Epicardial HDAC3 Promotes Myocardial Growth Through a Novel MicroRNA Pathway.

BACKGROUND: Establishment of the myocardial wall requires proper growth cues from nonmyocardial tissues. During heart development, the epicardium and epicardium-derived cells instruct myocardial growth by secreting essential factors including FGF (fibroblast growth factor) 9 and IGF (insulin-like growth factor) 2. However, it is poorly understood how the epicardial secreted factors are regulated, in particular by chromatin modifications for myocardial formation. The current study is to investigate whether and how HDAC (histone deacetylase) 3 in the developing epicardium regulates myocardial growth. METHODS: Various cellular and mouse models in conjunction with biochemical and molecular tools were employed to study the role of HDAC3 in the developing epicardium. RESULTS: We deleted Hdac3 in the developing murine epicardium, and mutant hearts showed ventricular myocardial wall hypoplasia with reduction of epicardium-derived cells. The cultured embryonic cardiomyocytes with supernatants from Hdac3 knockout (KO) mouse epicardial cells also showed decreased proliferation. Genome-wide transcriptomic analysis revealed that Fgf9 and Igf2 were significantly downregulated in Hdac3 KO mouse epicardial cells. We further found that Fgf9 and Igf2 expression is dependent on HDAC3 deacetylase activity. The supplementation of FGF9 or IGF2 can rescue the myocardial proliferation defects treated by Hdac3 KO supernatant. Mechanistically, we identified that microRNA (miR)-322 and miR-503 were upregulated in Hdac3 KO mouse epicardial cells and Hdac3 epicardial KO hearts. Overexpression of miR-322 or miR-503 repressed FGF9 and IGF2 expression, while knockdown of miR-322 or miR-503 restored FGF9 and IGF2 expression in Hdac3 KO mouse epicardial cells. CONCLUSIONS: Our findings reveal a critical signaling pathway in which epicardial HDAC3 promotes compact myocardial growth by stimulating FGF9 and IGF2 through repressing miR-322 or miR-503, providing novel insights in elucidating the etiology of congenital heart defects and conceptual strategies to promote myocardial regeneration.

Baseline extracellular vesicle TGF-ß is a predictive biomarker for response to immune checkpoint inhibitors and survival in non-small cell lung cancer.

BACKGROUND: Immune-checkpoint inhibitors (ICIs) are an effective therapeutic strategy, improving the survival of patients with lung cancer compared with conventional treatments. However, novel predictive biomarkers are needed to stratify which patients derive clinical benefit because the currently used and highly heterogenic histological PD-L1 has shown low accuracy. Liquid biopsy is the analysis of biomarkers in body fluids and represents a minimally invasive tool that can be used to monitor tumor evolution and treatment effects, potentially reducing biases associated with tumor heterogeneity associated with tissue biopsies. In this context, cytokines, such as transforming growth factor-ß (TGF-ß), can be found free in circulation in the blood and packaged into extracellular vesicles (EVs), which have a specific delivery tropism and can affect in tumor/immune system interaction. TGF-ß is an immunosuppressive cytokine that plays a crucial role in tumor immune escape, treatment resistance, and metastasis. Thus, we aimed to evaluate the predictive value of circulating and EV TGF-ß in patients with non-small-cell lung cancer receiving ICIs. METHODS: Plasma samples were collected in 33 patients with advanced non-small-cell lung cancer before and during treatment with ICIs. EV were isolated from plasma by serial ultracentrifugation methods and circulating and EV TGF-ß expression levels were evaluated by enzyme-linked immunosorbent assay. RESULTS: Baseline high expression of TGF-ß in EVs was associated with nonresponse to ICIs as well as shorter progression-free survival and overall survival, outperforming circulating TGF-ß levels and tissue PD-L1 as a predictive biomarker. CONCLUSION: If validated, EV TGF-ß could be used to improve patient stratification, increasing the effectiveness of treatment with ICIs and potentially informing combinatory treatments with TGF-ß blockade. PLAIN LANGUAGE SUMMARY: Treatment with immune-checkpoint inhibitors (ICIs) has improved the survival of some patients with lung cancer. However, the majority of patients do not benefit from this treatment, making it essential to develop more reliable biomarkers to identify patients most likely to benefit. In this pilot study, the expression of transforming growth factor-ß (TGF-ß) in blood circulation and in extracellular vesicles was analyzed. The levels of extracellular vesicle TGF-ß before treatment were able to determine which patients would benefit from treatment with ICIs and have a longer survival with higher accuracy than circulating TGF-ß and tissue PD-L1, which is the currently used biomarker in clinical practice.

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 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.

The current status and future of cardiac stem/progenitor cell therapy for congenital heart defects from diabetic pregnancy.

Pregestational maternal diabetes induces congenital heart defects (CHDs). Cardiac dysfunction after palliative surgical procedures contributes to the high mortality of CHD patients. Autologous or allogeneic stem cell therapies are effective for improving cardiac function in animal models and clinical trials. c-kit+ cardiac progenitor cells (CPCs), the most recognized CPCs, have the following basic properties of stem cells: self-renewal, multicellular clone formation, and differentiation into multiple cardiac lineages. However, there is ongoing debate regarding whether c-kit+ CPCs can give rise to sufficient cardiomyocytes. A new hypothesis to address the beneficial effect of c-kit+ CPCs is that these cells stimulate endogenous cardiac cells through a paracrine function in producing a robust secretome and exosomes. The values of other cardiac CPCs, including Sca1+ CPCs and cardiosphere-derived cells, are beginning to be revealed. These cells may be better choices than c-kit+ CPCs for generating cardiomyocytes. Adult mesenchymal stem cells are considered immune-incompetent and effective for improving cardiac function. Autologous CPC therapy may be limited by the observation that maternal diabetes adversely affects the biological function of embryonic stem cells and CPCs. Future studies should focus on determining the mechanistic action of these cells, identifying new CPC markers, selecting highly effective CPCs, and engineering cell-free products.

Maternal diabetes and high glucose in vitro trigger Sca1+ cardiac progenitor cell apoptosis through FoxO3a.

Recent controversies surrounding the authenticity of c-kit+ cardiac progenitor cells significantly push back the advance in regenerative therapies for cardiovascular diseases. There is an urgent need for research in characterizing alternative types of cardiac progenitor cells. Towards this goal, in the present study, we determined the effect of maternal diabetes on Sca1+ cardiac progenitor cells. Maternal diabetes induced caspase 3-dependent apoptosis in Sca1+ cardiac progenitor cells derived from embryonic day 17.5 (E17.5). Similarly, high glucose in vitro but not the glucose osmotic control mannitol triggered Sca1+ cardiac progenitor cell apoptosis in a dose- and time-dependent manner. Both maternal diabetes and high glucose in vitro activated the pro-apoptotic transcription factor, Forkhead O 3a (FoxO3a) via dephosphorylation at threonine 32 (Thr-32) residue. foxo3a gene deletion abolished maternal diabetes-induced Sca1+ cardiac progenitor cell apoptosis. The dominant negative FoxO3a mutant without the transactivation domain from the C terminus blocked high glucose-induced Sca1+ cardiac progenitor cell apoptosis, whereas the constitutively active FoxO3a mutant with the three phosphorylation sites, Thr-32, Ser-253, and Ser-315, being replaced by alanine residues mimicked the pro-apoptotic effect of high glucose. Thus, maternal diabetes and high glucose in vitro may limit the regenerative potential of Sca1+ cardiac progenitor cells by inducing apoptosis through FoxO3a activation. These findings will serve as the guide in optimizing the autologous therapy using Sca1+ cardiac progenitor cells in cardiac defect babies born exposed to maternal diabetes.

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!

Summary of the 2015 International Paediatric Heart Failure Summit of Johns Hopkins All Children's Heart Institute.

In the United States alone, ∼14,000 children are hospitalised annually with acute heart failure. The science and art of caring for these patients continues to evolve. The International Pediatric Heart Failure Summit of Johns Hopkins All Children's Heart Institute was held on February 4 and 5, 2015. The 2015 International Pediatric Heart Failure Summit of Johns Hopkins All Children's Heart Institute was funded through the Andrews/Daicoff Cardiovascular Program Endowment, a philanthropic collaboration between All Children's Hospital and the Morsani College of Medicine at the University of South Florida (USF). Sponsored by All Children's Hospital Andrews/Daicoff Cardiovascular Program, the International Pediatric Heart Failure Summit assembled leaders in clinical and scientific disciplines related to paediatric heart failure and created a multi-disciplinary "think-tank". The purpose of this manuscript is to summarise the lessons from the 2015 International Pediatric Heart Failure Summit of Johns Hopkins All Children's Heart Institute, to describe the "state of the art" of the treatment of paediatric cardiac failure, and to discuss future directions for research in the domain of paediatric cardiac failure.

Neonatal Cardiac Mesenchymal Stromal Cells Promote Recovery of Infarcted Myocardium through CD44 Mediated FoxP3+ T-Regulatory Cells after Vascular Infusion.

Intravenous infusion has been used as the method of cell delivery in many preclinical studies as well as in some early clinical trials. Among its advantages are broad distribution, ability to handle a large-volume infusion, and ease of access. Progenitor cells used in cell-based therapy act through their secretomes, rather than their ability to differentiate into lineage-specific cell type. Since not all progenitor cells have similar secretome potency, the innate abilities of the secretome of cells used in clinical trials will obviously dictate their effectiveness. We previously found that cardiac neonatal mesenchymal stromal cells (nMSCs) are more effective in repairing the infarcted myocardium compared to adult mesenchymal stromal cells (aMSCs) due to their robust secretome (Sharma et al Circulation Research 120:816-834, 2017). In this study, we explored the efficacy of intravenous (IV) delivery of nMSCs for myocardial recovery. Six-week-old male Brown Norway rats underwent acute MI by ligation of the left anterior descending artery, followed by IV infusion of cell dose 5 × 106 nMSCs/rat body weight (kg) or saline on days 0 and 5. We found that cardiac parameters in the rodent ischemia model improved 1 month after nMSCs infusion, and the result is comparable with the intramyocardial injection of nMSCs. Tracking the infused cells in target organ revealed that their movement after IV delivery was mediated by the cell surface receptor CD44. Systemic injection of nMSCs stimulated immunomodulatory responses specifically by increasing FoxP3+ T-regulatory cell influenced anti-inflammatory macrophages (M2) in heart. These data demonstrate that nMSCs promote immunogenic tolerance via CD44-driven T-reg/M2 stimulation that helps nMSCs for longer viability in the injured myocardium for better functional recovery. Our data also demonstrate a rationale for a clinical trial of IV infusion of nMSCs to promote cardiac function improvement in the ischemic patients.

Small Molecule Activators of Mitochondrial Fusion Prevent Congenital Heart Defects Induced by Maternal Diabetes.

Most congenital heart defect (CHD) cases are attributed to nongenetic factors; however, the mechanisms underlying nongenetic factor-induced CHDs are elusive. Maternal diabetes is one of the nongenetic factors, and this study aimed to determine whether impaired mitochondrial fusion contributes to maternal diabetes-induced CHDs and if mitochondrial fusion activators, teriflunomide and echinacoside, could reduce CHD incidence in diabetic pregnancy. We demonstrated maternal diabetes-activated FoxO3a increases miR-140 and miR-195, which in turn represses Mfn1 and Mfn2, leading to mitochondrial fusion defects and CHDs. Two mitochondrial fusion activators are effective in preventing CHDs in diabetic pregnancy.

Genetic deletion or pharmacologic inhibition of histone deacetylase 6 protects the heart against ischaemia/reperfusion injury by limiting tumour necrosis factor alpha-induced mitochondrial injury in experimental diabetes.

AIMS: The histone deacetylase 6 (HDAC6) inhibitor, tubastatin A (TubA), reduces myocardial ischaemia/reperfusion injury (MIRI) in type 1 diabetic rats. It remains unclear whether HDAC6 regulates MIRI in type 2 diabetic animals. Diabetes augments the activity of HDAC6 and the generation of tumour necrosis factor alpha (TNF-α) and impairs mitochondrial complex I (mCI). Here, we examined how HDAC6 regulates TNF-α production, mCI activity, mitochondria, and cardiac function in type 1 and type 2 diabetic mice undergoing MIRI. METHODS AND RESULTS: HDAC6 knockout, streptozotocin-induced type 1 diabetic, and obese type 2 diabetic db/db mice underwent MIRI in vivo or ex vivo in a Langendorff-perfused system. We found that MIRI and diabetes additively augmented myocardial HDAC6 activity and generation of TNF-α, along with cardiac mitochondrial fission, low bioactivity of mCI, and low production of adenosine triphosphate. Importantly, genetic disruption of HDAC6 or TubA decreased TNF-α levels, mitochondrial fission, and myocardial mitochondrial nicotinamide adenine dinucleotide levels in ischaemic/reperfused diabetic mice, concomitant with augmented mCI activity, decreased infarct size, and improved cardiac function. Moreover, HDAC6 knockout or TubA treatment decreased left ventricular dilation and improved cardiac systolic function 28 days after MIRI. H9c2 cardiomyocytes with and without HDAC6 knockdown were subjected to hypoxia/reoxygenation injury in the presence of high glucose. Hypoxia/reoxygenation augmented HDAC6 activity and TNF-α levels and decreased mCI activity. These negative effects were blocked by HDAC6 knockdown. CONCLUSION: HDAC6 is an essential negative regulator of MIRI in diabetes. Genetic deletion or pharmacologic inhibition of HDAC6 protects the heart from MIRI by limiting TNF-α-induced mitochondrial injury in experimental diabetes.

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.

Stem cell therapy for pediatric dilated cardiomyopathy.

Dilated cardiomyopathy is a serious and life-threatening disorder in children. It is the most common form of pediatric cardiomyopathy. Therapy for this condition has varied little over the last several decades and mortality continues to be high. Currently, children with dilated cardiomyopathy are treated with pharmacological agents and mechanical support, but most require heart transplantation and survival rates are not optimal. The lack of common treatment guidelines and inadequate survival rates after transplantation necessitates more therapeutic clinical trials. Stem cell and cell-based therapies offer an innovative approach to restore cardiac structure and function towards normal, possibly reducing the need for aggressive therapies and cardiac transplantation. Mesenchymal stem cells and cardiac stem cells may be the most promising cell types for treating children with dilated cardiomyopathy. The medical community must begin a systematic investigation of the benefits of current and novel treatments such as stem cell therapies for treating pediatric dilated cardiomyopathy.