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a Department of Internal Medicine,
b Research Center for Genetic Engineering and Cell Transplantation, and
c Department of Pediatrics, Tokai University School of Medicine, Kanagawa, Japan; and
d Division of Hematology, Pharmaceutical Frontier Research Laboratory, JT Inc., Japan
Key Words. Stem cell • Peripheral blood • Bone marrow • HESS-5 • Stromal cell • SRC
Kiyoshi Ando, M.D., Division of Hematology, Department of Internal Medicine, Tokai University School of Medicine, Bohseidai, Isehara, Kanagawa, 259-1193, Japan. Telephone: 81-463-93-1121, ext. 2230; Fax: 81-463-4511; e-mail: andok{at}keyaki.cc.u-tokai.ac.jp
Human bone marrow (BM) or mobilized peripheral blood (mPB) CD34+ cells have been shown to loose their stem cell quality during culture period more easily than those from cord blood (CB). We previously reported that human umbilical CB stem cells could effectively be expanded in the presence of human recombinant cytokines and a newly established murine bone marrow stromal cell line HESS-5. In this study we assessed the efficacy of this xenogeneic coculture system using human BM and mPB CD34+ cells as materials. We measured the generation of CD34+CD38– cells and colony-forming units, and assessed severe-combined immunodeficient mouse-repopulating cell (SRC) activity using cells five days after serum-free cytokine-containing culture in the presence or the absence of a direct contact with HESS-5 cells. As compared with the stroma-free culture, the xenogeneic coculture was significantly superior on expansion of CD34+CD38– cells and colony-forming cells and on maintenance of SRC activity. The PKH26 study demonstrated that cell division was promoted faster in cells cocultured with HESS-5 cells than in cells cultured without HESS-5 cells. These results indicate that HESS-5 supports rapid generation of primitive progenitor cells (PPC) and maintains reconstituting ability of newly generated stem cells during ex vivo culture irrespective of the source of samples. This xenogeneic coculture system will be useful for ex vivo manipulation such as gene transduction to promote cell division and the generation of PPC and to prevent loss of stem cell quality.
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