NR ALUQ
AU Tumas,D.B.; Spangrude,G.J.; Brooks,D.M.; Williams,C.D.; Chesebro,B.
TI High-frequency cell surface expression of a foreign protein in murine hematopoietic stem cells using a new retroviral vector
QU Blood 1996 Jan 15; 87(2): 509-17
PT journal article
AB A retroviral vector (pSFF) derived from murine Friend spleen focus forming virus was used to transduce murine hematopoietic stem cells and express a cell surface marker protein, mutated murine prion protein, in vitro and in vivo after transplantation. To enhance retroviral vector integration in bone marrow cells, mice were treated with 5-fluorouracil (5-FU) to increase stem cell mitotic activity, which peaked on day 8 post-5-FU. The infectivity titer of the vector, pSFF-mPrP-3F4, was determined by a novel assay in which antigen-positive foci of infected cells were detected after replication and spread of the vector in cultures of mixed packaging cell lines. Infection of Sca-1+/Lineageneg-low cells with pSFF-mPrP-3F4 resulted in marker protein expression in 40% of the progeny cells after 7 days of culture. Transplantation of marrow cells or sorted Sca-1+/Lineageneg-low cells transduced with vector resulted in 3F4-positive mPrP expression in 11% to 37% of donor-derived peripheral blood leukocytes at 2 weeks. Though the percentage of 3F4-positive blood cells gradually declined, at 28 weeks 23% of recipient mice still maintained expression of the marker gene. Expression was observed in lymphoid, myeloid, and erythroid lineages and was detected in Sca-1+/Lineageneg-low marrow cells. The multilineage, high-frequency expression observed suggests that pSFF may be useful in gene therapy directed at hematopoietic stem cells and their differentiated progeny.
MH Animal; Bone Marrow/drug effects; Cell Lineage; Defective Viruses/*genetics; Fluorouracil/pharmacology; Friend murine leukemia virus/*genetics; Gene Expression Regulation, Viral; Genes, Reporter; *Genetic Vectors; Helper Viruses/*genetics; *Hematopoietic Stem Cell Transplantation; Hematopoietic Stem Cells/*metabolism/virology; Immunophenotyping; Mice; Mice, Inbred C57BL; PrPc Proteins/biosynthesis/*genetics; Recombinant Fusion Proteins/*biosynthesis; Repetitive Sequences, Nucleic Acid; Spleen Focus-Forming Viruses/*genetics; Transfection
AD Laboratories of Persistent Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, MT 59840, USA
SP englisch
PO USA