LACK OF C EBP-ALPHA GENE-EXPRESSION RESULTS IN INCREASED DNA-SYNTHESIS AND AN INCREASED FREQUENCY OF IMMORTALIZATION OF FRESHLY ISOLATED RAT HEPATOCYTES/

The CCAAT/enhancer binding protein alpha (C/EBP alpha) binds to specif ic promoter sequences and directs transcription of many genes expresse d in the liver, Overexpression of C/EBP alpha in established cell line s inhibits cell proliferation. Primary hepatocytes from newborn C/EBP alpha(-/-) mice and normal littermates were used to determine whether the absence of C/EBP alpha increased proliferation and/or transformati on of these cells in vitro. DNA synthesis, as measured by bromodeoxyur idine (BrdU) incorporation 24 hours postharvest, was fourfold higher i n cells from C/EBP alpha(-/-) pups. Established cell lines were derive d from 7 of 8 hepatocyte cultures initiated from null mutants, 4 of 23 cultures from heterozygotes, and 0 of 12 cultures from wild-type anim als, C/EBP alpha(-/-) cultures had epithelial morphology, showed bile canaliculi, and expressed albumin messenger RNA (mRNA). When cultured on Matrigel, which promotes differentiation, cell lines derived from C /EBP alpha(-/-) mice formed cords and increased albumin mRNA expressio n by 1.7- to 3.8-fold. C/EBP alpha(-/-) cell lines exhibited rapid gro wth and rapid accumulation of chromosomal abnormalities, and were capa ble of forming nodules when inoculated into the abdominal subcutaneous tissue of nude mice. Our data show that C/EBP alpha is an important r egulator of hepatocyte proliferation and participates in the maintenan ce of the nontransformed hepatic phenotype in vitro.