Influence of GSTM1, GSTT1, GSTP1, NAT1, NAT2, EPHX1, MTR and MTHFR polymorphism on chromosomal aberration frequencies in human lymphocytes.

We have studied the influence of genetic polymorphisms in the xenobiotic-metabolizing genes GSTM1, GSTP1, GSTT1, EPHX1, NAT1 and NAT2 and the folate-metabolizing genes MTR and MTHFR on the frequencies of cells with chromosomal aberrations (CAs) in peripheral lymphocytes of Norwegian men. Log-linear Poisson regression models were applied on 357 subjects of whom data on all the polymorphisms examined were available. Total CAs and chromosome-type aberrations (CSAs) were significantly increased by higher age alone, whereas chromatid-type aberrations (CTAs) were elevated by the GSTT1-null genotype and MTHFR codon 222 variant allele and chromatid gaps (CTGs) by EPHX1 high activity genotype and occupational exposure. Stratification by smoking and age (<40 and ≥40 years) showed that the effect of the GSTT1 null and EPHX1 high activity genotypes only concerned (older) smokers, in agreement with the roles of the respective enzymes in detoxification and metabolic activation. The MTHFR codon 222 variant allele was associated with high CTGs in smokers, the MTR codon 919 variant allele with high CTAs in older smokers and the NAT2 fast acetylator genotype with high CTGs in older subjects. Among younger nonsmokers, however, carriers of the MTHFR codon 222 and MTR codon 919 variant alleles showed a decrease in the level of CTGs and total CAs, respectively. In conclusion, polymorphisms of GSTT1, EPHX1, MTHFR, MTR and NAT2 differentially affect the frequency of CTAs, CSAs and CTGs, showing interaction with smoking and age. It appears that CA subtypes rather than total CAs should be considered in this type of studies.

Influence of DNA repair gene polymorphisms of hOGG1, XRCC1, XRCC3, ERCC2 and the folate metabolism gene MTHFR on chromosomal aberration frequencies.

We have studied the effect of genetic polymorphisms in the DNA repair genes hOGG1, XRCC1, XRCC3, ERCC2 and the MTHFR gene in the folate metabolism on the frequencies of cells with chromosomal aberrations (CA), chromosome-type aberrations (CSA), chromatid-type aberrations (CTA), chromatid breaks (CTB) and chromatid gaps (CTG) scored in peripheral blood lymphocytes from 651 Norwegian subjects of Caucasian descendant. DNA was extracted from fixed cell suspensions. The log-linear Poisson regression model was used for the combined data which included age, smoking, occupational exposure and genotype for 449 subjects. Our results suggest that individuals carrying the hOGG1 326Cys or the XRCC1 399Gln allele have an increased risk of chromosomal damage, while individuals carrying the XRCC1 194Trp or the ERCC2 751Gln allele have a reduced risk regardless of smoking habits and age. Individuals carrying the XRCC1 280His allele had an increased risk of CSA which was only apparent in non-smokers. This was independent of age. A protective effect of the XRCC3 241Met allele was only found in the older age group in non-smokers for CA, CSA and CTA, and in smokers for CSA. In the youngest age group, the opposite effect was found, with an increased risk for CA, CTA and CTG in smokers. Carrying the MTHFR 222Val allele gave an increased risk for chromosome and chromatid-type aberrations for both non-smokers and smokers, especially for individuals in the older age group, and with variable results in the youngest age group. The variables included in the different regression models accounted, however, for only 4-10% of the variation. The frequency ratio for CTG was significantly higher than for CTA and CTB for only 7 of the 43 comparisons performed. Some of the gap frequencies diverge from the trend in the CA, CSA, CTA and CTB results.

Single-track sequencing for genotyping of multiple SNPs in the N-acetyltransferase 1 (NAT1) gene.

BACKGROUND: Fast, cheap and reliable methods are needed to identify large populations, which may be at risk in relation to environmental exposure. Polymorphisms in NAT1 (N-acetyl transferase) may be suitable markers to identify individuals at risk. RESULTS: A strategy allowing to address simultaneously 24 various genetic variants in the NAT1 gene using the single sequencing reaction method on the same PCR product is described. A modified automated DNA sequencing using only one of the sequence terminators was used to genotype PCR products in single-track sequencing reactions of NAT1 and was shown to be universal for both DNA sequencing using labeled primers and labeled nucleotides. By this method we detected known SNPs at site T640G, which confers the NAT1*11 allele with frequency of 0.036, further T1088A and C1095A with frequency of 0.172 and 0.188, respectively and a deletion of TAATAATAA in the poly A signal area with a frequency 0.031. All observed frequencies were in Hardy Weinberg equilibrium and comparable to those in Caucasian population. The single-track signatures of the variant genotypes were verified on samples previously genotyped by RLFP. CONCLUSIONS: The method could be of great help to scientists in the field of molecular epidemiology of screening of large populations for known informative biomarkers of susceptibility, such as NAT1.

Genetic diagnosis of Charcot-Marie-Tooth disease in a population by next-generation sequencing.

Charcot-Marie-Tooth (CMT) disease is the most prevalent inherited neuropathy. Today more than 40 CMT genes have been identified. Diagnosing heterogeneous diseases by conventional Sanger sequencing is time consuming and expensive. Thus, more efficient and less costly methods are needed in clinical diagnostics. We included a population based sample of 81 CMT families. Gene mutations had previously been identified in 22 families; the remaining 59 families were analysed by next-generation sequencing. Thirty-two CMT genes and 19 genes causing other inherited neuropathies were included in a custom panel. Variants were classified into five pathogenicity classes by genotype-phenotype correlations and bioinformatics tools. Gene mutations, classified certainly or likely pathogenic, were identified in 37 (46%) of the 81 families. Point mutations in known CMT genes were identified in 21 families (26%), whereas four families (5%) had point mutations in other neuropathy genes, ARHGEF10, POLG, SETX, and SOD1. Eleven families (14%) carried the PMP22 duplication and one family carried a MPZ duplication (1%). Most mutations were identified not only in known CMT genes but also in other neuropathy genes, emphasising that genetic analysis should not be restricted to CMT genes only. Next-generation sequencing is a cost-effective tool in diagnosis of CMT improving diagnostic precision and time efficiency.

Cytogenetic effects of 18.0 and 16.5 GHz microwave radiation on human lymphocytes in vitro.

BACKGROUND: There are few cell studies on the direct genotoxic effects of microwave radiation. In this study, cytogenetic effects of microwave radiation alone or in combination with mitomycin C (MMC) were investigated. MATERIALS AND METHODS: Lymphocytes from two smoking and four non-smoking donors were exposed for 53 hours in vitro to 1.0 W/m(2) continuous-wave radiation at 18.0 GHz or 10 W/m(2) pulsed-wave at 16.5 GHz, alone or in combination with MMC. DNA synthesis and repair were inhibited in vitro in some cultures. RESULTS: No synergistic effect was observed in cells exposed to combinations of microwave radiation and in vitro exposure to MMC, or to cells pre-exposed in vivo to tobacco smoke. For the 16.5 GHz pulsed exposure, a non-significant trend consisting of an increase in aberration frequencies with microwave radiation was shown for the DNA synthesis and repair inhibited cultures both with and without MMC. CONCLUSION: Neither 18.0 GHz continuous-wave nor 16.5 GHz pulsed-wave exposure to human lymphocytes in vitro induced statistically significant increases in chromosomal aberration frequencies. 16.5 GHz pulsed-wave exposure requires further documentation before a true negative conclusion can be drawn.

Cytogenetic effects of exposure to 2.3 GHz radiofrequency radiation on human lymphocytes in vitro.

BACKGROUND: No previous in vitro studies have tested radio frequency radiation for at least one full cell cycle in culture. The aim was to test if exposure used in mobile phones and wireless network technologies would induce DNA damage in cultured human lymphocytes with and without a known clastogen. MATERIALS AND METHODS: Lymphocytes from six donors were exposed to 2.3 GHz, 10 W/m(2) continuous waves, or 2.3 GHz, 10 W/m(2) pulsed waves (200 Hz pulse frequency, 50% duty cycle). Mitomycin C was added to half of the cultures. DNA synthesis and repair were inhibited in one experiment. RESULTS: No statistically significant differences were observed between control and exposed cultures. A weak trend for more chromosomal damage with the interaction of pulsed fields with mitomycin C compared to a constant field was observed. CONCLUSION: Exposure during the whole cell cycle in inhibited cultures did not resulted in significant differences in chromosomal aberrations as compared to controls.