P286Matrix production and proteolitic activity of cardiomyocyte progenitor cells during in vitro differentiation

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Human cardiomyocyte progenitor cells (CMPCs) reside in the heart and are able to differentiate into cardiomyocytes. In addition, these cells have been shown to restore myocardial damage after injury in an animal model. To repair myocardial damage, not only the lost cardiomyocytes need to be replaced but also the scar tissue needs to be remodeled and replaced with "healthy" extracellular matrix (ECM). Interestingly, the role of CMPCs in this facet of cardiac repair is not yet known. Therefore, the expression and production of ECM proteins and activity of ECM remodeling by metalloproteinases (MMPs) and their inhibitors (TIMPs) was investigated in undifferentiated CMPCs and CMPCs at different time points of the differentiation process into the cardiomyogenic lineage in an in vitro model. In this study, human CMPCs were cultured on coverslips and differentiation was induced by addition of 5-azacytidine followed by TGF-β1 treatment along with addition of vitamin C. Before the onset of differentiation and at weeks 1, 2, 3 and 4 in the differentiation process samples were taken to examine the expression of cardiac differentiation markers. Furthermore, the expression the ECM proteins collagen I, III, elastin and fibronectin and ECM remodeling proteins MMPs and TIMPs were studied at the gene level using real time PCR. Moreover, the expression and localization of ECM proteins inside or outside the cell was examined using confocal microscopy and MMP activity was determined using zymography. The undifferentiated CMPCs expressed ECM proteins at a low level, with the exception of elastin, which was not expressed. During differentiation the expression of collagen I and III and fibronectin increased and interestingly also elastin was induced. MMP-1 expression was downregulated during differentiation, while MMP-2 was upregulated during the whole period of differentiation and MMP-9 expression was only upregulated in the first week of differentiation. The expression of TIMP-2 and -4 was induced during differentiation. These data suggest that CMPCs are able to modulate their own environment, especially upon differentiation. This study provides new insights into matrix production and remodeling capacity of human CMPCs and hence opens new ways for cardiac repair.

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