin is highly expressed in the heart, an organ that is rich in mitochondria and therefore susceptible to cell death caused by mitochondrial dysfunction. The functional role of Parkin in the heart remains largely unexplored. While characterizing the cardiac phenotype of Parkin knockout (Parkin-/-) mice, I found that Parkin was not essential for the normal turnover of mitochondria in the heart. Instead, I determined that Parkin was a critical component in the response to acute cardiac stress such as a myocardial infarction (MI). I found that Parkin supported the adaption to MI stress by enabling mitophagy in the infarction border zone for the clearance of dysfunctional mitochondria. Further, I determined that the mechanism of Parkin function in the heart was dependent on the activity of the mitochondrial fission protein Drp1, but did not depend on the mitochondrial kinase PINK1. Lastly, I investigated the role of Parkin in response to chronic pathophysiological stresses that cause cardiac hypertrophy. Parkin-/- mice had impairments in both physiological hypertrophy in response to exercise and pathological hypertrophy in response to transverse aortic constriction. In contrast, cardiac-specific Parkin transgenic mice (Parkin-TG) had a normal hypertrophic response to pressure overload, but rapidly developed heart failure. Thus, I conclude that Parkin is crucial for the development of cardiac hypertrophy in response to chronic stress in addition to its role in adaptation to acute stress. Parkin may be a viable therapeutic target to promote autophagic clearance of dysfunctional mitochondria, but excessive Parkin may be detrimental and can result in a predisposition to heart failure in response to chronic stress

Background

Aberrant Ca²⁺ handling is a prominent feature of heart failure. Elucidation of the molecular mechanisms responsible for aberrant Ca²⁺ handling is essential for the development of strategies to blunt pathological changes in calcium dynamics. The peptidyl-prolyl cis-trans isomerase peptidyl-prolyl isomerase 1 (Pin1) is a critical mediator of myocardial hypertrophy development and cardiac progenitor cell cycle. However, the influence of Pin1 on calcium cycling regulation has not been explored. On the basis of these findings, the aim of this study is to define Pin1 as a novel modulator of Ca²⁺ handling, with implications for improving myocardial contractility and potential for ameliorating development of heart failure.

Methods and results

Pin1 gene deletion or pharmacological inhibition delays cytosolic Ca²⁺ decay in isolated cardiomyocytes. Paradoxically, reduced Pin1 activity correlates with increased sarco(endo)plasmic reticulum calcium ATPase (SERCA2a) and Na²⁺/Ca²⁺ exchanger 1 protein levels. However, SERCA2a ATPase activity and calcium reuptake were reduced in sarcoplasmic reticulum membranes isolated from Pin1-deficient hearts, suggesting that Pin1 influences SERCA2a function. SERCA2a and Na²⁺/Ca²⁺ exchanger 1 associated with Pin1, as revealed by proximity ligation assay in myocardial tissue sections, indicating that regulation of Ca²⁺ handling within cardiomyocytes is likely influenced through Pin1 interaction with SERCA2a and Na²⁺/Ca²⁺ exchanger 1 proteins.

Conclusions

Pin1 serves as a modulator of SERCA2a and Na²⁺/Ca²⁺ exchanger 1 Ca²⁺ handling proteins, with loss of function resulting in impaired cardiomyocyte relaxation, setting the stage for subsequent investigations to assess Pin1 dysregulation and modulation in the progression of heart failure.

Senescence-associated dysfunction deleteriously affects biological activities of human c-Kit⁺ cardiac progenitor cells (hCPCs), particularly under conditions of in vitro culture. In comparison, preservation of self-renewal and decreases in mitochondrial reactive oxygen species (ROS) are characteristics of murine CPCs in vivo that reside within hypoxic niches. Recapitulating hypoxic niche oxygen tension conditions of ∼1% O₂ in vitro for expansion of hCPCs rather than typical normoxic cell culture conditions (21% O₂ ) could provide significant improvement of functional and biological activities of hCPCs. hCPCs were isolated and expanded under permanent hypoxic (hCPC-1%) or normoxic (hCPC-21%) conditions from left ventricular tissue explants collected during left ventricular assist device implantation. hCPC-1% exhibit increased self-renewal and suppression of senescence characteristics relative to hCPC-21%. Oxidative stress contributed to higher susceptibility to apoptosis, as well as decreased mitochondrial function in hCPC-21%. Hypoxia prevented accumulation of dysfunctional mitochondria, supporting higher oxygen consumption rates and mitochondrial membrane potential. Mitochondrial ROS was an upstream mediator of senescence since treatment of hCPC-1% with mitochondrial inhibitor antimycin A recapitulated mitochondrial dysfunction and senescence observed in hCPC-21%. NAD⁺ /NADH ratio and autophagic flux, which are key factors for mitochondrial function, were higher in hCPC-1%, but hCPC-21% were highly dependent on BNIP3/NIX-mediated mitophagy to maintain mitochondrial function. Overall, results demonstrate that supraphysiological oxygen tension during in vitro expansion initiates a downward spiral of oxidative stress, mitochondrial dysfunction, and cellular energy imbalance culminating in early proliferation arrest of hCPCs. Senescence is inhibited by preventing ROS through hypoxic culture of hCPCs. Stem Cells 2019;37:555-567.

Cardiomyocyte ploidy has been described but remains obscure in cardiac interstitial cells. Ploidy of c-kit+ cardiac interstitial cells was assessed using confocal, karyotypic, and flow cytometric technique. Notable differences were found between rodent (rat, mouse) c-kit+ cardiac interstitial cells possessing mononuclear tetraploid (4n) content, compared to large mammals (human, swine) with mononuclear diploid (2n) content. In-situ analysis, confirmed with fresh isolates, revealed diploid content in human c-kit+ cardiac interstitial cells and a mixture of diploid and tetraploid content in mouse. Downregulation of the p53 signaling pathway provides evidence why rodent, but not human, c-kit+ cardiac interstitial cells escape replicative senescence. Single cell transcriptional profiling reveals distinctions between diploid versus tetraploid populations in mouse c-kit+ cardiac interstitial cells, alluding to functional divergences. Collectively, these data reveal notable species-specific biological differences in c-kit+ cardiac interstitial cells, which could account for challenges in extrapolation of myocardial from preclinical studies to clinical trials.