James J.H. Chong

Human embryonic-stem-cell-derived cardiomyocytes regenerate non-human primate hearts

Pluripotent stem cells provide a potential solution to current epidemic rates of heart failure by providing human cardiomyocytes to support heart regeneration. Studies of human embryonic-stem-cell-derived cardiomyocytes (hESC-CMs) in small-animal models have shown favourable effects of this treatment. However, it remains unknown whether clinical-scale hESC-CM transplantation is feasible, safe or can provide sufficient myocardial regeneration. Here we show that hESC-CMs can be produced at a clinical scale (more than one billion cells per batch) and cryopreserved with good viability. Using a non-human primate model of myocardial ischaemia followed by reperfusion, we show that cryopreservation and intra-myocardial delivery of one billion hESC-CMs generates extensive remuscularization of the infarcted heart. The hESC-CMs showed progressive but incomplete maturation over a 3-month period. Grafts were perfused by host vasculature, and electromechanical junctions between graft and host myocytes were present within 2 weeks of engraftment. Importantly, grafts showed regular calcium transients that were synchronized to the host electrocardiogram, indicating electromechanical coupling. In contrast to small-animal models, non-fatal ventricular arrhythmias were observed in hESC-CM-engrafted primates. Thus, hESC-CMs can remuscularize substantial amounts of the infarcted monkey heart. Comparable remuscularization of a human heart should be possible, but potential arrhythmic complications need to be overcome.

Adult Cardiac-Resident MSC-like Stem Cells with a Proepicardial Origin

Colony-forming units - fibroblast (CFU-Fs), analogous to those giving rise to bone marrow (BM) mesenchymal stem cells (MSCs), are present in many organs, although the relationship between BM and organ-specific CFU-Fs in homeostasis and tissue repair is unknown. Here we describe a population of adult cardiac-resident CFU-Fs (cCFU-Fs) that occupy a perivascular, adventitial niche and show broad trans-germ layer potency in vitro and in vivo. CRE lineage tracing and embryo analysis demonstrated a proepicardial origin for cCFU-Fs. Furthermore, in BM transplantation chimeras, we found no interchange between BM and cCFU-Fs after aging, myocardial infarction, or BM stem cell mobilization. BM and cardiac and aortic CFU-Fs had distinct CRE lineage signatures, indicating that they arise from different progenitor beds during development. These diverse origins for CFU-Fs suggest an underlying basis for differentiation biases seen in different CFU-F populations, and could also influence their capacity for participating in tissue repair.

Progenitor cells identified by PDGFR-alpha expression in the developing and diseased human heart.

Platelet-derived growth factors (PDGFs) and their tyrosine kinase receptors play instrumental roles in embryonic organogenesis and diseases of adult organs. In particular, platelet-derived growth factor receptor-alpha (PDGFRα) is expressed by multipotent cardiovascular progenitors in mouse and human embryonic stem cell systems. Although cardiac PDGFRα expression has been studied in multiple species, little is known about its expression in the human heart. Using immunofluorescence, we analyzed PDGFRα expression in both human fetal and diseased adult hearts, finding strong expression in the interstitial cells of the epicardium, myocardium, and endocardium, as well as the coronary smooth muscle. Only rare endothelial cells and cardiomyocytes expressed PDGFRα. This pattern was consistent for both the fetal and adult diseased hearts, although more PDGFRα+ cardiomyocytes were noted in the latter. In vitro differentiation assays were then performed on the PDGFRα+ cell fraction isolated from the cardiomyocyte-depleted human fetal hearts. Protocols previously reported to direct differentiation to a cardiomyocyte (5-azacytidine), smooth muscle (PDGF-BB), or endothelial cell fates (vascular endothelial growth factor [VEGF]) were used. Although no significant cardiomyocyte differentiation was observed, PDGFRα+ cells generated significant numbers of smooth muscle cells (smooth muscle-α-actin+ and smooth muscle myosin+) and endothelial cells (CD31+). These data suggest that a subfraction of the cardiac PDGFRα+ populations are progenitors contributing predominantly to the vascular and mesenchymal compartments of the human heart. It may be possible to control the fate of these progenitors to promote vascularization or limit fibrosis in the injured heart.

Cardiac regeneration using pluripotent stem cells--progression to large animal models.

Pluripotent stem cells (PSCs) have indisputable cardiomyogenic potential and therefore have been intensively investigated as a potential cardiac regenerative therapy. Current directed differentiation protocols are able to produce high yields of cardiomyocytes from PSCs and studies in small animal models of cardiovascular disease have proven sustained engraftment and functional efficacy. Therefore, the time is ripe for cardiac regenerative therapies using PSC derivatives to be tested in large animal models that more closely resemble the hearts of humans. In this review, we discuss the results of our recent study using human embryonic stem cell derived cardiomyocytes (hESC-CM) in a non-human primate model of ischemic cardiac injury. Large scale remuscularization, electromechanical coupling and short-term arrhythmias demonstrated by our hESC-CM grafts are discussed in the context of other studies using adult stem cells for cardiac regeneration.

Comparison of Human Embryonic Stem Cell-Derived Cardiomyocytes, Cardiovascular Progenitors, and Bone Marrow Mononuclear Cells for Cardiac Repair

Summary Cardiomyocytes derived from human embryonic stem cells (hESC-CMs) can improve the contractility of injured hearts. We hypothesized that mesodermal cardiovascular progenitors (hESC-CVPs), capable of generating vascular cells in addition to cardiomyocytes, would provide superior repair by contributing to multiple components of myocardium. We performed a head-to-head comparison of hESC-CMs and hESC-CVPs and compared these with the most commonly used clinical cell type, human bone marrow mononuclear cells (hBM-MNCs). In a nude rat model of myocardial infarction, hESC-CMs and hESC-CVPs generated comparable grafts. Both similarly improved systolic function and ventricular dilation. Furthermore, only rare human vessels formed from hESC-CVPs. hBM-MNCs attenuated ventricular dilation and enhanced host vascularization without engrafting long-term or improving contractility. Thus, hESC-CMs and CVPs show similar efficacy for cardiac repair, and both are more efficient than hBM-MNCs. However, hESC-CVPs do not form larger grafts or more significant numbers of human vessels in the infarcted heart.

Developmental origins and lineage descendants of endogenous adult cardiac progenitor cells.

Mammalian hearts carry a number of primitive stem cell-like populations, although the magnitude of their contribution to tissue homeostasis and repair remains controversial. Recent CRE recombinase-based lineage tracing experiments suggest only a minor contribution to the formation of new cardiomyocytes from such cells, albeit one that might be augmented therapeutically. As the field explores clinical translation of cardiac stem cells, it will be important to understand the biology of these cells in great detail. In this review we document the various reported stem and progenitor cell populations in mammalian hearts and discuss the current state of knowledge on their origins and lineage capabilities.

Stem cell therapy for cardiac dysfunction

Following significant injury, the heart undergoes induced compensation and gradually deteriorates towards impending heart failure. Current therapy slows but does not halt the resultant adverse remodeling. Stem cell therapy, however, has the potential to regenerate or repair infarcted heart tissue and therefore is a promising therapeutic strategy undergoing intensive investigation. Due to the wide range of stem cells investigated, it is difficult to navigate this field. This review aims to summarize the main types of stem cells (both of cardiac and extra-cardiac origin) that possess promising therapeutic potential. Particular focus is placed on clinical trials supporting this therapeutic strategy.

Cell Therapy for Left Ventricular Dysfunction: An Overview for Cardiac Clinicians

Cell therapies specifically targeting heart failure could greatly decrease morbidity and burgeoning health care costs worldwide. Due to the great number of cell types being investigated, navigating the cardiovascular regeneration field can be difficult. This brief review gives an overview of the main cell types being explored for cardiac cell therapy. These include populations from extra-cardiac sources (skeletal myoblasts, bone marrow derived mononuclear cells, endothelial progenitor cells, bone marrow or adipose derived mesenchymal stem cells and embryonic or induced pluripotent stem cells as well as newly discovered cardiac stem cell populations (isl1(+), c-kit(+), sca1(+), sca1(+)/pdgfrα(+), cardiosphere derived, cardiac side-population and epicardium derived cells). Although clinical trials using both groups of cell sources have been performed, the vast majority of studies have used bone marrow mononuclear cells. The current wave of clinical trials includes large studies refining specifics of bone marrow mononuclear cell therapy and early phase trials of mesenchymal stem cell and cardiac stem cell populations. Embryonic stem cell derived therapies are being studied in large animal models with the aim of swift progression to clinical trials. Lessons learnt from the intense investigation in this infant field have resulted in rapid translational progress and it is likely that several clinical products/protocols for cardiac repair will be available in the not too distant future.

Comparison of Left Ventricular Ejection Fraction and Inducible Ventricular Tachycardia in ST-Elevation Myocardial Infarction Treated by Primary Angioplasty Versus Thrombolysis

Electrophysiologic studies predict the risk for sudden death after myocardial infarction (MI). Although primary angioplasty has become the preferred method of treatment for ST-elevation MI, intravenous thrombolysis remains the first-line treatment in 30% to 70% of cases worldwide. Rates of ventricular tachyarrhythmias may vary according to type of reperfusion treatment. This study was undertaken to examine the hypothesis that the left ventricular ejection fraction (LVEF) and rates of inducible ventricular tachycardia may be more favorable in treatment with primary angioplasty rather than thrombolysis. Consecutive patients receiving primary angioplasty (n = 225) or thrombolysis (n = 195) for ST-elevation MI were included. The mean LVEF was 48 +/- 12% for the primary angioplasty group and 46 +/- 13% for the thrombolysis group (p = 0.30). The proportion of patients with LVEFs <40% was 30% in the primary angioplasty group and 30% in the thrombolysis group (p = 0.98). Patients with LVEFs <40% underwent electrophysiologic studies. Ventricular tachycardia was inducible in 23 of 66 primary angioplasty patients (34.8%) compared with 21 of 55 (38.1%) thrombolysis patients (p = 0.69). Implantable cardiac defibrillators were inserted in 30 patients, of whom 8 (27%) had appropriate device activations. The mean time from MI to first spontaneous activation was 387 +/- 458 days. In conclusion, patients treated with thrombolysis or primary angioplasty for ST-elevation MIs had similar resultant LVEFs and rates of inducible ventricular tachycardia. There was a surprisingly high rate of spontaneous defibrillator activations, often occurring late after MI.

Evaluation of lesion and thermodynamic characteristics of Symplicity and EnligHTN renal denervation systems in a phantom renal artery model.

AIMS Radiofrequency renal artery denervation has been used effectively to treat resistant hypertension. However, comparison of lesion and thermodynamic characteristics for different systems has not been previously described. We aimed to assess spatiotemporal lesion growth and ablation characteristics of Symplicity and EnligHTN systems. METHODS AND RESULTS A total of 39 ablations were performed in a phantom renal artery model using Symplicity (n=17) and EnligHTN (n=22) systems. The phantom model consisted of a hollowed gel block surrounding a thermochromic liquid crystal (TLC) film, exhibiting temperature sensitivity of 50-78°C. Flow was simulated using 37°C normal saline with impedance equal to blood. Radiofrequency ablations with each system were delivered with direct electrode tip contact to the TLC. Lesion size was interpreted from the TLC as the maximum dimensions of the 51°C isotherm. Mean lesion depth was 3.82 mm±0.04 versus 3.44 mm±0.03 (p<0.001) for Symplicity and EnligHTN, respectively. Mean width was 7.17 mm±0.08 versus 6.23 mm±0.07 (p<0.001), respectively. With EnligHTN, steady state temperature was achieved 20 sec earlier, and was 15°C higher than Symplicity. CONCLUSIONS In this phantom model, Symplicity formed larger lesions compared to EnligHTN with lower catheter-tip temperature. The clinical significance of our findings needs to be explored further.