Sheikh Raisuddin

Oxidative stress biomarkers of exposure to deltamethrin in freshwater fish, Channa punctatus Bloch

The pyrethroid class of insecticides, including deltamethrin, are being used as substitutes for organochlorines and organophosphates in pest-control programs because of their low environmental persistence and toxicity. Ecotoxicological consequences of deltamethrin, particularly its effects on antioxidants in fish and other aquatic organisms, are not well understood. We investigated the effect of deltamethrin (0.75 microg/L) on antioxidants in a freshwater fish, Channa punctatus Bloch, using standard laboratory conditions. A single exposure for 48 h caused induction of various antioxidant enzymes and nonenzymatic antioxidants in kidney and liver. The induction of these antioxidants was not very prominent in gills. In fact, certain antioxidants were found to be depleted in gills. Catalase activity was decreased in all the tissues. Deltamethrin also induced lipid peroxidation in all the tissues, gills showing the highest levels. Glutathione, which is an established nonenzymatic antioxidant in fish, was significantly (P<0.001) increased in all the tissues. Ascorbic acid content increased in kidney and liver while it decreased in gills. The findings of the present investigation show that deltamethrin has oxidative-stress-inducing potential in fish, and gills are the most sensitive organs. It is also interesting to note that gills are the primary sites of deltamethrin absorption and their antioxidant potential is also very poor. The various parameters studied in this investigation can also be used as biomarkers of exposure to deltamethrin. It is suggested that appropriate ecotoxicological risk assessment should be made in the areas where deltamethrin is proposed to be used in pest control activities.

Biomarkers of oxidative stress: a comparative study of river Yamuna fish Wallago attu (Bl. & Schn.)

Various oxidative stress biomarkers in gill, kidney and liver tissues in the Indian freshwater fish Wallago attu (Bl. & Schn.) were investigated. Fish were collected from two sites along the river Yamuna, which differ in their extent and type of pollution load. A comparison was made between the biomarker responses and general water chemistry at the two sites. The oxidative stress biomarkers that were analyzed included superoxide dismutase (SOD), catalase (CAT), xanthine oxidase (XOD) and glutathione redox cycle enzymes viz., glutathione peroxidase (GPx), glutathione reductase (GR) and glucose 6-phosphate dehydrogenase (G6PD). Levels of reduced glutathione (GSH) and lipid peroxidation (LPO) were also evaluated. All biomarkers; SOD (P<0.001 in liver, kidney and gill), XOD (P<0.01 in kidney and P<0.001 in liver and gill), GR (P<0.01 in liver, P>0.05 in kidney and P<0.001 in gill), G6PD (P<0.001 in liver, P>0.05 in kidney and P<0.01 in gill), GSH (P<0.001 in liver, kidney and gill) and LPO (P>0.05 in liver, kidney and gill) were found to be substantially higher in the fish collected from Panipat when compared with values in tissues of fish collected from Agra site. GPx and CAT showed a varied response. GPx activity was higher (P<0.001) in gills and kidney of the fish collected at Panipat site. However, liver showed significant low values (P<0.01) when compared with Agra site values. CAT activity was found to be significantly low, in both liver (P<0.01) and kidney (P<0.001) whereas in gills non-significant (P>0.05) low values were observed. Water chemistry data at two sites indicated that Panipat site with higher biochemical oxygen demand, chemical oxygen demand, pH and low dissolved oxygen was comparatively more polluted than Agra site. Industrial activity profile of both the sites also indicates that Panipat has vigorous industrial activity coupled with intensive use of chemicals in agricultural practices in Haryana state. The findings of the present investigation provide a rational use of oxidative stress biomarkers in aquatic ecosystem pollution biomonitoring. This is also the first such attempt reported from India.

Induction of hepatic antioxidants in freshwater catfish (Channa punctatus Bloch) is a biomarker of paper mill effluent exposure

Enzymatic and non-enzymatic antioxidants serve as an important biological defense against environmental oxidative stress. Information on antioxidant defense in fish is meager despite that fish are constantly exposed to a myriad of environmental stress including the oxidants. This study, therefore, assesses the activities of antioxidant enzymes viz., glutathione peroxidase, catalase and glutathione S-transferase and the non-enzymatic antioxidants viz., glutathione and metallothionein in various tissues of freshwater fish Channa punctatus (Bloch), in response to short-term and long-term exposures to paper mill effluent. The fish were exposed to the effluent at a concentration of 1.0% (v/v) for 15, 30, 60 and 90 days. The exposure caused a time-dependent increase in glutathione level (P < 0.001), activities of glutathione peroxidase (P < 0.05 to P < 0.001), glutathione S-transferase (P < 0.001) and a marginal initial decrease in catalase activity in the liver (P < 0.01 to P < 0.001). Metallothionein was induced in liver after 60 days of exposure. Two isoforms of metallothionein were detected. Catalase activity also increased 60 days afterwards. Antioxidant pattern was different in gill and kidney showing that liver was more resistant to oxidative damage as compared to gills and kidney. Our results demonstrate a pollutant-induced adaptive response in fish. In addition, levels of enzymatic and non-enzymatic tissue antioxidants may serve as surrogate markers of exposure to oxidant pollutants in fish.

Effects of exposure to multiple trace metals on biochemical, histological and ultrastructural features of gills of a freshwater fish, Channa punctata Bloch

The trace metals are frequently encountered as mixtures of essential and non-essential elements. Therefore, evaluation of their toxic effects individually does not offer a realistic estimate of their impact on biological processes. We studied effects of a mixture of four essential and toxic metals (Cu, Cd, Fe and Ni) on biochemical and morphological characteristics of the gills of a biomarker freshwater fish Channa punctata (Bloch) using environmentally relevant concentrations. Fish were exposed to metal mixture through tank water for 7, 15 and 30 days. Biochemical studies as well as light microscopy (LM) and scanning electron microscopy (SEM) revealed significant metal exposure-induced alterations in gills. Besides ultastructural changes, activities of antioxidant enzymes such catalase (CAT), glutathione S-transferase (GST) and superoxide dismutase (SOD) were significantly altered in the gills of exposed fish. The reduced glutathione (GSH) was significantly (p<0.001) decreased, while lipid peroxidation (LPO) was significantly (p<0.001) increased. The main alterations in general morphology of fish gills included spiking and fusion of secondary lamellae, formation of club-shaped filaments, and vacuolization and necrosis of filament epithelium in the interlamellar regions. SEM studies showed gradual increase of the density and apical surface area of the chloride cells and transformation of the surface structure of the pavement cells. The results of this study indicate adaptive as well a toxic responses in fish gills exposed to mixture of trace metals. Low concentrations of trace metal appear to compromise the antioxidant defense of gills. Lesions in the gill morphology caused by the effect of low concentrations of trace metals could lead to functional alterations and interference with fundamental processes such as maintenance of osmoregulation, gas exchange and xenobiotic metabolism in the exposed fish populations.

Protein carbonyls: novel biomarkers of exposure to oxidative stress-inducing pesticides in freshwater fish Channa punctata (Bloch).

It has been established in mammalian system including humans that direct damage to proteins or chemical modification of amino acids in proteins during oxidative stress can give rise to protein carbonyls. Protein carbonyl induction, as a biomarker of oxidative stress was used in laboratory studies to assess the toxic effects of pesticides in freshwater fish, Channa punctata (Bloch), exposed to deltamethrin, endosulfan and paraquat. Protein carbonyls were measured in gills, kidney and liver. Significant (P<0.05-0.001) increase in protein carbonyls was observed in response to single 48h exposure to various pesticides in all the tissues. The time kinetics study involving deltamethrin (0.75μg/L) also showed a significant (P<0.05-0.001) induction of protein carbonyls in all the organs. The induction was significant (P<0.05-0.001) in all the durations of exposure (12h, 96h, 7 days, 14 days, 28 days). However, relatively pronounced induction was observed during shorter duration of exposure. The findings of the present investigation showed that deltamethrin had the maximum oxidative stress-inducing potential among the three pesticides used and gills are the most sensitive organs prone to oxidative damage. It is suggested that measurement of carbonyl groups may provide a convenient technique for detecting and quantifying oxidative modification of proteins during oxidative stress. The induction of protein carbonyl in fish was identified as a potentially useful biomarker of oxidative stress that warrants its application in the field investigations.

IMMUNOMODULATORY EFFECTS OF FENUGREEK (TRIGONELLA FOENUM GRAECUM L.) EXTRACT IN MICE

Immunomodulatory activity of aqueous extract of Trigonella foenum graecum L., a widely used medicinal and dietary herb, was evaluated in male Swiss albino mice. Mice were treated with three doses of extract (50, 100 and 250 mg/kg body weight per os) for 10 days. Body weight, relative organ weight, cellularity of lymphoid organs, delayed type of hypersensitivity (DTH) response, plaque-forming cell (PFC) assay, haemagglutination titre (HT), quantitative haemolysis of SRBC (QHS) assay, phagocytosis, and lymphoproliferation were studied in various groups of animals. At doses of 50 and 100 mg/kg, a significant increase (p < 0.05) in relative organ weight of thymus was observed but there was no effect on kidney and spleen weights. Liver weight also increased significantly at doses of 100 and 250 mg/kg. However, no elevation in the levels of liver function test (LFT) enzymes was observed. As regards lymphoid organ cellularity, spleen recorded no significant increase at any dose, whereas cellularities of thymus and bone marrow were significantly increased. T. foenum graecum extract elicited a significant (p < 0.001) increase in the DTH response at doses of 50 and 100 mg/kg, but the change at higher dose of 250 mg/kg was not statistically significant. Humoral immunity as measured by PFC showed an elevated response at a dose of 100 mg/kg, but at 50 and 250 mg/kg, no significant effect was observed. In the HT test, plant extract also showed modulatory effect at all the doses. Plant extract elicited a significant increase in phagocytic index and phagocytic capacity of macrophages. Stimulatory response of plant extract was also observed in lymphoproliferation assay but the response was weak. Overall, T. foenum graecum showed a stimulatory effect on immune functions in mice. As it is used for a variety of medicinal purposes, its immunostimulatory effect, as reported in this study, strengthens the rationale of its use in several Ayurvedic and Unani drugs.

Beneficial effects of lamivudine in hepatitis B virus-related decompensated cirrhosis

BACKGROUND/AIMS HBV-related chronic liver disease patients often present with hepatic decompensation and are not eligible for interferon therapy. Whether long-term lamivudine is effective in these patients was prospectively evaluated. METHODS Eighteen patients with HBV-related decompensated cirrhosis, all with quantitative DNA +ve and 10 HBeAg +ve, were given lamivudine 150 mg/d. RESULTS Each patient received at least 9 months (mean 17.9) of lamivudine. Three HBeAg+ve patients (30%) seroconverted to anti-HBe and one lost HBsAg during the follow-up. An improvement from baseline in the aspartate aminotransferase (130 vs. 72 IU/l, p<0.04); alanine aminotransferase (111 vs. 58 IU/l, p<0.01) and Child-Pugh score (8.3 vs 6.7, p<0.013) was seen. Lamivudine had no significant side-effects. HBV DNA became undetectable in all patients by 8 weeks of therapy. In three (17%) patients, HBV DNA again became positive at 9, 9 and 27 months. YMDD mutant was, however, detected in only one (6%). A significant reduction was noted in the morbidity and hospitalizations for complications of liver disease before and after starting lamivudine (1.5+/-0.7 vs. 0.6+/-0.7, p<0.002). CONCLUSIONS In decompensated HBV-related cirrhosis, lamivudine: i) is effective in suppressing HBV DNA and seroconversion to anti-HBe (30%), ii) can achieve significant improvement in clinical and biochemical status of liver functions.

Pollutant-induced over-activation of phagocytes is concomitantly associated with peroxidative damage in fish tissues

Pollutant-induced abnormal functioning of phagocytes and associated consequences were studied in freshwater catfish Heteropneustes fossilis (Bloch). Fish were exposed to effluent collected from bleached kraft type of paper mill at the concentration levels of 0.5, 1 and 2% for 15, 30, 60 and 90 days. Respiratory burst activity of peritoneal and head kidney phagocytes of exposed fish was measured by nitroblue tetrazolium reduction assay. Lipid peroxidation (LPO) was estimated in liver, gill and kidney of fish by measuring thiobarbituric acid reaction substances. It was observed that the phagocyte-activating xenobiotics of effluent induced an increase in the respiratory burst activity in phagocytes. The induction of respiratory burst activity was concomitantly associated with an increase in the peroxidative damage of tissues. The tissues most affected were kidney and gills. The change in LPO values in the gills of exposed fish was concentration- and time-dependent, showing significant increases (P<0.05 to <0.001) in all the exposed groups as compared with control fish. An almost similar pattern of LPO was observed in head kidney tissue (P<0.05 to <0.001). As regards liver, increase in LPO was not widespread, except at 0.5% for 90 days (P<0.05). In fact, reduced rates of LPO were observed in the livers of some groups. The results of respiratory burst corroborate with the phagocytic activation as well as with the extent of lipid peroxidation in the tissues, showing high population of circulatory phagocytes. Our results demonstrate that fish of polluted water are subjected to oxidative stress of multifarious dimensions.

Heat shock protein (Hsp) gene responses of the intertidal copepod Tigriopus japonicus to environmental toxicants.

The induction of heat shock proteins (Hsps) is considered as an important protective, ecophysiologically adaptive, and genetically conserved response to environmental stress in all organisms. Among the Hsps, the heat shock protein 70 (Hsp70) family members are most extensively studied for their characterization and induction in response to environmental stressors in a range of species. We studied expression of ten Hsp transcripts in response to heat treatment in an intertidal marine copepod Tigriopus japonicus and observed that expression of Hsp70 was more pronounced than other Hsps. Subsequently, cDNA and genomic sequences of T. japonicus Hsp70 (TJ-Hsp70) were worked out by molecular cloning techniques and phylogenetic relationship was analyzed. The bacterial expression of TJ-Hsp70 and its expression in response to metal and endocrine-disrupting chemical (EDC) exposures were also studied. The TJ-Hsp70 transformed bacteria showed increased thermotolerance compared to bacteria with vector only. All the trace metals (i.e. copper, silver, and zinc) caused a concentration-dependent increase in the expression of Hsp70 transcripts. Effect of EDCs on Hsp70 expression was differential. While 4-nonylphenol (NP) and 4-t-octylpheno (OP) caused downregulation, bisphenol A (BPA) caused upregulation. The promoter region of the genomic Hsp70 sequence contained putative xenobiotic response elements (XREs) indicating that TJ-Hsp70 regulation not only by temperature but also by xenobiotics. These findings suggest that in T. japonicus, Hsp70 has a conserved role of thermotolerance and its expression in response to xenobiotics exposure appears to be a protective response.

Expression of glutathione S-transferase (GST) genes in the marine copepod Tigriopus japonicus exposed to trace metals.

The intertidal copepod, Tigriopus japonicus has been recognized as a potential model species for marine pollution toxicity testing. Toxicity ranges of several biocides, endocrine-disrupting chemicals (EDCs), and trace metals are known in T. japonicus. A large number of expressed sequence tags (ESTs) and genomic DNA are also sequenced from T. japonicus. In this study, expression of ten glutathione S-transferase (GST) genes was studied in the copepods exposed to trace metals. Expression of these genes was also studied against exposure to hydrogen peroxide (H(2)O(2)) used as a positive control with prooxidant activity. Of all genes, expression of GST-Sigma (GSTS) was highly upregulated in H(2)O(2) as well as trace metal-exposed copepods. In the time-course study, expression of GSTS mRNA was more consistent compared to other GSTs such as GST-Omega, GST-Delta1, GST-Theta3 or microsomal GST1 (mGST1). GSTS is predominantly reported from the insects. Coupled with the previous study of the in vitro antioxidant role of T. japonicus GSTS, these findings imply an antioxidant role for GSTS and highlight its importance as a biomarker of exposure to trace metals in T. japonicus. However, further validation and field trials would be necessary to propose GSTS gene expression as biomarker of exposure to trace metals, as for some trace metals such as silver the response was not consistent in concentration and time-series exposure experiments.