Expression of the Human β -globin Gene Following Retroviral-Mediated Transfer into Multipotential Hematopoietic Progenitors of Mice

• Published in: Proceedings of the National Academy of Sciences - PNAS 1988-08, Vol.85 (16), p.6062-6066
• Main Authors: Karlsson, Stefan, Bodine, David M., Perry, Leland, Papayannopoulou, Thalia, Nienhuis, Arthur W.
• Format: Article
• Language: English
• Subjects: 3T3 cells; 550401 - Genetics- Tracer Techniques; 560152 - Radiation Effects on Animals- Animals; ANIMAL CELLS; ANIMALS; ANTIBODIES; BASIC BIOLOGICAL SCIENCES; BETA DECAY RADIOISOTOPES; BETA-MINUS DECAY RADIOISOTOPES; BIOLOGICAL MATERIALS; BLOOD; BLOOD CELLS; BODY FLUIDS; BONE MARROW CELLS; Cell lines; CFU; CONNECTIVE TISSUE CELLS; DAYS LIVING RADIOISOTOPES; ELECTROPHORESIS; ERYTHROCYTES; Erythrocytes - analysis; EXTERNAL IRRADIATION; Female; GENE RECOMBINATION; GENE REGULATION; GENES; Genetic Vectors; GLOBIN; Globins - analysis; Globins - genetics; Hematopoietic Stem Cells - metabolism; Humans; Infections; IRRADIATION; ISOTOPES; LETHAL IRRADIATION; LIGHT NUCLEI; MAMMALS; MAN; MATERIALS; MICE; Mice, Inbred C57BL; MICROORGANISMS; MONOCLONAL ANTIBODIES; NUCLEI; ODD-ODD NUCLEI; PARASITES; PHOSPHORUS 32; PHOSPHORUS ISOTOPES; PRIMATES; RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT; RADIOISOTOPES; Retroviridae - genetics; RNA; RODENTS; SOMATIC CELLS; Spleen; SPLEEN CELLS; Stem cells; TRANSCRIPTION; Transfection; TRANSPLANTS; VERTEBRATES; VIRUSES; WHOLE-BODY IRRADIATION
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.85.16.6062

Efficient transfer of the β -globin gene into primitive hematopoietic progenitors was achieved with consistent and significant expression in the progeny of those cells. Retroviral vectors containing the intact genomic human β -globin gene and the neomycin (G418)-resistance (neoR) gene were constructed. These gave titers of 106 or more neoR colony-forming units/ml when packaged in ψ 2 cells. Mouse bone marrow cells were infected by coculture with producer cells and injected into lethally irradiated animals. Several parameters were varied to enhance infection frequency of colony-forming units, spleen (CFU-S); overall 41% of 116 foci studied contained an intact proviral genome. The human β -globin gene was expressed in 31 of 35 CFU-S-derived spleen colonies that contained the intact vector genome at levels ranging from 1% to 5% of that of the mouse β -globin genes. Infected bone marrow cells were also injected into genetically anemic W/Wν recipients without prior irradiation. Human β -globin chains were detected in circulating erythrocytes by immunofluorescent staining with a specific monoclonal antibody. All animals injected with donor cells that had been cultured in G418 (1 mg/ml) for 48 hr after retroviral infection had circulating erythrocytes containing human β -globin chains between 3 and 8 weeks after transplantation.