INFIDELITY OF DNA REPAIR AND HEALTH RISK

我们打算开发细胞遗传学（广泛性）和分子（特异性）

We intend to develop cytogenetic (broad based) and molecular (specific) assays for documentation of pollutant-induced DNA repair problem in cigarette smokers and in benzene-exposed workers. In the cytogenetic assay, blood lymphocytes from exposed individuals, matched controls and positive controls (cell line from zeroderma pigmentosum patients) will be irradiated with UV-light in vitro. These cells are challenged to repair the UV-light induced DNA damage. Abnormal/insufficient repair of the damage will cause the pollutant exposed cells and positive control cells to have more chromosome aberrations than those from normal controls. The repair defect will be confirmed biochemically as reduced uptake of labeled nucleotides. Three molecular assays will be developed for use in population studies to confirm and detect specific DNA repair problems. In one assay, cell homogenates from the same individuals will be used to repair UV-irradiated non-replicating plasmids, pSV2catSVgpt. Repair activities will be quantitated by measuring the uptake of labeled dAMP in recovered plasmids. In addition, alteration in the level of expression of two DNA repair enzymes, b-polymerase and O6-methylguanine-DNA methyltransferase, will be documented by using monoclonal antibodies in Western blot analyses. We expect that chronic exposure to toxic pollutants will affect most DNA repair mechanisms and the defect should be detected by our combination of assays. In addition, our protocols using unique and state of the art techniques should provide us with the sensitivities to detect the problem. This study will allow us to evaluate and recommend the most appropriate biomarker(s) for identification of potential human health risk from exposure to toxic pollutants.