Mayukh Banerjee

High arsenic in rice is associated with elevated genotoxic effects in humans

Arsenic in drinking water may cause major deleterious health impacts including death. Although arsenic in rice has recently been demonstrated to be a potential exposure route for humans, there has been to date no direct evidence for the impact of such exposure on human health. Here we show for the first time, through a cohort study in West Bengal, India, involving over 400 human subjects not otherwise significantly exposed to arsenic through drinking water, elevated genotoxic effects, as measured by micronuclei (MN) in urothelial cells, associated with the staple consumption of cooked rice with >200 μg/kg arsenic. Further work is required to determine the applicability to populations with different dietary and genetic characteristics, but with over 3 billion people in the world consuming rice as a staple food and several percent of this rice containing such elevated arsenic concentrations, this study raises considerable concerns over the threat to human health.

Mechanism of erythrocyte death in human population exposed to arsenic through drinking water.

Arsenic contamination in drinking water is one of the biggest natural calamities, which has become an imperative threat to human health throughout the world. Abbreviation of erythrocyte lifespan leading to the development of anemia is a common sequel in arsenic exposed population. This study was undertaken to explore the mechanism of cell death in human erythrocytes during chronic arsenic exposure. Results revealed transformation of smooth discoid red cells into evaginated echinocytic form in the exposed individuals. Further distortion converted reversible echinocytes to irreversible spheroechinocytes. Arsenic toxicity increased membrane microviscosity along with an elevation of cholesterol/phospholipid ratio, which hampered the flexibility of red cell membrane and made them less deformable. Significant increase in the binding of merocyanine 540 with erythrocyte membrane due to arsenic exposure indicated disruption of lipid packing in the outer leaflet of the cell membrane resulting from altered transbilayer phospholipid asymmetry. Arsenic induced eryptosis was characterized by cell shrinkage and exposure of phosphatidylserine at the cell surface. Furthermore, metabolic starvation with depletion of cellular ATP triggered apoptotic removal of erythrocytes from circulation. Significant decrease in reduced glutathione content indicating defective antioxidant capacity was coupled with enhancement of malondialdehyde and protein carbonyl levels, which pointed to oxidative damage to erythrocyte membrane. Arsenic toxicity intervened into red cell membrane integrity eventually leading to membrane destabilization and hemoglobin release. The study depicted the involvement of both erythrophagocytosis and hemolysis in the destruction of human erythrocytes during chronic arsenic exposure.

Role of genomic instability in arsenic-induced carcinogenicity. A review.

Exposure to chronic arsenic toxicity is associated with cancer. Although unstable genome is a characteristic feature of cancer cells, the mechanisms leading to genomic instability in arsenic-induced carcinogenesis are poorly understood. While there are excellent reviews relating to genomic instability in general, there is no comprehensive review presenting the mechanisms involved in arsenic-induced genomic instability. This review was undertaken to present the current state of research in this area and to highlight the major mechanisms that may involved in arsenic-induced genomic instability leading to cancer. Genomic instability is broadly classified into chromosomal instability (CIN), primarily associated with mitotic errors; and microsatellite instability (MIN), associated with DNA level instability. Arsenic-induced genomic instability is essentially multi-factorial in nature and involves molecular cross-talk across several cellular pathways, and is modulated by a number of endogenous and exogenous factors. Arsenic and its metabolites generate oxidative stress, which in turn induces genomic instability through DNA damage, irreversible DNA repair, telomere dysfunction, mitotic arrest and apoptosis. In addition to genetic alteration; epigenetic regulation through promoter methylation and miRNA expression alters gene expression profiling leading to genome more vulnerable and unstable towards cancer risk. Moreover, mutations or silencing of pro-apoptotic genes can lead to genomic instability by allowing survival of damaged cells that would otherwise die. Although a large body of information is now generated regarding arsenic-induced carcinogenesis; further studies exploring genome-wide association, role of environment and diet are needed for a better understanding of the arsenic-induced genomic instability.

Comparison of health effects between individuals with and without skin lesions in the population exposed to arsenic through drinking water in West Bengal, India

A study was conducted to explore the effect of arsenic causing conjunctivitis, neuropathy and respiratory illness in individuals, with or without skin lesions, as a result of exposure through drinking water, contaminated with arsenic to similar extent. Exposed study population belongs to the districts of North 24 Parganas and Nadia, West Bengal, India. A total of 725 exposed (373 with skin lesions and 352 without skin lesions) and 389 unexposed individuals were recruited as study participants. Participants were clinically examined and interviewed. Arsenic content in drinking water, urine, nail and hair was estimated. Individuals with skin lesion showed significant retention of arsenic in nail and hair and lower amount of urinary arsenic compared to the group without any skin lesion. Individuals with skin lesion also showed higher risk for conjunctivitis ((odds ratio) OR: 7.33, 95% CI: 5.05–10.59), peripheral neuropathy (OR: 3.95, 95% CI: 2.61–5.93) and respiratory illness (OR: 4.86, 95% CI: 3.16–7.48) compared to the group without any skin lesion. The trend test for OR of the three diseases in three groups was found to be statistically significant. Again, individuals without skin lesion in the exposed group showed higher risk for conjunctivitis (OR: 4.66, 95% CI: 2.45–8.85), neuropathy (OR: 3.99, 95% CI: 1.95–8.09), and respiratory illness (OR: 3.21, 95% CI: 1.65–6.26) when compared to arsenic unexposed individuals. Although individuals with skin lesions were more susceptible to arsenic-induced toxicity, individuals without skin lesions were also subclinically affected and are also susceptible to arsenic-induced toxicity and carcinogenicity when compared to individuals not exposed to arsenic.

Toxicogenomics of arsenic: classical ideas and recent advances.

Exposure to arsenic, a toxic metalloid distributed widely in nature, has been known to result in hazardous health outcomes including cancer. Incidence of arsenic toxicity, mostly from usage of underground water, has been reported from different corners of the world spanning more than 21 countries. Recent studies have radically influenced our knowledge on the level of toxicity imparted by different chemical forms of the metalloid. To understand the basic biology for arsenic metabolism different species have been studied at the molecular level, which has unraveled a wealth of information. However, there is no ideal animal model that can be used to understand arsenic toxicity in humans prompting the investigators to undertake the study directly in human cell lines and on affected individuals in areas of exposure. It is interesting to note that only a subset of exposed individuals demonstrate pathological features, i.e. skin lesions sometimes followed by cancer. Also, the extent of affectedness varies remarkably between individuals exposed to similar levels of arsenic. These observations led to investigation of genomic variation in exposed individuals as a potential contributing factor for differential susceptibility. An impressive number of studies have been published on variation in global gene expression and the potential association of candidate genes with arsenic-induced pathology. While there are excellent reviews relating to arsenic metabolism in general, there is no comprehensive review presenting the studies described in humans related to the metalloid toxicity. Our goal in this article has been to present the current state of research on this area to help formulate strategies for future studies.

Arsenic-induced mitochondrial instability leading to programmed cell death in the exposed individuals.

In West Bengal, India, more than 6 million people in nine districts are exposed to arsenic through drinking water. It is regarded as the greatest arsenic calamity in the world. Arsenic is a well-documented human carcinogen, which does not induce cancer in any other animal model. Interestingly, at lower concentrations, arsenic is known to induce apoptosis in various cancer cell lines in vitro. We have studied apoptosis in human peripheral blood mononuclear cells (PBMC) of 30 arsenic exposed skin lesion individuals by annexin V-FITC staining and compared with 28 unexposed individuals. The percentage of apoptotic cells in individuals with skin lesions was significantly higher (p<0.001) in comparison to unexposed individuals. In the exposed individuals with skin lesions, there were elevated levels of intracellular reactive oxygen species (ROS), mitochondrial membrane permeability and increased cytochrome c release, leading to increased downstream caspase activity. Arsenic-induced DNA damage was confirmed by DNA ladder formation and confocal microscopy. We also observed that chronic arsenic exposure reduced Bcl-2/Bax ratio and also resulted in cell cycle arrest of PBMC in G0/G1 phase. All these observations indicate that mitochondria-mediated pathway may be responsible for arsenic-induced apoptosis.

DNA Repair Deficiency Leads to Susceptibility to Develop Arsenic-Induced Premalignant Skin Lesions

In West Bengal, India, although more than 6 million people are exposed to arsenic through drinking water, only 15–20% showed arsenic‐induced skin lesions, including premalignant hyperkeratosis. This indicates toward some factors that confer susceptibility to arsenic‐induced carcinogenicity. In this work, we wanted to explore whether differences in DNA repair capacity could impart arsenic‐induced carcinogenicity, through Comet assay, chromosomal aberration (CA) assay and challenge assay. Sixty arsenic exposed (30 individuals with arsenic‐induced premalignant hyperkeratosis and 30 without skin lesion, but drinking similar arsenic contaminated water) and 30 arsenic unexposed individuals were recruited as study participants. Alkaline comet assay, and challenge assay were carried out in whole blood and CA study in lymphocytes to find out the DNA damage and DNA repair capacity in both hyperkeratotic and without skin lesion individuals. DNA damage as well as CA were found to be significantly higher in the arsenic‐exposed individuals compared to unexposed individuals (p < 0.001). Within the exposed group, there was no significant difference as far as the level of DNA damage is concerned (p > 0.05), but CA was significantly higher in exposed individuals with hyperkeratosis than exposed individuals without hyperkeratosis (p < 0.01). Challenge assay showed that upon induction of DNA damage, the repair capacity in the exposed individuals with premalignant hyperkeratosis is significantly less (p < 0.001) than that of individuals without skin lesion, although the basal level of DNA damage was similar in both. Thus, the deficiency in DNA repair capacities in the hyperkeratotic individuals emerges as a prime contender for arsenic carcinogenicity.

Arsenic-induced toxicity and carcinogenicity: a two-wave cross-sectional study in arsenicosis individuals in West Bengal, India

In the state of West Bengal in India, over 26 million individuals are exposed to arsenic via drinking water. Dermatological, non-dermatological disorders and cancers are associated with arsenic toxicity. Of late, there has been a decrease in the arsenic concentration in drinking water owing to governmental efforts, raising the possibility of remediation. A cross-sectional study was conducted, where 189 arsenicosis and 171 unexposed individuals were recruited at two time points, (2005–06 and 2010–11) with concomitant decrease in the level of arsenic exposure via drinking water in the arsenicosis group in 2010–11. Parameters studied included dermatological, non-dermatological health status and cytogenetic damage. Decrease of arsenic exposure (190.1 μg/l to 37.94 μg/l) resulted in significant decline in the number of individuals having dermatological disorders (P<0.01) and in the severity of each dermatological outcome (P<0.0001). Micronucleus formation in urothelial cells and lymphocytes decreased significantly (P<0.001). However, there was a significant (P<0.001) rise in the incidence of each of the non-dermatological diseases, that is, peripheral neuropathy, conjunctivitis and respiratory distress over the period. Thirteen (6.87%) of the initially recruited arsenicosis individuals died of cancer, in this period. Remediation by arsenic-safe drinking water can reduce dermatological manifestations and cytogenetic insult; but is unable to counter the non-dermatological symptoms.

Functional compensation of glutathione S-transferase M1 (GSTM1) null by another GST superfamily member, GSTM2

The gene for glutathione-S-transferase (GST) M1 (GSTM1), a member of the GST-superfamily, is widely studied in cancer risk with regard to the homozygous deletion of the gene (GSTM1 null), leading to a lack of corresponding enzymatic activity. Many of these studies have reported inconsistent findings regarding its association with cancer risk. Therefore, we employed in silico, in vitro, and in vivo approaches to investigate whether the absence of a functional GSTM1 enzyme in a null variant can be compensated for by other family members. Through the in silico approach, we identified maximum structural homology between GSTM1 and GSTM2. Total plasma GST enzymatic activity was similar in recruited individuals, irrespective of their GSTM1 genotype (positive/null). Furthermore, expression profiling using real-time PCR, western blotting, and GSTM2 overexpression following transient knockdown of GSTM1 in HeLa cells confirmed that the absence of GSTM1 activity can be compensated for by the overexpression of GSTM2.

Evaluation of the serum catalase and myeloperoxidase activities in chronic arsenic-exposed individuals and concomitant cytogenetic damage

Chronic arsenic exposure through contaminated drinking water is a major environmental health issue. Chronic arsenic exposure is known to exert its toxic effects by a variety of mechanisms, of which generation of reactive oxygen species (ROS) is one of the most important. A high level of ROS, in turn, leads to DNA damage that might ultimately culminate in cancer. In order to keep the level of ROS in balance, an array of enzymes is present, of which catalase (CAT) and myeloperoxidase (MPO) are important members. Hence, in this study, we determined the activities of these two enzymes in the sera and chromosomal aberrations (CA) in peripheral blood lymphocytes in individuals exposed and unexposed to arsenic in drinking water. Arsenic in drinking water and in urine was used as a measure of exposure. Our results show that individuals chronically exposed to arsenic have significantly higher CAT and MPO activities and higher incidence of CA. We found moderate positive correlations between CAT and MPO activities, induction of CA and arsenic in urine and water. These results indicate that chronic arsenic exposure causes higher CAT and MPO activities in serum that correlates with induction of genetic damage. We conclude that the serum levels of these enzymes might be used as biomarkers of early arsenic exposure induced disease much before the classical dermatological symptoms of arsenicosis begin to appear.