Evaluation of arsenic induced genotoxicity and its impact on life processes of Daphnia magna.

Heavy metals like arsenic is ubiquitously present in the environment. Geologic and anthropogenic activities are the root cause behind high concentration of arsenic in natural water bodies demanding strict monitoring of water quality prior to human consumption and utilization. In the present study, we have employed Daphnia magna for studying the biological effects of environmentally relevant high concentration of arsenic in water. In acute toxicity study, the LC50 value for 24hr exposure was found to be 2.504 mg/L, which gradually decreased with increase in time period (24hr- 96hr) to 2.011 mg/ L at 96hr. Sub-chronic toxicity was evaluated over 12 days using sub-lethal concentrations (5 %, 10 %, 15 %, and 20 % of the 24-hr LC50). Survivability in Daphnia showed a decreasing trend from 96 % to 91 % with increase in arsenic concentrations from 5 % of LC50 24 hr value to 20 % of LC 50 24hr value respectively. Alongside decreased survivability, there was a significant reduction in body size, with organisms exposed to the highest concentration of arsenic measuring 0.87±0.01 mm compared to 1.51±0.10 mm in the control group. Reproductive potential declined concentration dependently with exposure, with the highest reduction observed at 20 % of LC50 24hr value, where offspring numbers decreased to 7±1 from 23±5 in the control. Heart rate decreased in concentration and time-dependent manners, with the lowest rates observed at the highest arsenic concentration (279±16 bpm after 24hr and 277±27 bpm after 48hr). Comet assay and micronucleus assay conducted after 48 hrs of exposure revealed concentration-dependent genotoxic effects in Daphnia magna. Our results indicate negative impact on physiology and reproduction of Daphnia magna at environmentally existent concentration of arsenic. Also Daphnia magna could serve as a sensitive test system for investigating arsenic contamination in water bodies.

Effects of changed water regime on the toxicity of silver nanoparticles (AgNPs) in tadpoles of Fejervarya limnocharis.

Climate change is viewed as one of the important causes of the amphibian population decline. Aspects of climate change like increase in water temperature and drying up of habitats have been underrepresented. The expanding production and usage of metal nanoparticles like silver nanoparticles (AgNPs) make them likely to end up in aquatic ecosystems. To arrive at a realistic assessment of the impact of AgNPs in a warming world, we have investigated the effects of temperature on the acute toxicity of AgNPs in tadpoles of Fejervarya limnocharis at 24, 48, 72 and 96 h of exposure. The various aspects of sub-lethal toxicities of AgNPs with increase in temperature were also investigated. Besides, the effects of habitat desiccation on the sub-lethal toxicities of AgNPs in the tadpoles were analysed. The LC50 values of AgNPs at four different time points were found to be significantly different between the two different temperatures. Alterations in survival pattern, life history traits, amplifications in genotoxic potential and oxidative stress were observed with increased water temperature following AgNP exposure. The phenomenon of habitat desiccation was also found to significantly affect the toxicity of AgNPs with respect to alterations in mortality rate, time to metamorphosis and morphometric parameters of metamorphosed tadpoles. The findings suggest that changed water regime such as increased water temperature as well as reduction in water level accelerated the toxic effects of AgNPs in F. limnocharis tadpoles which is likely to affect their natural populations.

Toxic effects of fenvalerate in tadpoles of Fejervarya limnocharis: a multiparametric study.

Pyrethroids are among the most widely used insecticides. Fenvalerate (FEN), a synthetic pyrethroid, is frequently used in domestic and agricultural settings to control insects which ultimately find its way into the aquatic ecosystems. The larval stages of amphibians, which are experiencing a rapid population decline, are spent in aquatic habitats, thus making them vulnerable to FEN exposure. The potential toxic effects of pyrethoids in general and FEN in particular are not well understood. The present study was carried out to assess the toxicity of FEN in tadpoles of Fejervarya limnocharis. FEN at different concentrations (0, 4, 5, 6, 7, and 8 mg/L) induced substantial lethal effects. The estimated LC50 values were 8.54, 6.73, 5.44, and 4.44 mg/L at 24, 48, 72, and 96 h respectively. Exposure to environmentally relevant sub-lethal concentrations delayed metamorphosis and reduced survivality. FEN was found to be genotoxic in erythrocyte micronucleus and comet assay. Further, sub-lethal concentrations of FEN adversely affected the antioxidant defense mechanism of the exposed individuals with parallel increase oxidative damage to membrane lipids. The swimming behavior in the form of startle response, swirl response, and total movements was decreased with a concomitant decrease in AChE activity. In addition, FEN exhibited significant cardiotoxicity by decreasing the cardiac rate of the exposed individuals. The present findings clearly indicate that FEN can cause significant toxicity to the tadpoles of F. limnocharis affecting their survival and fitness in the natural environment.

A comparative account of phototoxicity of anthracene and pyrene in the tadpoles of the anuran amphibian Fejervarya limnocharis using multiple toxicological end points.

Anthracene (Anth) and pyrene (Pyr), two of the priority polycyclic aromatic hydrocarbons (PAHs), being lipophilic in nature, not only accumulate in animals, but also settle in the sediment of water bodies leading to continuous exposure for animals. Anth and Pyr when exposed to sunlight can be photoactivated and have harmful effects on aquatic organisms. A comparative analysis was carried out to assess the acute, sub-chronic, genetic and biochemical toxicity of Anth and Pyr in F. limnocharis tadpoles following short exposures to sunlight on a daily basis. In the bioaccumulation studies, it was found that both Anth and Pyr accumulated in the tadpole tissues in a concentration and time dependent manner. The LC50 values for Anth (under 15 min of daily sunlight exposure) were found to be 2.87, 2.59, 2.28, 1.80 mg/L at 24, 48, 72 and 96 h of the exposures. The corresponding LC50 values for Pyr were 1.03, 0.80, 0.62, 0.42 mg/L. Sublethal exposure of Anth and Pyr affected the survivality, time to metamorphosis as well as morphometric parameters under sunlight exposure. In the genotoxicity assessment studies, particularly the micronucleus test and comet assay, it was found that Pyr led to a higher incidence of micronucleus formation and DNA damage in comparison to Anth. The exposure to PAHs resulted in significant changes in the activity of antioxidant-mediated protective response, specifically the SOD activity, which varied between the groups treated with Anth and Pyr. On the other hand, Pyr treated group showed a higher level of GSH as compared to Anth treated groups. Moreover, the elevation in MDA level in the Anth and Pyr treated groups suggests an increase in lipid peroxidation. Future research should focus on understanding the ecotoxicological risk faced by anuran amphibia due to PAHs that frequently occur in aquatic environments and developing strategies to mitigate these risks.

Assessment of toxicity, genotoxicity and oxidative stress in Fejervarya limnocharis exposed to tributyltin.

Tributyltin (TBT) is widely used in various commercial applications due to its biocidal properties. Toxicological and genotoxicological data on TBT exposure to amphibians is insufficient. Our study aimed to determine the acute toxicity and genotoxic potential of TBT in Fejervarya limnocharis tadpoles. Furthermore, oxidative stress was also investigated in TBT-treated tadpoles. Tadpoles of Gosner stage (26-30) were screened and subjected to increasing concentrations of TBT (0, 3, 7, 11, 15, 19, 23 µg/L) for determining the LC50 values for 24 h, 48 h, 72 h, and 96 h. LC50 values of TBT for 24 h, 48 h, 72 h, and 96 h were found to be 19.45, 15.07, 13.12, and 11.84 µg/L respectively. Based on the 96 h LC50 value (11.84 µg/L), tadpoles were exposed to different sub-lethal concentrations of TBT for the evaluation of its genotoxic potential and effects on oxidative balance. The role of TBT on survivability, growth, and time to metamorphosis was also assessed. TBT exposure significantly altered the life history traits measured, increased mortality, and delayed the time taken to metamorphosis. Results indicated significant induction of micronucleus (MN, p < 0.001) and other erythrocytic nuclear aberrations (ENA, p < 0.01) in the TBT-treated groups. Significant alterations in comet parameters and oxidative balance were also observed in the treated groups. The present study findings might add to the cause of the gradual population decline seen in the amphibians. This study also demonstrates the alteration of the life-history traits, oxidative balance, and DNA damage upon TBT exposure which can have long-term consequences for the anuran amphibian F. limnocharis.

Cytotoxicity and genotoxicity of tributyltin in the early embryonic chick, Gallus gallus domesticus.

Tributyltin (TBT) is used in many commercial applications, including pesticides and antifouling paints, due to its biocidal properties. We examined the cytotoxicity and genotoxicity of TBT in the early chick embryo (Gallus gallus domesticus). Chick embryos (11 days) were treated with various doses of TBT to measure LD50 values for 24, 48, and 72 h exposures, which were determined to be 110, 54, and 18 µg/egg, respectively. The embryos were exposed to sub-lethal doses of TBT for evaluation of cytotoxicity and genotoxicity. An increase in the incidence of micronuclei (MN) was observed but it was not statistically significant. Induction of other nuclear abnormalities (ONA) after 72 h TBT exposure was significant. A significant increase in comet assay tail DNA content was also detected in TBT-exposed embryos. Cytotoxicity was also evidenced by alteration in the polychromatic erythrocytes (PCE) to normochromatic erythrocytes (NCE) ratio and by an increase in the erythroblast population in treated organisms. The cytotoxicity and genotoxicity of TBT may have long-term complications in later stages of the life cycle.

Heavy metal bioaccumulation triggers oxystress, genotoxicity and immunomodulation in head kidney macrophages of Channa punctatus Bloch.

Studies on heavy metal induced toxicity have been conducted in many water bodies across the globe and such effects have been evaluated in various fish species. The present study was designed to determine the load of some heavy metals in select sites in Southern Assam, India, along with estimating their concentration in tissues of Channa punctatus Bloch. inhabiting those niches. The effect of heavy metals in oxystress generation, genotoxicity and subsequent immune response in fish was also evaluated. In all of these sites, the concentration of Hg, Cd, Pb and Cr were above the permissible ranges while their concentrations were several folds higher in the piscine tissues due to bioaccumulation and possible biomagnification. Kidney showed the highest metal pollution index followed by liver and gills. Generation of ROS was significantly elevated and that in turn triggered oxystress, as is evident from enhanced lipid peroxidation, protein carbonylation and respiratory burst activity. These were in association with the compromised antioxidant enzyme levels with concomitant damage to DNA as evident from Comet parameters. The innate immune potential was significantly impaired as evident from the compromised cell adhesion, phagocytosis, intracellular killing activity in head kidney macrophages (HKM) along with decreased release of nitric oxide (NO) and myeloperoxidase (MPO). Immunosuppression was further validated at protein levels where compromised release of cytokines viz. TNF-α, IL-1ß, IL-6, IL-10 and IL-12 and cell signaling molecules iNOS and NF-κß were noted. Thus the present study indicates genotoxicity along with a compromise in immune status of Channa punctatus Bloch. living in a habitat laden with heavy metals.

Areca nut and smokeless tobacco exposure induces micronucleus, other nuclear abnormalities and cytotoxicity in early chick embryo.

Areca nut (AN) and smokeless tobacco (SLT) are indiscriminately consumed among the populations of Southeast and South Asian countries, even by women during the gestational period. This study aimed to investigate the genotoxic and cytotoxic potentials of AN and Sadagura (SG), a unique homemade SLT preparation, alone and in combination in early chick embryos. Fertile white leghorn chicken eggs were randomly divided into five treatment groups: vehicle control, positive control (Mitomycin C, 20 µg/egg), AN, SG, and AN+SG. AN, SG, and AN+SG were given at dosages of 0.125, 0.25, and 0.5 mg/egg. The hen's egg test for micronucleus induction (HET-MN) was performed in chick embryos to evaluate the genotoxic potential of the test agents. Furthermore, the cytotoxic potential was assessed by studying erythroblast cell populations and the polychromatic erythrocytes (PCEs) to normochromatic erythrocytes (NCEs) ratio. Our results indicated a significant increase (p < .001) in MN frequency and other nuclear abnormalities, suggesting the potential of AN and SG to cause genotoxicity. Also, AN and SG exposure alone and in combination considerably altered the erythroblast cell population (%) and the PCE to NCE ratio in all the treatment periods. Our findings established the genotoxic and cytotoxic potential of both AN and SG alone and in combination during early embryonic development in the chick embryo.

GCMS analysis of sadagura (smokeless tobacco), its enhanced genomic instability causing potential due to arsenic co-exposure, and vitamin-C supplementation as a possible remedial measure: a study involving multiple model test systems.

North-eastern states of India, including Assam, have a high prevalence of head and neck cancer cases. In these regions, Sadagura is a unique form of smokeless tobacco (SLT). There are fewer reports regarding the effects of simultaneous sadagura and arsenic co-exposure. Analysis of chemical compounds present in sadagura aqueous extract was done using gas chromatography-mass spectrometry. Estimation of arsenic contamination in groundwater and bioaccumulation in human tissues was performed by using atomic absorption spectroscopy. Buccal micronucleus cytome (BMCyt) assay and analysis of various peripheral blood parameters were performed among study volunteers. Chronic exposure (90 days) experiments were performed in mice test system in vivo to determine any possible protective potential of vitamin C (Vit-C) supplementation against sadagura and arsenic co-exposure. BMCyt assay results revealed a higher incidence of micronucleated cells (p < 0.001), and cell death biomarker among sadagura consumers residing in arsenic affected areas. Comet assay of mice femur bone marrow cells following chronic exposure of the test substances revealed a reduction in DNA damage due to Vit-C supplementation. Histological examination of the hepatic and renal tissues revealed marked improvement due to Vit-C supplementation in mice against sadagura and arsenic chronic co-exposure. Indiscriminate consumption, presence of various harmful compounds in sadagura along with arsenic co-exposure might be a vital link for the higher incidence of oral cancer in the region. Chronic Vit-C supplementation study results in mice show its effective remedial potential against combined sadagura and arsenic co-mediated genotoxicity and ultrastructural changes in major organs.

Altered expression of junctional proteins as a potential biomarker in oral precancerous and cancerous patients.

Due to a lower survival rate in patients with advanced clinical stages of oral cancer, discovering a biomarker that could diagnose and predict disease progression is vital. Cell-cell junctional proteins play a crucial role in the maintenance of tissue architecture but are often deregulated in different cancer. The present study investigates the expression of cell-cell junctional proteins viz: e-cadherin (E-cad) and zonula occludens-1 (ZO-1) in oral precancerous (OED) and cancerous (OSCC) patients to monitor if they can serve as practicable molecular markers. The ultrastructural junctional complex was studied by transmission electron microscopy, and the expression of proteins was performed by immunohistochemistry. The relationship between the expression of protein and clinicopathological features of the patients was checked by Pearson's correlation test. Furthermore, the survival curve of the follow-up data was estimated by the Kaplan-Meier method. We observed a disrupted junctional complex and a significantly decreased immunoexpression of E-cad and ZO-1 in OED and OSCC when compared to the adjacent non-cancerous tissues. The expression of ZO-1 was associated with TNM stages, whereas E-cad was associated with histological grades as well as TNM stages. A positive correlation was observed between the expression of ZO-1 and E-cad proteins in OED and OSCC. Further, follow-up studies revealed that high ZO-1 and E-cad expressing patients survived longer than their low expressed counterparts. The present study shows disruption of junctional complex and alteration of junctional proteins expression that could draw the attention of health professionals to explore junctional proteins as a possible therapeutic target in oral cancer.

JunD accentuates arecoline-induced disruption of tight junctions and promotes epithelial-to-mesenchymal transition by association with NEAT1 lncRNA.

Head and neck cancers are highly prevalent in south-east Asia, primarily due to betel nut chewing. Arecoline, the primary alkaloid is highly carcinogenic; however its role in promoting tumorigenesis by disrupting junctional complexes and increasing risk of metastasis is not well delineated. Subsequently, the effects of low and high concentrations of arecoline on the stability of tight junctions and EMT induction were studied. A microarray analysis confirmed involvement of a MAPK component, JunD, in regulating tight junction-associated genes, specifically ZO-1. Results established that although arecoline-induced phosphorylation of JunD downregulated expression of ZO-1, JunD itself was modulated by the lncRNA-NEAT1 in presence of arecoline. Increased NEAT1 in tissues of HNSCC patients significantly correlated with poor disease prognosis. Here we show that NEAT1-JunD complex interacted with ZO-1 promoter in the nuclear compartment, downregulated expression of ZO-1 and destabilized tight junction assembly. Consequently, silencing NEAT1 in arecoline-exposed cells not only downregulated the expression of JunD and stabilized expression of ZO-1, but also reduced expression of the EMT markers, Slug and Snail, indicating its direct regulatory role in arecoline-mediated TJ disruption and disease progression.

Consumption pattern and genotoxic potential of various smokeless tobacco products in Assam, India: A public health concern.

Smokeless tobacco (SLT) consumption is presumed to be one of the major causes of high incidence of oral cancer in India. The present study aimed to document various types of SLT products consumed and their potential impact on the genome instability on the population from Assam state in Northeast India. A cross-sectional study (n = 5000) showed that 60.56 % of the study population consumed at least one of the three forms (sadagura, zarda and khaini) of SLT of which 52.0 % were only sadagura users. Genotoxicity assessment using buccal cytome assay in 240 age and sex matched volunteers revealed that except for zarda, other forms of SLT induced significantly higher incidence micronuclei in the buccal epithelial cells compared to the control individuals. Similar effects were also observed in other cytome parameters related to cell proliferation, cytokinesis defects and cell death. Significantly higher incidence of micronucleus was observed among sadagura and khaini users in lymphocyte cytokinesis-blocked micronucleus assay. The addition of lime in sadagura increased the pH and anion levels which possibly result in higher absorption and may lead to the development of cellular anomalies.

Aqueous Extract of Moringa oleifera Exhibit Potential Anticancer Activity and can be Used as a Possible Cancer Therapeutic Agent: A Study Involving In Vitro and In Vivo Approach.

OBJECTIVES: New cases of cancers are increasing at an alarming rate globally. It has been hypothesized that modern cancer treatment is associated with lots of side effects and thus evoking the need to develop safer treatment measures. Thus, the present study was undertaken to evaluate the anti-carcinogenic potential of a highly nutricious plant "Moringa oleifera" (MO) in vitro and in vivo. METHODS: GC-MS analysis of aqueous extract of Moringa oleifera (AEMO) was employed to identify the bioactive compound present. Anti-tumor activity of AEMO was assessed in EAC (Ehrlich acites carcinoma) induced solid tumor bearing mice by analyzing tumor weight (TW) and Tumor volume (TV). To assess AEMO induced cytotoxicity, EAC and HEp-2 (Human laryngeal carcinoma) cells were treated with AEMO (0.05, 0.1, 0.25, 0.5 and 1 mg/ml) for both 48 h and 72 h and trypan blue, MTT and LDH released assay was done. Further, cell cycle assay and apoptosis assay was done in EAC cells to understand the mechanism of AEMO induced tumor regression. RESULTS: GC-MS analysis revealed the presence of quinic acid, octadecanoic acid, hexadecanoic acid (palmitic acid), α-tocopherol (Vitamin-E) and É£-sitosterol as major bioactive compounds. AEMO administration reduced the TV and TW of tumor-bearing mice and increases the life span. Side effect analysis showed that AEMO treatment did not induce significant alterations of liver and kidney function and hematological parameters. Further, in vitro cytotoxicity assays revealed that AEMO treatment induced dose and time-dependent toxicity in both the cell lines tested. Flow cytometric analysis confirmed significant induction of apoptotic cells by changing the mitochondrial membrane potential in EAC cell line. CONCLUSION: The results of the present study suggest that AEMO has immense potential to inhibit the tumor progression without affecting the normal physiology and functioning of the body and thus can be used as a cancer therapeutic agent.

Arsenic and smokeless tobacco exposure induces DNA damage and oxidative stress in reproductive organs of female Swiss albino mice.

Arsenic contamination in the groundwater of Southern Assam, India is well-documented. A specific type of smokeless tobacco (sadagura, SG) is highly prevalent among the local population. Thus, the present study is aimed to evaluate the toxicological implications of arsenic and smokeless tobacco co-exposure on the reproductive health of female mice. The estrous cycle of experimental animals was monitored for 30 days. Histopathological studies and comet assay of ovarian and uterine tissues were performed after 30 days of exposure to SG and arsenic (sodium arsenite, SA). Oxidative stress was estimated biochemically by taking tissue glutathione, lipid peroxidation (LPO), and superoxide dismutase activity as endpoints. Our findings indicated a prolonged diestrus phase in the SG + L + SA group (p < 0.001). Histopathological study revealed abnormal tissue architecture in treated groups. Comet assay study showed that SG + SA exposure significantly induced DNA damage in test animals. The elevated LPO level in the SG + SA group indicated oxidative stress generation in the reproductive tissues. The present study suggests that female reproductive organs are vulnerable to SA and SG and oxidative stress generation may be the possible mechanism behind DNA damage, impaired follicular growth, atresia, and altered estrous cycle in the mice test system.

A Review on Role of Arecoline and Its Metabolites in the Molecular Pathogenesis of Oral Lesions with an Insight into Current Status of Its Metabolomics.

Areca nut consumption is a popular habit in Southeast Asian countries. One of the important biologically active alkaloids of areca nut is arecoline, which plays a role in mediating the development of several pathologies of the primary exposure site, the oral cavity. Studies on the metabolism of arecoline revealed the formation of several metabolites which themselves might be toxic. Moreover, polymorphisms in genes encoding enzymes involved in the metabolism of arecoline might predispose an organism towards the development of oral cancer. The present review tries to accumulate all the relevant existing literature and then elucidate the molecular mechanism by which arecoline plays a role in the development of oral submucous fibrosis and oral cancer. Existing information regarding arecoline metabolism, enzymes involved in the metabolic process and biological effects of the metabolites of arecoline have also been compiled and compared to study the toxicity of metabolites with its parent compound arecoline and whether they play any role in the pathogenesis of oral cancer mediated by areca nut consumption. A repertoire of molecular targets has come up in the discussion whose expression profile is perturbed by arecoline. Construction of induction cascade from existing literature has given an idea about the process of molecular pathogenesis. The summarized and analysed data can help to determine the molecular mechanism and drug targets, which in turn could be helpful in the prevention or treatment of these pathological conditions. It also brings into light areas where further research needs to be directed.

Phenanthrene alters oxidative stress parameters in tadpoles of Euphlyctis cyanophlyctis (Anura, Dicroglossidae) and induces genotoxicity assessed by micronucleus and comet assay.

Phenanthrene (PHE), a tricyclic polycyclic aromatic hydrocarbon (PAH), is ubiquitously found in aquatic environments. It is one of the major components in PAH mixtures. It has been identified as one of the 16 priority PAHs for toxicological evaluations. PHE is reported to induce lethal and sub-lethal toxicity in various aquatic indicator organisms. However, no toxicological data of PHE in anuran amphibians could be found. Amphibian larvae (tadpoles) develop in aquatic habitats. Therefore, exposure to PHE could negatively impact their development and fitness in later periods as they move in to the terrestrial habitat following metamorphosis. In the present study, we have analyzed the effects of PHE in Euphlyctis cyanophlyctis tadpoles. PHE induced concentration-dependent lethal effects in the tadpoles. The estimated LC50 values were 16.52, 15.29, 13.69, and 12.28 mg/L at 24, 48, 72, and 96 h of exposure respectively. These LC50 values are significantly higher than the reported environmental concentration of PHE. However, the strong negative correlation (R2 = 0.997, p < 0.001) between the LC50 value and exposure time indicates that longer exposure to lower concentration may cause significant lethal effects. Besides, PHE at environmentally relevant concentrations induced significant sub-lethal toxicities. Exposure to sub-lethal concentrations was found to be genotoxic in erythrocyte micronucleus as well as comet assays. Sub-lethal concentrations of PHE significantly increased superoxide dismutase activity and tissue glutathione level as well as induced lipid peroxidation. The present findings clearly indicate that PHE is a potential threat to the early life stages of amphibians. Further investigations are necessary to ascertain the implications of these early effects during adult life stages.

Joint detection of claudin-1 and junctional adhesion molecule-A as a therapeutic target in oral epithelial dysplasia and oral squamous cell carcinoma.

Abnormal expression of claudin-1 (CLDN-1) and junctional adhesion molecule-A (JAM-A) has been described in certain malignancies but their clinical relevance is poorly understood. The present study aims to elucidate the role of CLDN-1 and JAM-A in oral epithelial dysplasia (OED) and oral squamous cell carcinoma (OSCC). Changes in the expression of these proteins were identified immunohistochemically on tissue sections from patients with OED and OSCC and compared with control. A correlation between the expression level of proteins and clinicopathological features was analyzed by Pearson's correlation χ2 test. The survival curve of the follow-up data was estimated by the Kaplan-Meier method followed by the log-rank test. CLDN-1 and JAM-A were highly expressed in OED and OSCC tissues when compared to control. Also, delocalization of CLDN-1 from the membrane to the cytoplasm to the nucleus was observed as the cell proceeds from normal to malignancy. Increased expression of CLDN-1 and JAM-A in both OED and OSCC were concomitant with histological grades. In addition, increased JAM-A was associated with perineural invasion of cancer cells. A positive correlation between the expression level of proteins was observed in OED (r = 0.733) and OSCC (r = 0.577). Kaplan-Meier analysis in patients with OSCC showed that the survival rate was lower in patients with high CLDN-1 and high JAM-A expression compared to low expressed patients. To conclude, the elevated level and delocalization of CLDN-1 and JAM-A suggest their use as tumor markers. A positive correlation between CLDN-1 and JAM-A suggests joint detection of these proteins as a future diagnostic tool in oral precancerous and cancerous conditions.

Correlation of serum lactate dehydrogenase and alkaline phosphatase in different histological grades of head and neck squamous cell carcinoma and premalignant lesions.

CONTEXT: Head neck cancer (HNCA) in North-East India accounts for 54.48% cancers at all sites, one of the highest in the country. Increased serum lactate dehydrogenase (LDH) activity is considered as a marker of cellular necrosis while serum alkaline phosphatase (ALP) is recognized as an important marker of induction of tumor cell differentiation. Considering the importance and need of biomarker in HNCA and there being no previous study on tumor markers from this highly prevalent region, we intended to examine the role of serum LDH and ALP in head and neck squamous cell carcinoma (HNSCC) and epithelial precursor lesions (EPLs) and also to find their correlation with the different histological grades of tumor. MATERIALS AND METHODS: This is a prospective observational study on patients with HNSCC and precancerous lesions attending Department of ENT of a Teaching Hospital. Serum LDH and ALP was measured in HNSCC, EPL and control groups and the results were statistically analyzed and compared using analysis of variance, Pearson correlation coefficient, and sensitivity-specificity analysis. RESULTS: Serum LDH was found to be a significant marker of HNSCC. Total serum LDH level was high in both premalignant and HNSCC cases. There was a significant correlation between serum LDH and grades of HNSCC showing highest levels of expression in moderately differentiated SCC. Elevated serum LDH was also seen in erythroleukoplakia, leukoplakia, and verrucous lesion, but serum ALP levels were not significant. CONCLUSIONS: This is the first study from this highly prevalent region of HNCA showing that serum LDH could be regarded as a biomarker for malignant and premalignant conditions of the head and neck.

Melatonin attenuates radiofrequency radiation (900 MHz)-induced oxidative stress, DNA damage and cell cycle arrest in germ cells of male Swiss albino mice.

Increasing male infertility of unknown aetiology can be associated with environmental factors. Extensive use of mobile phones has exposed the general population to unprecedented levels of radiofrequency radiations (RFRs) that may adversely affect male reproductive health. Therefore, the present study investigated the effect of RFR Global System for Mobile communication (GSM) type, 900 MHz and melatonin supplementation on germ cell development during spermatogenesis. Swiss albino mice were divided into four groups. One group received RFR exposure for 3 h twice/day for 35 days and the other group received the same exposure but with melatonin ( N-acetyl-5-methoxytryptamine) (MEL; 5 mg/kg bw/day). Two other groups received only MEL or remain unexposed. Sperm head abnormality, total sperm count, biochemical assay for lipid peroxides, reduced glutathione, superoxide dismutase activity and testis histology were evaluated. Additionally, flow cytometric evaluation of germ cell subtypes and comet assay were performed in testis. Extensive DNA damage in germ cells of RFR-exposed animals along with arrest in pre-meiotic stages of spermatogenesis eventually leading to low sperm count and sperm head abnormalities were observed. Furthermore, biochemical assays revealed excess free radical generation resulting in histological and morphological changes in testis and germ cells morphology, respectively. However, these effects were either diminished or absent in RFR-exposed animals supplemented with melatonin. Hence, it can be concluded that melatonin inhibits pre-meiotic spermatogenesis arrest in male germ cells through its anti-oxidative potential and ability to improve DNA reparative pathways, leading to normal sperm count and sperm morphology in RFR-exposed animals.

Effect of nutritional status on arsenic and smokeless tobacco induced genotoxicity, sperm abnormality and oxidative stress in mice in vivo.

BACKGROUND: Recently, high concentrations of arsenic have been documented in ground waters of Southern Assam, India. Indiscriminate smokeless tobacco consumption is a common practice in this region. Correlation between nutritional status and arsenic and smokeless tobacco-induced health effects has not been taken up in humans or other test systems. METHODS: Mice were divided into groups based on protein (casein) content in the diet: High protein (40%), optimum protein (20%), and low protein (5%). Simultaneous chronic exposure (90 days) to arsenic and smokeless tobacco (sadagura) orally was given to evaluate the extent of the cytological and genotoxicological damage. Micronucleus assay and Comet assay of the femur bone marrow cells were conducted. Germ cell toxicity was evaluated by recording the sperm head abnormalities and total sperm count. Cell cycle analysis was performed in femur bone marrow cells using flow cytometer. Hepatic, renal, and intestinal tissues were analyzed for various oxidative stress evaluations. Histological examination of liver and kidney was performed. RESULTS: Notably, high protein diet groups had lower arsenic and sadagura induced genotoxicity, germ cell abnormalities and oxidative stress as compared to optimum protein and low protein diet counterparts. CONCLUSION: Our study indicates that sufficient levels of dietary protein appear to reduce the long-term arsenic and smokeless tobacco-induced toxicity in mice test system, as compared to lower or deficient amount of protein in the diet. This observation has implications and invites further studies especially epidemiological studies in the human population exposed to arsenic in South East Asian countries. Environ. Mol. Mutagen. 59:386-400, 2018. © 2018 Wiley Periodicals, Inc.