In a meta-analysis using the database of the International Project on Genetic Susceptibility to Environmental Carcinogens (GSEC) the allele and genotype frequencies for many of the more commonly studied metabolic genes (CYP1A1, CYP2E1, CYP2D6, GSTM1, GSTT1, NAT2, GSTP1, and EPHX1) in the human population have been determined . If we take together the genotype frequencies of cases and controls of our study we observed the same genotype frequency of GSTM1, GSTP1, GSTT1, and NAT2 as reported by . These results indicate that the genotype frequencies in our study population did not deviate from published data and that the distribution of genotypes was not influenced by our study design. If individuals were divided in cases and controls using the scores from self-reported chemical-related sensitivity, we observed significant differences for distribution of genotype frequencies of GSTM1, GSTT1 and the NAT2 gene. Our results suggest that individuals with a deletion of GSTM1 or GSTT1 or slow acetyltators are at higher risk for developing self-reported chemical-related sensitivity.
The glutathione S-transferases are known to inactivate exogenous chemicals by glutathione conjugation. It is suggested that individuals with decreased glutathione conjugation are more prone to be unable to metabolize chemicals in the environment. It is also possible that both enzymes, GSTM1 and GSTT1, are metabolizing endogenous substrates less effectively due to gene deletion leading to an increased level of the parent compound. Although we did not differentiate between heterozygous and homozygous carriers of GSTM1 or GSTT1 by means of PCR, each of the homozygous deletions alone supported a direct role of the enzyme being associated with increased risk to self-reported chemical-related sensitivity. According to other authors, glutathione S-transferases play an important role in the detoxification of toxic chemicals. Nakajima et al. analysed GSTM1 genotype and total GST activity using 1-chloro-2,4-dinitrobenzene as a substrate and observed a greater GST activity in patients with the GSTM1 gene . An interaction between GSTM1 genotypes and benzo [a]pyrene DNA adducts through air pollution in urban and rural areas was investigated in 120 healthy non-smoking residents indicating that the deletion of the GSTM1 gene may be an important step in the early onset of diseases . It is also consistent with cases of our study group that homozygous GSTM1 deletion may be associated with an enhanced chemical-related sensitivity.
For GSTT1, there was a significant difference (OR: 2.80; p < 0.0001) between cases and controls in genotype frequency. This over-representation of cases with a homozygous GSTT1 deletion suggests that the GSTT1 enzyme plays an important role in glutathione conjugation of exogenous and/or endogenous substrates. This might result in cellular damage leading to an increased sensitivity if exposed to environmental chemicals. As described by other publications, individuals with a homozygous deletion of GSTT1 lack the possibility for enzymatic conjugation of environmental carcinogens such as 1,3-butadiene, ethylene oxide, epoxybutanes, methyl bromide, dichloromethane, and monohalomethanes [5–7]. In vitro experiments showed that the GSTT1 null genotype increased the sensitivity for sister chromatid exchange after exposure to diepoxybutane . Genotoxic effects have been observed after exposure of lymphocytes to styrene or the metabolite styrene-7,8-oxide using a sister chromatid exchange assay [42, 43]. These findings provide evidence that frequently used chemicals increase the susceptibility to develop chemical-related diseases. In addition, the GSTT1 null genotype conferred a 2.8-fold reduction in risk of relapse in childhood acute lymphoblastic leukemia indicating a more cytotoxic effect of chemotherapy . In our study, we observed an increased chemical-related sensitivity in subjects with homozygous GSTT1 deletion leading to the assumption that the deletion of this gene augments the susceptibility to environmental chemicals.
No case-control differences were observed in genotype or allelic frequencies of GSTP1. Common chemicals are metabolized by GSTP1 and have been associated with risk to develop diseases like non-Hodgkin's lymphoma, hepatocellular and prostate carcinoma, as well as Alzheimer. Furthermore, Gilliland et al. reported a diesel exhaust particle enhancement in patients with GSTP1 Ile105Ile genotype . However, we cannot exclude GSTP1 as a candidate gene for chemical-induced sensitivity since we did not analyse for promotor hypermethylation. Interestingly, the homoyzgous GSTP1*D/*D and the heterozygous GSTP1*C/*D genotype were not observed in cases and controls.
The GST enzymes, GSTM1, GSTT1 and GSTP1, are described to protect cells and organs from oxidative stress by conjugation of glutathione . They detoxify a variety of electrophilic compounds, including oxidized lipid, DNA and catechol products generated by reactive oxygen species-induced damage to intracellular molecules. Therefore we can conclude that the deletion of GSTM1 and/or GSTT1 gene in individuals with chemical-related sensitivity lead to the loss of protection against oxidative stress. At the endpoint of this cellular process, individuals with chemical-related sensitivity may be more prone to symptoms like muscular pain, cardiovascular diseases, gastrointestinal disorders and several other symptoms that are described by patients suffering from MCS [2, 3].
The results of our study show that cases were more frequently slow acetylators. The over-representation of the homozygous rapid acetylator genotype (NAT2*4/*4) reported by the study of McKeown-Eyssen et al. has been associated with the role of NAT2 in bioactivating arylamines to protein-binding metabolites . The results of our study indicate that inactivation of arylamines through N-acetylation is an important mechanism. Wormhoudt et al. described that the slow acetylator genotype NAT2*6/*6 leads to a significant decreased acetylation capacity (11% vs. 100% compared to NAT2*4/*4) . This genotype was observed more frequent in cases compared to controls in our study. Nevertheless, our results are not in contrast to the study of McKeown-Eyssen et al. since our study subjects were identified by a questionnaire asking for chemical hypersensitivity and not for symptoms of MCS. McKeown-Eyssen et al. drew up female patients from a larger study where participants were identified by symptoms described in six previously published MCS case definitions [2, 3]. We also calculated our study subjects stratified by gender and observed that female slow acetylators were more prone to report chemical-related sensitivity than male subjects. We cannot exclude that some individuals of our study fit into one of the published MCS case definitions but since the results of the MCS study of McKeown-Eyssen et al. are symptoms-related, the results of our study are not comparable with those from McKeown-Eyssen's study. In addition, since the area of McKeown-Eyssen's study corresponds to the area where we did our study in regard to industrialisation, we suppose that slow acetylation and lack of glutathione conjugation is an important step to increase chemical-related sensitivity.