Delineating involvement of MAPK/NF-κB pathway during mitigation of permethrin-induced oxidative damage in fish gills by melatonin.

Present study evaluated involvement of transcription factors during permethrin-induced gill toxicity and its amelioration by melatonin. First, adult Notoptertus notopterus females were exposed to permethrin at nominal concentrations [C: 0.0, P1: 0.34, P2: 0.68 µg/L] for 15 days followed by intramuscular melatonin administration (100 µg/kg body weight) for 7 days. Gill MDA, XO, LDH levels increased, while Na+-K+-ATPase, SDH, cytochrome C oxidase levels decreased with increasing permethrin concentrations. Glutathione, SOD, CAT, GST, GRd levels increased in P1 than C, but decreased in P2 than P1, C. Melatonin administration restored gill enzyme and antioxidant levels in P1, P2. Next, isolated gill tissues were exposed to permethrin at 25, 50 µM doses along with melatonin administration (100 µg/mL). NF-κB, NRF2, Keap1, ERK, Akt, caspases protein expression changed significantly during permethrin-induced gill damage. Melatonin administration amended permethrin-induced molecular imbalance through modulation of caspase proteins and MAPK/NF-κB signal transduction pathway via melatonin receptor 1.

Cardiac RGS7 and RGS11 drive TGFß1-dependent liver damage following chemotherapy exposure.

Off target damage to vital organ systems is an unfortunate side effect of cancer chemotherapy and remains a major limitation to the use of these essential drugs in the clinic. Despite decades of research, the mechanisms conferring susceptibility to chemotherapy driven cardiotoxicity and hepatotoxicity remain unclear. In the livers of patients with a history of chemotherapy, we observed a twofold increase in expression of G protein regulator RGS7 and a corresponding decrease in fellow R7 family member RGS11. Knockdown of RGS7 via introduction of RGS7 shRNA via tail vein injection decreased doxorubicin-induced hepatic collagen and lipid deposition, glycogen accumulation, and elevations in ALT, AST, and triglycerides by approximately 50%. Surprisingly, a similar result could be achieved via introduction of RGS7 shRNA directly to the myocardium without impacting RGS7 levels in the liver directly. Indeed, doxorubicin-treated cardiomyocytes secrete the endocrine factors transforming growth factor ß1 (TGFß1) and TGFß superfamily binding protein follistatin-related protein 1 (FSTL1). Importantly, RGS7 overexpression in the heart was sufficient to recapitulate the impacts of doxorubicin on the liver and inhibition of TGFß1 signaling with the receptor blocker GW788388 ameliorated the effect of cardiac RGS7 overexpression on hepatic fibrosis, steatosis, oxidative stress, and cell death as well as the resultant elevation in liver enzymes. Together these data demonstrate that RGS7 controls both the release of TGFß1 from the heart and the profibrotic and pro-oxidant actions of TGFß1 in the liver and emphasize the functional significance of endocrine cardiokine signaling in the pathogenesis of chemotherapy drive multiorgan damage.

Abiotic factors and heavy metals defining eco-physiological niche in fish.

Ecosystem dynamics undergoing alterations in structure and function highlights the need to look into the relations between ecological parameters and organismal fitness and tolerance. Ecophysiological studies are used to understand how organisms adapt to and cope up with environmental stress. Current study uses a process-based approach to model physiochemical parameters regarding seven different fish species. Species respond to climatic variations via acclimation or adaptation through physiological plasticity. Four sites are differentiated into two types based on the water quality parameters and metal contamination. Seven fish species are clustered into two groups, each group depicting separate pattern of response in similar habitat. In this manner, biomarkers from three different physiological axes- stress, reproduction, and neurology were taken to determine the organism's ecological niche. Cortisol, Testosterone, Estradiol, and AChE are the signature molecules estimated for the said physiological axes. The ordination technique, nonmetric multidimensional scaling, has been utilized to visualize the differentiated physiological response to changing environmental conditions. Then, Bayesian Model Averaging (BMA) was used to identify the factors that play a key role in refining the stress physiology and determining the niche. Current study confirms different species belonging to similar habitats respond to various environmental and physiological factors in a different manner as various biomarkers respond in a species-specific pattern that induces the choice of habitat preference controlling its ecophysiological niche. In the present study, it is quite apparent that adaptive mechanism of fish to environmental stress is achieved through modification of physiological mechanisms through a panel of biochemical markers. These markers organize a cascade of physiological event at various levels including reproduction.

RGS11-CaMKII complex mediated redox control attenuates chemotherapy-induced cardiac fibrosis.

Dose limiting cardiotoxicity remains a major limiting factor in the clinical use of several cancer chemotherapeutics including anthracyclines and the antimetabolite 5-fluorouracil (5-FU). Prior work has demonstrated that chemotherapeutics increase expression of R7 family regulator of G protein signaling (RGS) protein-binding partner Gß5, which drives myocyte cytotoxicity. However, though several R7 family members are expressed in heart, the exact role of each protein in chemotherapy driven heart damage remains unclear. Here, we demonstrate that RGS11, downregulated in the human heart following chemotherapy exposure, possesses potent anti-apoptotic actions, in direct opposition to the actions of fellow R7 family member RGS6. RGS11 forms a direct complex with the apoptotic kinase CaMKII and stress responsive transcription factor ATF3 and acts to counterbalance the ability of CaMKII and ATF3 to trigger oxidative stress, mitochondrial dysfunction, cell death, and release of the cardiokine neuregulin-1 (NRG1), which mediates pathological intercommunication between myocytes and endothelial cells. Doxorubicin triggers RGS11 depletion in the murine myocardium, and cardiac-specific OE of RGS11 decreases doxorubicin-induced fibrosis, myocyte hypertrophy, apoptosis, oxidative stress, and cell loss and aids in the maintenance of left ventricular function. Conversely, RGS11 knockdown in heart promotes cardiac fibrosis associated with CaMKII activation and ATF3/NRG1 induction. Indeed, inhibition of CaMKII largely prevents the fibrotic remodeling resulting from cardiac RGS11 depletion underscoring the functional importance of the RGS11-CaMKII interaction in the pathogenesis of cardiac fibrosis. These data describe an entirely new role for RGS11 in heart and identify RGS11 as a potential new target for amelioration of chemotherapy-induced cardiotoxicity.

Melatonin ameliorates lipopolysaccharide induced brain inflammation through modulation of oxidative status and diminution of cytokine rush in Danio rerio.

Lipopolysaccharide (LPS) is known to induce inflammation and immunonomodulation in a piscine model of Danio rerio. Present study aimed to explore the ability of melatonin in attenuating LPS-induced oxidative damages using this model. In LPS-exposed fish, activation of stress marker MDA was observed in brain with corresponding augmentation of multiple pro-inflammatory cytokines (IL1ß, IL6, IL10 and TNFα). In addition, it also showed marked increase in the levels of heat shock factor (HSF) and heat shock proteins (HSPs) in association with transcription factors (NF-kB and NRF2) and mitogen-activated protein kinases (MAPKs). The changes in the levels of these mediators are highly correlated with the induction of pro-inflammatory cytokines. In melatonin-treated fishes, significant amelioration of oxidative stress was observed with reduced levels of MDA and pro-inflammatory cytokines. Melatonin also modulated expression of HSPs that facilitated the brain to overcome inflammation-induced stress by directly initiating NFkB/NRF2 translocation. In summary, melatonin effectively functions to reduce stress induced inflammatory signalling through modulation of oxidative stress and protects the brain from the neuropathological insult.

Effects of salinity stress on antioxidant status and inflammatory responses in females of a "Near Threatened" economically important fish species Notopterus chitala: a mechanistic approach.

In the present study, acute stress responses of adult female Notopterus chitala were scrutinized by antioxidant status and inflammation reaction in the gill and liver at five different salinity exposures (0, 3, 6, 9, 12 ppt). Oxidative defense was assessed by determining superoxide dismutase (SOD), catalase (CAT), glutathione S-transferase, and glutathione reductase activities, while malondialdehyde (MDA), glutathione, and xanthine oxidase levels were determined as indicators of oxidative load. Pro-inflammatory cytokines (IL-1ß, IL-6, IL-10, and TNFα) and caspase 1 levels were also analyzed. Expression levels of transcription factors (NRF2 and NF-κB) and molecular chaperons (HSF, HSP70, and HSP90) were estimated to evaluate their relative contribution to overcome salinity stress. MDA showed a significant (P < 0.05) increase (gill, + 25.35-90.14%; liver, + 23.88-80.59%) with salinity; SOD (+ 13.72-45.09%) and CAT (+ 12.73-33.96%) exhibited a sharp increase until 9 ppt, followed by a decrease at the highest salinity (12 ppt) (gill, - 3.92%; liver, - 2.18%). Levels of cytokines were observed to increase (+ 52.8-127.42%) in a parallel pattern with increased salinity. HSP70 and HSP90 expressions were higher in gill tissues than those in liver tissues. NRF2 played pivotal role in reducing salinity-induced oxidative load in both the liver and gills. Serum cortisol and carbonic anhydrase were measured and noted to be significantly (P < 0.05) upregulated in salinity stressed groups. Gill Na+-K+-ATPase activity decreased significantly (P < 0.05) in fish exposed to 6, 9, and 12 ppt compared to control. Present study suggests that a hyperosmotic environment induces acute oxidative stress and inflammation, which in turn causes cellular death and impairs tissue functions in freshwater fish species such as Notopterus chitala.

Evaluating the role of dietary plant extracts to allow adaptation to thermal stress in a cold stream ornamental fish, Botia rostrata (Günther, 1868).

Increases in ambient temperature affect the biochemical status of fish, and dietary supplementation with bioactive phytoconstituents may promote resilience against environmental stress. This study evaluated the impact of three plant extracts on the biochemical status of a cold stream fish Botia rostrata (Günther, 1868) under high temperatures. After 1 month dietary supplementation separately with Mucuna pruriens methanol extract (0.25 g/kg feed), Tribulus terrestris ethanol extract (0.5 g/kg feed) and Basella alba ethanol extract (1.0 g/kg feed), juvenile fish (Wt. 4.3 ± 0.5g) were exposed to different sublethal heat stress [28 ± 0.5 °C (T1), 32 ± 0.5 °C (T2), 36 ± 0.5 °C (T3)]. Control fish were fed a diet without any plant extract and maintained at 24 ± 0.5 °C. Serum and muscle tissues were collected to measure different biochemical parameters, muscle metabolic enzymes and molecular chaperons before and after heat stress. Before stress, the group fed the Mucuna diet showed significant (P < 0.05) increases in serum glucose [+10.92%], protein [+18.93%], muscle heat shock protein (HSP) 90 [+8.6%] compared to the control group. No significant change (P > 0.05) of stress parameters was observed between control, Tribulus and Basella fed fish. The control group exposed to T3 showed significant differences (P < 0.05) in protein [-26.19%], lactate dehydrogenase [+93.69%], fructose 1,6 bisphosphate [-35.19%], phosphorylase 'a' [+35.72%], HSP60 [+69.54%], HSP70 [+84.85%], HSP90 [+92.07%], heat shock factor (HSF) 1 [+88.48%] suggesting susceptibility of Botia to this temperature. Among the three plant extracts, Mucuna methanol extract was effective to enhance resistance against temperature-induced biochemical alterations. After exposure to T3, only the fish fed Mucuna diet showed no mortality. Fish fed Mucuna diet exposed to 36 ± 0.5 °C showed significantly higher (P < 0.05) glucose [+42.82%], protein [+11.98%], citrate synthase [+59.81%], phosphorylase 'a' [+14.96%], Glucose 6 phosphate dehydrogenase [+60.87%], HSP60 [+34.13%], HSP70 [+41.42%], HSP90 [+65.91%], HSF1 [+61.32%] compared to those in Mucuna fed fish maintained at 24 ± 0.5 °C. These results highlight temperature-induced biochemical alterations in Botia and point towards the potential use of Mucuna in overcoming such adverse high thermal stress.

Organometallic Folate Gold Nanoparticles Ameliorate Lipopolysaccharide-Induced Oxidative Damage and Inflammation in Zebrafish Brain.

Synthesized organometallic gold-based folate nanoparticles (FAuNPs) were characterized, and its defense against lipopolysaccharide (LPS)-induced brain inflammation in Zebra fish was proven. Vitamin entrapment efficiency of these particles was found to be nearly 70%. The in vitro pH-dependent drug release dialysis study of FAuNPs confirmed a slow, sustained, and gradual release of folate for a period of 24 h. Both AuNPs and FAuNPs did not cause any marked changes in food intake, body weight, color, behavioral pattern, blood parameters, and hepatotoxicity. Histology of liver showed no changes between treated and control groups of fishes. The ex vivo study showed significant uptake of FAuNPs to free folate in folate receptor negative Hek293 cells, confirming a strategy to overcome folate deficiency in the brain. Antioxidant status and activities of few crucial brain enzymes were also measured to assess the brain function and found to be returned to the basal level, following FAuNP treatment. The transcription factor NRF2-Keap 1 expression pattern was also noted, and a prominent modulation was observed in the LPS-treated and FAuNP-administered group. Decisive brain enzymes like AChE and Na+K+ATPase were decreased significantly after LPS treatment, which is restored with FAuNP treatment. Caspases increased sharply after LPS treatment and diminished following FAuNP treatment. We conclude that FAuNP due to its high physical stability and uptake could be utilized against severe brain inflammation, leading to brain injury and neurodegeneration.

Crosstalk between heat shock proteins, NRF2, NF-κB and different endogenous antioxidants during lead-induced hepatotoxicity in Puntius ticto.

Present study aims to investigate interaction of molecular chaperons (heat shock protein 70, heat shock protein 90) with transcriptional factors (nuclear factor kappa B/nuclear factor E2-related factor 2/Kelch-like ECH-associated protein 1) to evaluate their role during metal induced stress in fish hepatocytes. Adult Puntius ticto were exposed to lead nitrate at 0 mg/l (control), 1/50th (0.04 mg/l) and 1/20th (0.12 mg/l) of LC50 for 30 days and sacrificed to collect liver tissues. Activity of selected liver enzymes, antioxidants and metallothionein were analyzed. Levels of heat shock protein 70, heat shock protein 90, nuclear factor kappa B, nuclear factor E2-related factor 2 and Kelch-like ECH-associated protein 1 were also measured. Liver enzymes showed a significant increase (p < 0.05) in both Pb exposed groups indicating that the liver might be at risk of damage. Increased level of lipid peroxidation due to metal stress was marked by significant increase (p < 0.05) in malondialdehyde level in fish exposed to the higher Pb concentration compared to control (+ 13.7 %). Significant increase (p < 0.05) in gluthathione reductase (+ 35 %, + 39.2 %), glutathione s-transferase (+ 22.4 %, + 50.4 %) activities and decrease in reduced glutathione level (- 6.75 %, - 12.25 %) in fish exposed to both lower and higher Pb concentration compared to control also indicated metal induced oxidative damage in fish liver. Super oxide dismutase and catalase activities increased significantly (p < 0.05) during exposure to lower Pb concentration, while decreased significantly (p < 0.05) during exposure to higher Pb concentration compared to those in control. Significant (P < 0.05) increase (+ 52.63 %, + 89.47 %) in metallothionein in Pb exposed groups confirmed its role in detoxification process of the metal. Heat shock protein 70 and heat shock protein 90 expression levels increased significantly (p < 0.05) during metal exposure indicating their role as modulator of stress-induced antioxidant protein remodelling. A positive correlation between nuclear factor kappa B/nuclear factor E2-related factor 2/Kelch-like ECH-associated protein 1 with gluthathione regulatory enzymes (gluthathione reductase and glutathione s-transferase) was noted. Current study effectively illuminates the critical role of different factors (heat shock proteins/nuclear factor kappa B/nuclear factor E2-related factor 2/Kelch-like ECH-associated protein 1) to influence the expression and synthesis of antioxidants and other functional enzymes in lead-exposed fish liver.

Consequences of sodium dodecyl sulfate exposure on the antioxidant status and steroidogenesis in fish gonad.

A native Indian catfish (Heteropneustes fossilis) and an Indian major carp (Cirrhinus cirrhosus) were treated with sodium dodecyl sulfate (SDS) to elucidate its consequences on antioxidant status and sex steroid synthesis. Fishes were exposed to 1/10th of LC50 of SDS for 30 days. Different enzymatic and non-enzymatic antioxidants, major sex steroids, steroidogenic enzymes, gonadosomatic index (GSI), and serum cortisol level were measured in both male and female fishes. Levels of enzymatic and non-enzymatic antioxidants altered significantly in both the fish species. Steroid hormones in both male and female fishes decreased significantly in the SDS-treated group. Profiles of steroidogenic enzymes (hydroxysteroid dehydrogenase; 3ß-HSD and 17ß-HSD) show significant decrease in the SDS treatment group in both carp and catfish than that of the control group irrespective of sexes. GSI also decreases significantly in the SDS-treated group when compared with the control group in spawning phase of carp and catfish irrespective of sexes. Serum cortisol level was increased significantly after SDS exposure. Higher level of cortisol was noted in the spawning phase than that of pre-spawning phase. Current study proves that SDS could interrupt the normal steroidogenesis process in fish, leading to impaired sex hormone production. Detrimental effect of SDS was noted to be more prominent on female steroidogenesis than that of male fish. A species or season-specific variation in steroidogenesis was also observed which indicates that SDS-induced toxicity depends on the sexual status of the specific fish species.

Enzymatic, non enzymatic antioxidants and glucose metabolism enzymes response differently against metal stress in muscles of three fish species depending on different feeding niche.

Current study aims to determine difference in metal accumulation pattern in muscle of Liza parsia (pelagic, omnivore), Amblypharyngodon mola (surface feeder, herbivore) and Mystus gulio (benthic, carnivore) depending on their niche and feeding habit and how it affects the endogenous antioxidants and glucose metabolism in fish muscle. Fishes were collected from Malancha, Diamond Harbour and Chandanpiri, West Bengal, India. Concentrations of lead, zinc, cadmium, chromium were measured in water, sediment and fish muscle. Metal pollution index (MPI) and bioconcentration factor (BCF) was calculated to evaluate the ability of fish to accumulate specific metals in muscle tissue from the aquatic environment. Metal concentrations were found significantly higher (P < 0.05) in water, sediment, fish muscles from Malancha than Chandanpiri and Diamond Harbour. L. parsia (MPI: 0.4-1.6) showed highest metal deposition in their muscle followed by A. mola (MPI: 0.37-1.38) and M. gulio (MPI: 0.2-1.2). Malondealdehyde, superoxide dismutase, catalase, glutathione S transferase, glutathione reductase and cortisol levels increased in case of L. parsia from Malancha and Chandanpiri. Succinate dehydrogenase, lactate dehydrogenase, Ca+2 ATPase and cytochrome C oxidase levels were significantly (P < 0.05) lower at Malancha and Chandanpiri than Diamond Harbour. Heat shock protein (HSP70) expression was significantly (P < 0.05) higher in all fish species at Malancha followed by Chandanpiri and Diamond Harbour. Glucose, glycogen, hexokinase, phosphofructokinase and glycogen phosphorylase levels varied between sites and selected fish species. Serum cortisol level was measured and found to be the highest in L. parsia from Malancha (2.94 ± 0.12 ng/ml) and the lowest in M. gulio from Diamond Harbour (0.7 ± 0.05 ng/ml). The results indicate that metal toxicity alters antioxidant levels, oxidative status and energy production in fish in species specific manner. Our results also indicate that Mystus has the highest degree of adaptability in response to metal toxicity possibly due to its specific food habit and niche position. Therefore, it can be concluded that maintenance of oxidative and metabolic status to combat metal-induced oxidative load will be helpful for the fishes to acquire better resistance under such eco-physiological stress. Alteration of niche and interactive segregation in aquatic organism may be one of the key modulator of resistance against such stress.

Effectiveness of melatonin to restore fish brain activity in face of permethrin induced toxicity.

Present study demonstrates permethrin induced oxidative damage in fish brain and explores effectiveness of melatonin to ameliorate brain function. Adult female Notopterus notopterus were exposed to nominal permethrin concentrations at 1/20th (0.34 µg/l) and 1/10th (0.68 µg/l) of LC50 for 15 days. The measured permethrin concentrations using gas chromatography (GC-ECD) were 0.28 µg/l and 0.57 µg/l, respectively. Some fish were sacrificed to collect brain tissue after 15 days of exposure. Remaining fish from both groups were administered exogenous melatonin (50 µg/kg, 100 µg/kg body weight) for 7 days and brain tissues were collected. Brain enzymes, ntioxidant factors, HSP70, HSP90, nuclear factor-kappa binding (NFkB), melatonin receptor (MT1R) proteins were measured. Permethrin treatment significantly (P < 0.05) decreased the levels of glutathione and brain enzymes. Malondialdehyde (MDA), xanthine oxidase (XO), HSPs increased at each concentration of permethrin. However, superoxide dismutase, glutathione s-transferase levels increased at low permethrin concentration followed by sharp decrease at higher concentration. Expression of NFkB and MT1R increased significantly (P < 0.05). Melatonin administration reinstated activity of brain enzymes, reduced MDA, XO levels and modulated HSPs. Melatonin also increased expression of NFkB and MT1R. Exogenous melatonin improves oxidative status in permethrin stressed fish brain. Melatonin modulates expression of HSPs that enables brain to become stress tolerant and survive by initiating NFkB translocation. Melatonin could act through melatonin receptor protein to induce synthesis of antioxidant proteins. Therefore the study successfully evaluates the potential of melatonin application for better culture and management of fish against pesticide toxicity.

Enzymatic, Non-enzymatatic Antioxidant Levels and Heat Shock Protein Expression as Indicators of Metal Induced Toxicity and Reproductive Modulation in Female Indian Major Carp Cirrhinus cirrhosus.

Effects of zinc (Zn) and lead (Pb) exposure on oxidative biomarkers and heat shock protein (HSP) expression, and their possible relation to ovarian steroidogenesis in fish were investigated. Female Cirrhinus cirrhosus were exposed to 1/10th of LC50 of metals for 30 days, and kept for another 30 days without metal exposure. Metal concentrations, antioxidants, HSPs, 17ß-estradiol and steroidogenic enzymes were analysed in brain and ovary after 15 and 30 days of exposure and a 15 and 30 day recovery period. Activities of enzymatic and non-enzymatic antioxidants showed duration dependent variation in both exposure and recovery period. HSP70 and HSP90 expressions increased following metal exposure, with the expression being higher in brain than ovary. 17ß-Estradiol, steroidogenic enzymes decreased significantly (p < 0.05) after metal exposure. The present study suggests that metals have differential and tissue specific influence on oxidative status and manipulate ovarian steroidogenesis probably through the modulation of HSPs.

Physiological response of fish under variable acidic conditions: a molecular approach through the assessment of an eco-physiological marker in the brain.

The current study demonstrates oxidative damage and associated neurotoxicity following pH stress in two freshwater carp Labeo rohita and Cirrhinus cirrhosus. Carp (n = 6, 3 replicates) were exposed to four different pH (5.5, 6, 7.5, and 8) against control (pH 6.8 ± 0.05) for 7 days. After completion of treatment, levels of enzymatic (superoxide dismutase [SOD], catalase [CAT], glutathione reductase [GRd]) and non-enzymatic antioxidants (malondialdehyde [MDA], glutathione [GSH]), brain neurological parameters (Na+-K+ATPase, acetylcholinesterase [AcHE], monoamine oxidase [MAO], and nitric oxide [NO]), xanthine oxidase (XO), heat shock proteins (HSP70 and HSP90), and transcription factor NFkB were measured in carp brain. Variation in the pH caused a significant alteration in the glutathione system (glutathione and glutathione reductase), SOD-CAT system, and stress marker malondialdehyde (MDA). Xanthine oxidase was also induced significantly after pH exposure. Brain neurological parameters (MAO, NO, AChE, and Na+-K+ATPase) were significantly reduced at each pH-treated carp group though inhibition was highest at lower acidic pH (5.5). Cirrhinus cirrhosus was more affected than that of Labeo rohita. Molecular chaperon HSP70 expression was induced in all pH-treated groups though such induction was more in acid-stressed fish. HSP90 was found to increase only in acid-stressed carp brain. Expression of NFkB was elevated significantly at each treatment group except for pH 7.5. Finally, both acidic and alkaline pH in the aquatic system was found to disturb oxidative balance in carp brain which ultimately affects the neurological activity in carp. However, acidic environment in the aquatic system was more detrimental than the alkaline system regarding oxidative damage and subsequent neurotoxicity in carp brain.

Physiological and behavioural responses to acid and osmotic stress and effects of Mucuna extract in Guppies.

Variation in pH (acidification) and salinity conditions have severe impact at different levels of biological organization in fish. Present study focused to assess the effects of acidification and salinity changes on physiological stress responses at three different levels of function: i) hormonal and oxidative response, ii) osmoregulation and iii) reproduction, in order to identify relevant biomarkers. Second objective of the study was to evaluate the efficacy of plant (Mucuna pruriens) extract for alleviating pH and salinity related stress. Guppies (Poecilia reticulata) were exposed to different pH (6.0, 5.5, 5.0) and salinity (1.5, 3.0, 4.5 ppt) for 7, 14 and 21 days. Following exposure to stress for respective duration, fish were fed diet containing methanol extract of Mucuna seeds (dose 0.80 gm/kg feed) for 7, 14 and 21 days to measure their possible recovery response. Stress hormone (cortisol), hepatic oxidative stress parameters [superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GRd), glutathione peroxidise (GPx), glutathione S-transferase (GST), malondialdehyde (MDA), glutathione (GSH)], gill osmoregulatory response (Na+-K+ATPase activity), sex steroid profiles and mating behaviours (gonopodial thrust and gestation period) were estimated. Cortisol and MDA levels increased with dose and duration of acid and salinity stress, and cortisol levels were higher in males than in females. Effect on Na+-K+ATPase activity was more intense by salinity stress rather than pH induced stress. Both acid and salinity stress reduced sex steroid levels, and mating response was highly affected by both stresses in a dose- and duration-dependent manner. Mucuna treatment reduced stress-induced alteration of cortisol, MDA, Na+-K+ATPase activity and reproductive parameters. Dietary administration of Mucuna seed extract decreased the intensity of environmental stressors at all three functional levels. Mucuna treatment was more effective against salinity stress than acid stress. Thus, cortisol, oxidative stress marker MDA and Na+-K+ATPase could be effective indicators for acid and salinity stress in wild and domestic fish populations. Dietary administration of Mucuna extract may limit the detrimental effects of acidification and salinity variations that are the inevitable outcomes expected under global climate change conditions.

Melatonin ameliorates H2O2-induced oxidative stress through modulation of Erk/Akt/NFkB pathway.

BACKGROUND: Improper control on reactive oxygen species (ROS) elimination process and formation of free radicals causes tissue dysfunction. Pineal hormone melatonin is considered a potent regulator of such oxidative damage in different vertebrates. Aim of the current communication is to evaluate the levels of oxidative stress and ROS induced damage, and amelioration of oxidative status through melatonin induced activation of signaling pathways. Hepatocytes were isolated from adult Labeo rohita and exposed to H2O2 at three different doses (12.5, 25 and 50 µM) to observe peroxide induced damage in fish hepatocytes. Melatonin (25, 50 and 100 µg/ml) was administered against the highest dose of H2O2. Enzymatic and non-enzymatic antioxidants such as malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) was measured spectrophotometrically. Expression level of heat shock proteins (HSP70 and HSP90), HSPs-associated signaling molecules (Akt, ERK, cytosolic and nuclear NFkB), and melatonin receptor was also measured by western blotting analysis. RESULTS: H2O2 induced oxidative stress significantly altered (P < 0.05) MDA and GSH level, SOD and CAT activity, and up regulated HSP70 and HSP90 expression in carp hepatocytes. Signaling proteins exhibited differential modulation as revealed from their expression patterns in H2O2-exposed fish hepatocytes, in comparison with control hepatocytes. Melatonin treatment of H2O2-stressed fish hepatocytes restored basal cellular oxidative status in a dose dependent manner. Melatonin was observed to be inducer of signaling process by modulation of signaling molecules and melatonin receptor. CONCLUSIONS: The results suggest that exogenous melatonin at the concentration of 100 µg/ml is required to improve oxidative status of the H2O2-stressed fish hepatocytes. In H2O2 exposed hepatocytes, melatonin modulates expression of HSP70 and HSP90 that enable the hepatocytes to become stress tolerant and survive by altering the actions of ERK, Akt, cytosolic and nuclear NFkB in the signal transduction pathways. Study also confirms that melatonin could act through melatonin receptor coupled to ERK/Akt signaling pathways. This understanding of the mechanism by which melatonin regulates oxidative status in the stressed hepatocytes may initiate the development of novel strategies for hepatic disease therapy in future.

Consequences of oxidative damage and mitochondrial dysfunction on the fatty acid profile of muscle of Indian Major Carps considering metal toxicity.

Current study aims to find interrelation between mitochondrial enzyme function and fatty acid profile in fish muscle and role of antioxidant agents to maintain their balance in response to metal accumulation. Fishes (Labeo rohita, Catla catla, Cirrhinus cirrhosus) were collected from two sites (Nalban Bheri and Diamond Harbour, India). Concentrations of metals (lead, cadmium, copper, nickel, zinc), enzymatic and non-enzymatic antioxidant activity (malondialdehyde, superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, glutathione S-transferase), muscle enzyme activity (acetylcholinesterase, succinate dehydrogenase, lactate dehydrogenase, Ca2+ATPase, AMP-deaminase, lipoamide reductase, cytochrome C oxidase, aldolase) and fatty acid composition in muscle tissues were analyzed. Metal concentrations were significantly higher (P < 0.05) in fish muscles from Nalban compared to those in Diamond Harbour. Increased activity of antioxidant enzymes was noted with diminished mitochondrial enzymes activity and altered fatty acid composition in response to higher metal accumulation. Higher metal concentration in fish muscle of Nalban seems to significantly (P < 0.05) affect poly and monounsaturated fatty acid content, possibly due to oxidative damage and accumulation of hazardous reactive oxygen species (ROS) molecules. Changes in fatty acid contents following metal accumulation were observed to be species specific. Current study is the first correlative study to illuminate the level of oxidative damage and possible consequences on muscle cellular integrity, mitochondrial functionality and flesh quality against bioaccumulation of different metals in carps. Future studies are needed to quantify the relative contributions of enzymatic and low-molecular-mass antioxidants in protecting mitochondrial function and maintenance of proper fatty acid oxidation during acclimation to long term metal exposure.

Impact of Metal Toxicity on Oxidative Balance and Mitochondrial Enzyme Function in Muscle of Tilapia.

Present study investigates the effect of metal accumulation on antioxidant level and mitochondrial enzymes function in muscle of Oreochromis mossambicus. Metal accumulation in muscle upregulated stress marker malondialdehyde and the activity of different antioxidant enzymes with no significant alteration in glutathione system. Metal exposure to fish muscle decreased the activity of mitochondrial enzymes. AMP deaminase, aldolase, cytochrome C oxidase and lipoamide reductase showed positive correlation with acetylcholinesterase, glutathione reductase, reduced glutathione and glutathione peroxidase, but negative correlation with superoxide dismutase, catalase, glutathione S-transferase and thiobarbituric acid reactive substance. Analysis of these biomarkers clearly indicates the change in oxidative load in muscle tissues and provides insight to muscle response to the metal exposure. Therefore, the study outlines the potential use of biomarkers in context of muscle mitochondrial enzymes relating to oxidative processes that take place in the fish muscle following metal exposure and toxicity.

Melatonin ameliorates H2O2-induced oxidative stress through modulation of Erk/Akt/NFkB pathway

BACKGROUND: Improper control on reactive oxygen species (ROS) elimination process and formation of free radicals causes tissue dysfunction. Pineal hormone melatonin is considered a potent regulator of such oxidative damage in different vertebrates. Aim of the current communication is to evaluate the levels of oxidative stress and ROS induced damage, and amelioration of oxidative status through melatonin induced activation of signaling pathways. Hepatocytes were isolated from adult Labeo rohita and exposed to H2O2 at three different doses (12.5, 25 and 50 µM) to observe peroxide induced damage in fish hepatocytes. Melatonin (25, 50 and 100 µg/ml) was administered against the highest dose of H2O2. Enzymatic and non-enzymatic antioxidants such as malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) was measured spectrophotometrically. Expression level of heat shock proteins (HSP70 and HSP90), HSPs-associated signaling molecules (Akt, ERK, cytosolic and nuclear NFkB), and melatonin receptor was also measured by western blotting analysis. RESULTS: H2O2 induced oxidative stress significantly altered (P < 0.05) MDA and GSH level, SOD and CAT activity, and up regulated HSP70 and HSP90 expression in carp hepatocytes. Signaling proteins exhibited differential modulation as revealed from their expression patterns in H2O2-exposed fish hepatocytes, in comparison with control hepatocytes. Melatonin treatment of H2O2-stressed fish hepatocytes restored basal cellular oxidative status in a dose dependent manner. Melatonin was observed to be inducer of signaling process by modulation of signaling molecules and melatonin receptor. CONCLUSIONS: The results suggest that exogenous melatonin at the concentration of 100 µg/ml is required to improve oxidative status of the H2O2-stressed fish hepatocytes. In H2O2 exposed hepatocytes, melatonin modulates expression of HSP70 and HSP90 that enable the hepatocytes to become stress tolerant and survive by altering the actions of ERK, Akt, cytosolic and nuclear NFkB in the signal transduction pathways. Study also confirms that melatonin could act through melatonin receptor coupled to ERK/Akt signaling pathways. This understanding of the mechanism by which melatonin regulates oxidative status in the stressed hepatocytes may initiate the development of novel strategies for hepatic disease therapy in future.

Change in redox state and heat shock protein expression in an Indian major carp Cirrhinus cirrhosus exposed to zinc and lead.

Fish are exposed to different heavy metals that may induce numerous physiological changes. In the present study, we examined the redox state in response to a severe stress resulting from two heavy metals (Zinc and Lead) contamination in carp Cirrhinus cirrhosus. Fish were exposed to 1/10th of LC50 of the respective metals [zinc chloride (2.72 mg/L) and lead acetate (2.53 mg/L)] for 30 days and allowed to recover for another 30 days without any metal exposure. Concentration of metals, different enzymatic and non-enzymatic antioxidant agents and expression levels of heat shock protein (HSP) 70 and 90 were measured in the liver and the kidney of fish. The lipid peroxide levels in fish tissues gradually increased with duration of treatment for both metals. After 15 days of treatment, glutathione (GSH) levels had increased, but decreased as the treatment continued for 30 days and returned to basal levels after a 30-day recovery period. Activities of all the anti-oxidant enzymes, except glutathione peroxidase, in stressed fish were significantly increased compared to those in the control at 15 days and continued till the 30th day of treatment, showing a tendency to return to basal levels after the recovery period. Expression levels of HSP70 and HSP90 gradually increased after zinc and lead treatment, respectively. The expression of HSP was higher in the liver. The results suggest that different heavy metals may have differential effects on the redox state and induction of oxidative stress in carp, in vivo.