Altered expression of hepatic CAR-dependent genes leads to Increase, cell proliferation (hepatocytes)

The altered expression of mouse and rat genes that are related to increased cell proliferation will, by definition, produce measurable changes in cell proliferation in hepatocytes (Elcombe et al., 2014; Yang and Wang, 2014). Hepatocytes have the ability to regenerate when properly stimulated, such as following partial hepatectomy. The CAR-responsive genes such as Gadd45b, Ki67 and the Cdc20 are necessary gene targets that are part of this synchronized response that results in progress out of the quiescent cell cycle stage (G0), resulting in DNA replication during S-phase (a measurable marker of cell proliferation), and eventual cell division.

The Moderate quantitative understanding supporting the linkage between CAR-responsive gene expression changes and measurable changes in cell proliferation (increased BrdU labeling or Ki67 labelling index) has been established in a limited number of studies with ample dose-response data (Deguchi et al., 2009; Geter et al., 2014; Jones et al., 2009). Typically, these dose-response studies show that effects on gene expression (e.g. increased Cyp2b10 mRNA) and associated events related to liver metabolism (e.g. increased PROD activity) can occur at lower, non-tumorigenic dose levels, and changes in markers of cell proliferation including cell cycle pathway changes and BrdU or Ki67 labelling index are seen at somewhat higher dose levels. For example the study of Geter et al. (2014) with phenobarbital showed that in male CD-1 mice, the most sensitive parameters measured was PROD activity (BMD = 0.2 – 2.4 mg/kg/day), cell proliferation by BrdU labeling index occurred at higher dose levels (BMD = 13 – 14 mg/kg/day) and in general for this particular study, significant changes in gene expression pathways were only seen at dose levels slightly higher than these apical markers.

The pathway leading from CAR-mediated gene expression changes to an increase in cell proliferation (BrdU, PCNA or Ki67 labeling index) has strong evidence indicating it is selectively observed in mice and rats, but not in other mammalian species (Deguchi et al., 2009; Foster, 2000; LeBaron et al., 2013; Parzefall et al., 1991; Plant et al., 1998). Recently a chimeric mouse model that had >90% replacement of mouse hepatocytes with human hepatocytes was used to examine the comparative in vivo effects of 3-5 dietary concentrations of phenobarbital in CD-1 mouse hepatocytes vs. the chimeric human hepatocytes (Yamada et al., 2014). In the liver cells of CD-1 mice after 1 week of treatment, increased mRNA levels of Cyp2b10, Cyp3a11, Ki67 and Gadd45b were observed, along with increased BrdU labeling index. In the human liver cells of chimeric mice, increased mRNA levels of CYP2B and CYP3A were observed, but there were no differences from control for Ki67 and GADD45B mRNA levels nor any increases in BrdU labeling index (Yamada et al., 2014). In summary, a large amount of experimental data including both in vitro and in vivo studies has demonstrated that the downstream key event of increased cell proliferation following treatment with CAR activators occurs selectively in mice and rats, and is correlated with CAR-mediated gene expression changes that reflect pathways of cell cycle control and cell proliferation.