Different cardiac stem/progenitor cells have been recently identified in the post-natal heart. We describe here the identification, clonal expansion and characterization of self-renewing progenitors that differ from those previously described for high spontaneous cardiac differentiation. Unique coexpression of endothelial and pericyte markers identify these cells as cardiac mesoangioblasts and allow prospective isolation and clonal expansion from the juvenile mouse ventricle. Cardiac mesoangioblasts express many cardiac transcription factors and spontaneously differentiate into beating cardiomyocytes that assemble mature sarcomeres and express typical cardiac ion channels. Cells similarly isolated from the atrium do not spontaneously differentiate. When injected into the ventricle after coronary artery ligation, cardiac mesoangioblasts efficiently generate new myocardium in the peripheral area of the necrotic zone, as they do when grafted in the embryonic chick heart. These data identify cardiac mesoangioblasts as committed progenitors, downstream of earlier stem/progenitor cells and suitable for the cell therapy of a subset of juvenile cardiac diseases. Cell Death and Differentiation (2008) Several acute or chronic cardiac diseases are characterized by progressive expansion of the left ventricular chamber, with replacement by fibrous deposition in the ventricular wall. One approach proposed for reverse myocardial remodeling is regeneration of cardiac myocytes using stem cells.1 On the basis of distinct cell surface markers such as Sca-1 or c-Kit, different cardiac stem-like cells have been isolated that can restore cardiac function after ischemic injury.2,3 None of these cells shows spontaneous cardiac differentiation and they also differentiate into other tissue types of the heart.2-5 On the other hand, Isl-1 expressing progenitors appear to be committed to cardiac differentiation but still require interactions with other cells for both proliferation and differentiation. 4 It is also becoming clear that a significant part of the beneficial effect that most of these cells exert on the infarcted heart is due to the secretion of factors that increase survival of residual myocardium and/or favor angiogenesis. 6 This was for example the case of embryonic mesoangioblasts whose transplantation resulted in a 50% recovery of cardiac function but whose differentiation into new cardiomyocytes was rare. 7Our recent work on mesoangioblasts isolated from postnatal skeletal muscle, 8,9 indicated that these cells, possibly because of a local commitment, exhibit efficient differentiation into skeletal muscle. On this basis, we isolated mesoangioblast-like cells from different regions of the post-natal mouse heart.Here we describe the isolation, through a specific explant culture method, of self-renewing committed cardiac progenitors from different regions of the juvenile heart. These cells, operationally termed 'cardiac mesoangioblasts', show a unique phenotype and high spontaneous cardiomyocyte differentiation; they can be e...
Rationale: A cell-based biological pacemaker is based on the differentiation of stem cells and the selection of a population displaying the molecular and functional properties of native sinoatrial node (SAN) cardiomyocytes. So far, such selection has been hampered by the lack of proper markers. CD166 is specifically but transiently expressed in the mouse heart tube and sinus venosus, the prospective SAN. Objective:We have explored the possibility of using CD166 expression for isolating SAN progenitors from differentiating embryonic stem cells. Methods and Results:
The efficacy of cardiac repair by stem cell administration relies on a successful functional integration of injected cells into the host myocardium. Safety concerns have been raised about the possibility that stem cells may induce foci of arrhythmia in the ischemic myocardium. In a previous work ( 36 ), we showed that human cord blood CD34+ cells, when cocultured on neonatal mouse cardiomyocytes, exhibit excitation-contraction coupling features similar to those of cardiomyocytes, even though no human genes were upregulated. The aims of the present work are to investigate whether human CD34+ cells, isolated after 1 wk of coculture with neonatal ventricular myocytes, possess molecular and functional properties of cardiomyocytes and to discriminate, using a reporter gene system, whether cardiac differentiation derives from a (trans)differentiation or a cell fusion process. Umbilical cord blood CD34+ cells were isolated by a magnetic cell sorting method, transduced with a lentiviral vector carrying the enhanced green fluorescent protein (EGFP) gene, and seeded onto primary cultures of spontaneously beating rat neonatal cardiomyocytes. Cocultured EGFP+/CD34+-derived cells were analyzed for their electrophysiological features at different time points. After 1 wk in coculture, EGFP+ cells, in contact with cardiomyocytes, were spontaneously contracting and had a maximum diastolic potential (MDP) of −53.1 mV, while those that remained isolated from the surrounding myocytes did not contract and had a depolarized resting potential of −11.4 mV. Cells were then resuspended and cultured at low density to identify EGFP+ progenitor cell derivatives. Under these conditions, we observed single EGFP+ beating cells that had acquired an hyperpolarization-activated current typical of neonatal cardiomyocytes (EGFP+ cells, −2.24 ± 0.89 pA/pF; myocytes, −1.99 ± 0.63 pA/pF, at −125 mV). To discriminate between cell autonomous differentiation and fusion, EGFP+/CD34+ cells were cocultured with cardiac myocytes infected with a red fluorescence protein-lentiviral vector; under these conditions we found that 100% of EGFP+ cells were also red fluorescent protein positive, suggesting cell fusion as the mechanism by which cardiac functional features are acquired.
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