Phenobarbital at low dose exerts hormesis in rat hepatocarcinogenesis by reducing oxidative DNA damage, altering cell proliferation, apoptosis and gene expression

Abstract

Our recent research indicated that phenobarbital (PB) may inhibit the development of N-diethylnitrosamine (DEN)-initiated pre-neoplastic lesions at low doses in a rat liver medium-term bioassay (Ito test), while high doses exhibit promoting activity. This raises the question of whether treatment with low doses of PB might reduce cancer risk. For clarification, male 6-week-old F344 rats were treated with PB at doses of 0, 2, 15 and 500 p.p.m. in the diet for 10 or 33 weeks after initiation of hepatocarcinogenesis with DEN. In a second, short-term experiment, animals were given PB at doses of 2, 4, 15, 60 and 500 p.p.m. for 8 days. Formation of glutathione S-transferase placental form (GST-P) positive foci and liver tumors was inhibited at 2 p.p.m. Generation of oxidative DNA damage marker, 8-hydroxy-2'-deoxyguanosine (8-OHdG), cellular proliferation within the areas of GST-P positive foci and apoptosis in background liver parenchyma were suppressed. Suppression of 8-OHdG formation by PB at low dose might be related to the enhanced mRNA expression of 8-OHdG repair enzyme, oxoguanine glycosylase 1 (Ogg1). Moreover, as detected by cDNA microarray analysis, PB treatment at low dose enhanced mRNA expression of glutamic acid decarboxylase (GAD65), an enzyme involved in the synthesis of gamma-aminobutyric acid (GABA), and suppressed MAP kinase p38 and other intracellular kinases gene expression. On the contrary, when PB was applied at a high dose, GST-P positive foci numbers and areas, tumor multiplicity, hydroxyl radicals and 8-OHdG levels were greatly elevated with the increase in CYP2B1/2 and CYP3A2 mRNA, protein, activity and gene expression of GST, nuclear tyrosine phosphatase, NADPH- cytochrome P-450 reductase and guanine nucleotide binding protein G(O) alpha subunit. These results indicate that PB exhibits hormetic effect on rat hepatocarcinogenesis initiated with DEN by differentially altering cell proliferation, apoptosis and oxidative DNA damage at high and low doses.

Low dose effects of dichlorodiphenyltrichloroethane (DDT) on gene transcription and DNA methylation in the hypothalamus of young male rats: implication of hormesis-like effects

“…This reducalthough the detailed mechanism underlying this response is obscure. Kinoshita et al (2003) reported that low-dose treatment with phenobarbital, which is a nongenotoxic carcinogen like DDT (Harada et al, 2003), induced a and enzyme induction.…”

Liver tumor promoting effect of orphenadrine in rats and its possible mechanism of action including CAR activation and oxidative stress

“…It has been reported that treatment with PB increased mRNA expression and protein levels of CYP2B in a twostage liver carcinogenesis bioassay in rats (Kinoshita et al, 2003;Waxman and Azaroff, 1992). The nuclear translocation of CAR induced by PB stimulates the expression of various genes that enhance hepatic tumor promotion (Deguchi et al, 2009;Kawamoto et al, 1999).…”

Enhanced liver tumor promotion activity in rats subjected to combined administration of phenobarbital and orphenadrine

“…The CYP family generates ROS as byproducts of microsomal oxidation, and PB, a CYP2B1/2 inducer, has been shown to increase hydroxyl radical levels in the livers of rats (Kinoshita et al, 2003;Waxman and Azaroff, 1992). Imaoka et al (2004) reported that CYP2B1 induction by PB induces ROS production and genomic DNA oxidation that may contribute to tumor promotion by PB.…”

“…PB induces a large spectrum of drug metabolizing enzymes, including cytochrome P450 family 2 subfamily B (CYP2B) in the liver of rats (Kinoshita et al, 2003;Waxman and Azaroff, 1992). In addition, it has been shown that microsomal reactive oxygen species (ROS) production and production of thiobarbituric acidreactive substances (TBARS) induced with increasing Cyp2b induction due to PB treatment are involved in the liver tumor-promoting effect in rats (Morita et al, 2011).…”