Background Human cardiac progenitor cells (hCPCs) may promote myocardial regeneration in adult ischemic myocardium. The regenerative capacity of hCPCs in young patients with nonischemic congenital heart defects for potential use in congenital heart defect repair warrants exploration. Methods and Results Human right atrial specimens were obtained during routine congenital cardiac surgery across 3 groups: neonates (age, <30 days), infants (age, 1 month to 2 years), and children (age, >2 to ≤13 years). C-kit+ hCPCs were 3-fold higher in neonates than in children >2 years of age. hCPC proliferation was greatest during the neonatal period as evidenced by c-kit+ Ki67+ expression but decreased with age. hCPC differentiation capacity was also greatest in neonatal right atrium as evidenced by c-kit+, NKX2–5+, NOTCH1+, and NUMB+ expression. Despite the age-dependent decline in resident hCPCs, we isolated and expanded right atrium–derived CPCs from all patients (n = 103) across all ages and diagnoses using the cardiosphere method. Intact cardiospheres contained a mix of heart-derived cell subpopulations that included cardiac progenitor cells expressing c-kit+, Islet-1, and supporting cells. The number of c-kit+–expressing cells was highest in human cardiosphere-derived cells (hCDCs) grown from neonatal and infant right atrium. Furthermore, hCDCs could differentiate into diverse cardiovascular lineages by in vitro differentiation assays. Transplanted hCDCs promoted greater myocardial regeneration and functional improvement in infarcted myocardium than transplanted cardiac fibroblasts. Conclusions Resident hCPCs are most abundant in the neonatal period and rapidly decrease over time. hCDCs can be reproducibly isolated and expanded from young human myocardial samples regardless of age or diagnosis. hCPCs are functional and have potential in congenital cardiac repair.

Rationale Cardiac progenitor cells are an attractive cell type for tissue regeneration but their mechanism for myocardial remodeling is still unclear. Objective This investigation determines how chronological age influences the phenotypic characteristics and the secretome of human cardiac progenitor cells (CPCs), as well as their potential to recover injured myocardium. Methods and Results Adult (aCPCs) and neonatal (nCPCs) cells were derived from patients more than 40 years or less than one month of age, respectively, and their functional potential was determined in a rodent myocardial infarction (MI) model. A more robust in vitro proliferative capacity of nCPCs, compared to aCPCs, correlated with significantly greater myocardial recovery mediated by nCPCs in vivo. Strikingly, a single injection of nCPC-derived total conditioned media (nTCM) was significantly more effective than nCPCs, aCPC-derived TCM (aTCM), or nCPC-derived exosomes in recovering cardiac function, stimulating neovascularization, and promoting myocardial remodeling. High resolution accurate mass spectrometry (HRAMS) with reverse phase liquid chromatography fractionation and mass spectrometry (LC-MS/MS) was employed to identify proteins in the secretome of aCPCs and nCPCs, and literature-based networking software identified specific pathways affected by the secretome of CPCs in the setting of MI. Examining the TCM, we quantified changes in the expression pattern of 804 proteins in nTCM and 513 proteins in aTCM. Literature-based proteomic network analysis identified that 46 and 6 canonical signaling pathways were significantly targeted by nTCM and aTCM, respectively. One leading candidate pathway is heat shock factor-1 (HSF-1), potentially affecting 8 identified pathways for nTCM but none for aTCM. To validate this prediction, we demonstrated that modulation of HSF-1 by knockdown in nCPCs or overexpression in aCPCs significantly altered the quality of their secretome. Conclusions In conclusion, a deep proteomic analysis revealed both detailed and global mechanisms underlying the chronological age-based differences in the ability of CPCs to promote myocardial recovery via the components of their secretome.

Background Human stem cells (CSCs) promote myocardial regeneration in adult ischemic myocardium. The regenerative capacity of CSCs in the very young patients with non-ischemic congenital heart defects has not been explored. We hypothesized that isolated neonatal-derived CSCs may have a higher regenerative ability than adult-derived CSCs and might address the structural deficiencies of congenital heart disease. Methods and Results Human specimens were obtained during routine cardiac surgical procedures from right atrial appendage tissue discarded from two age groups: neonates and adults patients. We developed a reproducible isolation method that generated cardiosphere derived cells (CDCs), regardless of starting tissue weight or age. Neonatal-derived CDCs demonstrated increased number of cardiac progenitor cells expressing c-kit+, flk-1 and Islet-1 by flow cytometry and immunofluorescence. When transplanted into infarcted myocardium, neonatal-derived CDCs had a significantly higher ability to preserve myocardial function, prevent adverse remodeling and enhance blood vessel preservation and/or formation when compared to adult CDCs. Lastly, neonatal-derived CDCs were more cardiomyogenic than adult CDCs when co-cultured with neonatal cardiomyocytes and displayed enhanced angiogenic function compared to adult CDCs. Conclusions Neonatal-derived CDCs have a strong regenerative ability when compared to adult-derived CDCs that may depend on angiogenic cytokines and an increase prevalence of stem cells. This has important implications in the potential use of CDCs in future clinical trials.