title: Cytokine transgene expression and promoter usage in primary CD34+ cells using particle-mediated gene delivery.
abstract: Induction or short-term transgenic expression of specific cytokines, growth factors, or other candidate therapeutic genes in hematopoietic progenitor or stem cells is potentially applicable to gene therapy for cancer. In this study, we explored the application of a gene gun technique, as an alternative to viral vectors, for ex vivo gene transfer into and transient gene expression in highly enriched CD34+ cells derived from human umbilical cord blood. Twenty-four hours posttransfection, 32.6 to 1500 pg/l x 10(6) CD34+ cells of transient gene expression was routinely obtained for specific cytokine and reporter genes. Transgene expression at the single-cell level was revealed by X-Gal staining of lacZ cDNA-transfected CD34+ cells. Expression of four candidate therapeutic genes, namely human granulocyte-macrophage colony-stimulating factor, tumor necrosis factor alpha, interleukin 2, and interferon gamma, was detectable for 4 to 7 days in CD34+ cells. A human elongation factor 1alpha promoter/intron 1 transcription unit was identified as a strong cellular promoter for CD34+ cells, exhibiting strength similar to that of the commonly employed cytomegalovirus immediate-early promoter. These results suggest that the nonviral, gene gun technique offers an efficient alternative approach for transient transgenic studies of hematopoietic cells and may provide new possibilities for certain cancer gene therapy strategies using CD34+ cells.
authors.affiliation: Department of Pediatrics, University of Wisconsin-Madison, 53792, USA.
authors.forename: Z Q
authors.initials: ZQ
authors.lastname: Ye
authors.forename: P
authors.initials: P
authors.lastname: Qiu
authors.forename: J K
authors.initials: JK
authors.lastname: Burkholder
authors.forename: J
authors.initials: J
authors.lastname: Turner
authors.forename: J
authors.initials: J
authors.lastname: Culp
authors.forename: T
authors.initials: T
authors.lastname: Roberts
authors.forename: N T
authors.initials: NT
authors.lastname: Shahidi
authors.forename: N S
authors.initials: NS
authors.lastname: Yang
mesh_terms: D018952:Antigens, CD34 / Q000032:analysis*; D019470:Biolistics; D002470:Cell Survival; D002478:Cells, Cultured; D016207:Cytokines / Q000096:biosynthesis / Q000235:genetics*; D005312:Fetal Blood / Q000166:cytology; D005434:Flow Cytometry; D015870:Gene Expression; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods; D006412:Hematopoietic Stem Cells / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D011401:Promoter Regions, Genetic; D013997:Time Factors; D014162:Transfection; D019076:Transgenes
chemical_list: D018952:Antigens, CD34; D016207:Cytokines; D008156:Luciferases