Although numerous preclinical investigations have consistently demonstrated salubrious effects of c-kitpos cardiac cells administered after myocardial infarction, the mechanism of action remains highly controversial. We and others have found little or no evidence that these cells differentiate into mature functional cardiomyocytes, suggesting paracrine effects. In this review, we propose a new paradigm predicated on a comprehensive analysis of the literature, including studies of cardiac development; we have dubbed this conceptual construct “string theory of c-kitpos cardiac cells” because it reconciles multifarious and sometimes apparently discrepant results. There is strong evidence that, during development, the c-kit receptor is expressed in different pools of cardiac progenitors (some capable of robust cardiomyogenesis and others with little or no contribution to myocytes). Accordingly, c-kit positivity, in itself, does not define the embryonic origins, lineage capabilities, or differentiation capacities of specific cardiac progenitors. C-kitpos cells derived from the first heart field (FHF) exhibit cardiomyogenic potential during development, but these cells are likely depleted shortly before or after birth. The residual c-kitpos cells found in the adult heart are probably of proepicardial origin, possess a mesenchymal phenotype, and are capable of contributing significantly only to non-myocytic lineages (fibroblasts, smooth muscle cells, endothelial cells). If these two populations (FHF and proepicardium) express different levels of c-kit, the cardiomyogenic potential of FHF progenitors might be reconciled with recent results of c-kitpos cell lineage tracing studies. The concept that c-kit expression in the adult heart identifies epicardium-derived, non-cardiomyogenic precursors with a mesenchymal phenotype helps to explain the beneficial effects of c-kitpos cell administration to ischemically damaged hearts despite the observed paucity of cardiomyogenic differentiation of these cells.

A recent phase I clinical trial (SCIPIO) has shown that autologous c-kit+ cardiac progenitor cells (CPCs) improve cardiac function and quality of life when transplanted into patients with ischemic heart disease. Although c-kit is widely used as a marker of resident CPCs, its role in the regulation of the cellular characteristics of CPCs remains unknown. We hypothesized that c-kit plays a role in the survival, growth, and migration of CPCs. To test this hypothesis, human CPCs were grown under stress conditions in the presence or absence of SCF, and the effects of SCF-mediated activation of c-kit on CPC survival/growth and migration were measured. SCF treatment led to a significant increase in cell survival and a reduction in cell death under serum depletion conditions. In addition, SCF significantly promoted CPC migration in vitro. Furthermore, the pro-survival and pro-migratory effects of SCF were augmented by c-kit overexpression and abrogated by c-kit inhibition with imatinib. Mechanistically, c-kit activation in CPCs led to activation of the PI3K and the MAPK pathways. With the use of specific inhibitors, we confirmed that the SCF/c-kit-dependent survival and chemotaxis of CPCs are dependent on both pathways. Taken together, our findings suggest that c-kit promotes the survival/growth and migration of human CPCs cultured ex vivo via the activation of PI3K and MAPK pathways. These results imply that the efficiency of CPC homing to the injury site as well as their survival after transplantation may be improved by modulating the activity of c-kit.

Background Although c-kitpos cardiac stem cells (CSCs) preserve left ventricular (LV) function and structure after myocardial infarction (MI), CSC doses have been chosen arbitrarily and the dose-effect relationship is unknown. Methods and Results Rats underwent a 90-min coronary occlusion followed by 35 days of reperfusion. Vehicle or CSCs at 5 escalating doses (0.3 ×106, 0.75 ×106, 1.5 ×106, 3.0 ×106, and 6.0 ×106 cells/heart) were given intracoronarily 4 h after reperfusion. The lowest dose (0.3 ×106) had no effect on LV function and morphology, whereas 0.75, 1.5, and 3.0 ×106 significantly improved regional and global LV function (echocardiography and hemodynamic studies). These three doses had similar effects on echocardiographic parameters (infarct wall thickening fraction, LV end-systolic and end-diastolic volumes, LV ejection fraction) and hemodynamic variables (LV end-diastolic pressure, LV dP/dtmax, preload adjusted maximal power, end-systolic elastance, preload recruitable stroke work), and produced similar reductions in apoptosis, scar size, infarct wall thinning, and LV expansion index and similar increases in viable myocardium in the risk region (morphometry). Infusion of 6.0 ×106 CSCs markedly increased post-procedural mortality. GFP and BrdU staining indicated that persistence of donor cells and formation of new myocytes were negligible with all doses. Conclusions Surprisingly, in this rat model of acute MI, the dose-response relationship for intracoronary CSCs is flat. A minimal dose between 0.3 and 0.75 ×106 is necessary for efficacy; above this threshold, a four-fold increase in cell number does not produce greater improvement in LV function or structure. Further increases in cell dose are harmful.

Histone deacetylase (HDAC) regulation is an essential process in myogenic differentiation. Inhibitors targeting the activity of specific HDAC family members have been shown to enhance the cardiogenic differentiation capacity of discrete progenitor cell types; a key property of donor cell populations contributing to their afforded benefits in cardiac cell therapy applications. The influence of HDAC inhibition on cardiac-derived mesenchymal stromal cell (CMC) transdifferentiation or the role of specific HDAC family members in dictating cardiovascular cell lineage specification has not been investigated. In the current study, the consequences of HDAC inhibition on patient-derived CMC proliferation, cardiogenic program activation, and cardiovascular differentiation/cell lineage specification were investigated using pharmacologic and genetic targeting approaches. Here, CMCs exposed to the pan-HDAC inhibitor sodium butyrate (NaBu) exhibited induction of a cardiogenic transcriptional program and heightened expression of myocyte and endothelial lineage-specific markers when coaxed to differentiate in vitro. Further, shRNA knockdown screens revealed CMCs depleted of HDAC1 to promote the induction of a cardiogenic transcriptional program characterized by enhanced expression of cardiomyogenic- and vasculogenic-specific markers, a finding which depended upon and correlated with enhanced acetylation and stabilization of p53. Cardiogenic gene activation and elevated p53 expression levels observed in HDAC1-depleted CMCs were associated with improved aptitude to assume a cardiomyogenic/vasculogenic cell-like fate in vitro. These results suggest that HDAC1 depletion-induced p53 expression alters CMC cell fate decisions and identify HDAC1 as a potential exploitable target to facilitate CMC-mediated myocardial repair in ischemic cardiomyopathy.