In situ localization and distribution of xenobiotic-activating enzymes and aryl hydrocarbon hydroxylase activity in lungs of untreated rats.
The present investigation was undertaken to more precisely establish where xenobiotics can be oxidatively metabolized and bioactivated within the lung. To accomplish this, antibodies raised against NADPH-cytochrome P-450 reductase (EC 1.6.2.4) and cytochromes P-450 BNF-B, PB-B, and PCN-E (the major forms of cytochrome P-450 induced by beta-naphthoflavone, phenobarbital, and pregnenolone-16 alpha-carbonitrile, respectively) that had been purified to apparent homogeneity from rat liver microsomes were used to determine the localizations and distributions of these enzymes immunohistochemically at the light microscopic level within lungs of untreated rats. Additionally, the intrapulmonary sites at which benzo(alpha)pyrene undergoes hydroxylation were identified in situ by means of fluorescence histochemistry. Immunohistochemical staining for NADPH-cytochrome P-450 reductase and cytochromes P-450 BNF-B, PB-B, and PCN-E was detected in bronchial epithelial cells, both ciliated and nonciliated (Clara) bronchiolar epithelial cells, and type II pneumocytes as well as other cells in the alveolar wall. Results of microfluorometric analyses of the immunofluorescence staining intensities of bronchial epithelial cells, Clara cells, and type II pneumocytes demonstrated further that Clara cells bound the antibodies raised to NADPH-cytochrome P-450 reductase and cytochrome P-450 PB-B to significantly greater extents than did bronchial epithelial cells and type II pneumocytes. Thus, in lungs of untreated rats, Clara cells contain the greatest amounts of these two enzymes. In marked contrast, the antibodies directed against cytochromes P-450 BNF-B and PCN-E were each bound to similar extents by bronchial epithelial cells, Clara cells, and type II pneumocytes. In agreement with immunohistochemical observations on the intrapulmonary localizations of NADPH-cytochrome P-450 reductase and cytochromes P-450 BNF-B, PB-B, and PCN-E in untreated rats, benzo(alpha)pyrene was hydroxylated in situ by bronchial and bronchiolar epithelial cells and alveolar wall cells, especially type II pneumocytes. These immunohistochemical and histochemical findings, thus, demonstrate that bronchial epithelial cells, Clara and ciliated bronchiolar epithelial cells, and type II pneumocytes as well as other alveolar wall cells represent sites for the in vivo oxidative metabolism and bioactivation of xenobiotics in lungs of untreated rats.
Voigt
J M
JM
Department of Pharmacology, University of Iowa, Iowa City 52242.
Kawabata
T T
TT
Burke
J P
JP
Martin
M V
MV
Guengerich
F P
FP
Baron
J
J
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000032:analysis*; D001579:Benzopyrene Hydroxylase / Q000032:analysis*; D001711:Biotransformation; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis*; D005455:Fluorescent Antibody Technique; D007124:Immunoenzyme Techniques; D008168:Lung / Q000166:cytology / Q000201:enzymology*; D008297:Male; D009251:NADPH-Ferrihemoprotein Reductase / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains; D015262:Xenobiotics / Q000378:metabolism*
D015262:Xenobiotics; D003577:Cytochrome P-450 Enzyme System; D001579:Benzopyrene Hydroxylase; D001189:Aryl Hydrocarbon Hydroxylases; D009251:NADPH-Ferrihemoprotein Reductase