Immunocytochemical analysis of soluble epoxide hydrolase and catalase in mouse and rat hepatocytes demonstrates a peroxisomal localization before and after clofibrate treatment.
The intracellular localization of soluble epoxide hydrolase and catalase was investigated in hepatocytes from untreated and clofibrate-treated male C57B1/6 mice and from untreated male Sprague-Dawley rats. Polyclonal rabbit antibodies directed against purified mouse liver cytosolic epoxide hydrolase and rat liver catalase were used and their specificity ascertained by Ouchterlony immunodiffusion and immunoblotting. The IgG fraction was purified and incubated with cryosections of isolated hepatocytes or liver tissue, priorly fixed in 4% paraformaldehyde, and protein-A gold conjugates were used to visualize the antigen-antibody reaction. The soluble form(s) of epoxide hydrolase was found to be localized in the matrix of peroxisomes in hepatocytes from normal and clofibrate-treated mice and normal rats. No significant reactivity was found against plasma membrane, nuclei, mitochondria, the Golgi apparatus, endoplasmic reticulum, lysosomes, or cytosol. Catalase was also localized to peroxisomes in all samples investigated. Accordingly, both the catalase and the epoxide hydrolase activities routinely recovered in the high-speed supernatant after subfractionation of rat and mouse liver tissue mostly seemed to be due to extensive matrix leakage from peroxisomes, and this phenomenon may also be found in other species. Rat hepatocytes contained less epoxide hydrolase than mouse hepatocytes, as judged by both immunocytochemical labeling and biochemical data. Clofibrate treatment of mice decreased the labeling density of epoxide hydrolase and catalase in hepatocytes peroxisomes, as expected, and more unlabeled peroxisomes were observed.
Hollinshead
M
M
European Molecular Biology Laboratory, Heidelberg/Federal Republic of Germany.
Meijer
J
J
D000818:Animals; D002374:Catalase / Q000378:metabolism*; D002994:Clofibrate / Q000494:pharmacology*; D004851:Epoxide Hydrolases / Q000378:metabolism*; D007150:Immunohistochemistry; D008099:Liver / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008830:Microbodies / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D008854:Microscopy, Electron; D051381:Rats; D011919:Rats, Inbred Strains
D002374:Catalase; D004851:Epoxide Hydrolases; D002994:Clofibrate