Abstract
Adult cardiac progenitor/stem cells (CPC/CSC) are multipotent resident populations involved in cardiac homeostasis and heart repair. Assisted by complementary RNAseq analysis, we defined the fraction of the CPC proteome associable with specific functions by comparison with human bone marrow mesenchymal stem cells (MSC), the reference population for cell therapy, and human dermal fibroblasts (HDF), as a distant reference. Label-free proteomic analysis identified 526 proteins expressed differentially in CPC. iTRAQ analysis confirmed differential expression of a substantial proportion of those proteins in CPC relative to MSC, and systems biology analysis defined a clear overrepresentation of several categories related to enhanced angiogenic potential. The CPC plasma membrane compartment comprised 1,595 proteins, including a minimal signature of 167 proteins preferentially or exclusively expressed by CPC. CDH5 (VE-cadherin), OX2G (OX-2 membrane glycoprotein; CD200), GPR4 (G protein-coupled receptor 4), CACNG7 (calcium voltage-gated channel auxiliary subunit gamma 7) and F11R (F11 receptor; junctional adhesion molecule A; JAM-A; CD321) were selected for validation. Their differential expression was confirmed both in expanded CPC batches and in early stages of isolation, particularly when compared against cardiac fibroblasts. Among them, GPR4 demonstrated the highest discrimination capacity between all cell lineages analyzed.
Highlights
Adult cardiac progenitor/stem cells (CPC/cardiac stem cells (CSC)) are multipotent resident populations involved in cardiac homeostasis and heart repair
As a first approach to define specific CPC functions, we used mRNAseq to compare human CPC from three independent donors (CPC1–3) with human bone marrow mesenchymal stem cells (MSC) (n = 3; aiming to identify putative genes related to multipotency), and with human dermal fibroblasts (HDF) as a distant reference (n = 3; to discard genes expressed in all cell types)
The strong pro-angiogenic www.nature.com/scientificreports activity of CPC compared with MSC has been recently confirmed by comparative secretome analysis[29], demonstrating an important role for CXCL6, which we found upregulated by RNAseq (Fig. 1d)
Summary
Adult cardiac progenitor/stem cells (CPC/CSC) are multipotent resident populations involved in cardiac homeostasis and heart repair. CDH5 (VE-cadherin), OX2G (OX-2 membrane glycoprotein; CD200), GPR4 (G protein-coupled receptor 4), CACNG7 (calcium voltage-gated channel auxiliary subunit gamma 7) and F11R (F11 receptor; junctional adhesion molecule A; JAM-A; CD321) were selected for validation Their differential expression was confirmed both in expanded CPC batches and in early stages of isolation, when compared against cardiac fibroblasts. This controversy prompted a more precise study of c-kit + populations, which concluded that the evident differences seem to be related to the intrinsic limitations of the technique used[11,12] Current thoughts on these issues are more conciliatory and ckit-expression is considered necessary but not sufficient to define CSC13, and the limitations of most lineage-tracing mouse models using c-kit promoter seem evident[11]. Low turnover based on resident CSC/CPC is, compatible with a degree of transient dedifferentiation and limited proliferation of pre-existing cardiomyocytes in response to specific signals[15]
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