Resistance of primary cultured mouse hepatic tumor cells to cellular senescence despite expression of p16(Ink4a), p19(Arf), p53, and p21(Waf1/Cip1)

Primary cultured mouse hepatic cells become senescent within a short period , although rare cells form colonies from which continuously proliferating c ell lines can be established. In contrast, hepatic tumor (HT) cells show li ttle senescence and higher colony-forming capacity. To assess this differen ce, we investigated p16(Ink4a)/p19(Arf)/p53/p21(Waf1/Cip1) expression in pr imary normal and FIT cells, together with cell lines established from both. In primary normal cells, p16(Ink4a)/p19(Arf) were expressed only in associ ation with senescence and disappeared at later stages of colony formation. In contrast, primary HT cells showed sustained p16(Ink4a)/p19(Arf) expressi on from the beginning. No p16(Ink4a)/ p19(Arf) alterations, such as deletio n, mutations, or hypermethylation, were detected in the primary HT cells, a lthough most cell lines derived from either normal or HIT cell colonies los t p16(Ink4a), or p19(Arf) expression owing to hypermethylation or homozygou s deletion of p16(Ink4a)/p19(Arf), On the other hand, primary normal and HT cells and most cell lines showed constitutively elevated expression of p53 /p21(Waf1/Cip1), with a further increment after ultraviolet ir radiation, i ndicating a functionally normal p53 pathway. These results indicate that pr imary HT cells are resistant to senescence despite retaining p16(Ink4a)/p19 (Arf)/p53/p21(Waf1/ciP1) expression and that loss of p16(Ink4a)/p19(Arf) fu nction is associated only with establishment of the cell lines. (C) 2001 Wi ley-Liss, Inc.