Human CD34+ hematopoietic cells transduced by retrovirus-mediated interferon alpha gene maintains regeneration capacity and engraftment in NOD/SCID mice.
To achieve long-term expression of human interferon alpha-5 (IFNalpha) gene in the bone marrow (BM) hematopoietic microenvironment, replication-deficient retroviral vector LSN-IFNalpha was used to deliver the IFNalpha gene into human BM CD34+ cells. After fibronectin-facilitated transduction, a fraction of CD34+ cells was plated in methylcellulose medium with or without G418 to assess transduction efficiency and the effect of IFNalpha gene transfer on colony formation. Colony-forming assay in the presence of G418 (400 microg/mL) revealed that 41% CFU-GM colonies are G418 resistant after infection with LSN-IFNalpha retrovirus. There was no significant difference in CFU-GM/BFU-E colony formation among IFNalpha gene-transduced CD34+ cells, control vector (LXSN) transduced-CD34+ cells and nontransduced CD34+ cells. Another portion of CD34+ cells was grown in liquid medium to measure IFNalpha production. RIA revealed that IFNalpha gene-transduced CD34+ cells produced 72.2 +/- 15.4 U/mL (10(6) cells/24 hours) of IFNalpha compared with 8.3 +/- 2.1 U/mL and 4.3 +/- 1.2 U/mL in LXSN-transduced or nontransduced CD34+ cells, respectively. The remaining portion of transduced CD34+ cells was transplanted into immunodeficient (NOD/SCID) mice to allow analysis of long-term expression of IFNalpha. Transplantation of 1x10(6) CD34+ cells into sublethally irradiated NOD/SCID mice showed that IFNalpha and neo(r) mRNA were detectable in engrafted mouse BM cells for up to 6 months. We conclude that continual local expression of IFNalpha in transduced CD34+ cells does not impair either CD34+ cell growth and differentiation or engraftment and long-term survival in NOD/SCID mice.
Quan
S
S
Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA.
Seiter
K
K
Feldman
E
E
Yang
L
L
Argani
I
I
Farley
T J
TJ
Abraham
N G
NG
Ahmed
T
T
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D002454:Cell Differentiation / Q000235:genetics / Q000276:immunology; D002470:Cell Survival; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D005260:Female; D015870:Gene Expression; D005822:Genetic Vectors; D006098:Granulocytes / Q000166:cytology; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D016898:Interferon-alpha / Q000235:genetics* / Q000276:immunology*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D011863:Radioimmunoassay; D012190:Retroviridae / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D018952:Antigens, CD34; D016898:Interferon-alpha