INDUCTION OF REPLICATIVE DNA-SYNTHESIS AND PPAR-ALPHA-DEPENDENT GENE-TRANSCRIPTION BY WY-14-643 IN PRIMARY RAT HEPATOCYTE AND NONPARENCHYMAL CELL COCULTURES

Peroxisome proliferators (PP) are known hepatocarcinogens in rats and mice. We have investigated the ability of Wyeth-14 643 (Wy), a PP and potent rodent carcinogen, to induce replicative DNA synthesis and to m odulate the levels of peroxisome proliferator activated receptor-alpha (PPAR alpha) transcriptionally-dependent genes in primary rat hepatoc yte (HPC) cultures and hepatocyte/non-parenchymal cell (HPC/NPC) co-cu ltures maintained on Matrigel, Four days after plating, cells were tre ated with Wy and replicative DNA synthesis was quantitated using [H-3] thymidine incorporation and specific mRNA transcript levels were deter mined by reverse-transcriptase polymerase chain reaction (RT-PCR). An increase in HPC replicative DNA synthesis was detected at 48 h in both Wy-treated HPC and HPC/NPC co-cultures relative to controls, This inc rease was approximately 3- and 6-fold in HPC and HPC/NPC cultures resp ectively, and was Wy concentration-dependent, The levels of PPAR alpha -transcriptionally dependent genes [cytochrome P4504A1, acyl-CoA oxida se (AOxase), and liver-fatty acid binding protein (L-FABP)] transcript s were determined as indicators of PPAR alpha activation, These transc ripts increased dose-dependently at 48 h in HPC/NPC cultures up to 10 mu M Wy, Similarly, RT-PCR product levels were also increased in HPC c ultures with 10 mu M Wy at 48 h, In conclusion, we have investigated t he transcription of PPAR alpha-dependent genes and HPC replicative DNA synthesis by Wy in HPC/NPC co-cultures. Results of this work are clea rly more reflective of the known in vivo effects of PP and suggest tha t HPC/NPC cocultures are more appropriate than HPC cultures for such s tudies, The effect of PP on human HPC/NPC co-cultures is currently bei ng investigated in our laboratory in an attempt to assess human risks to these chemicals more directly.