Quantitative RT-PCR measurement of cytochromes p450 1A1, 1B1, and 2B7, microsomal epoxide hydrolase, and NADPH oxidoreductase expression in lung cells of smokers and nonsmokers.
Bronchial epithelial cells (BEC) are the progenitors of bronchogenic carcinomas and are exposed to polycyclic aromatic hydrocarbon (PAH) procarcinogens through inhalation of combustion products. PAH are converted to carcinogenic molecules through a combination of monoxygenation by cytochrome p450 (CYP) enzymes in the presence of NADPH oxidoreductase (OR) and hydrolysis by microsomal epoxide hydrolase (mEH). In artificial systems, the relative expression of these genes determines whether carcinogenic or noncarcinogenic species are generated during metabolism. This relationship was explored in humans by using quantitative competitive reverse transcriptase polymerase chain reaction amplification to determine the range of expression of CYP1A1, CYP1B1, mEH, and NADPH OR in BEC recovered from 10 nonsmokers and 9 smokers. CYP2B7 expression was evaluated because, although little is known of its substrate specificity, it is expressed at high levels in human lung tissue. CYP1A1 and CYP1B1 were expressed in BEC at significantly different levels (P < 0.05) in the 9 smokers at 1.4 +/- 2.3 x 10(4) and 2.4 +/- 3.2 x 10(3) molecules/10(6) beta-actin molecules (mean +/- STD), respectively, but each was measurable in only one of the 10 nonsmokers. There was significant inter-individual variation (P < 0.05) in both CYP1A1 and CYP1B1 expression among the subjects for whom sufficient data were obtained. The inducibility of human BEC CYP1A1 gene by PAH exposure was confirmed in vitro by incubating cultured immortalized human BEC with beta-naphthoflavone and observing a > 6-fold induction of CYP1A1 after 24 h. In contrast to BEC, alveolar macrophages expressed CYP1A1 at low (30-70 molecules/10(6) beta-actin molecules) to unmeasurable levels in both smokers and nonsmokers. There was no significant difference in expression of mEH, CYP2B7, or NADPH OR in smokers compared with nonsmokers. The inter-individual variation in absolute and relative expression of PAH metabolism enzymes in BEC reported here supports the hypothesis that inter-individual variation in ability to activate/inactivate inhaled PAH carcinogens accounts for at least some of the inter-individual variation in risk for bronchogenic carcinoma.
Willey
J C
JC
Department of Medicine, Medical College of Ohio, Toledo 13699-0008, USA. jwilley@opus.mco.edu
Coy
E L
EL
Frampton
M W
MW
Torres
A
A
Apostolakos
M J
MJ
Hoehn
G
G
Schuermann
W H
WH
Thilly
W G
WG
Olson
D E
DE
Hammersley
J R
JR
Crespi
C L
CL
Utell
M J
MJ
D000328:Adult; D001189:Aryl Hydrocarbon Hydroxylases; D001999:Bronchoscopy; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis; D065633:Cytochrome P-450 CYP1B1; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis*; D000072467:Cytochrome P450 Family 2; D017931:DNA Primers; D004851:Epoxide Hydrolases / Q000096:biosynthesis*; D005260:Female; D006801:Humans; D008168:Lung / Q000201:enzymology*; D008297:Male; D008861:Microsomes / Q000201:enzymology; D019255:NADPH Oxidases / Q000096:biosynthesis*; D016133:Polymerase Chain Reaction / Q000379:methods; D012016:Reference Values; D012907:Smoking / Q000378:metabolism*
D017931:DNA Primers; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585496:CYP1B1 protein, human; C107082:CYP2B7P protein, human; D019363:Cytochrome P-450 CYP1A1; D065633:Cytochrome P-450 CYP1B1; D000072467:Cytochrome P450 Family 2; D019255:NADPH Oxidases; D004851:Epoxide Hydrolases