Objective-CD34 + cells, present within the bone marrow, have previously been shown to possess pancreatic endocrine potential. Based on this observation, we explored the capacity of CD34 + cells derived in culture from the differentiation of human embryonic stem cells (hESC), for their in-vivo pancreatic endocrine capacity.Methods-Sheep were transplanted with hESC-derived CD34 + cells, as well as non-sorted differentiated cultures. Transplantations were carried out with in-utero intraperitoneal injections prior to the development of the immune system in the fetus so that tolerance towards foreign antigens was acquired during gestation and persisted in the adult.Results-All cell populations that were tested demonstrated human cellular activity and long-term presence up to 5 years. However, the in-vivo beta-cell-like activity achieved from the transplantation of the sorted CD34 + cell population was not augmented by transplanting the entire cell population from which the CD34 + cells were isolated. Human DNA and insulin mRNA were detected in sheep pancreases. An average of 1.51 ng/mL human C-peptide was detected in serum from 8 animals transplanted with differentiated cell populations and assayed up to 55 months post-transplantation. Transplantation of as few as 23,500 cells resulted in long-term sustainable beta-cell like activity. Teratomas were absent in the transplanted animals.Conclusion-Our data suggest that hESC-derived CD34 + cells have a potential for long term invivo endocrine cellular activity which could prove useful in regenerative medicine. Since the same cell population has previously been shown to contain hematopoietic potential, it could be used for the induction of immunological tolerance and bone marrow chimerism prior to cellular therapy for diabetes. KeywordsHuman embryonic stem cells; Pancreas; Transplantation; Beta-cells; C-peptide Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. One potential therapy for diabetics is the restoration of beta-cell mass for the endogenous control of glycemia [1,2]. To this end, human embryonic stem cells (hESC) which have the capacity to form tissues of all three germ layers cells [3], are being evaluated for cellular transplantation therapy. hESC differentiation cultures have been developed for the derivation of progenitor cells that engraft and differentiate in-vivo into beta-cell-like cells [4,5]. Such research has had to occur in the absence of knowledge of a single or a group of cell-surface markers that may be used to unequivocally identify and isolate pancreatic stem cells [6]. Hence, focus has been placed ...

Background Inadequate engraftment of hematopoietic stem cells (HSCs) following in-utero HSC transplantation (IUHSCT) remains a major obstacle for the prenatal correction of numerous hereditary disorders. HSCs express CXCR4 receptors that allow homing and engraftment in response to SDF1 ligand present in the bone marrow (BM) stromal niche. Plerixafor, a mobilization drug, works through the interruption of the CXCR4-SDF1 axis. Methods We used the fetal sheep large animal model to test our hypotheses that: a) by administering plerixafor in-utero before performing IUHSCT to release fetal HSCs and thus vacating recipient HSC niches, b) by using human mesenchymal stromal/stem cells (MSCs) to immunomodulate and humanize the fetal BM niches, and c) by increasing the CXCR4+ fraction of CD34+ HSCs, we could improve engraftment. Human cord blood-derived CD34+ cells and human bone marrow-derived MSCs were used for these studies. Results When MSCs were transplanted one week prior to CD34+ cells with plerixafor treatment, we observed 2.80% donor hematopoietic engraftment. Combination of this regimen with additional CD34+ cells at the time of MSC infusion increased engraftment levels to 8.77%. Next, increasing the fraction of CXCR4+ cells in the CD34+ population albeit transplanting at a late gestation age was not beneficial. Our results show engraftment of both lymphoid and myeloid lineages. Discussion Prior MSC and HSC cotransplantation followed by manipulation of the CXCR4-SDF1 axis in IUHSCT provides an innovative conceptual approach for conferring competitive advantage to donor HSCs. Our novel approach could provide a clinically relevant approach for enhancing engraftment early in the fetus.