Retroviral mediated expression of CD18 in normal and deficient human bone marrow progenitor cells.
The CD18 gene encodes the beta subunit of leukocyte integrins, and autosomal recessive deficiency of CD18 in humans causes a life-threatening abnormality of granulocyte migration. A high titer amphotropic retrovirus encoding CD18 was used to infect bone marrow cells from normal and CD18-deficient human donors. Infected cells were maintained in a long-term culture system and analyzed (1) using PCR to detect the provirus in granulocyte or macrophage colonies (CFU-GMs) derived from the culture, (2) using reverse transcription-PCR (RT-PCR) to detect transcripts in the floating cells in the culture, and (3) using immunostaining of the floating cells to analyze expression of human CD18. By both cocultivation and supernatant infection, a significant fraction (10-82%) of the CFU-GMs were positive for the provirus after five weeks in long-term culture, and as high as 10-15% of the floating cells were positive by immunostaining after nine weeks in the long-term culture. In all cases, cocultivation showed higher infection efficiency than supernatant infection. This is the first report of the introduction of human CD18 cDNA into the bone marrow progenitor cells of patients with leukocyte adhesion deficiency.
Yorifuji
T
T
Institute for Molecular Genetics, Baylor College of Medicine, Houston, TX 77030.
Wilson
R W
RW
Beaudet
A L
AL
D016475:3T3 Cells; D000818:Animals; D015703:Antigens, CD / Q000235:genetics*; D001483:Base Sequence; D018821:CD18 Antigens; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors; D006412:Hematopoietic Stem Cells / Q000276:immunology* / Q000382:microbiology; D006801:Humans; D007153:Immunologic Deficiency Syndromes / Q000235:genetics / Q000276:immunology / Q000628:therapy; D018370:Leukocyte-Adhesion Deficiency Syndrome; D051379:Mice; D008969:Molecular Sequence Data; D011533:Proviruses / Q000235:genetics; D012326:RNA Splicing; D016028:Receptors, Leukocyte-Adhesion / Q000235:genetics*; D012190:Retroviridae / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection
D015703:Antigens, CD; D018821:CD18 Antigens; D017931:DNA Primers; D016028:Receptors, Leukocyte-Adhesion