Hepatic transcriptomic alterations for N,N-dimethyl-p-toluidine (DMPT) and p-toluidine after 5-day exposure in rats

Abstract

N,N-dimethyl-p-toluidine (DMPT), an accelerant for methyl methacrylate monomers in medical devices, was a liver carcinogen in male and female F344/N rats and B6C3F1 mice in a 2-year oral exposure study. p-Toluidine, a structurally related chemical, was a liver carcinogen in mice but not in rats in an 18-month feed exposure study. In this current study, liver transcriptomic data were used to characterize mechanisms in DMPT and p-toluidine liver toxicity and for conducting benchmark dose (BMD) analysis. Male F344/N rats were exposed orally to DMPT or p-toluidine (0, 1, 6, 20, 60 or 120 mg/kg/day) for 5 days. The liver was examined for lesions and transcriptomic alterations. Both chemicals caused mild hepatic toxicity at 60 and 120 mg/kg and dose-related transcriptomic alterations in the liver. There were 511 liver transcripts differentially expressed for DMPT and 354 for p-toluidine at 120 mg/kg/day (false discovery rate threshold of 5 %). The liver transcriptomic alterations were characteristic of an anti-oxidative damage response (activation of the Nrf2 pathway) and hepatic toxicity. The top cellular processes in gene ontology (GO) categories altered in livers exposed to DMPT or p-toluidine were used for BMD calculations. The lower confidence bound benchmark doses for these chemicals were 2 mg/kg/day for DMPT and 7 mg/kg/day for p-toluidine. These studies show the promise of using 5-day target organ transcriptomic data to identify chemical-induced molecular changes that can serve as markers for preliminary toxicity risk assessment.

Keywords: Liver toxicity; Molecular markers; N,N-dimethyl-p-toluidine; p-Toluidine.

Figure 1

DMPT and p-toluidine structures

Figure 2

Postulated intermediate metabolites of DMPT and p-toluidine

Figure 3

Control and DMPT liver lesions A. Centrilobular region of liver from a young, male F344/NTac rat exposed to vehicle control corn oil by gavage for 5 days. There is one mitotic figure (arrow) and no evidence of individual cell death. CV = central vein. H&E. B. Centrilobular region of liver from a young, male F344/NTac rat exposed to 120 mg/kg DMPT by gavage for 5 days. There are numerous shrunken, eosinophilic bodies, several of which appear to be in clear vacuoles in the cytoplasm of hepatocytes (arrows). A larger hepatocyte with brightly eosinophilic cytoplasm and karyorhectic nuclear debris is also present (arrowhead). CV = central vein. H&E. Same magnification as Fig. 3a. C. Centrilobular region of liver from a young, male F344/NTac rat exposed to 60 mg/kg DMPT by gavage for 5 days. In the centrilobular region, there are numerous mitotic figures present (arrows). CV = central vein. H&E. Same magnification as Fig. 3a

Figure 4

Heat map showing the unsupervised hierarchical clustering analysis illustrating prominent gene expression patterns at the high dose (120 mg/kg) in (A) significant DMPT liver transcripts and (B) significant p-Toluidine liver transcripts determined at a false discovery rate threshold of 0.05.

Figure 5

Activation of Nrf2 pathway by DMPT and p-toluidine