Effect Of Co-exposure To PCBs And BaP On DNA Damage Repair And Genetic Stability

Both of polychlorinated biphenyls (PCBs) and benzoapyrene (BaP) are ubiquitous and important environmental pollutants, can lead to various adverse effects to human being which including carcinogenesis and mutagenesis. Previous studies confirmed that PCBs can effectively induce the bioactivation of BaP to its ultimate carinogenic, and finally enhance the genotoxicity of BaP exposure. But, the adverse effects discovered by toxicology research were few confirmed by epidemiology studies in normal population. For reasons that these end point genotoxic effects can only happen in human after a long period of PCBs and BaP exposure. Base on the fact, to evaluate and predict the adverse effects of exposure of these two environmental pollutants on human health, we need to explore the early effects of damage leading by PCBs and BaP.ObjectAlthough, environmental hazards react with human being in different approach, in most case they can directly or indirectly lead express change of related genes. Compared with the traditional toxicological methods, the research of expression changes of toxicity related genes can be a more sensitive indicator to reflect the characteristics of environmental factors. In the present study, to find out the early response genes, we employed expression microarray to investigate the gene expression changes on several important pathways after co-exposure to PCBs and BaP. Then base on the result of microarray, we focus our research on three important pathways:DNA damage repair, cell cycle and apoptosis. By. these studies we try to explain the toxic mechanism of PCBs and BaP co-exposure, and finally provide technological support for early evaluation of the health hazard leading by PCBs and BaP.MethodsIn present study, HepG2 was used as the target cells, PCB126 and PCB153 were chosen as the representative congers of PCBs. Cells were treated with PCB126 (lnmol/L), PCB153 (10μmol/L) and BaP (50μmol/L) for 72h alone, or pretreated with PCB126 and PCB153 for 48h then co-treated with BaP and PCBs, DMSO (10ml/L) was used as solvent control. Gene expression changes on important pathways were evaluated by gene expression microarray. Base on the results of microarray, we chose PCB126 (0.01,0.1,1, and 10nmol/L)and BaP (50μmol/L) as research compounds, to evaluate their effect on DNA damage repair, cell cycle and apoptosis. The protein expression levels of nucleotide excision repair (NER) factors XPA, XPB and XPC were detected; this experiments were carried out again with the BaP bioactivation inhibitor ANF, to evaluate the relationship between DNA damage and repair capability. Cell cycle and apoptosis change were detected by flow cytometry. For following studies to explore the role of aryl hydrocarbon receptor (AhR) in PCBs and BaP synergism, we developed stably transfected HepG2 cell line with low expression of AhR.ResultExposure to BaP mainly lead to expression change of DNA damage repair genes, cell cycle and apoptosis regulation genes, all of these changes will benefit the cells to maintain their genetic integrity. Exposure to PCB126 or PCB153 alone could induce the expression of genes related with metabolism, and meanwhile the inductive capability of PCB126 is higher than PCB153. The expression profiling of PCB126 and BaP co-exposure was quite different with BaP alone, it led a complex adverse expression effect on damage repair genes, cell cycle and apoptosis regulation genes.Exposure to BaP or PCB126 alone induced the expression of XPA, compared with solvent control respectively (P<0.01). Compared with BaP alone, co-exposure to PCB126 and BaP inhibited the expression of XPA (P<0.01) except for the co-exposure to 0.01-nM PCB126 and BaP (P>0.05). After add ANF, except for the co-exposure to 10-nM PCB126 and BaP could inhibited the expression of XPA, other groups were recovery to level of solvent control.Exposure to BaP alone inhibited the expression of XPB, compare with solvent control (P<0.01). Exposure to PCB126 alone induce the expression of XPB, at the concentration of 1-nM and 10-nM (P<0.01). Compared with BaP alone, co-exposure to PCB126 and BaP induced the expression of XPB (P<0.01). After add ANF, except for the exposure to 1-nM and 10-nM PCB126 alone could induce the expression of XPB, other groups were recovery to level of solvent control.No statistically significant change of XPC expression was observed in cells exposed to BaP or PCB126 alone, compared with solvent control respectively (P>0.05). Compared with BaP alone, co-exposure to PCB126 and BaP inhibited the expression of XPC (P<0.05), except for the co-exposure to 0.01-nM PCB126 and BaP (P>0.05). After add ANF, except for the co-exposure to 10-nM PCB126 and BaP could inhibited the expression of XPC, other groups were recovery to level of solvent control.Exposure to BaP alone caused a significant increase of the percentage of cells at G1 phase but decrease it at S, compared with DMSO solvent control (P<0.01). Exposure to PCB126 alone caused a significant decrease of the percentage of cells at G1 but increase it at G2, compared with DMSO solvent control (P<0.01). Compared with BaP alone, co-exposure to PCB126 and BaP could effectively decrease the percentage of cells at G0/G1 phase, but increase the percentage of cells at S phase (P<0.01).Exposure to BaP or PCB126 alone caused a significant increase of apoptosis, compared with DMSO solvent control (P<0.05). Compared with BaP alone., co-exposure to PCB126 and BaP could effectively increase the apoptosis of HepG2 (P<0.01).Compare with the normal cell, the protein expression of AhR in RNA inference cells was decreased by 18.3%.Conclusion(1) PCBs could affect the ordered response of organism to the damage leading by BaP exposure, which may be caused by inhibit the damage repair capability, interfere the cell cycle and apoptosis process.(2) All of mechanisms mentioned above are important factors, which can interfere the outcome of BaP exposure and provide technological support for evaluation of the health hazard leading by PCBs and BaP.(3) As the represent of coplanar PCBs, PCB126 can enhance the genotoxicity of BaP, which caused by inhibiting the DNA damage repair capability, interfering the normal cell cycle and apoptosis process.