Lack of C/EBPα gene expression results in increased DNA synthesis and an increased frequency of immortalization of freshly isolated rat hepatocytes

Abstract

The CCAAT/enhancer binding protein α (C/EBPα) binds to specific promoter sequences and directs transcription of many genes expressed in the liver. Overexpression of C/EBPα in established cell lines inhibits cell proliferation. Primary hepatocytes from newborn C/EBPα(-/-) mice and normal littermates were used to determine whether the absence of C/EBPα increased proliferation and/or transformation of these cells in vitro. DNA synthesis, as measured by bromodeoxyuridine (BrdU) incorporation 24 hours postharvest, was fourfold higher in cells from C/EBPα(-/-) pups. Established cell lines were derived from 7 of 8 hepatocyte cultures initiated from null mutants, 4 of 23 cultures from heterozygotes, and 0 of 12 cultures from wild-type animals. C/EBPα(-/-) 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/EBPα(- /-) mice formed cords and increased albumin mRNA expression by 1.7- to 3.8- fold. C/EBPα(-/-) cell lines exhibited rapid growth and rapid accumulation of chromosomal abnormalities, and were capable of forming nodules when inoculated into the abdominal subcutaneous tissue of nude mice. Our data show that C/EBPα is an important regulator of hepatocyte proliferation and participates in the maintenance of the nontransformed hepatic phenotype in vitro.