Metabolism and bioactivation of 3-methylindole by Clara cells, alveolar macrophages, and subcellular fractions from rabbit lungs.
3-Methylindole (3MI), a fermentation product of tryptophan produced by intestinal and ruminal microflora, has been shown to cause pneumotoxicity in several species subsequent to cytochrome P450-mediated biotransformation. Among several species studied, rabbits are comparatively resistant to 3MI-induced pneumotoxicity, especially when compared to goats or mice. In this study, rabbit pulmonary cells and subcellular fractions were used to examine the metabolism and bioactivation of 3MI. A covalent-binding metabolite was produced in 3MI incubations by both Clara cells and macrophages. The addition of the cytochrome P450 inhibitor, 1-aminobenzotriazole, to these incubations inhibited the production of covalent-binding metabolite(s) by 94% in Clara cells and only 24% in macrophages. In incubations of Clara cells or macrophages with 3MI and N-acetylcysteine (NAC), a polar conjugate was detected and tentatively identified as an adduct of 3-hydroxy-3-methylindolenine (3H3MIN). Also identified were 3[(N-glutathione-S-yl)-methyl]-indole (3MI-GSH) and 3-methyloxindole (3MOI). In rabbit lung microsomal incubations with 3MI and glutathione (GSH), 3MI-GSH, 3MOI, indole-3-carbinol, and a GSH adduct of 3H3MIN were identified. The addition of cytosol to the microsomal incubations with GSH did not increase the rate of formation of the GSH adducts, indicating that cytosolic GSH-S-transferases are not essential in the formation of these metabolites. GSH significantly decreased the covalent binding of an electrophilic metabolite in microsomal incubations. These data suggest that GSH may be important in the mitigation of 3MI toxicity. Furthermore, the comparison of 3MI bioactivation to electrophilic intermediates in Clara cells and alveolar macrophages suggests that 3MI is metabolized by different oxidative pathways in the two different cell types, although the same metabolites were produced by the two cell types. This study shows that rabbit pulmonary enzymes are capable of bioactivating 3MI to reactive intermediates which become covalently bound to cellular macromolecules. This indicates that the relative resistance of rabbits to 3MI-induced pneumotoxicity is probably not due to differences in metabolic enzymes which convert 3MI to reactive intermediates.
Thornton-Manning
J R
JR
Department of Pharmacology and Toxicology, University of Utah, Salt Lake City 84112.
Nichols
W K
WK
Manning
B W
BW
Skiles
G L
GL
Yost
G S
GS
D000111:Acetylcysteine / Q000378:metabolism; D000818:Animals; D001711:Biotransformation; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D066298:In Vitro Techniques; D008168:Lung / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D016676:Macrophages, Alveolar / Q000378:metabolism; D008297:Male; D008861:Microsomes / Q000378:metabolism; D011817:Rabbits; D012862:Skatole / Q000378:metabolism* / Q000633:toxicity
D012862:Skatole; D005978:Glutathione; D000111:Acetylcysteine