FIG. 2.

Ability of keratinocytes and fibroblasts to be immortalized by hTERT expression early versus late in their life spans. The primary oral keratinocyte line OKF4 and the primary dermal fibroblast line R2F were transduced with the pBABE(hTERT)puro vector at either an early (a and c) or late (b and d) stage of their respective life spans. Downward arrows indicate the time of transduction. The OKF4 parent line and hTERT transductants were cultured in the fibroblast feeder cell system until drug selection was complete, after which half the cells were subsequently cultured in the feeder system and half in K-sfm medium. ∗ indicates the PD level at which cell populations were completely senescent. Note that R2F fibroblasts were immortalized by hTERT expression at any time during their life span, whereas OKF4 keratinocytes were not immortalized if hTERT was expressed late in the life span in either culture system. OKF4 cells yielded immortalized lines when hTERT was expressed earlier in the life span, but a long period of initially slow population doubling rates preceded the appearance of more rapidly dividing immortalized cells for OKF4/TERT (early) cells cultured in K-sfm, and a modest inflection of the growth curve and progressive selection of more rapidly dividing cells were also evident for OKF4/TERT (early) cells cultured with feeder cells. Small arrowheads at the ends of some lines in the graphs indicate immortalization.