IN-VITRO GROWTH-CHARACTERISTICS OF EMBRYO FIBROBLASTS ISOLATED FROM P53-DEFICIENT MICE

Fibroblast cultures were derived from mouse embryos containing either one (p53+/-) or two (p53-/-) inactivated p53 alleles and compared to n ormal embryo fibroblasts for a number of growth parameters. Early pass age p53-deficient embryo fibroblasts (p53-/-) divided faster than norm al embryo fibroblasts, achieved higher confluent densities, and had a higher fraction of division-competent cells under conditions of low ce ll density. Flow cytometry studies of early passage embryo fibroblasts showed that the percent of p53-deficient cells in G0/G1 was lower tha n in normal cells, consistent with the argument that p53 mediates a GI block. When p53-deficient and normal cells were passaged for long per iods of time, the homozygote (p53-/-) fibroblasts grew at a high rate for over 50 passages and never entered a non-growing senescent phase c haracteristic of the heterozygote (p53+/-) and normal (p53+/+) cells. The p53-deficient fibroblasts were genetically unstable during passagi ng, with the p53-/- cells showing a high degree of aneuploidy and the p53+/- cells displaying a moderate level of chromosomal abnormalities by passage 25. Surprisingly, the heterozygote cells lost their single wild type allele very early during culturing and in spite of this loss most heterozygote lines entered into senescence. We conclude that the loss of p53 by itself is insufficient to confer immortality on a cell , but does confer a growth advantage. Taken together, the findings con firm that the absence of p53 promotes genomic instability, which in tu rn may result in genetic alterations which directly produce immortalit y.