IMMORTALIZATION OF A HUMAN-DIPLOID FIBROBLAST CELL STRAIN - A DT-DIAPHORASE PARADOX

Transfection of a normal human diploid fibroblast cell strain, GM38, w ith a simian virus 40 (SV40) large T antigen containing plasmid, yield ed an immortal cell line, G38-8X, which had a similar sensitivity as t he parental cell strain to the quinone-containing chemotherapeutic age nt mitomycin C (MMC), under both aerobic and hypoxic exposure conditio ns. The activity level of DT-diaphorase was similar in both the parent al GM38 and G38-8X cells. Although DT-diaphorase could be detected by Western blot analysis, using two mouse anti-human monoclonal antibodie s, in GM38 cells, it was not detected in the G38-8X cells. C38-8X cell s have a slightly increased P450R activity (2-fold), and have elevated P-glycoprotein levels compared with the parental GM38 cell strain. Th e immortal G38-8X cell line is 2-fold more resistant to ionising radia tion than the parental GM38 cell strain (D-10 similar to 5 Gy). Althou gh these SV40 large T antigen immortalised human diploid fibroblasts b ehaved similarly to their parental cell strain in terms of MMC sensiti vity and DT-diaphorase activity, careful characterisation revealed tha t these cells had enhanced P-glycoprotein activity and had a decreased sensitivity to ionising radiation.