Detection and localization of CYP1A1/CYP1A 2 in murine liver, electrophoresis, immunoblotting and imniunocytochemistr y

Abstract

Multiple forms of cytochrome P450 exist some of which are selectively inducible by exposure of the organism to a variety of foreign compounds. In this study, a monoclonal antibody specific for 3-methyl-cholanthrene-inducible cytochrome P450, Mab 1-7-1, was used to detect, localize and quantify CYP1A1/ CYP1A2 in livers of C57BL/6 mice. Mab 1-7-1 recognized a faint band in the range Ibetween 45- 66 Kd in Western immunoblots of liver microsomes from control mice, and a strong band in the same range, in liver microsomes from p-naphthoflavone-treated mice. Microsome from control liver contained minimal levels of CYP1A1/CYP1A2; pretreatment with p-naphtho flavone caused an increase in their expression. Immunoelectron microscopy was used to demonstrate the cellular localization and quantification of these isozymes in the liver. The immunolabeling procedure confinncd the endoplasmic reticulum as the primary site of CYP1A1/CYP1A2 induction in hepatocytes. This organelle showed the highest labeling density after treatment with p-naphthoflavone. Increase in CYP1A1/CYP1A2 was 33.4-fold by morphometric analysis in induced hepatocytes in comparison to non-induced cells. In conclusion, CYP1A1/CYP1A2 is highly induced by p-naphthoflavone in C57BL/6 mouse liver, and the cellular site of expression is the endoplasmic reticulum.