Reconstitution of human haematopoiesis in non-obese diabetic/severe combined immunodeficient mice by clonal cells expanded from single CD34+CD38-cells expressing Flk2/Flt3

Yasuhiro Ebihara, Mika Wada, Takahiro Ueda, Mingjiang Xu, Atsushi Manabe, Ryuhei Tanaka, Mamoru Ito, Hideo Mugishima, Shigetaka Asano, Tatsutoshi Nakahata, Kohichiro Tsuji

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

In the present study, we examined the expression of Flk2/Flt3, a tyrosine kinase receptor, on human cord blood CD34+ haematopoietic progenitor/stem cells. In flow cytometric analysis, Flk2/Flt3 was expressed on 80% of CD34+ cells and their immature subpopulations, CD34+CD33- and CD34+CD38- cells. Methycellulose clonal culture of sorted CD34+Flk2/Flt3+ and CD34+Flk2/Flt3- cells showed that most of myelocytic progenitors expressed Flk2/Flt3, but erythroid and haematopoietic multipotential progenitors were shared by both fractions. When 1 × 104 lineage marker-negative (Lin-)CD34+Flk2/Flt3- cells were transplanted into non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice, none of the recipients possessed human CD45+ cells in bone marrow 11-12 weeks after the transplantation. In contrast, all recipients transplanted with 1 × 104 Lin-CD34+Flk2/Flt3+ cells showed successful engraftment. Furthermore, clonal cells expanded from single Lin-CD34+CD38-Flk2/Flt3+ cells in the culture with Flk2/Flt3 ligand, stem cell factor, thrombopoietin, and a complex of interleukin 6/soluble interleukin 6 receptor were individually transplanted into NOD/SCID mice. At 20 to 21 weeks after the transplantation, three out of 10 clones harvested at d 7 of culture, and three out of six clones at d 14 could reconstitute human haematopoiesis in recipient marrow. These results demonstrated that Flk2/Flt3 was expressed on a wide variety of human haematopoietic cells including long-term-repopulating haematopoietic stem cells.

Original languageEnglish (US)
Pages (from-to)525-534
Number of pages10
JournalBritish Journal of Haematology
Volume119
Issue number2
DOIs
StatePublished - 2002
Externally publishedYes

Fingerprint

SCID Mice
Hematopoiesis
Hematopoietic Stem Cells
Clone Cells
Transplantation
Thrombopoietin
Interleukin-6 Receptors
Stem Cell Factor
Receptor Protein-Tyrosine Kinases
Fetal Blood
Bone Marrow Cells
Interleukin-6
Cell Culture Techniques
Bone Marrow

Keywords

  • CD34
  • Cord blood
  • Flk2/Flt3
  • Long-term-repopulating haematopoietic stem cells (LTR-HSC)
  • NOD/SCID mice

ASJC Scopus subject areas

  • Hematology

Cite this

Reconstitution of human haematopoiesis in non-obese diabetic/severe combined immunodeficient mice by clonal cells expanded from single CD34+CD38-cells expressing Flk2/Flt3. / Ebihara, Yasuhiro; Wada, Mika; Ueda, Takahiro; Xu, Mingjiang; Manabe, Atsushi; Tanaka, Ryuhei; Ito, Mamoru; Mugishima, Hideo; Asano, Shigetaka; Nakahata, Tatsutoshi; Tsuji, Kohichiro.

In: British Journal of Haematology, Vol. 119, No. 2, 2002, p. 525-534.

Research output: Contribution to journalArticle

Ebihara, Yasuhiro ; Wada, Mika ; Ueda, Takahiro ; Xu, Mingjiang ; Manabe, Atsushi ; Tanaka, Ryuhei ; Ito, Mamoru ; Mugishima, Hideo ; Asano, Shigetaka ; Nakahata, Tatsutoshi ; Tsuji, Kohichiro. / Reconstitution of human haematopoiesis in non-obese diabetic/severe combined immunodeficient mice by clonal cells expanded from single CD34+CD38-cells expressing Flk2/Flt3. In: British Journal of Haematology. 2002 ; Vol. 119, No. 2. pp. 525-534.
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abstract = "In the present study, we examined the expression of Flk2/Flt3, a tyrosine kinase receptor, on human cord blood CD34+ haematopoietic progenitor/stem cells. In flow cytometric analysis, Flk2/Flt3 was expressed on 80{\%} of CD34+ cells and their immature subpopulations, CD34+CD33- and CD34+CD38- cells. Methycellulose clonal culture of sorted CD34+Flk2/Flt3+ and CD34+Flk2/Flt3- cells showed that most of myelocytic progenitors expressed Flk2/Flt3, but erythroid and haematopoietic multipotential progenitors were shared by both fractions. When 1 × 104 lineage marker-negative (Lin-)CD34+Flk2/Flt3- cells were transplanted into non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice, none of the recipients possessed human CD45+ cells in bone marrow 11-12 weeks after the transplantation. In contrast, all recipients transplanted with 1 × 104 Lin-CD34+Flk2/Flt3+ cells showed successful engraftment. Furthermore, clonal cells expanded from single Lin-CD34+CD38-Flk2/Flt3+ cells in the culture with Flk2/Flt3 ligand, stem cell factor, thrombopoietin, and a complex of interleukin 6/soluble interleukin 6 receptor were individually transplanted into NOD/SCID mice. At 20 to 21 weeks after the transplantation, three out of 10 clones harvested at d 7 of culture, and three out of six clones at d 14 could reconstitute human haematopoiesis in recipient marrow. These results demonstrated that Flk2/Flt3 was expressed on a wide variety of human haematopoietic cells including long-term-repopulating haematopoietic stem cells.",
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T1 - Reconstitution of human haematopoiesis in non-obese diabetic/severe combined immunodeficient mice by clonal cells expanded from single CD34+CD38-cells expressing Flk2/Flt3

AU - Ebihara, Yasuhiro

AU - Wada, Mika

AU - Ueda, Takahiro

AU - Xu, Mingjiang

AU - Manabe, Atsushi

AU - Tanaka, Ryuhei

AU - Ito, Mamoru

AU - Mugishima, Hideo

AU - Asano, Shigetaka

AU - Nakahata, Tatsutoshi

AU - Tsuji, Kohichiro

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AB - In the present study, we examined the expression of Flk2/Flt3, a tyrosine kinase receptor, on human cord blood CD34+ haematopoietic progenitor/stem cells. In flow cytometric analysis, Flk2/Flt3 was expressed on 80% of CD34+ cells and their immature subpopulations, CD34+CD33- and CD34+CD38- cells. Methycellulose clonal culture of sorted CD34+Flk2/Flt3+ and CD34+Flk2/Flt3- cells showed that most of myelocytic progenitors expressed Flk2/Flt3, but erythroid and haematopoietic multipotential progenitors were shared by both fractions. When 1 × 104 lineage marker-negative (Lin-)CD34+Flk2/Flt3- cells were transplanted into non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice, none of the recipients possessed human CD45+ cells in bone marrow 11-12 weeks after the transplantation. In contrast, all recipients transplanted with 1 × 104 Lin-CD34+Flk2/Flt3+ cells showed successful engraftment. Furthermore, clonal cells expanded from single Lin-CD34+CD38-Flk2/Flt3+ cells in the culture with Flk2/Flt3 ligand, stem cell factor, thrombopoietin, and a complex of interleukin 6/soluble interleukin 6 receptor were individually transplanted into NOD/SCID mice. At 20 to 21 weeks after the transplantation, three out of 10 clones harvested at d 7 of culture, and three out of six clones at d 14 could reconstitute human haematopoiesis in recipient marrow. These results demonstrated that Flk2/Flt3 was expressed on a wide variety of human haematopoietic cells including long-term-repopulating haematopoietic stem cells.

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