Dataset Information

Safety and Efficacy of Megakaryocytes Induced from Hematopoietic Stem Cells in Murine and Nonhuman Primate Models.

ABSTRACT: Because of a lack of platelet supply and a U.S. Food and Drug Administration-approved platelet growth factor, megakaryocytes have emerged as an effective substitute for alleviating thrombocytopenia. Here, we report the development of an efficient two-stage culture system that is free of stroma, animal components, and genetic manipulations for the production of functional megakaryocytes from hematopoietic stem cells. Safety and functional studies were performed in murine and nonhuman primate models. One human cryopreserved cord blood CD34⁺ cell could be induced ex vivo to produce up to 1.0 × 10⁴ megakaryocytes that included CD41a⁺ and CD42b⁺ cells at 82.4% ± 6.1% and 73.3% ± 8.5% (mean ± SD), respectively, yielding approximately 650-fold higher cell numbers than reported previously. Induced human megakaryocytic cells were capable of engrafting and producing functional platelets in the murine xenotransplantation model. In the nonhuman primate model, transplantation of primate megakaryocytic progenitors increased platelet count nadir and enhanced hemostatic function with no adverse effects. In addition, primate platelets were released in vivo as early as 3 hours after transplantation with autologous or allogeneic mature megakaryocytes and lasted for more than 48 hours. These results strongly suggest that large-scale induction of functional megakaryocytic cells is applicable for treating thrombocytopenic blood diseases in the clinic. Stem Cells Translational Medicine 2017;6:897-909.

SUBMITTER: Guan X

PROVIDER: S-EPMC5442772 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Safety and Efficacy of Megakaryocytes Induced from Hematopoietic Stem Cells in Murine and Nonhuman Primate Models.

Guan Xin X Qin Meng M Zhang Yu Y Wang Yanan Y Shen Bin B Ren Zhihua Z Ding Xinxin X Dai Wei W Jiang Yongping Y

Stem cells translational medicine 20161007 3

Because of a lack of platelet supply and a U.S. Food and Drug Administration-approved platelet growth factor, megakaryocytes have emerged as an effective substitute for alleviating thrombocytopenia. Here, we report the development of an efficient two-stage culture system that is free of stroma, animal components, and genetic manipulations for the production of functional megakaryocytes from hematopoietic stem cells. Safety and functional studies were performed in murine and nonhuman primate mode ...[more]

PMID: 28297572

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Project description:BackgroundHematopoietic stem cell (HSC) transplantation has been widely applied to the treatment of malignant blood diseases. However, limited number of functional HSCs hinders successful transplantation. The purpose of our current study is to develop a new and cost-efficient medium formulation that could greatly enhance the expansion of HSCs while retaining their long-term repopulation and hematopoietic properties for effective clinical transplantation.MethodsEnriched human CD34+ cells and mobilized nonhuman primate peripheral blood CD34+ cells were expanded with a new, cost-efficient expansion medium formulation, named hematopoietic expansion medium (HEM), consisting of various cytokines and nutritional supplements. The long-term repopulation potential and hematologic-lineage differentiation ability of expanded human cells were studied in the non-obese diabetic/severe combined immunodeficiency mouse model. Furthermore, the efficacy and safety studies were performed by autologous transplantation of expanded primate cells in the nonhuman primate model.ResultsHEM could effectively expand human CD34+ cells by up to 129 fold within 9 days. Expanded HSCs retained long-term repopulation potential and hematologic-lineage differentiation ability, as indicated by (1) maintenance (over unexpanded HSCs) of immunophenotypes of CD38-CD90+CD45RA-CD49f+ in CD34+ cells after expansion; (2) significant presence of multiple human hematopoietic lineages in mouse peripheral blood and bone marrow following primary transplantation; (3) enrichment (over unexpanded HSCs) in SCID-repopulating cell frequency measured by limiting dilution analysis; and (4) preservation of both myeloid and lymphoid potential among human leukocytes from mouse bone marrow in week 24 after primary transplantation or secondary transplantation. Moreover, the results of autologous transplantation in nonhuman primates demonstrated that HEM-expanded CD34+ cells could enhance hematological recovery after myelo-suppression. All primates transplanted with the expanded autologous CD34+ cells survived for over 18 months without any noticeable abnormalities.ConclusionsTogether, these findings demonstrate promising potential for the utility of HEM to improve expansion of HSCs for clinical application.

Project description:Parthenogenesis is the biological phenomenon by which embryonic development is initiated without male contribution. Whereas parthenogenesis is a common mode of reproduction in lower organisms, the mammalian parthenote fails to produce a successful pregnancy. We herein describe in vitro parthenogenetic development of monkey (Macaca fascicularis) eggs to the blastocyst stage, and their use to create a pluripotent line of stem cells. These monkey stem cells (Cyno-1 cells) are positive for telomerase activity and are immunoreactive for alkaline phosphatase, octamer-binding transcription factor 4 (Oct-4), stage-specific embryonic antigen 4 (SSEA-4), tumor rejection antigen 1-60 (TRA 1-60), and tumor rejection antigen 1-81 (TRA 1-81) (traditional markers of human embryonic stem cells). They have a normal chromosome karyotype (40 + 2) and can be maintained in vitro in an undifferentiated state for extended periods of time. Cyno-1 cells can be differentiated in vitro into dopaminergic and serotonergic neurons, contractile cardiomyocyte-like cells, smooth muscle, ciliated epithelia, and adipocytes. When Cyno-1 cells were injected into severe combined immunodeficient mice, teratomas with derivatives from all three embryonic germ layers were obtained. When grown on fibronectin/laminin-coated plates and in neural progenitor medium, Cyno-1 cells assume a neural precursor phenotype (immunoreactive for nestin). However, these cells remain proliferative and express no functional ion channels. When transferred to differentiation conditions, the nestin-positive precursors assume neuronal and epithelial morphologies. Over time, these cells acquire electrophysiological characteristics of functional neurons (appearance of tetrodotoxin-sensitive, voltage-dependent sodium channels). These results suggest that stem cells derived from the parthenogenetically activated nonhuman primate egg provide a potential source for autologous cell therapy in the female and bypass the need for creating a competent embryo.

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Safety and Efficacy of Megakaryocytes Induced from Hematopoietic Stem Cells in Murine and Nonhuman Primate Models.

Publications

Safety and Efficacy of Megakaryocytes Induced from Hematopoietic Stem Cells in Murine and Nonhuman Primate Models.

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