Genetic Damage Levels Of Peripheral Blood Lymphocytes And Their Association With Single Nucleotide Polymorphisms Of Nucleotide Excision Repair Genes In Workers Exposed To Formaldehyde

BACKGROUND:The International Agency for Research on Cancer reclassified formaldehyde (FA) as a known human carcinogen (Group 1) in June 2004, and concluded that there is sufficient evidence that FA causes nasopharyngeal cancer and myeloid leukemia. Evaluation on the early health effects of occupational exposure to FA on exposed workers and development of safty programs and of preventive measures may have important implications for public health. Early genetic lesions caused by FA, a genotoxic xenobiotics and carcinogen, include DNA strand breaks, chromosome damage, DNA-protein crosslinks (DPCs). However, evidence for the association between FA exposure and genetic damage in peripheral blood (PB) lymphocytes is limited and conflicting. Further evaluating the genotoxicity of FA on PB lymphocytes may help to elucidate the leukemogenicity of FA. Of the multiple pathways in cells participating in the repair of FA-caused genetic damage sequencially or collaboratively, nucleotide excision repair (NER) pathway is the most versatile and important, and is proposed to be involed in the repair of chromosome damage, DPCs, and DNA interstrand crosslinks. Single-nucleotide polymorphisms (SNPs) in nucleotide excision repair genes may result in differential expression or repair capacity of corresponding proteins. Investigations on the modifying effects of these SNPs on the association between FA exposure and genentic damage levels may help to discover risk alleles and to screen risk-allele carriers who are susceptible to FA exposure.OBJECTIVE:We sought to clarify the association of FA exposure levels and of the number of years of exposure with DNA strand breaks, chromosome damage, and DPCs. We also studied the potential modifying effects of the SNPs in the NER genes on FA exposure-genetic damage relationships.METHODS:In the first part, we monitored ambient FA concentrations in five different workplaces of a plywood factory in Shandong province, China, and determinded DNA strand breaks by comet assay (presented as Olive tail moment values, OTM values), chromosome breakage or loss by cytokinesis-block micronucleus (CBMN) assay (presented as CBMN frequencies), and DPC rates by KC1-SDS assay, in PB lymphocytes of 178 workers in the five workplaces. We examined the association of each endpoint with current FA exposure levels and with the number of years of FA exposure in the 178 workers. In ordre to validate the exposure-effect relationship, we further measured genetic damage levels in an additional 62 workers before and after an 8-hour occupational exposure to FA, and monitored FA exposure levels of representative workers with different job titiles. We analyzed the association of FA exposure levels during work hours and of the number of years of FA exposure with each endpoint before and after work exposure and with the dynamic changes of genetic damage levels across 8-hour FA exposure in the 62 workers.In the second part, we genotyped 24 tagSNPs in 8 NER genes using TaqMan probes, and analyzed their associations with genetic damage levels in the 178 FA-exposed workers with generalized regression models after controlling for exposure and potential confounders. We further validated the association between positive SNPs and genentic damage levels in 128 independent FA-exposed workers and in the population combining the 128 workers and the previous 178 workers. Thereafter, we used the dual-luciferase reporter assay to study the potential function of the positive SNPs from the previous multiple-stage population study in HepG2 and 293 cell lines.RESULTS:In the 178 workers, OTM values of the PB lymphocytes increased in a dose-dependent manner with increasing levels of current FA exposure (Ptrend=0.002), and CBMN frequencies increased with an increase in the number of years of exposure (Ptrend< 0.001), but DPC rates were not associated with FA exposure levels or with number of years of exposure. In the dynamic study of the 62 workers, OTM values and DPC rates were increased after an 8-hour FA exposure compared with those before exposure (P<0.001, P=0.019, respectively), whereas CBMN frequencies after the work exposure were not significantly different from those before the exposure. Furthermore, the increase in OTM values, but not in DPC rates, was associated with FA exposure levels during work hours (P=0.005). CBMN frequencies before and after the 8-hour work were associated with number of years of FA exposure (P=0.029, P<0.001, respectively).In the 178 workers, ERCC1 rs3212961 CA and AA genotype carriers had significantly lower OTM values than did CC genotype carriers (P=0.022, P=0.038, respectively), rs2336219 GA plus AA genotype carriers had lower OTM values than did GG genotype carriers (P=0.030). ERCC5 rs2094258 AA genotype carriers and DDB2 rs901746 AA genotype carriers had significantly higher OTM values than did corresponding GG genotype carriers (P=0.012, P=0.029, respectively). ERCC5 rs4150348 TT genotype carriers and rs2296147 CC genotype carriers had significantly higher CBMN frequencies than did corresponding GG and TT genotype carriers (P=0.041,P=0.045, respectively).In the replication study of the 128 workers, different genotype carriers of rs2336219 and of rs2094258 had comparative OTM values. rs4150348 TT genotype carriers and rs2296147 TC plus CC genotype carriers had significantly higher CBMN frequencies than did corresponding GG and TT genotype carriers (P=0.005, P<0.001, respectively).In the study of the combined population, workers carrying different genotypes of rs2336219 and of rs2094258 had similar OTM values. Workers carring TT genotype of rs4150348 and carring CC genotype of rs2296147 had significantly higher CBMN frequencies than did workers carring GG and TT genotype correspondingly (P=0.007, P=0.001, respectively).The HepG2 and 293 cells containing constructed plasmids with rs2296147 C alleles had significantly lower relative luciferase activity than did those containing plasmids with T alleles (P=0.002, P<0.001, respectively).CONCLUSIONS:We found a dose-response relationship between current FA exposure levels and DNA strand breaks, as well as between number of years of FA exposure and chromosome damage. Our findings confirm that FA exposure causes genetic damage in PB lymphocytes of plywood workers. We also found that the SNPs in the NER genes had no modifying effects on levels of DNA strand breaks, while rs4150348 and rs2296147 in ERCC5 may influence the chromosome damage levels of FA-exposed workers. Different alleles of rs2296147 may lead to differential transcriptional levels of ERCC5.