Abstract
BackgroundHuman bone marrow-derived mesenchymal stem/stromal cells (hBM MSCs) have multiple functions, critical for skeletal formation and function. Their functional heterogeneity, however, represents a major challenge for their isolation and in developing potency and release assays to predict their functionality prior to transplantation. Additionally, potency, biomarker profiles and defining mechanisms of action in a particular clinical setting are increasing requirements of Regulatory Agencies for release of hBM MSCs as Advanced Therapy Medicinal Products for cellular therapies. Since the healing of bone fractures depends on the coupling of new blood vessel formation with osteogenesis, we hypothesised that a correlation between the osteogenic and vascular supportive potential of individual hBM MSC-derived CFU-F (colony forming unit-fibroblastoid) clones might exist.MethodsWe tested this by assessing the lineage (i.e. adipogenic (A), osteogenic (O) and/or chondrogenic (C)) potential of individual hBM MSC-derived CFU-F clones and determining if their osteogenic (O) potential correlated with their vascular supportive profile in vitro using lineage differentiation assays, endothelial-hBM MSC vascular co-culture assays and transcriptomic (RNAseq) analyses.ResultsOur results demonstrate that the majority of CFU-F (95%) possessed tri-lineage, bi-lineage or uni-lineage osteogenic capacity, with 64% of the CFU-F exhibiting tri-lineage AOC potential. We found a correlation between the osteogenic and vascular tubule supportive activity of CFU-F clones, with the strength of this association being donor dependent. RNAseq of individual clones defined gene fingerprints relevant to this correlation.ConclusionsThis study identified a donor-dependent correlation between osteogenic and vascular supportive potential of hBM MSCs and important gene signatures that support these functions that are relevant to their bone regenerative properties.
Highlights
Human bone marrow-derived mesenchymal stem/stromal cells have multiple functions, critical for skeletal formation and function
Since the healing of bone defects in larger bone grafts or fractures depends on the coupling of new blood vessel formation with osteogenesis [55,56,57,58,59], we examined the potential of these individual hBM Mesenchymal stem and stromal cells (MSCs) clones, with differential osteogenic differentiation capacities, to support endothelial networks in co-culture assays in vitro
Since therapeutic doses of hBM MSCs require their expansion in vitro, we isolated a total of 133 colony-forming unit-fibroblast (CFU-F) colonies (P0) and expanded these into T25 tissue culture grade flasks (P1)
Summary
Human bone marrow-derived mesenchymal stem/stromal cells (hBM MSCs) have multiple functions, critical for skeletal formation and function. Mesenchymal stem and stromal cells (MSCs), known as mesenchymal or skeletal stem and progenitor cells (MSPCs or SSCs), are critical components of the post-natal bone marrow (BM) When derived from this tissue, they have the potential to differentiate into multiple lineages (including osteogenic (O), chondrogenic (C) and adipogenic (A)), and to form niche cells which support haematopoietic and blood vessel formation and function (reviewed in [1,2,3,4,5,6,7]). Both plastic adherent CFU-F and those derived after enrichment on density gradients and with the a variety of cell surface markers (e.g. CD271, CD146, MSCA-1, CD105, STRO-1, PDGFR-β (CD140b), ErbB2 (CD340), Frizzled-9 (CD349), CD90, CD295 (LepR), CD106, VCAM-1
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