DOSE-RESPONSES IN RAT HEPATIC PROTEIN MODIFICATION AND EXPRESSION FOLLOWING EXPOSURE TO THE RAT HEPATOCARCINOGEN METHAPYRILENE

Dose-related effects of methapyrilene (MP) on protein modification and expression were examined using two-dimensional gel electrophoresis (2 -D PAGE) coupled with computer analysis. Methapyrilene was administere d ad libitum at doses of 0, 62.5, 125, 250 and 1000 p.p.m. to male F-3 44 rats for 12 weeks beginning at 8 weeks of age. Following treatment, livers were removed and frozen for 2-D PAGE analysis. Separation of h epatic proteins was conducted using ISO-DALT(R) technology. Changes in abundance and modification of hepatic proteins were determined using the Kepler(TM) software package. Covalent modifications of three speci fic mitochondrial proteins were quantified using a charge modification index. Dose-response relationships were analyzed using Tukey's trend test. Results demonstrated that covalent modification of the three mit ochondrial proteins was linearly related to dose and that a dose effec t could be found at all dose levels in 2 out of 3 proteins. Two forms of change in protein expression were observed: (i) a dose-dependent ch ange with effects at all doses and (ii) a change only at the toxic dos e of 1000 p.p.m. MP. These results demonstrate a molecular effect of M P at doses that do not produce cellular responses including toxicity o r increases in cell replication suggesting that these specific mitocho ndrial modifications are molecular dosimeters but are probably not dir ect factors and/or sufficient factors in carcinogenesis. This study al so demonstrates the potential use of 2-D PAGE electrophoresis to delin eate the effect of dose on expression of specific proteins.