title: Ex vivo expansion of human umbilical cord hematopoietic progenitor cells using a coculture system with human telomerase catalytic subunit (hTERT)-transfected human stromal cells.
abstract: We developed a new human stromal cell line that could expand human hematopoietic progenitor/stem cells. Primary human bone marrow stromal cells were infected with retrovirus containing the human telomerase catalytic subunit (hTERT) gene, resulting in increased population doubling and the acquisition of cell immortalization. Characteristics of the hTERT-transduced stromal (hTERT-stromal) cells were identical with those of the primary stromal cells in terms of morphologic appearance and expression of surface antigens. Human cord blood (CB) CD34(+) cells were expanded by coculture with primary stromal or hTERT-stromal cells in the presence of stem cell factor, thrombopoietin, and Flk-2/Flt-3 ligand under serum-free condition. The degree of expansion of CD34(+) cells and total number of colony-forming units in culture (CFU-Cs) after 2 weeks' coculture with the hTERT-stromal cells were nearly the same as those after 2 weeks' coculture with primary stromal cells (CD34(+) cells, 118-fold +/- 8-fold versus 117-fold +/- 13-fold; CFU-Cs, 71-fold +/- 5-fold versus 67-fold +/- 5-fold of initial cell number). CB expansion on hTERT-stromal cells occurred at a similar rate through 7 weeks. In contrast, the rate of CB expansion on primary stromal cells had drastically declined at 7 weeks. In nonobese diabetic/severe combined immunodeficiency (SCID) mice, the degree of engraftment of SCID-repopulating cells that had been cocultured with hTERT-stromal cells for 4 weeks was significantly higher than that of precocultured CB cells. These results indicate that this hTERT-stromal cell line could be useful for ex vivo expansion of hematopoietic progenitor/stem cells and for analyzing the microenvironment of human bone marrow.
authors.affiliation: Department of Molecular Medicine, Sapporo Medical University, Japan.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Kawano
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Kobune
authors.forename: Miki
authors.initials: M
authors.lastname: Yamaguchi
authors.forename: Kiminori
authors.initials: K
authors.lastname: Nakamura
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Ito
authors.forename: Katsunori
authors.initials: K
authors.lastname: Sasaki
authors.forename: Sho
authors.initials: S
authors.lastname: Takahashi
authors.forename: Takafumi
authors.initials: T
authors.lastname: Nakamura
authors.forename: Hiroki
authors.initials: H
authors.lastname: Chiba
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Sato
authors.forename: Takuya
authors.initials: T
authors.lastname: Matsunaga
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Azuma
authors.forename: Kenji
authors.initials: K
authors.lastname: Ikebuchi
authors.forename: Hisami
authors.initials: H
authors.lastname: Ikeda
authors.forename: Junji
authors.initials: J
authors.lastname: Kato
authors.forename: Yoshiro
authors.initials: Y
authors.lastname: Niitsu
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Hamada
mesh_terms: D000818:Animals; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D004268:DNA-Binding Proteins; D005312:Fetal Blood / Q000166:cytology; D006410:Hematopoiesis / Q000187:drug effects; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism*; D019098:Telomerase / Q000235:genetics* / Q000494:pharmacology; D014162:Transfection; D014183:Transplantation, Heterologous; D018631:ras Proteins / Q000235:genetics / Q000494:pharmacology
chemical_list: D000957:Antigens, Viral, Tumor; D004268:DNA-Binding Proteins; D019098:Telomerase; D018631:ras Proteins