Retrovirus-mediated transfer of the herpes simplex virus thymidine kinase and enhanced green fluorescence protein genes in primary T lymphocytes.
The EGFP-tk retroviral vector, encoding enhanced green fluorescent protein (EGFP) and the herpes simplex virus thymidine kinase (HSV-tk) packaged in a Phoenix amphotropic cell line, was used to transduce healthy donor T lymphocytes. Infection yielded a mean of 41.8 +/- 9.3% SD (range 31.1-48.4%) EGFP-positive cells and a mean of 92 +/- 2% SD (range 90-94%) after cell sorting. EGFP expression remained stable for 30 d after infection. The entire gene transfer procedure had no significant effect on lymphocyte subsets and slightly reduced clonogenicity. Ganciclovir (gcv) treatment (1 microg/ml x 10 d) killed all EGFP-positive cells in the transduced and transduced/sorted populations, but had no effect on untransduced controls. Our results show that primary T lymphocytes can be transduced using an EGFP-tk vector that yields a homogeneous infected population without affecting lymphocyte subsets, function and clonogenicity.
Di Florio
S
S
Department of Clinical and Experimental Medicine, Perugia University, Perugia, Italy.
Sebastiani
C
C
Fagioli
M
M
Di Ianni
M
M
Alfonsi
D
D
Venditti
G
G
Pelicci
P G
PG
Tabilio
A
A
D000998:Antiviral Agents / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D005434:Flow Cytometry; D015774:Ganciclovir / Q000494:pharmacology; D005822:Genetic Vectors / Q000494:pharmacology*; D049452:Green Fluorescent Proteins; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology; D008164:Luminescent Proteins / Q000235:genetics; D018139:Simplexvirus / Q000201:enzymology*; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D013937:Thymidine Kinase / Q000235:genetics*; D013997:Time Factors; D014162:Transfection / Q000379:methods*
D000998:Antiviral Agents; D007376:Interleukin-2; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D013937:Thymidine Kinase; D015774:Ganciclovir