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A precise identification of adult human hemangioblast is still lacking. To identify circulating precursors having the developmental potential of the hemangioblast, we established a new ex vivo long-term culture model supporting the differentiation of both hematopoietic and endothelial cell lineages. We identified from peripheral blood a population lacking the expression of CD34, lineage markers, CD45 and CD133 (CD34-Lin-CD45-CD133- cells), endowed with the ability to differentiate after a 6-week culture into both hematopoietic and endothelial lineages. The bilineage potential of CD34-Lin-CD45-CD133- cells was determined at the single-cell level in vitro and was confirmed by transplantation into NOD/SCID mice. In vivo, CD34-Lin-CD45-CD133- cells showed the ability to reconstitute hematopoietic tissue and to generate functional endothelial cells that contribute to new vessel formation during tumor angiogenesis. Molecular characterization of CD34-Lin-CD45-CD133- cells unveiled a stem cell profile compatible with both hematopoietic and endothelial potentials, characterized by the expression of c-Kit and CXCR4 as well as EphB4, EphB2, and ephrinB2. Further molecular and functional characterization of CD34-Lin-CD45-CD133- cells will help dissect their physiologic role in blood and blood vessel maintenance and repair in adult life. [ABSTRACT FROM AUTHOR]

Abstract: The identity of biochemical players which underpin the commitment of CD34+ hematopoietic stem cells to immunogenic or tolerogenic dendritic cells is largely unknown. To explore this issue, we employed a previously established cell-based system amenable to shift dendritic cell differentiation from the immunogenic into the tolerogenic pathway upon supplementation with a conventional cytokine cocktail containing thrombopoietin (TPO) and IL-16. We show that stringent regulation of cathepsins S and D, two proteases involved in antigen presentation, is crucial to engage cell commitment to either route. In response to TPO+IL-16-dependent signaling, both cathepsins undergo earlier maturation and down-regulation. Additionally, cystatin C orchestrates cathepsin S expression through a tight but reversible interaction that, based on a screen of adult stem cells from disparate origins, CD14+ cells, primary fibroblasts and the MCF7 cell line, appears unique to CD34+ stem cells from peripheral and cord blood. As shown by CD4+ T cell proliferation in mixed-lymphocyte reactions, cell commitment to either pathway is disrupted upon cathepsin knockdown by RNAi. Surprisingly, similar effects were also observed upon gene overexpression, which prompts atypically accelerated maturation of cathepsins S and D in cells of the immunogenic pathway, similar to the tolerogenic route. Furthermore, RNAi studies revealed that cystatin C is a proteolytic target of cathepsin D and has a direct, causal impact on cell differentiation. Together, these findings uncover a novel biochemical cluster that is subject to time-controlled and rigorously balanced expression to mediate specific stem cell commitment at the crossroads towards tolerance or immunity. [Copyright &y& Elsevier]

Tissue engineering is one of the major challenges of orthopedics and trauma surgery for bone regeneration. Biomaterials filled with mesenchymal stem cells (MSCs) are considered the most promising approach in bone tissue engineering. Furthermore, our previous study showed that the multi-phase poly [ε-caprolactone]/thermoplastic zein-hydroxyapatite (PCL/TZ-HA) biomaterials improved rabbit (r) MSCs adhesion and osteoblast differentiation, thus demonstrating high potential of this bioengineered scaffold for bone regeneration. In the recent past, CD271 has been applied as a specific selective marker for the enrichment of MSCs from bone marrow (BM-MSCs). In the present study, we aimed at establishing whether CD271-based enrichment could be an efficient method for the selection of rBM-MSCs, displaying higher ability in osteogenic differentiation than non-selected rBM-MSCs in an in vitro system. CD271(+) cells were isolated from rabbit bone marrow and were compared with rMSCs in their proliferation rate and osteogenic differentiation capability. Furthermore, rCD271(+) cells were tested in their ability to adhere, proliferate and differentiate into osteogenic lineage, while growing on PCL/TZ-HA scaffolds, in comparison to rMSCs. Our result demonstrate that rCD271(+) cells were able to adhere, proliferate and differentiate into osteoblasts when cultured on PCL/TZ-HA scaffolds in significantly higher levels as compared to rMSCs. Based on these findings, CD271 marker might serve as an optimal alternative MSCs selection method for the potential preclinical and clinical application of these cells in bone tissue regeneration.