In Vitro Assessment of the Efficacy of a Macrocyclic Chelator in Reversing Methylmercury Toxicity.

Methylmercury (MeHg) is a highly neurotoxic compound to which human populations are exposed via fish consumption. Once in cells, MeHg actively binds thiols and selenols, interfering with the activity of redox enzymes such as thioredoxin (Trx) and the selenoenzyme thioredoxin reductase (TrxR) which integrate the thioredoxin system. In fact, it has been shown that inhibition of this system by MeHg is a critical step in the unfolding of cell death. Current clinical approaches to mitigate the toxicity of MeHg rely on the use of chelators, such as meso-2,3-dimercaptosuccinic acid (DMSA) which largely replaced British anti-Lewisite or 2,3-dimercapto-1-propanol (BAL) as the prime choice. However, therapeutic efficacy is limited and therefore new therapeutic options are necessary. In this work, we evaluated the efficacy of a macrocyclic chelator, 1-thia-4,7,10,13-tetraazacyclopentadecane ([15]aneN4S), in preventing MeHg toxicity, namely by looking at the effects over relevant molecular targets, i.e., the thioredoxin system, using both purified enzyme solutions and cell experiments with human neuroblastoma cells (SH-SY5Y). Results showed that [15]aneN4S had a similar efficacy to DMSA and BAL in reversing the inhibition of MeHg over purified TrxR and Trx by looking at both the 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) reduction assay and insulin reduction capability. In experiments with cells, none of the chelating agents could reverse the inhibition of TrxR by MeHg, which corroborates the high affinity of MeHg to the selenol in TrxR active site. [15]aneN4S and BAL, unlike DMSA, could prevent inhibition of Trx, which allows the maintenance of downstream functions, although BAL showed higher toxicity to cells. Overall these findings highlight the potential of using [15]aneN4S in the treatment of MeHg poisoning and encourage further studies, namely in vivo.

Risk assessment of methylmercury in pregnant women and newborns in the island of Madeira (Portugal) using exposure biomarkers and food-frequency questionnaires.

Methylmercury (MeHg) is a contaminant present in fish which exerts a severe impact on health predominantly exhibiting neurotoxicity that might irreversibly affect fetal neurodevelopment. Fish consumption in Portugal is the third highest in the world, particularly high in regions with fishing tradition such as the Madeira Archipelago. Therefore, this study aimed at assessing the risk of exposure to MeHg in a population of pregnant women residing in Madeira. Blood samples from pregnant women (533) and umbilical cord (194) were collected from volunteer participants collected at primary health services in Madeira (Portugal) and analyzed for total mercury (HgT) level. A food-frequency questionnaire was used to estimate exposure and indices of risk while HgT in blood were correlated with estimated exposure. Analysis of HgT levels in blood indicated that 30% of pregnant women surpassed the maximum safe level of 10 µg/L recommended by the WHO, which was derived from the consumption of predatory fish, rich in MeHg. In addition, HgT levels in cord blood were 1.3 fold higher than in maternal blood, indicating the high risk of exposure to MeHg in this population. It is thus important to provide nutritional advice concerning fish consumption as a food choice in order to reduce fetal exposure and potential neurologic damage.

Exposure assessment of pregnant Portuguese women to methylmercury through the ingestion of fish: cross-sectional survey and biomarker validation.

Methylmercury (MeHg) contamination is a critical public health problem in Portugal, where fish is an important component of the daily diet. The Portuguese are the third largest consumers in the world (after Japan and Iceland) but first in Europe. Prenatal exposure to MeHg is believed to be linked to fetal/child neurodevelopment and behavioral impairments due to the neurotoxicity of the compound. The objective of this study was to assess the exposure of pregnant Portuguese women to mercury (Hg) due to fish consumption, calculating the indices of risk and confirming exposure through analyses of a biomarker of exposure. The study consisted of a cross-sectional evaluation of 343 pregnant women recruited at their visit to two antenatal care units in Lisbon, Portugal. A food frequency questionnaire was used to estimate prenatal exposure. Total Hg levels in hair were analyzed by atomic absorption, in samples from 186 women. The average fish consumption was 3.1 meals per week. Median Hg level in the hair was 1.26 µg/g (range: 0.07-5.3 µg/g). The mean calculated risk index was 0.81; however, 28% of the pregnant women ingested levels above the provisional tolerable weekly intake (PTWI) level recommended by the World Health Organization (WHO; 1.6 mg/kg per body weight), indicating the possibility of risk due to MeHg exposure. Multiple linear regression analysis showed the risk index was reliably predicted from predatory fish species and number of fish meals consumed per week. Ingestion of black and silver scabbard fish as well as mixed predatory fish cooked in traditional dishes enhanced the toxicity risk. In conclusion, some exposure levels exceeded the reference value; therefore, nutritional counseling needs to be provided to populations at risk.

Isolation and characterization of mercury-resistant bacteria from sediments of Tagus Estuary (Portugal): implications for environmental and human health risk assessment.

Mercury (Hg) contamination of aquatic systems has been recognized as a global and serious problem affecting both human and environmental health. In the aquatic ecosystems, mercurial compounds are microbiologically transformed with methylation responsible for generation of methylmercury (MeHg) and subsequent biomagnification in food chain, consequently increasing the risk of poisoning for humans and wildlife. High levels of Hg, especially MeHg, are known to exist in Tagus Estuary as a result of past industrial activities. The aim of this study was to isolate and characterize Hg-resistant bacteria from Tagus Estuary. Mercury-resistant (Hg-R) bacteria were isolated from sediments of two hotspots (Barreiro and North Channel) and one reserve area (Alcochete). Mercury contamination in these areas was examined and bacterial susceptibility to Hg compounds evaluated by determination of minimal inhibitory concentrations (MIC). The isolates characterization was based on morphological observation and biochemical testing. Bacteria characteristics, distribution, and Hg resistance levels were compared with metal levels. Barreiro and North Channel were highly contaminated with Hg, containing 126 and 18 µg/g total Hg, respectively, and in Alcochete, contamination was lower at 0.87 µg/g total Hg. Among the isolates there were aerobic and anaerobic bacteria, namely, sulfate-reducing bacteria, and Hg resistance levels ranged from 0.16 to 140 µg/ml for Hg(2+) and from 0.02 to 50.1 µg/ml for MeHg. The distribution of these bacteria and the resistance levels were consistent with Hg contamination along the depth of the sediments. Overall, results show the importance of the characterization of Tagus Estuary bacteria for ecological and human health risk assessment.

Genetic polymorphisms of cytochrome P450cl7alpha (CYP17) and progesterone receptor genes (PROGINS) in the assessment of endometriosis risk.

We designed the present study in order to evaluate the eventual role of polymorphisms in the genes encoding cytochrome P450c17alpha (CYP17) and the progesterone receptor (PROGINS) as risk factors for endometriosis development. Eligible cases consisted of 121 women with surgically confirmed endometriosis who underwent treatment in a hospital in São Paulo, Brazil during the period from September 2003 to September 2005. The 281 controls were participants with normal gynecological as well as pelvic ultrasound evaluation, who did not have any gynecological conditions during their reproductive lives such as pelvic pain and/or dyspareunia nor infertility history. Genomic DNA was obtained from buccal cells and processed for DNA extraction using the GFX DNA extraction kit (GE Healthcare). The CYP17 (-34T-->C) polymerase chain reaction-restriction fragment length polymorphism assay has been described previously, as has the progesterone receptor polymorphism (PROGINS) detection assay. PROGINS heterozygosis genotype frequencies were shown to be statistically higher in endometriosis cases compared with controls. On the other hand, differences in the CYP17 polymorphism (-34T-->C) frequencies were not even close to significance (p = 0.278) according to our findings.