Highly efficient ex vivo expansion of human hematopoietic stem cells using Delta1-Fc chimeric protein

¹ Department of Regeneration Medicine for Hematopoiesis, Graduate School of Medicine, University of Tokyo, Tokyo, Japan; Division of Tissue Engineering, University of Tokyo Hospital, Tokyo, Japan
² Department of Hematology and Oncology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan
³ Department of Cell Therapy and Transplantation Medicine, University of Tokyo Hospital, Tokyo, Japan
⁴ The Japanese Red Cross Tokyo Blood Center, Tokyo, Japan
⁵ Department of Perinatal Medicine, Graduate School of Medicine, University of Tokyo, Tokyo, Japan
⁶ Division of Tissue Engineering, University of Tokyo Hospital, Tokyo, Japan; Department of Oral and Maxillofacial Surgery, Graduate School of Medicine, University of Tokyo, Tokyo, Japan
⁷ Department of Pediatrics, Graduate School of Medicine, Kyoto University, Kyoto, Japan
⁸ Pharmaceutical Division, Kirin Brewery Co., LTD, Tokyo, Japan
⁹ Corporate Research & Development and Central Research Laboratory, Asahi Kasei Corporation, Tokyo, Japan

^* To whom correspondence should be addressed. E-mail: schiba-tky{at}umin.ac.jp.

	Abstract

Ex vivo expansion of hematopoietic stem cells (HSCs) has been explored in the fields of stem cell biology, gene therapy and clinical transplantation. Here we demonstrate efficient ex vivo expansion of HSCs measured by long-term SCID-repopulating cells (SRCs) from human cord blood CD133-sorted cells using a soluble form of Delta1. After three-week culture on immobilized Delta1 supplemented with stem cell factor, thrombopoietin, Flt-3 ligand, IL-3 and IL-6/soluble IL-6 receptor chimeric protein (FP6) in a serum- and stromal cell-free condition, we achieved approximately 6-fold expansion of SRCs when evaluated by limiting dilution/transplantation assays. The maintenance of full multipotency and self-renewal capacity during culture was confirmed by transplantation to NOD/SCID/c^null mice, which showed myeloid, B, T and NK cells as well as CD133⁺CD34⁺ cells, and hematopoietic reconstitution in the secondary recipients. Interestingly, the CD133-sorted cells contained approximately 4.5 times more SRCs than the CD34-sorted cells. The present study provides a promising method to expand HSCs and encourages future trials on clinical transplantation.

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