Transplanted allogeneic cardiac progenitor cells secrete GDF-15 and stimulate an active immune remodeling process in the ischemic myocardium.

BACKGROUND: Despite promising results in clinical studies, the mechanism for the beneficial effects of allogenic cell-based therapies remains unclear. Macrophages are not only critical mediators of inflammation but also critical players in cardiac remodeling. We hypothesized that transplanted allogenic rat cardiac progenitor cells (rCPCs) augment T-regulatory cells which ultimately promote proliferation of M2 like macrophages by an as-yet undefined mechanism. METHODS AND RESULTS: To test this hypothesis, we used crossover rat strains for exploring the mechanism of myocardial repair by allogenic CPCs. Human CPCs (hCPCs) were isolated from adult patients undergoing coronary artery bypass grafting, and rat CPCs (rCPCs) were isolated from male Wistar-Kyoto (WKY) rat hearts. Allogenic rCPCs suppressed the proliferation of T-cells observed in mixed lymphocyte reactions in vitro. Transplanted syngeneic or allogeneic rCPCs significantly increased cardiac function in a rat myocardial infarct (MI) model, whereas xenogeneic CPCs did not. Allogeneic rCPCs stimulated immunomodulatory responses by specifically increasing T-regulatory cells and M2 polarization, while maintaining their cardiac recovery potential and safety profile. Mechanistically, we confirmed the inactivation of NF-kB in Treg cells and increased M2 macrophages in the myocardium after MI by transplanted CPCs derived GDF15 and it's uptake by CD48 receptor on immune cells. CONCLUSION: Collectively, these findings strongly support the active immunomodulatory properties and robust therapeutic potential of allogenic CPCs in post-MI cardiac dysfunction.

Autologous Cardiac Stem Cell Injection in Patients with Hypoplastic Left Heart Syndrome (CHILD Study).

Mortality in infants with hypoplastic left heart syndrome (HLHS) is strongly correlated with right ventricle (RV) dysfunction. Cell therapy has demonstrated potential improvements of RV dysfunction in animal models related to HLHS, and neonatal human derived c-kit+ cardiac-derived progenitor cells (CPCs) show superior efficacy when compared to adult human cardiac-derived CPCs (aCPCs). Neonatal CPCs (nCPCs) have yet to be investigated in humans. The CHILD trial (Autologous Cardiac Stem Cell Injection in Patients with Hypoplastic Left Heart Syndrome) is a Phase I/II trial aimed at investigating intramyocardial administration of autologous nCPCs in HLHS infants by assessing the feasibility, safety, and potential efficacy of CPC therapy. Using an open-label, multicenter design, CHILD investigates nCPC safety and feasibility in the first enrollment group (Group A/Phase I). In the second enrollment group, CHILD uses a randomized, double-blinded, multicenter design (Group B/Phase II), to assess nCPC efficacy based on RV functional and structural characteristics. The study plans to enroll 32 patients across 4 institutions: Group A will enroll 10 patients, and Group B will enroll 22 patients. CHILD will provide important insights into the therapeutic potential of nCPCs in patients with HLHS.Clinical Trial Registration https://clinicaltrials.gov/ct2/home NCT03406884, First posted January 23, 2018.

Stem Cell Therapy in Single-Ventricle Physiology: Recent Progress and Future Directions.

Current surgical and medical treatment options for single ventricle physiology conditions remain palliative. On the long term, despite treatment, the systemic ventricle has a significant risk of developing failure. There are unmet needs to develop novel treatment modalities to help ameliorate the ventricular dysfunction. Advances in the field of stem cell therapy have been promising for the treatment of heart failure. Numerous stem cell populations have been identified. Preclinical studies in small and large animal models provide evidence for effectiveness of this treatment modality and reveal several mechanisms of action by which stem cells exert their effect. Many clinical trials have been designed to further investigate the therapeutic potential that stem cell therapy may hold for pediatric populations with single ventricle physiology. In this review, we discuss the stem cell types used in these populations, some preclinical studies, and the clinical trials of stem cell therapy in single ventricle patients.

Stem Cell Therapy for Hypoplastic Left Heart Syndrome: Mechanism, Clinical Application, and Future Directions.

Hypoplastic left heart syndrome is a type of congenital heart disease characterized by underdevelopment of the left ventricle, outflow tract, and aorta. The condition is fatal if aggressive palliative operations are not undertaken, but even after the complete 3-staged surgical palliation, there is significant morbidity because of progressive and ultimately intractable right ventricular failure. For this reason, there is interest in developing novel therapies for the management of right ventricular dysfunction in patients with hypoplastic left heart syndrome. Stem cell therapy may represent one such innovative approach. The field has identified numerous stem cell populations from different tissues (cardiac or bone marrow or umbilical cord blood), different age groups (adult versus neonate-derived), and different donors (autologous versus allogeneic), with preclinical and clinical experience demonstrating the potential utility of each cell type. Preclinical trials in small and large animal models have elucidated several mechanisms by which stem cells affect the injured myocardium. Our current understanding of stem cell activity is undergoing a shift from a paradigm based on cellular engraftment and differentiation to one recognizing a primarily paracrine effect. Recent studies have comprehensively evaluated the individual components of the stem cells' secretomes, shedding new light on the intracellular and extracellular pathways at the center of their therapeutic effects. This research has laid the groundwork for clinical application, and there are now several trials of stem cell therapies in pediatric populations that will provide important insights into the value of this therapeutic strategy in the management of hypoplastic left heart syndrome and other forms of congenital heart disease. This article reviews the many stem cell types applied to congenital heart disease, their preclinical investigation and the mechanisms by which they might affect right ventricular dysfunction in patients with hypoplastic left heart syndrome, and finally, the completed and ongoing clinical trials of stem cell therapy in patients with congenital heart disease.

A Deep Proteome Analysis Identifies the Complete Secretome as the Functional Unit of Human Cardiac Progenitor Cells.

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), and their potential to recover injured myocardium. METHODS AND RESULTS: Adult (aCPCs) and neonatal (nCPCs) cells were derived from patients aged >40 years or <1 month, respectively, and their functional potential was determined in a rodent myocardial infarction model. A more robust in vitro proliferative capacity of nCPCs, compared with aCPCs, correlated with significantly greater myocardial recovery mediated by nCPCs in vivo. Strikingly, a single injection of nCPC-derived total conditioned media was significantly more effective than nCPCs, aCPC-derived TCM, or nCPC-derived exosomes in recovering cardiac function, stimulating neovascularization, and promoting myocardial remodeling. High-resolution accurate mass spectrometry with reverse phase liquid chromatography fractionation and mass spectrometry was used to identify proteins in the secretome of aCPCs and nCPCs, and the literature-based networking software identified specific pathways affected by the secretome of CPCs in the setting of myocardial infarction. Examining the TCM, we quantified changes in the expression pattern of 804 proteins in nCPC-derived TCM and 513 proteins in aCPC-derived TCM. The literature-based proteomic network analysis identified that 46 and 6 canonical signaling pathways were significantly targeted by nCPC-derived TCM and aCPC-derived TCM, respectively. One leading candidate pathway is heat-shock factor-1, potentially affecting 8 identified pathways for nCPC-derived TCM but none for aCPC-derived TCM. To validate this prediction, we demonstrated that the modulation of heat-shock factor-1 by knockdown in nCPCs or overexpression in aCPCs significantly altered the quality of their secretome. CONCLUSIONS: 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.

Study design and rationale for ELPIS: A phase I/IIb randomized pilot study of allogeneic human mesenchymal stem cell injection in patients with hypoplastic left heart syndrome.

Despite advances in surgical technique and postoperative care, long-term survival of children born with hypoplastic left heart syndrome (HLHS) remains limited, with cardiac transplantation as the only alternative for patients with failing single ventricle circulations. Maintenance of systemic right ventricular function is crucial for long-term survival, and interventions that improve ventricular function and avoid or defer transplantation in patients with HLHS are urgently needed. We hypothesize that the young myocardium of the HLHS patient is responsive to the biological cues delivered by bone marrow-derived mesenchymal stem cells (MSCs) to improve and preserve right ventricle function. The ELPIS trial (Allogeneic Human MEsenchymal Stem Cell Injection in Patients with Hypoplastic Left Heart Syndrome: An Open Label Pilot Study) is a phase I/IIb trial designed to test whether MSC injection will be both safe and feasible by monitoring the first 10 HLHS patients for new major adverse cardiac events. If our toxicity stopping rule is not activated, we will proceed to the phase IIb component of our study where we will test our efficacy hypothesis that MSC injection improves cardiac function compared with surgery alone. Twenty patients will be enrolled in a randomized phase II trial with a uniform allocation to MSC injection versus standard surgical care (no injection). The 2 trial arms will be compared with respect to improvement of right ventricular function, tricuspid valve annulus size, and regurgitation determined by cardiac magnetic resonance and reduced mortality, morbidity, and need for transplantation. This study will establish the safety and feasibility of allogeneic mesenchymal stem cell injection in HLHS patients and provide important insights in the emerging field of stem cell-based therapy for congenital heart disease patients.

Mesenchymal stem cells preserve neonatal right ventricular function in a porcine model of pressure overload.

Limited therapies exist for patients with congenital heart disease (CHD) who develop right ventricular (RV) dysfunction. Bone marrow-derived mesenchymal stem cells (MSCs) have not been evaluated in a preclinical model of pressure overload, which simulates the pathophysiology relevant to many forms of CHD. A neonatal swine model of RV pressure overload was utilized to test the hypothesis that MSCs preserve RV function and attenuate ventricular remodeling. Immunosuppressed Yorkshire swine underwent pulmonary artery banding to induce RV dysfunction. After 30 min, human MSCs (1 million cells, n = 5) or placebo (n = 5) were injected intramyocardially into the RV free wall. Serial transthoracic echocardiography monitored RV functional indices including 2D myocardial strain analysis. Four weeks postinjection, the MSC-treated myocardium had a smaller increase in RV end-diastolic area, end-systolic area, and tricuspid vena contracta width (P < 0.01), increased RV fractional area of change, and improved myocardial strain mechanics relative to placebo (P < 0.01). The MSC-treated myocardium demonstrated enhanced neovessel formation (P < 0.0001), superior recruitment of endogenous c-kit+ cardiac stem cells to the RV (P < 0.0001) and increased proliferation of cardiomyocytes (P = 0.0009) and endothelial cells (P < 0.0001). Hypertrophic changes in the RV were more pronounced in the placebo group, as evidenced by greater wall thickness by echocardiography (P = 0.008), increased cardiomyocyte cross-sectional area (P = 0.001), and increased expression of hypertrophy-related genes, including brain natriuretic peptide, ß-myosin heavy chain and myosin light chain. Additionally, MSC-treated myocardium demonstrated increased expression of the antihypertrophy secreted factor, growth differentiation factor 15 (GDF15), and its downstream effector, SMAD 2/3, in cultured neonatal rat cardiomyocytes and in the porcine RV myocardium. This is the first report of the use of MSCs as a therapeutic strategy to preserve RV function and attenuate remodeling in the setting of pressure overload. Mechanistically, transplanted MSCs possibly stimulated GDF15 and its downstream SMAD proteins to antagonize the hypertrophy response of pressure overload. These encouraging results have implications in congenital cardiac pressure overload lesions.

Cardiosphere-derived cells from pediatric end-stage heart failure patients have enhanced functional activity due to the heat shock response regulating the secretome.

We have demonstrated that human neonatal cardiosphere-derived cells (CDCs) derived from the young are more regenerative due to their robust secretome. However, it is unclear how the decompensated pediatric heart impacts the functional activity of their CDCs. Our aim was to characterize the potency of pediatric CDCs derived from normal functioning myocardium of control heart disease (CHD) patients to those generated from age-matched end stage heart failure (ESHF) patients and to determine the mechanisms involved. ESHF-derived CDCs contained a higher number of c-kit(+) , Islet-1(+) , and Sca-1(+) cells. When transplanted into an infarcted rodent model, ESHF-derived CDCs significantly demonstrated higher restoration of ventricular function, prevented adverse remodeling, and enhanced angiogenesis when compared with CHD patients. The superior functional recovery of the ESHF-derived CDCs was mediated in part by increased SDF-1α and VEGF-A secretion resulting in augmented recruitment of endogenous stem cells and proliferation of cardiomyocytes. We determined the mechanism is due to the secretome directed by the heat shock response (HSR), which is supported by three lines of evidence. First, gain of function studies demonstrated that increased HSR induced the lower functioning CHD-derived CDCs to significantly restore myocardial function. Second, loss-of function studies targeting the HSR impaired the ability of the ESHF-derived CDCs to functionally recover the injured myocardium. Finally, the native ESHF myocardium had an increased number of c-kit(+) cardiac stem cells. These findings suggest that the HSR enhances the functional activity of ESHF-derived CDCs by increasing their secretome activity, notably SDF-1α and VEGF-A.

Stem cell therapy for CHD: towards translation.

Stem cell therapy has the optimistic goal of regenerating the myocardium as defined by re-growth of lost or destroyed myocardium. As applied to patients with heart failure, many confuse or limit the regenerative definition to just improving myocardial function and/or decreasing myocardial scar formation, which may not be the most important clinical outcome to achieve in this promising field of molecular medicine. Many different stem cell-based therapies have been tested and have demonstrated a safe and feasible profile in adult patients with heart failure, but with varied efficacious end points reported. Although not achieved as of yet, the encompassing goal to regenerate the heart is still believed to be within reach using these cell-based therapies in adult patients with heart failure, as the first-generation therapies are now being tested in different phases of clinical trials. Similar efforts to foster the translation of stem cell therapy to children with heart failure have, however, been limited. In this review, we aim to summarise the findings from pre-clinical models and clinical experiences to date that have focussed on the evaluation of stem cell therapy in children with heart failure. Finally, we present methodological considerations pertinent to the design of a stem cell-based trial for children with heart failure, as they represent a population of patients with very different sets of issues when compared with adult patients. As has been taught by many learned clinicians, children are not small adults!

A strong regenerative ability of cardiac stem cells derived from neonatal hearts.

BACKGROUND: Human cardiac stem cells (CSCs) promote myocardial regeneration in adult ischemic myocardium. The regenerative capacity of CSCs in very young patients with nonischemic 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 2 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 with adult-derived CDCs. Last, neonatal-derived CDCs were more cardiomyogenic than adult-derived CDCs when cocultured with neonatal cardiomyocytes and displayed enhanced angiogenic function compared with adult-derived CDCs. CONCLUSIONS: Neonatal-derived CDCs have a strong regenerative ability when compared with 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.

Hypoxia-inducible factor 1-alpha enhances the secretome to rejuvenate adult cardiosphere-derived cells.

OBJECTIVE: After cardiac injury, endogenous repair mechanisms are ineffective. However, cell-based therapies provide a promising clinical intervention based on their ability to restore and remodel injured myocardium due to their paracrine factors. Recent clinical trials have demonstrated that adult cardiosphere-derived cell therapy is safe for the treatment of ischemic heart failure, although with limited regenerative potential. The limited efficiency of cardiosphere-derived cells after myocardial infarction is due to the inferior quality of their secretome. This study sought to augment the therapeutic potential of cardiosphere-derived cells by modulating hypoxia-inducible factor-1α, a regulator of paracrine factors. METHODS: Cardiosphere-derived cells were isolated and expanded from the right atrial appendage biopsies of patients undergoing cardiac surgery. To study the effect of hypoxia-inducible factor-1α on the secretome, cardiosphere-derived cells were transduced with hypoxia-inducible factor-1α-overexpressing lentivirus, and various cardioprotective factors within the secretome were quantified using enzyme-linked immunosorbent assays. Comparative analysis of the regenerative potential of cardiosphere-derived cells was performed in a rat myocardial infarction model. RESULTS: Mechanistically, overexpression of hypoxia-inducible factor-1α in adult cardiosphere-derived cells led to the enrichment of the secretome with vascular endothelial growth factor A, angiopoietin 1, stromal cell-derived factor 1α, and basic fibroblast growth factor. Intramyocardial administration of cardiosphere-derived cells transduced with hypoxia-inducible factor-1α after myocardial infarction significantly improved left ventricular ejection fraction, fractional shortening, left ventricular end-systolic volume, and cardiac output. Functional improvement of the rat heart correlated with improved adaptive remodeling of the infarcted myocardium by enhanced angiogenesis and decreased myocardial fibrosis. We also showed that hypoxia-inducible factor-1α expression in cardiosphere-derived cells was adversely affected by aging. CONCLUSIONS: Hypoxia-inducible factor-1α improves the functional potency of cardiosphere-derived cells to preserve myocardial function after myocardial infarction by enriching the cardiosphere-derived cells' secretome with cardioprotective factors. This strategy may be useful for improving the efficacy of allogeneic cell-based therapies in future clinical trials.

Exosomes mediated fibrogenesis in dilated cardiomyopathy through a MicroRNA pathway.

Cardiac fibrosis is a hallmark in late-stage familial dilated cardiomyopathy (DCM) patients, although the underlying mechanism remains elusive. Cardiac exosomes (Exos) have been reported relating to fibrosis in ischemic cardiomyopathy. Thus, we investigated whether Exos secreted from the familial DCM cardiomyocytes could promote fibrogenesis. Using human iPSCs differentiated cardiomyocytes we isolated Exos of angiotensin II stimulation conditioned media from either DCM or control (CTL) cardiomyocytes. Of interest, cultured cardiac fibroblasts had increased fibrogenesis following exposure to DCM-Exos rather than CTL-Exos. Meanwhile, injecting DCM-Exos into mouse hearts enhanced cardiac fibrosis and impaired cardiac function. Mechanistically, we identified the upregulation of miRNA-218-5p in the DCM-Exos as a critical contributor to fibrogenesis. MiRNA-218-5p activated TGF-ß signaling via suppression of TNFAIP3, a master inflammation inhibitor. In conclusion, our results illustrate a profibrotic effect of cardiomyocytes-derived Exos that highlights an additional pathogenesis pathway for cardiac fibrosis in DCM.

HSF-1 enhances cardioprotective potential of stem cells via exosome biogenesis and their miRNA cargo enrichment.

Stem cell therapy provides a hope to no option heart disease patient group. Stem cells work via different mechanisms of which paracrine mechanism is reported to justify most of the effects. Therefore, identifying the control arms for paracrine cocktail production is necessary to tailor stem cell functions in disease contextual manner. In this study, we describe a novel paracrine cocktail regulatory axis, in stem cells, to enhance their cardioprotective abilities. We identified that HSF1 knockout resulted in reduced cardiac regenerative abilities of mesenchymal stem cells (MSCs) while its overexpression had opposite effects. Altered exosome biognesis and their miRNA cargo enrichment were found to be underlying these altered regenerative abilities. Decreased production of exosomes by MSCs accompanied their loss of HSF1 and vice versa. Moreover, the exosomes derived from HSF1 depleted MSCs showed significantly reduced candidate miRNA expression (miR-145, miR-146, 199-3p, 199b and miR-590) compared to those obtained from HSF1 overexpressing MSCs. We further discovered that HSF1 mediates miRNAs' enrichment into exosomes via Y binding protein 1 (YBX1) and showed, by loss and gain of function strategies, that miRNAs' enrichment in mesenchymal stem cell derived exosomes is deregulated with altered YBX1 expression. It was finally demonstrated that absence of YBX1 in MSCs, with normal HSF1 expression, resulted in significant accumulation of candidate miRNAs into the cells. Together, our data shows that HSF1 plays a critical role in determining the regenerative potential of stem cells. HSF1 does that by affecting exosome biogenesis and miRNA cargo sorting via regulation of YBX1 gene expression.

Stem cell therapy for single ventricle congenital heart disease - current state and future directions.

Hypoplastic left heart syndrome (HLHS) is one of the most complex forms of congenital heart disease, characterized by an underdeveloped left ventricle, outflow tract and aorta. Current surgical and medical treatment for this disease remains palliative. As a result of the multi-step surgery, the right ventricle plays the role of the systemic ventricle, which inevitably leads to its failure. There is an urgent need to develop new treatments to ameliorate the right ventricle failure. Stem cell therapy may represent a new approach to single ventricle pathology. Great numbers of small and large animal studies have proven this therapy to be safe and effective in hypoplastic left heart syndrome. Several clinical trials have been designed to investigate the potential of mesenchymal stem cells in univentricular heart physiology. With increasing evidence, understanding of the mechanism of stem cells' action has shifted from the concept of differentiation into various heart cell types to paracrine activity playing the major role. The secretome of stem cells has been identified as their functional unit. In this review, we present different types of stem cells used in single ventricle diseases in children as well as their preclinical investigations. We also summarize clinical applications of stem cells in children with HLHS.

Safety and efficacy of transcoronary transfer of human neonatal stem cells to ischemic myocardium using a novel cell-delivery system (CIRCULATE catheter) in swine model of acute myocardial infarction.

Introduction: Stem cell-based therapies have shown promise in adults with ischemic cardiomyopathy and children with congenital heart diseases, especially those without available therapeutic options. Human neonatal mesenchymal stem cells (nMSCs) have greater regenerative potential than adult stem cells. Aim: To describe our experience with a novel catheter system for transcoronary delivery of cell-based therapies (CIRCULATE catheter) in the intra-coronary delivery of nMSCs in a swine acute myocardial infarct model. Material and methods: A newly developed catheter system (CIRCULATE catheter) with several unique features, including an expandable intra-coronary reservoir with spirally placed side holes of varying diameter, was used. nMSCs together with their secretome were used for the treatment. Pigs underwent myocardial infarction by inflating a 2.5 mm angioplasty balloon in the left anterior descending artery for 60 min. After reperfusion, stem cell therapy or placebo was administered via the novel catheter. TTE was performed at baseline, 1 h after the procedure, and before the euthanasia. Troponin blood concertation was evaluated at baseline, and after 48 h. The heart was harvested, sliced, and stained with triphenyl tetrazolium chloride (TTC). Infarct size to area-at-risk ratio was calculated. Troponin was assessed at baseline and after 48 h. Results: Thirty-nine pigs were operated with the mortality rate of 5.13% (exclusively malignant arrhythmia). Infarct size to area-at-risk ratio was significantly lower in the treatment group. Treated animals had higher ejection fraction than controls. Conclusions: Intra-coronary delivery of neonatal mesenchymal stem cells reduces the infarct size and restores myocardial function in a swine model. The novel catheter system (CIRCULATE catheter) tested in this study was safe and effective in transcoronary cell delivery of human neonatal mesenchymal stem cells.

Comparative efficacy and mechanism of action of cardiac progenitor cells after cardiac injury.

Successful cell therapy requires cells to resist the hostile ischemic myocardium, be retained to continue secreting cardioprotective growth factors/exosomes, and resist immunological host responses. Clinically relevant stem/progenitor cells in a rodent model of acute myocardial infarction (MI) demonstrated that neonatal cardiac mesenchymal stromal cells (nMSCs) provide the most robust cardiac functional recovery. Transplanted nMSCs significantly increased the number of tissue reparative macrophages and regulatory T-cells and decreased monocyte-derived inflammatory macrophages and neutrophils in the host myocardium. mRNA microarray and single-cell analyses combined with targeted depletion studies established CD47 in nMSCs as a key molecule responsible for cell retention in the myocardium through an antiphagocytic mechanism regulated by miR34a-5p. Gain and loss-of-function studies demonstrated that miR34a-5p also regulated the production of exosomes and cardioprotective paracrine factors in the nMSC secretome. In conclusion, miR34a-5p and CD47 play an important role in determining the composition of nMSCs' secretome and immune evasion, respectively.

Circulating exosomes derived from transplanted progenitor cells aid the functional recovery of ischemic myocardium.

The stem cell field is hindered by its inability to noninvasively monitor transplanted cells within the target organ in a repeatable, time-sensitive, and condition-specific manner. We hypothesized that quantifying and characterizing transplanted cell-derived exosomes in the recipient plasma would enable reliable, noninvasive surveillance of the conditional activity of the transplanted cells. To test this hypothesis, we used a human-into-rat xenogeneic myocardial infarction model comparing two well-studied progenitor cell types: cardiosphere-derived cells (CDCs) and c-kit+ cardiac progenitor cells (CPCs), both derived from the right atrial appendage of adults undergoing cardiopulmonary bypass. CPCs outperformed the CDCs in cell-based and in vivo regenerative assays. To noninvasively monitor the activity of transplanted CDCs or CPCs in vivo, we purified progenitor cell-specific exosomes from recipient total plasma exosomes. Seven days after transplantation, the concentration of plasma CPC-specific exosomes increased about twofold compared to CDC-specific exosomes. Computational pathway analysis failed to link CPC or CDC cellular messenger RNA (mRNA) with observed myocardial recovery, although recovery was linked to the microRNA (miRNA) cargo of CPC exosomes purified from recipient plasma. We further identified mechanistic pathways governing specific outcomes related to myocardial recovery associated with transplanted CPCs. Collectively, these findings demonstrate the potential of circulating progenitor cell-specific exosomes as a liquid biopsy that provides a noninvasive window into the conditional state of the transplanted cells. These data implicate the surveillance potential of cell-specific exosomes for allogeneic cell therapies.

Regenerative medicine therapy for single ventricle congenital heart disease.

One of the most complex forms of congenital heart disease (CHD) involving single ventricle physiology is hypoplastic left heart syndrome (HLHS), characterized by underdevelopment of the left ventricle (LV), mitral and aortic valves, and narrowing of the ascending aorta. The underdeveloped LV is incapable of providing long-term systemic flow, and if left untreated, the condition is fatal. Current treatment for this condition consists of three consecutive staged palliative operations: the first is conducted within the first few weeks of birth, the second between 4 to 6 months, and the third and final surgery within the first 4 years. At the conclusion of the third surgery, systemic perfusion is provided by the right ventricle (RV), and deoxygenated blood flows passively to the pulmonary vasculature. Despite these palliative interventions, the RV, which is ill suited to provide long-term systemic perfusion, is prone to eventual failure. In the absence of satisfying curative treatments, stem cell therapy may represent one innovative approach to the management of RV dysfunction in HLHS patients. Several stem cell populations from different tissues (cardiac and non-cardiac), different age groups (adult- vs. neonate-derived), and different donors (autologous vs. allogeneic), are under active investigation. Preclinical trials in small and large animal models have elucidated several mechanisms by which these stem cells affect the injured myocardium, and are driving the shift from a paradigm based upon cellular engraftment and differentiation to one based primarily on paracrine effects. Recent studies have comprehensively evaluated the individual components of the stem cells' secretomes, shedding new light on the intracellular and extracellular pathways at the center of their therapeutic effects. This research has laid the groundwork for clinical application, and there are now several trials of stem cell therapies in pediatric populations that will provide important insights into the value of this therapeutic strategy in the management of HLHS and other forms of CHD. This article reviews the many stem cell types applied to CHD, their preclinical investigation and the mechanisms by which they might affect RV dysfunction in HLHS patients, and finally, the completed and ongoing clinical trials of stem cell therapy in patients with CHD.

Cardiac Progenitor Cells Enhance Neonatal Right Ventricular Function After Pulmonary Artery Banding.

BACKGROUND: C-kit+ cardiac progenitor cells (CPCs) have been shown to be safe and effective in large-animal models and in an early-phase clinical trial for adult patients with ischemic heart disease. However, CPCs have not yet been evaluated in a preclinical model of right ventricular (RV) dysfunction, which is a salient feature of many forms of congenital heart disease. METHODS: Human c-kit+ CPCs were generated from right atrial appendage biopsy specimens obtained during routine congenital cardiac operations. Immunosuppressed Yorkshire swine (6 to 9 kg) underwent pulmonary artery banding to induce RV dysfunction. Thirty minutes after banding, pigs received intramyocardial injection into the RV free wall with c-kit+ CPCs (1 million cells, n = 5) or control (phosphate-buffered saline, n = 5). Pigs were euthanized at 30 days postbanding. RESULTS: Banding was calibrated to a consistent rise in the RV-to-systemic pressure ratio across both groups (postbanding: CPCs = 0.76 ± 0.06, control = 0.75 ± 0.03). At 30 days postbanding, the CPCs group demonstrated less RV dilatation and a significantly greater RV fractional area of change than the control group (p = 0.002). In addition, measures of RV myocardial strain, including global longitudinal strain and strain rate, were significantly greater in the CPCs group at 4 weeks relative to control (p = 0.004 and p = 0.01, respectively). The RV free wall in the CPCs group demonstrated increased arteriole formation (p < 0.0001) and less myocardial fibrosis compared with the control group (p = 0.02). CONCLUSIONS: Intramyocardial injection of c-kit+ CPCs results in enhanced RV performance relative to control at 30 days postbanding in neonatal pigs. This model is important for further evaluation of c-kit+ CPCs, including long-term efficacy.