Cord blood (CB) mononuclear cells (MNC) were demonstrated to contribute to organ/tissue regeneration, however, the identity of the specific cell type(s) involved remains unknown. Recently, it had been identified in adult bone marrow (BM) a mobile, SDF-1-, HGF/SF- and LIF-responsive population of CXCR4+ non-hematopoietic MNC that expresses markers (RQ-PCR, immunhistochemistry) of early pluripotent/tissue committed stem cells (TCSC) for skeletal muscles, heart, neural tissue, liver, pancreas, epidermis and intestinal epithelium (
). We hypothesized that a similar population of these rare cells may also be present in CB, however, their final yield may depend on the method of MNC preparation. We hypothesized that since these cells are very small (~3–5 μm), they may cross a Ficoll-Paque gradient or sedimentate more rapidly than other CB MNC and as result of this are lost during routine CB preparations. Thus, taking in consideration their small size, in order to evaluate if these cells are present in CB, we isolated small CB MNC by employing Percoll gradient (1.078–1.095). We found that this allows us to isolate from CB a population of small cells (2.5% of the total number of MNC) that is enriched in a population of TCSC/PSC(~0.002% of MNC) that we have originally identified in BM. Accordingly, these CB-derived TCSC/PSC CXCR4+ cells are very small (~3μm), posses large nuclei that contain embryonic stem cell type open chromatin (euchromatin), and express several markers for skeletal muscles, heart, neural tissue, liver, pancreas, epidermis and intestinal epithelium as well as pluripotent stem cell (PSC) transcription factors such as Oct-4, Nanog and Rex-1. In vitro cultures of CB-derived small TCSC/PSC are able to grow neurospheres that gave rise to neuronal (β-III tubulin+, nestin+) and macroglia (O4+, MBP+, GFAP+) lineages and cardiomyocytes (β-myosin heavy chain+, α-sarcomeric actin+). Based on this we conclude that CB contains embryonic-like stem cells which may be lost during routine procedures to isolate MNC. Thus, Percoll gradient centrifugation allows for optimal isolation of these small CXCR4+ PSC/TCSC and we postulate that the CB tissue/organ regenerating potential may be much higher than initially postulated and we are currently testing this hypothesis in vivo in animal models.
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