Immunoperoxidase quantitation of 4-aminobiphenyl- and polycyclic aromatic hydrocarbon-DNA adducts in exfoliated oral and urothelial cells of smokers and nonsmokers.
Immunoperoxidase methods using two antibodies were developed for detection and quantitation of DNA damage in single cells. A monoclonal antibody that recognizes 4-aminobiphenyl (4-ABP)-DNA adducts was initially tested on liver tissues of BALB/c mice treated with 4-ABP, then applied to the detection of adducts in oral mucosa and exfoliated urothelial cells of smokers and nonsmokers. Levels of 4-ABP-DNA in exfoliated urothelial cells were elevated in each of 20 smokers (mean relative staining intensity, 517 +/- 137) compared with age-, race-, and sex-matched nonsmokers (313 +/- 79; P < 0.0005). Significantly higher damage levels were also observed in oral mucosa cells of smokers compared with nonsmokers (552 +/- 157 versus 326 +/- 101; P < 0.0005). A polyclonal antiserum that recognizes benzo(a)pyrene and structurally related polycyclic aromatic hydrocarbon (PAH) diol epoxide-DNA adducts was also applied to the same study samples after validation by staining of 10T1/2 cells treated with (+/-)-trans-anti-benzo(a)pyrene diol epoxide. Smokers had higher levels of PAH-DNA damage in oral mucosa and exfoliated urothelial cells than nonsmokers (oral mucosa cells, 684 +/- 107 versus 370 +/- 83; P < 0.0005; urothelial cells, 689 +/- 72 versus 495 +/- 57; P < 0.0005). A similar 2-3-fold range in relative staining was found in smokers and nonsmokers for both 4-ABP- and PAH-DNA, suggesting the importance of individual differences in capacity to metabolize the carcinogens and/or repair damaged DNA. Significant correlations were found among the biomarkers in both cell types. This noninvasive method, requiring small numbers of cells and with a relatively low cost, will be useful for monitoring DNA damage in large-scale molecular epidemiology studies.
Hsu
T M
TM
Division of Environmental Health Sciences, Columbia School of Public Health, New York, New York, USA.
Zhang
Y J
YJ
Santella
R M
RM
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000032:analysis; D000611:Aminobiphenyl Compounds / Q000032:analysis*; D000818:Animals; D000911:Antibodies, Monoclonal; D001564:Benzo(a)pyrene / Q000032:analysis; D015415:Biomarkers / Q000032:analysis; D002273:Carcinogens / Q000032:analysis*; D016022:Case-Control Studies; D002452:Cell Count; D002478:Cells, Cultured; D004396:Coloring Agents; D018736:DNA Adducts / Q000032:analysis*; D004260:DNA Repair; D005260:Female; D006801:Humans; D007124:Immunoenzyme Techniques; D008099:Liver / Q000737:chemistry / Q000473:pathology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D017720:Molecular Epidemiology; D009061:Mouth Mucosa / Q000737:chemistry* / Q000473:pathology; D011083:Polycyclic Compounds / Q000032:analysis*; D012907:Smoking / Q000378:metabolism*; D019459:Urothelium / Q000737:chemistry* / Q000473:pathology
D000611:Aminobiphenyl Compounds; D000911:Antibodies, Monoclonal; D015415:Biomarkers; D002273:Carcinogens; D004396:Coloring Agents; D018736:DNA Adducts; D011083:Polycyclic Compounds; C006757:4-biphenylamine; D001564:Benzo(a)pyrene; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide