title: Support of human cord blood progenitor cells on human stromal cell lines transformed by SV40 large T antigen under the influence of an inducible (metallothionein) promoter.
abstract: We describe the development of a human bone marrow (BM) culture system which allows study of the interaction of stromal cell lines (SCL) and highly purified hematopoietic progenitor cells. Normal BM stromal cells were electroporated with a plasmid containing the simian virus 40 (SV40) large T antigen (SV40 T Ag) under the control of a synthetic metallothionein promoter (MT4); this construct is designated MT4 SV40 T Ag. SCL in which the rate of proliferation could be controlled by altering the zinc (Zn) concentration were characterized, demonstrating that the SCL were heterogeneous with respect to G-CSF and GM-CSF production. Suppression of SCL proliferation on removal of Zn made it possible to use these lines in coculture with purified CD34+ progenitor cells from umbilical cord blood. The ability to control proliferation of SCL has allowed us to maintain the survival and expansion of colony-forming cells in culture for up to 2 months. These lines have enabled us to test for stromal cell characteristics at a clonal level and provided us with a tool to analyze the events leading to lineage commitment and hematopoietic differentiation, as demonstrated by suppression of hematopoiesis by an antibody directed against the c-kit molecule.
authors.affiliation: Lions Clinical Cancer Research Laboratory, Walter and Eliza Hall Institute of Medical Research, PO Royal Melbourne Hospital, Victoria, Australia.
authors.forename: F M
authors.initials: FM
authors.lastname: Cicuttini
authors.forename: M
authors.initials: M
authors.lastname: Martin
authors.forename: E
authors.initials: E
authors.lastname: Salvaris
authors.forename: L
authors.initials: L
authors.lastname: Ashman
authors.forename: C G
authors.initials: CG
authors.lastname: Begley
authors.forename: J
authors.initials: J
authors.lastname: Novotny
authors.forename: D
authors.initials: D
authors.lastname: Maher
authors.forename: A W
authors.initials: AW
authors.lastname: Boyd
mesh_terms: D015703:Antigens, CD / Q000032:analysis; D018952:Antigens, CD34; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics; D000954:Antigens, Surface / Q000032:analysis; D001853:Bone Marrow / Q000276:immunology; D001854:Bone Marrow Cells; D002455:Cell Division / Q000187:drug effects; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000187:drug effects; D016179:Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000096:biosynthesis; D006410:Hematopoiesis; D016298:Hematopoietic Cell Growth Factors / Q000096:biosynthesis; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology; D006801:Humans; D066298:In Vitro Techniques; D015850:Interleukin-6 / Q000096:biosynthesis; D008668:Metallothionein / Q000235:genetics; D010982:Platelet-Derived Growth Factor / Q000235:genetics; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D019089:Stem Cell Factor; D015032:Zinc / Q000494:pharmacology
chemical_list: D015703:Antigens, CD; D018952:Antigens, CD34; D000952:Antigens, Polyomavirus Transforming; D000954:Antigens, Surface; D016298:Hematopoietic Cell Growth Factors; D015850:Interleukin-6; D010982:Platelet-Derived Growth Factor; D012333:RNA, Messenger; D019089:Stem Cell Factor; D016179:Granulocyte Colony-Stimulating Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D008668:Metallothionein; D015032:Zinc