title: Identification of long-term repopulating potential of human cord blood-derived CD34-flt3- severe combined immunodeficiency-repopulating cells by intra-bone marrow injection.
abstract: Recently, we have identified human cord blood (CB)-derived CD34-negative (CD34(-)) severe combined immunodeficiency (SCID)-repopulating cells (SRCs) using the intra-bone marrow injection (IBMI) method (Blood 2003;101:2924). In contrast to murine CD34(-) Kit(+)Sca-1(+)Lineage(-) (KSL) cells, human CB-derived Lin(-)CD34(-) cells did not express detectable levels of c-kit by flow cytometry. In this study, we have investigated the function of flt3 in our identified human CB-derived CD34(-) SRCs. Both CD34(+)flt3(+/-) cells showed SRC activity. In the CD34(-) cell fraction, only CD34(-)flt3(-) cells showed distinct SRC activity by IBMI. Although CD34(+)flt3(+) cells showed a rather weak secondary repopulating activity, CD34(+)flt3(-) cells repopulated many more secondary recipient mice. However, CD34(-)flt3(-) cells repopulated all of the secondary recipients, and the repopulating rate was much higher. Next, we cocultured CD34(-)flt3(-) cells with the murine stromal cell line HESS-5. After 1 week, significant numbers of CD34(+)flt3(+/-) cells were generated, and they showed distinct SRC activity. These results indicated that CB-derived CD34(-)flt3(-) cells produced CD34(+)flt3(-) as well as CD34(+)flt3(+) SRCs in vitro. The present study has demonstrated for the first time that CB-derived CD34(-) SRCs, like murine CD34(-) KSL cells, do not express flt3. On the basis of these data, we propose that the immunophenotype of very primitive long-term repopulating human hematopoietic stem cells is Lin(-)CD34(-)c-kit(-)flt3(-). Disclosure of potential conflicts of interest is found at the end of this article.
authors.affiliation: Department of Stem Cell Biology and Regenerative Medicine, Graduate School of Medical Science, Kansai Medical University, Moriguchi, Osaka 570-8506, Japan.
authors.forename: Takafumi
authors.initials: T
authors.lastname: Kimura
authors.forename: Rumiko
authors.initials: R
authors.lastname: Asada
authors.forename: Jianfeng
authors.initials: J
authors.lastname: Wang
authors.forename: Takashi
authors.initials: T
authors.lastname: Kimura
authors.forename: Miho
authors.initials: M
authors.lastname: Morioka
authors.forename: Kazuo
authors.initials: K
authors.lastname: Matsui
authors.forename: Katsuya
authors.initials: K
authors.lastname: Kobayashi
authors.forename: Kae
authors.initials: K
authors.lastname: Henmi
authors.forename: Shiro
authors.initials: S
authors.lastname: Imai
authors.forename: Masakazu
authors.initials: M
authors.lastname: Kita
authors.forename: Takashi
authors.initials: T
authors.lastname: Tsuji
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Sasaki
authors.forename: Susumu
authors.initials: S
authors.lastname: Ikehara
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Sonoda
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D001854:Bone Marrow Cells / Q000166:cytology; D049109:Cell Proliferation; D002478:Cells, Cultured; D036101:Cord Blood Stem Cell Transplantation / Q000379:methods*; D005312:Fetal Blood / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D017148:Infusions, Intraosseous; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism; D016511:Severe Combined Immunodeficiency / Q000378:metabolism / Q000473:pathology*; D014183:Transplantation, Heterologous; D051941:fms-Like Tyrosine Kinase 3 / Q000378:metabolism*
chemical_list: D018952:Antigens, CD34; C497970:FLT3 protein, human; D019009:Proto-Oncogene Proteins c-kit; D051941:fms-Like Tyrosine Kinase 3