title: Selective induction of peroxisomal enzymes by the hypolipidemic drug bezafibrate. Detection of modulations by automatic image analysis in conjunction with immunoelectron microscopy and immunoblotting.
abstract: Quantitative immunoelectron microscopy in conjunction with quantitative analysis of immunoblots have been used to study the effects of bezafibrate (BF), a peroxisome-proliferating hypolipidemic drug, upon six different enzyme proteins in rat liver peroxisomes (Po). Antibodies against following peroxisomal enzymes: catalase, urate oxidase, alpha-hydroxy acid oxidase, acyl-CoA oxidase, bifunctional enzyme (hydratase-dehydrogenase) and thiolase, were raised in rabbits, and their monospecificities were confirmed by immunoblotting. Female Sprague-Dawley rats were treated for 7 days with 250 mg/kg/day bezafibrate and liver sections were incubated with the appropriate antibodies followed by the protein A-gold complex. The labeling density for each enzyme was estimated by automatic image analysis. In parallel experiments immunoblots prepared from highly purified peroxisome fractions of normal and BF-treated rats were incubated with the same antibodies. The antigens were visualized by an improved protein A-gold method including an anti-protein A step and silver amplification. The immunoblots were also quantitated by an image analyzer. The results revealed a selective induction of beta-oxidation enzymes by bezafibrate with thiolase showing the most increase followed by bifunctional protein and acyl-CoA oxidase. The labeling density for catalase and alpha-hydroxy acid oxidase was reduced, confirming fully the quantitative analysis of immunoblots which in addition revealed reduction of uricase. These observations demonstrate that hypolipidemic drugs induce selectively the beta-oxidation enzymes while other peroxisomal enzymes are reduced. The quantitative immunoelectron microscopy with automatic image analysis provides a versatile, highly sensitive and efficient method for rapid detection of modulations of individual proteins in peroxisomes.
mesh_terms: D000101:Acetyl-CoA C-Acetyltransferase / Q000378:metabolism*; D000123:Acetyltransferases / Q000378:metabolism*; D000818:Animals; D001629:Bezafibrate / Q000494:pharmacology*; D002374:Catalase / Q000378:metabolism*; D005260:Female; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D008099:Liver / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D008830:Microbodies / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D008854:Microscopy, Electron; D010088:Oxidoreductases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
chemical_list: D010088:Oxidoreductases; D002374:Catalase; D000123:Acetyltransferases; D000101:Acetyl-CoA C-Acetyltransferase; D001629:Bezafibrate