Involvement of Cytochrome P450 3A in the Metabolism and Covalent Binding of 14C-Monocrotaline in Rat Liver Microsomes

M. J. Reid, M. W. Lamé, D. Morin, Dennis W Wilson, H. J. Segall

Research output: Contribution to journalArticle

57 Scopus citations

Abstract

The metabolism and covalent binding of 14C-monocrotaline in Sprague-Dawley (SD) rat liver microsomes was investigated using the inducers dexamethasone, clotrimazole, pregnenolone-16α-carbonitrile, and phenobarbital. Monocrotaline is a pyrrolizidine alkaloid (PA) that causes a syndrome in rats that is a model for human primary pulmonary hypertension. It has been documented that bioactivation of PAs (dehydrogenation to reactive pyrroles) in the liver by cytochromes P450 is required for their toxicity. Covalent binding of these reactive pyrroles to tissue macromolecules has been hypothesized to correspond to PA toxicosis. We correlated metabolism and total microsomal covalent binding of 14C-monocrotaline with cytochrome P450 3A using the aforementioned inducers, troleandomycin (a cytochrome P450 3A inhibitor), erythromycin N-demethylase assay of cytochrome P450 3A activity, and Western blots employing anti-rat cytochrome P450 3A antibodies. In addition, autoradiography of membranes electroblotted from SDS-PAGE demonstrated the formation of radiolabeled adducts with specific protein(s). The most intensely radiolabeled protein bands have an apparent molecular weight of ∼52 kDa, which was similar to the molecular weight detected by anti-rat cytochrome P450 3A antibodies in the Western blots. No radiolabeled proteins were detected in microsomes pretreated with troleandomycin.

Original languageEnglish (US)
Pages (from-to)157-166
Number of pages10
JournalJournal of Biochemical and Molecular Toxicology
Volume12
Issue number3
StatePublished - 1998

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Keywords

  • Autoradiography
  • Covalent Binding
  • Cytochrome P450 3A
  • Metabolism
  • Monocrotaline
  • Pyrrolizidine Alkaloid

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Health, Toxicology and Mutagenesis
  • Toxicology

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