title: Retroviral-mediated gene transfer of a mutant H-ras gene into normal human bone marrow alters myeloid cell proliferation and differentiation.
abstract: To investigate the effects of mutant ras expression on the growth and differentiation of normal human bone marrow, we used retrovirus-mediated gene transfer. A retrovirus (HR-1) containing a mutant ras gene (H12-ras) in addition to the selectable neo gene was transferred by cocultivation of a packaging cell line with long-term cultures of normal human bone marrow. Controls were established by cocultivating aliquots of the same bone marrow with a retrovirus (VSN-2) containing only neo. The efficiency of gene transfer, as determined by the percentage of G418-resistant colony-forming units-granulocyte/macrophage (CFU-GM) immediately after termination of cocultivation, was similar: 8 +/- 4% with HR-1 and 5 +/- 3% with VSN-2. After a further week in long-term culture, there was an increase in the number and percentage of G418-resistant CFU-GM in both the HR-1-infected and VSN-2-infected marrows. Thereafter, the numbers of G418-resistant CFU-GM declined, becoming undetectable at 4 weeks. The time course of the production of G418-resistant colonies was not significantly different in HR-1- and VSN-2-infected marrows, indicating that H12-ras did not alter the proliferation of normal CFU-GM. However, the total cellularity of HR-1-infected marrow cultures was significantly greater than that of VSN-2-infected marrow cultures. This was due to increased cellular proliferation of HR-1-infected cultured cells, since differential counts showed a significant increase in myeloid blast cells together with a slight reduction in mature myeloid cells in HR-1-infected marrow compared to baseline and to VSN-2-infected marrow. No leukemic blast cell colonies were grown from HR-1-infected marrows or control marrows, and HR-1 infection did not confer serum independence. These data show successful retroviral infection of normal bone marrow progenitor cells and suggest that expression of mutant H12-ras in such cells results in enhanced proliferation of early myeloid cells at the expense of differentiation.
authors.affiliation: Department of Haematology, Royal Postgraduate Medical School, Hammersmith Hospital, London, England.
authors.forename: J
authors.initials: J
authors.lastname: Maher
authors.forename: F
authors.initials: F
authors.lastname: Colonna
authors.forename: D
authors.initials: D
authors.lastname: Baker
authors.forename: L
authors.initials: L
authors.lastname: Luzzatto
authors.forename: I
authors.initials: I
authors.lastname: Roberts
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D001853:Bone Marrow / Q000502:physiology; D001854:Bone Marrow Cells; D002454:Cell Differentiation / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D018014:Gene Transfer Techniques; D011905:Genes, ras / Q000235:genetics* / Q000502:physiology*; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000502:physiology; D006801:Humans; D051379:Mice; D009154:Mutation / Q000235:genetics*; D015689:Oncogene Protein p21(ras) / Q000235:genetics / Q000502:physiology; D010641:Phenotype; D012190:Retroviridae / Q000235:genetics*; D013997:Time Factors
chemical_list: D015689:Oncogene Protein p21(ras)