Table 1.

Prolonged activation of the MAPK pathway inhibits DNA synthesis in cells expressing p21, but not in cells where p21 protein expression has been blocked

Infection/treatment	Wild-type hepatocyte	p21 null hepatocyte	Wild-type p21 antisense	Hep G2 hepatoma	Hep G2 hepatoma + p21 antisense
ΔB-Raf:ER + vehicle	10,150  ± 2,250	9,700  ± 1,600	9,950  ± 1,850	72,000  ± 5,100	94,700  ± 6,700
ΔB-Raf:ER + TAM	2,050  ± 300b	26,600  ± 2,100ca	25,250  ± 1,650ca	6,650  ± 850b	124,100  ± 8,500c
ΔB-Raf:ER + TAM + PD98059	7,300  ± 850bc	10,450  ± 1,550a	9,240  ± 1,100a	41,450  ± 3,050	52,800  ± 4,000
ΔB-Raf:ER + vehicle + PD98059	8,100  ± 950ba	8,200  ± 1,300ba	8,450  ± 1,200ba	45,250  ± 2,900	47,650  ± 4,200

Cells were infected with kinase active ΔB-Raf:ER poly-L-lysine adenovirus (at 250 m.o.i.), followed by culture as in Methods. In some experiments and in addition to ΔB-Raf:ER infection, cells were also infected with either null recombinant adenovirus or a p21 antisense recombinant adenovirus (at 100 m.o.i. each). After 24 h to allow protein expression, hepatocytes were treated with either vehicle control or with 100 nM 4-hydroxytamoxifen (TAM) for 36 h (total time in primary culture, 60 h). In some experiments, prior to 4-hydroxytamoxifen addition, cells were treated with 50 μM PD98059. Hepatocytes under all conditions were continually incubated with 4 μCi/ml ³H-thymidine throughout the final 36 h of culture, after which they were lysed and ³H-thymidine incorporation into DNA determined as in Methods. Addition of 4-hydroxytamoxifen did not alter the rate of DNA synthesis (data not shown). Data are the means (counts per minute ³H-thymidine incorporated into DNA ± SEM) of sextuplicate experiments from 4 separate animals. 

^a p < 0.05 increase compared to wild-type hepatocyte ΔB-Raf:ER + 4-hydroxytamoxifen cells. 

^b p < 0.05 decrease compared with ΔB-Raf:ER + vehicle control. 

^c p < 0.05 increase compared to ΔB-Raf:ER + vehicle control.