Dinitrotoluene isomer-specific enhancement of the expression of diethylnitrosamine-initiated hepatocyte foci.

Abstract

Technical grade dinitrotoluene (TDNT) was shown to be a potent hepatocarcinogen in rats; however, ambiguous results were obtained from bioassays which evaluated 2,4-DNT, the principal isomer in the technical mixture. The present studies were designed to determine the relative hepatocyte foci promoting activity of TDNT and the primary isomers present in the technical mixture, 2,4- and 2,6-DNT. A rat hepatic initiation-promotion protocol was used in which male Fischer-344 rats were initiated with a single dose of diethylnitrosamine (DEN) (150 mg/kg, i.p.) and permitted to recover for 2 weeks. Following the recovery period, the animals were fed diets containing TDNT,2,4-DNT,2,6-DNT or phenobarbital (PB). The TDNT animals were killed after 3 or 6 weeks of feeding and the 2,4-DNT-,2,6-DNT-and PB-treated animals were killed after 6 or 12 weeks of feeding. Appropriate DEN and DNT controls were also killed at each time point. Sections from three liver lobes of each animal were stained for gamma-glutamyl transferase (GGT) and the number of GGT+ foci per cm3 (Nv) was calculated using stereological methods. TDNT feeding produced a dose-dependent increase in GGT+ foci (Nv) at 3 and 6 weeks relative to both DEN and TDNT controls. Administration of 2,4-DNT and PB produced time-dependent increases in GGT+ foci (Nv), while 2,6-DNT produced dose- and time-dependent increases. Unlike 2,4-DNT and PB, the high dose (14 mg/kg) of 2,6-dNT produced an increase in GGT+ foci in the control groups and in mean foci volume relative to the DEN control. These results establish that TDNT, 2,4-DNT and 2,6-DNT have hepatocyte foci promoting activity and that 2,6-DNT is approximately 10 times more potent than 2,4-DNT. In addition, TDNT administration neither alters hepatocyte replication following partial hepatectomy nor acts as a 'growth selection' agent in the Solt-Farber 'growth selection' model suggesting that differential effects on hepatocyte replication are not responsible for the promoting activity of TDNT. Based on previous initiation study results from our laboratory and the present data, 2,6-DNT appears to be a complete hepatocarcinogen while under the conditions of these studies 2,4-DNT is an apparent 'pure promoter'. These results provide an explanation for the conflicting DNT bioassay results.