Hepatic immune response of Coilia nasus infected with Anisakidae during ovarian development.

Anisakidae parasitism is a prevalent disease in wild populations of Coilia nasus, and can result in a significant loss of germplasm resources. To elucidate the immune response mechanism of C. nasus livers to Anisakidae infection, we collected and analysed 18 parasitic and 18 non-parasitic livers at gonadal developmental stages II, III, and V using histopathology, molecular biology and transcriptome methods. The hepatic portal area of the parasitic group exhibited an increase in the fibrous stroma and thickened hepatic arteries with positive Ly-6G staining, indicating inflammation and immune responses in the liver. Hepatocyte cytokine levels and the expression of liver function-related genes indicated that fish livers responded similarly to Anisakidae parasitism across different gonadal developmental stages. Oxidative stress indices showed more intense changes in stage II samples, whereas gene expression levels of Nrf2 and C3 were significantly increased in parasitised livers during stage III and V. Liver transcriptome sequencing identified 2575 differentially expressed genes between the parasitic and non-parasitic groups at the three gonadal developmental stages. KEGG pathway analysis showed that natural killer cell-mediated cytotoxicity, the NOD-like receptor signaling pathway, neutrophil extracellular trap formation, and other immune pathways were significantly enriched. Expression patterns varied across developmental stages, suggesting that innate immunity was primarily responsible for the liver immune response to Anisakidae infection during C. nasus migration, possibly related to water temperature changes or shifts in the gonadal developmental stage. In summary, this study investigated the immune response of C. nasus to Anisakidae parasitism under natural conditions, focusing on reproductive aspects and environmental changes, thereby establishing a foundation for elucidating the molecular mechanisms underlying the immune response of Anisakidae in C. nasus.

Involvement of reactive oxygen species (ROS) in the hepatopancreatic cytotoxicity, oxidative stress, and apoptosis induced by microcystin-LR in Eriocheir sinensis.

There is limited knowledge about the toxicity of Microcystin-LR (MC-LR) in crustaceans, despite its high toxicity to aquatic organisms. This research aimed to explore the effects of MC-LR on cytotoxicity, oxidative stress, and apoptosis in the hepatopancreas of Eriocheir sinensis, as well as elucidate the involvement of reactive oxygen species (ROS) and potential mechanisms of toxicity. In vivo and in vitro exposures of crabs to MC-LR and N-acetylcysteine (NAC) were performed, followed by assessments of cell morphology, viability, tissue pathology, biochemical indicators, gene expression, and hepatopancreatic transcriptome. Results revealed that MC-LR facilitated the entry of the MC-LR transporter oatp3a into hepatopancreatic cells, leading to upregulated expression of phase I detoxification enzyme genes (cyp4c, cyp2e1, and cyp3) and downregulated the phase II enzyme genes (gst1, gpx, gsr2, gclc, and nqo1), resulting in increased ROS levels and cytotoxic effects. MC-LR exhibited cytotoxicity, reducing cell viability and inducing abnormal nuclear morphology with a 48 h-IC50 value of approximately 120 µm. MC-LR exposure caused biochemical changes indicative of oxidative stress damage and evident hepatopancreatic lesions. Additionally, MC-LR exposure regulated the levels of bax and bcl-2 expression, activating caspase 3 and 6 to induce cell apoptosis. Intervention with NAC attenuated MC-LR-induced ROS production and associated toxic effects. Transcriptome analysis revealed enrichment of differentially expressed genes in pathways related to cytochrome P450-mediated xenobiotic metabolism and the FoxO signaling pathway. These findings shed light on the potential mechanisms underlying MC-LR toxicity and provide valuable references for further research and conservation efforts regarding the health of aquatic animals.

Gap-free genome assembly of anadromous Coilia nasus.

The Chinese tapertail anchovy, Coilia nasus, is a socioeconomically important anadromous fish that migrates from near ocean waters to freshwater to spawn every spring. The analysis of genomic architecture and information of C. nasus were hindered by the previously released versions of reference genomes with gaps. Here, we report the assembly of a chromosome-level gap-free genome of C. nasus by incorporating high-coverage and accurate long-read sequence data with multiple assembly strategies. All 24 chromosomes were assembled without gaps, representing the highest completeness and assembly quality. We assembled the genome with a size of 851.67 Mb and used BUSCO to estimate the completeness of the assembly as 92.5%. Using a combination of de novo prediction, protein homology and RNA-seq annotation, 21,900 genes were functionally annotated, representing 99.68% of the total predicted protein-coding genes. The availability of gap-free reference genomes for C. nasus will provide the opportunity for understanding genome structure and function, and will also lay a solid foundation for further management and conservation of this important species.

Salvianolic Acid B Regulates Oxidative Stress, Autophagy and Apoptosis against Cyclophosphamide-Induced Hepatic Injury in Nile Tilapia (Oreochromis niloticus).

Salvianolic acid B (Sal B), as one of the main water-soluble components of Salvia miltiorrhizae, has significant pharmacological activities, including antioxidant, free radical elimination and biofilm protection actions. However, the protective effect of Sal B on Nile tilapia and the underlying mechanism are rarely reported. Therefore, the aim of this study was to evaluate the effects of Sal B on antioxidant stress, apoptosis and autophagy in Nile tilapia liver. In this experiment, Nile tilapia were fed diets containing sal B (0.25, 0.50 and 0.75 g·kg-1) for 60 days, and then the oxidative hepatic injury of the tilapia was induced via intrapleural injection of 50 g·kg-1 cyclophosphamide (CTX) three times. After the final exposure to CTX, the Nile tilapia were weighed and blood and liver samples were collected for the detection of growth and biochemical indicators, pathological observations and TUNEL detection, as well as the determination of mRNA expression levels. The results showed that after the CTX treatment, the liver was severely damaged, the antioxidant capacity of the Nile tilapia was significantly decreased and the hepatocyte autophagy and apoptosis levels were significantly increased. Meanwhile, dietary Sal B can not only significantly improve the growth performance of tilapia and effectively reduce CTX-induced liver morphological lesions, but can also alleviate CTX-induced hepatocyte autophagy and apoptosis. In addition, Sal B also significantly regulated the expression of genes related to antioxidative stress, autophagy and apoptosis pathways. This suggested that the hepatoprotective effect of Sal B may be achieved through various pathways, including scavenging free radicals and inhibiting hepatocyte apoptosis and autophagy.

Alleviative effects of astragaloside IV on cyclophosphamide-induced oxidative damage and immunosuppression in tilapia (Oreochromis niloticus).

Astragaloside IV (ASIV) has effects of antioxidation and immunologic enhancement. However, there are few reports on the application and potential mechanism of ASIV in aquaculture. In this study, we investigated the effect of ASIV on growth, antioxidation, and immune function of tilapia. Tilapia were fed a diet containing 0.1, 0.2, and 0.5 g·kg-1 ASIV for 60 days, followed by an intrapleural injection of 50 mg·kg-1 cyclophosphamide (CTX) to induce oxidative damage and immunosuppression. Then tilapia were weighed and blood, liver, spleen, kidney, and intestinal were collected. The results showed ASIV increased the final weight, relative weight rate, and specific growth rate of tilapia, reduce conversion ratio, and reduced the morphological lesions of tissues. Meanwhile, ASIV alleviated CTX-induced oxidative damage by improving antioxidant activity in serum and tissues and inhibiting lipid peroxidation. Additionally, ASIV attenuated the immunosuppression of tilapia caused by CTX, regulated immunochemical indexes in serum, increased the viability of peripheral blood leukocytes and head kidney macrophages, and restored respiratory burst activity (O2-) in head kidney macrophages and splenocytes. Furthermore, qPCR data showed ASIV up-regulated antioxidant-related gene expression of nrf2, ho-1, gpx3, and cat and immune-related gene expression including C3 and igm. In conclusion, ASIV as a feed additive can not only improve the growth performance but also enhance the antioxidant capacity and immune function of tilapia, which may be associated with the ability of ASIV to scavenge free radicals, reduce lipid peroxidation levels, and stabilize numbers of immune cells.

Solubility Variation and Prediction Model of CO2 in Water-Bearing Crude Oil.

The solubility of CO2 in water-bearing crude oil is of great significance for the calculation of crude oil reserves, the development of CO2-EOR (CO2-enhanced oil recovery), CO2-CCUS (carbon capture, utilization, and storage), and CO2 assisted steam huff-and-puff technology, and the optimization of the design of CO2 for heavy oil pipeline transportation. In order to determine the variation of the solubility of water-bearing crude oil by injecting CO2 into the formation, taking the Upper Wuerhe Formation reservoir in the 53 East Block as an example, the study of the dissolution characteristics of CO2 in water-bearing crude oil at different temperature and pressure conditions was carried out by using a high-temperature and high-pressure reaction kettle. At the same time, a new solubility prediction model of CO2 in water-bearing crude oil was proposed based on the existing solubility prediction models. The results show that, under the same water cut, the solubility of CO2 in water-bearing crude oil decreases with the increase of temperature and decreases with the decrease of pressure. At the same time, the solubility of CO2 in water-bearing crude oil is more sensitive to pressure. At the same temperature, the solubility of CO2 in water-bearing crude oil decreases with the increase of water cut, and the higher the pressure, the greater the effect of water cut on the solubility of CO2 in water-bearing crude oil. The newly established combined prediction model of CO2 solubility in water-bearing crude oil is convenient for calculation and has a wide range of applications. The average relative error is only 9.5%, which can meet the requirements of engineering calculation accuracy.

Protective Effects of Glycyrrhiza Total Flavones on Liver Injury Induced by Streptococcus agalactiae in Tilapia (Oreochromis niloticus).

Clinical studies have confirmed that Glycyrrhiza total flavones (GTFs) have good anti-hepatic injury, but whether they have a good protective effect on anti-hepatic injury activity induced by Streptococcus agalactiae in tilapia (Oreochromis niloticus) is unknown. The aims of this study were to investigate the protective effects of Glycyrrhiza total flavones on liver injury induced by S. agalactiae (SA) and its underlying mechanism in fish. A total of 150 tilapia were randomly divided into five groups, each with three replicates containing 10 fish: normal control group, S. agalactiae infection group, and three Glycyrrhiza total flavone treatment groups (addition of 0.1, 0.5, or 1.0 g of GTF to 1 kg of feed). The normal control group was only fed with basic diet, after 60 d of feeding, and intraperitoneal injection of the same volume of normal saline (0.05 mL/10 g body weight); the S. agalactiae infection group was fed with basic diet, and the S. agalactiae solution was intraperitoneally injected after 60 d of feeding (0.05 mL/10 g body weight); the three GTF treatment groups were fed with a diet containing 0.1, 0.5, or 1.0 g/kg of GTF, and the S. agalactiae solution was intraperitoneally injected after 60 d of feeding (0.05 mL/10 g body weight). After 48 h injection, blood and liver tissues were collected to measure biochemical parameters and mRNA levels to evaluate the liver protection of GTFs. Compared with the control group, the serum levels of glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), alkaline phosphatase (AKP) and glucose (GLU) in the streptococcal infection group increased significantly, while the levels of total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) decreased significantly; observations of pathological sections showed obvious damage to the liver tissue structure in response to streptococcal infection. S. agalactiae can also cause fatty liver injury, affecting the function of fatty acid ß-oxidation and biosynthesis in the liver of tilapia, and also causing damage to function of the immune system. The addition of GTFs to the diet could improve oxidative stress injury caused by S. agalactiae in tilapia liver tissue to different degrees, promote the ß-oxidation of fatty acids in the liver, accelerate the lipid metabolism in the liver, and repair the damaged liver tissue. GTFs have a good protective effect on liver injury caused by streptococcus.

A proteomics approach reveals digestive and nutritional responses to food intake in anadromous Coilia nasus.

The estuarine tapertail anchovy, Coilia nasus, is an anadromous fish that undertakes over a 600-km spawning migration along the Yangtze River of China. They generally cease feeding during this process, but we recently documented that a small proportion of them appear to feed. Research on proteomic responses is essential for understanding the phenomenon of C. nasus feeding. In this study, we used an iTRAQ-based proteomics approach to study the changes in protein expression in response to food intake in C. nasus following voluntary fasting. Coilia nasus in the feeding group (CSI) were fed shrimp or small fish, whereas those in the control group (CSN) were starved. We identified 3279 proteins in the gastric tissue/stomach, of which 279 were significantly differentially expressed. In all, 133 differentially expressed proteins (DEPs) were upregulated and 146 proteins were downregulated in CSI compared with those in CSN C. nasus. In addition to gastric acid secretion caused by gastric distention, a functional analysis suggested that a series of DEPs were involved mainly in the regulation of protein digestion (e.g., carboxypeptidase A1 and chymotrypsin A-like), immune response (e.g., lysozyme and alpha 2-macroglobulin), and nutrition metabolism (e.g., glyceraldehyde 3-phosphate dehydrogenase, glycogenin, long-chain acyl-CoA synthetase, and creatine kinase). Real-time PCR confirmed that the mRNA levels of the DEPs were similar those obtained using iTRAQ. These results indicate that the nutrients obtained through food were effectively utilized by C. nasus, thereby providing energy for swimming, gonadal maturation, primary metabolism, and an enhanced immune function to better resist pathogen interference. This research contributes to the elucidation of nutritional regulation mechanisms of C. nasus to better protect the wild population.

Anisakidae parasitism activated immune response and induced liver fibrosis in wild anadromous Coilia nasus.

Anisakidae nematode larvae is one of the most common parasites in wild anadromous Coilia nasus. This study aims to explore the mechanism of the C. nasus immune response to the parasitism of Anisakid nematode larvae. Results found that Anisakid nematode larvae parasitism caused liver injury as evidenced by histomorphology results as well as high levels of aminotransferase and aspertate aminotransferase. Furthermore, Anisakid nematode larvae parasitism induced an immune response in the host, which was characterized by the elevated populations of macrophages and neutrophils in the liver and head-kidney in the Anisakidae-infected group compared to the noninfected group. The expression of immunoglobulin IgM and IgD in the liver and head-kidney was also increased in the Anisakidae-infected group. The Anisakidae-infected group showed higher activity of antioxidant enzymes catalase and superoxide dismutase, which indicates severe oxidative stress, and increased production of pro-inflammatory cytokines, TNF-α, IL-6 as well as MCP-1 in the liver compared with the noninfected group. As a result of inflammation, livers of hosts in the Anisakidae-infected group showed fibrosis, and elevated expression of associated proteins including α-smooth muscle actin, fibronectin, collagen type I and type III compared with the noninfected group. We demonstrated that Anisakid nematode larvae parasitism results in injury and fibrosis in the liver, and triggers immune cell infiltration and inflammation in the liver and head-kidney of C. nasus. Altogether, the results provide a foundation for building an interaction between parasite and host, and will contribute to C. nasus population and fishery resource protection.

Simultaneous Pyrolysis of Coal and Biomass in a Drop-Tube-Fixed-Bed Reactor.

Copyrolysis of coal and biomass has been extensively studied to exploit its inherent synergistic effects; however, the different pyrolysis temperature zones of coal and biomass seriously affect the realization of these effects. Therefore, a new copyrolysis method (preheating the coal to a certain temperature and then adding the biomass in a drop-tube-fixed-bed reactor, denoted as M1) was designed herein to achieve "simultaneous" pyrolysis of coal and biomass. The yields of products and the characteristics of M1-produced tar were estimated and compared with those of tar obtained by fixed-bed-reactor (denoted as M2)-based copyrolysis. M1 achieved a higher tar yield and lower water content than M2. The M1-generated tar exhibited a lower free-radical concentration, higher H/C ratio, higher levels of uncondensed aromatic hydrogen, and shorter side-chains than that produced by M2. The temperature of HLBE coal at which the WSs were fed to the reactor in M1, denoted as T F, affects the "simultaneous" pyrolysis. T F values of 300, 400, and 500 °C were studied, and it was found that the tar yield obtained at a T F of 400 °C (the main pyrolysis temperature of coal) is the highest, the water yield is the lowest, and the free-radical concentration of the tar is also the lowest among the investigated samples.

Alteration of endoplasmic reticulum stress, inflammation and anti-oxidative status in cyclophosphamide-damaged liver of Nile tilapia (Oreochromis niloticus).

Cyclophosphamide (CTX) is a common immunosuppressant, and it can also results in liver injury in human and animals. In this study, the CTX-induced liver injury mechanism in tilapia (Oreochromis niloticus) was investigated by studying alteration of endoplasmic reticulum stress (ERS), inflammation and anti-oxidative status. Tilapia was intraperitoneally injected CTX at the doses of 10, 25, 50, 75 and 100 mg·kg-1, and the blood and liver tissues were collected. The results showed that CTX administration had a significant cytotoxicity on hepatocytes, and increased the liver index. The extensive vacuolar degeneration, unclear cell outline and other histological lesions were also observed. CTX administration markedly decreased the antioxidant ability and enhanced lipid peroxidation in liver. Furthermore, qPCR data showed that CTX administration at 50-100 mg·kg-1 up-regulated gene expressions of cyp1a, cyp2k1 and cyp3a, and inflammatory response-related genes including rel, relb, nfκb1, il-6, il-8, il-10 and tnf-α. CTX significantly promoted the mRNA levels of ERS-related genes (eif2α, crt, parp1, grp78, ire1, xbp1s and chop) in a dose dependent manner. Additionally, CTX injection at 75-100 mg·kg-1 could down-regulate gene expressions of anti-oxidative status including nrf2, ucp2, ho-1, gpx3, gstα and cat. Overall results suggested CTX injection induced liver damage which was related to the cytotoxic effect on hepatocytes, decrease of antioxidant capacity, inflammatory response and ERS.

Effects of cyclophosphamide on antioxidative and immune functions of Nile tilapia (Oreochromis Niloticus) via the TLR-NF-κB signaling pathway.

Intensive aquaculture often results in immunosuppression in fish, which may cause a series of diseases. In this study, to investigate the immunosuppressive mechanisms in fish, tilapia were intrapleural injected cyclophosphamide (CTX) at the doses of 10, 25, 50, 75 and 100 mg·kg-1 to induce immunosuppression. We determined the viability of immune cells, the content of lysozyme (LZM) and immunoglobulin M (IgM), the levels of nitric oxide (NO) and antioxidant parameters. Meanwhile, the mRNA levels of complement C3 (c3), igm and the genes associated with the TLR-NF-κB signaling pathway in the head kidney (HK) and spleen were also determined. The results showed that CTX had a significant cytotoxic effect on peripheral blood leukocytes, HK macrophages and spleen cells in a dose-dependent manner. The protein and mRNA levels of C3 and IgM were down-regulated with the increase of CTX concentrations in serum, HK and/or spleen. The NO and LZM contents decreased significantly in HK and spleen after CTX treatments with 75 and 100 mg·kg-1. CTX treatments with 50, 75 and/or 100 mg·kg-1 markedly decreased the antioxidant ability and enhanced lipid peroxidation in HK and spleen. Furthermore, qPCR data showed that CTX treatments with 50-100 mg·kg-1 clearly down-regulated the mRNA levels of tlr2, myd88, irak1, traf6, nfκb1, nfκb2, il-6, il-10 and tnf-α in the HK and/or spleen. Overall results suggested that CTX treatment had a cytotoxic effect on immune cells, induced lipid peroxidation, decreased the antioxidant capacity and inhibited immune function. The immunosuppressive mechanisms of CTX may be associated with the TLR-NF-κB signaling pathway.

Alleviative effects of total flavones of Glycyrrhiza uralensis Fisch on oxidative stress and lipid metabolism disorder induced by high-fat diet in intestines of Tilapia (Oreochromis niloticus).

Total flavones of Glycyrrhiza uralensis Fisch (GTF) are main components of Glycyrrhiza uralensis Fisch, which have anti-oxidation and lipid-lowering effects. However, its protective effects on the intestinal tissue of tilapia (Oreochromis niloticus) are unknown. The aims of the study were to evaluate the protective effects of GTF on the intestinal tissue of tilapia after high-fat diet (HFD) feeding. Tilapia (initial weight 30 ± 1 g) received diets containing four doses of GTF (0.05, 0.1, 0.5, and 1.0 g/kg diet) for 90 days. The intestinal tissues were collected to determine biochemical parameter, gene expression and protein level. The results showed that the HFD reduced antioxidant indexes and increased the fat level, lipid oxidation products in the intestinal tissue relative to the control. Adding GTF to the HFD resulted in an increase of antioxidant indexes, fat level and lipid oxidation products decreased after 60, 90 days. In the HFD group, mRNA level of fatty acid transport protein 1 (FATP1) was increased at 60 day and then decreased at 90 day. The mRNA levels of fatty acid binding protein 1 (FABP1) and sterol regulatory element binding protein 1c (SREBP 1c) were significantly increased at 60 or 90 day after HFD feeding. The mRNA levels of acetate coenzyme A carboxylase (ACCA) peroxisome proliferator-activated receptor γ (PPAR-γ) and PPAR-α were decreased significantly at 30, 60 and/or 90 days after HFD feeding. Western blotting results also showed that nuclear factor (NF)-κß C-Rel (NF-κß C-Rel) and mitogen-activated protein kinase 8 (MAPK8) protein expression in intestinal tissue increased after consumption of the HFD. However, adding GTF to the HFD reversed the changes of genes related to fatty acid synthesis and metabolism, and the level of NF-κß c-Rel and MAPK8 at different degrees. Overall, these results indicated that GTF promoted decomposition and metabolism of fatty acids in intestinal tissue, alleviated oxidative stress damage caused by the HFD, and had certain protective effects on the intestinal tissue of tilapia.

Application of transcriptome analysis to understand the adverse effects of hydrogen peroxide exposure on brain function in common carp (Cyprinus carpio).

Hydrogen peroxide (H2O2), as a common disinfectant, has been extensively used in aquaculture. The toxicity of high ambient H2O2 for gills and liver of fish has received attention from many researchers. However, whether H2O2 exposure induced brain injury and neurotoxicity has not been reported in fish. Therefore, this study aimed to explore the potential mechanism of H2O2 toxicity in brain of common carp via transcriptome analysis and biochemical parameter detection. The fish were exposed to 0 (control) and 1 mM of H2O2 for 1 h per day lasting 14 days. The results showed that H2O2 exposure caused oxidative damage in brain evidenced by decreased glutathione (GSH), total antioxidant capacity (T-AOC) and nicotinamide adenine dinucleotide (NAD+) levels, and increased formation of malondialdehyde (MDA) and 8-hydroxy-2'-deoxyguanosine (8-OHdG). Meanwhile, H2O2 exposure reduced 5-hydroxytryptamine (5-HT) level, and down-regulated tryptophan hydroxylase 1 (tph1a), tph2, 5-hydroxytryptamine receptor 1A-beta (htr1ab) and htr2b expression in brain. Transcriptome analysis showed that H2O2 exposure up-regulated 604 genes and down-regulated 1209 genes in brain. Go enrichment displayed that the differently expressed genes (DEGs) were enriched mainly in cellular process, single-organism process, metabolic process, and biological regulation in the biological process category. Further, KEGG enrichment indicated that H2O2 exposure led to dysregulation of neurotransmitter signals including depression of glutamatergic synapse, GABAergic synapse and endocannabinoid signaling. Also, we found the alteration of three key pathways including calcium, cAMP and HIF-1 in brain after H2O2 exposure. In conclusion, our data indicated that H2O2 exposure induced oxidative damage and neurotoxicity, possibly related to dysregulation of neurotransmitters and calcium, cAMP and HIF-1 signaling pathways, which may adversely affect learning, memory and social responses of common carp. This study provided novel insight into biological effects and underlying mechanism of H2O2 toxicity in aquatic animal, and contributed to proper application of H2O2 in aquaculture.

Alteration of lipid metabolism, autophagy, apoptosis and immune response in the liver of common carp (Cyprinus carpio) after long-term exposure to bisphenol A.

Bisphenol A (BPA), as a phenolic compound, is harmful to human health, and its residue in the aquatic environment also threatens the health of aquatic animals. In this research, the toxicity effects of BPA on liver tissues were evaluated in common carp (Cyprinus carpio) after long-term exposure. Fish were exposed to five concentrations of BPA (0, 0.01, 0.1, 0.5 and 2 mg/L) for 30 days. The blood and liver tissues were gathered to analyze biochemical indices and genes transcription levels. The data related to lipid metabolism showed that BPA exposure increased serum total cholesterol (TC), low density lipoprotein cholesterol (LDL-C) and high density lipoprotein cholesterol (HDL-C) levels, upregulated the expressions of fatp1, pparγ, fas, atgl, hsl, pparα, cpt1b, acox-1, and downregulated the expression of dgat1 in liver. Antioxidative parameters displayed a reduced antioxidant ability and increased lipid peroxidation after BPA exposure. Meanwhile, the upregulations of nrf2, ho-1, cyp1a and cyp1b genes revealed an adaptive response mechanism against oxidative stress-induced adverse effects. After 30 days of exposure, BPA induced apoptosis and endoplasmic reticulum stress (ERS) via upregulating the expression levels of apoptosis and ERS-related genes and increasing Ca2+ concentration in liver. Moreover, the downregulation of mtor and the upregulation of atg3, atg7, tfeb, uvrag and mcoln1 indicated that BPA could influence the normal process of autophagy. Furthermore, BPA exposure activated toll like receptors (TLRs) pathway to mediate the inflammatory response. Our results demonstrated that BPA exposure disturbed lipid metabolism, and induced oxidative stress, ERS, apoptosis, autophagy and inflammatory response in the liver of common carp. These findings contributed to the understanding of hepatotoxicity mechanism induced by BPA in fish.

Effects of chronic glyphosate exposure on antioxdative status, metabolism and immune response in tilapia (GIFT, Oreochromis niloticus).

Glyphosate (Gly) is an active ingredient of herbicide, its underlying toxicity on fish is still unclear. The aim of this study was to evaluate chronic toxicity of Gly on tilapia via determining antioxidative status, metabolism, inflammation and immune response. The fish were exposed to different concentrations of Gly (0, 0.2, 0.8, 4 and 16 mg/L) for 80 days. The blood, liver, gills and spleen were collected to assay biochemical parameters and genes expression after 80 days of exposure. The results showed that treatments with higher Gly (4 and/16 mg/L) significantly increased the levels of TC, TG, AST, ALT, LDL-C and MDA, and apparently decreased the levels of SOD, GSH, CAT, HDL-C, HK, G3PDH, FBPase and G6PD in serum, liver and/or gills. The gene expression data showed that the treatments with Gly adversely affected Nrf2 pathway in liver, gills and spleen, as shown by significant changes of nrf2, keap1, ho-1, nqo1 and gsta mRNA levels. Meanwhile, inflammatory response was activated via enhancing the mRNA levels of nf-κb2, rel, rela tnf-α, and il-1ß, and immunotoxicity was caused through downregulating the genes expression of c-lzm, hep, igm, hsp70 and c3 in liver, gills and/or spleen of tilapia after Gly exposure. Moreover, the mRNA levels of cyp1a and cyp3a were upregulated in 16 or 0.2 mg/kg Gly group in liver. Overall results suggested chronic Gly exposure reduced antioxidative ability, disturbed liver metabolism, promoted inflammation and suppressed immunity. Interestingly, the Nrf2 and NF-κB signaling pathways played key roles in Gly chronic toxicity.

Immune, inflammatory, autophagic and DNA damage responses to long-term H2O2 exposure in different tissues of common carp (Cyprinus carpio).

Hydrogen peroxide (H2O2) is a stable reactive oxygen species (ROS) in aquatic environment, and high concentration of ambient H2O2 may directly or indirectly affect aquatic animal health. However, the response mechanism of fish to ambient H2O2 has not been well studied yet. Therefore, the aim of the study was to investigate the immune, inflammatory, autophagic and DNA damage responses to long-term H2O2 exposure in different tissues of common carp. The results showed that H2O2 exposure induced a significant immune response, with alterations in the levels of immune parameters including AKP, ACP, LZM, C3, HSP90 and HSP70 in different tissues. The inflammatory response evoked by H2O2 exposure was associated with the activations of TLRs and NF-κB (P65) in the majority of tested tissues. The autophagy process was significantly affected by H2O2 exposure, evidenced by the upregulations of the autophagy-related genes in liver, gills, muscle, intestines, heart and spleen and the downregulations in kidney. Meanwhile, the mRNA level of atm, a primary transducer of DNA damage response, was upregulated in liver, gills, intestines and spleen, and the DNA damage was evidenced by increased 8-OHdG level in intestines after H2O2 exposure. Moreover, cell cycle regulation-related genes, including cyclin A1, B and/or E1, highly expressed in all tested tissues except heart after H2O2 exposure. Interestingly, IBR analysis exhibited that immune, inflammatory, autophagic and DNA damage responses to H2O2 exposure were in a dose-dependent and tissue-specific manner. These data may contribute to understanding H2O2 toxicity for fish and assessing potential risk of H2O2 in aquatic environment.

Effects of dietary baicalin supplementation on growth performance, antioxidative status and protection against oxidative stress-induced liver injury in GIFT tilapia (Oreochromis niloticus).

Baicalin, a main bioactive compound of Scutellaria baicalensis, has a variety of pharmacological activities including antioxidation, anti-inflammation and hepatoprotection. However, there are few reports on these biological activities in fish. Therefore, the aim of this study was to assess the effects of baicalin on growth performance, antioxidative status and hepatoprotection in tilapia. The fish were fed on different doses of baicalin (0, 0.4, 0.8 and 1.6 g/kg diet). After feeding 60 days, parts of fishes were netted, and the blood, liver, gills and muscle tissues were collected to analyze the antioxidative effect. The remaining fishes were injected with saline or hydrogen peroxide (H2O2) for challenge test. The results showed that the specific growth rate of fish was slightly increased in three baicalin treatments, and the feed efficiency was clearly improved in 0.4 g/kg baicalin treatment. Meanwhile, the antioxidative capacity in blood, liver and/or gill was enhanced in treatments with 0.4, 0.8 and/or 1.6 g/kg baicalin. After challenge test, the pre-treatments with baicalin effectively alleviated H2O2-induced liver injury. In serum and liver, pre-treatments with 0.8 and/or 1.6 g/kg baicalin suppressed the oxidative damage induced by H2O2, as evidenced by improvement of the levels of SOD, T-AOC and GSH and the decline of MDA level. More important, pre-treatments with 0.4, 0.8 and/or 1.6 g/kg baicalin blocked the upregulation of mRNA levels of tlr1, myd88, irak4, rela, tnf-α and il-1ß in H2O2-induced liver injury. In summary, dietary baicalin supplementation could improve feed efficiency, enhance antioxidative ability and alleviate oxidative stress-induced hepatotoxicity in tilapia.

Chronic exposure of hydrogen peroxide alters redox state, apoptosis and endoplasmic reticulum stress in common carp (Cyprinus carpio).

Hydrogen peroxide (H2O2) appears to be ubiquitous in natural water. Higher level of H2O2 can cause physiological stress, immunosuppression and even death in aquatic animals, but the physiological and molecular mechanisms of H2O2 toxicity are not well studied. Thus, the aim of the present study was to exposure potential toxic mechanisms of H2O2 via assessing the effects on redox state, apoptosis and endoplasmic reticulum (ER) stress in common carp. The fish were subjected to four concentrations of H2O2 (0, 0.25, 0.5 and 1 mM) for 14 days. And then, the tissues including blood, liver, muscle, gills, intestines, heart, kidney and spleen were collected to measure biochemical parameter and gene expression. The results showed that H2O2 exposure suppressed the majority antioxidative parameters in serum, liver, muscle and intestines, but enhanced T-SOD, CAT and T-AOC levels in gills. In all tested tissues, the MDA content was significantly promoted by H2O2 exposure. The oxidative stress-related genes including nrf2, gstα, sod, cat and/or gpx1 were upregulated in liver, gills, muscle, intestines, and/or kidney, but downregulated in heart after H2O2 exposure. Moreover, the ho-1 mRNA level was inhibited by H2O2 exposure in all tissues except intestines and spleen. After 14 days of exposure, H2O2 induced ER stress and initiated IRE1 and PERK pathways, which activated downstream genes, including chop, grp78 and/or xbp1s, to regulate UPR in liver, gills, muscle and/or heart. Meanwhile, H2O2 exposure activated MAPK pathway to regulate mitochondria-related genes including bcl-2, bax and cytc, which further triggered cas-8, cas-9 and cas-3, and accelerated apoptosis in liver, gills, muscle and heart. Importantly, in different tissues, the genes associated with oxidative stress, ER stress and apoptosis showed a different influence, and more significant influence was observed in the muscle, gills and liver. Overall results suggested that long-term H2O2 exposure induced oxidative stress, ER stress and apoptosis in the majority of tested tissues of common carp. The Nrf2, IRE1, PERK and MAPK pathways played important roles in H2O2-induced toxicity in fish. These data enriched the toxicity mechanism of H2O2 in fish, which might contribute to the risk assessment of H2O2 in aquatic environment.

Effects of high-fat diet on antioxidative status, apoptosis and inflammation in liver of tilapia (Oreochromis niloticus) via Nrf2, TLRs and JNK pathways.

Fatty liver injury (or disease) is a common disease in farmed fish, but its pathogenic mechanism is not fully understood. Therefore the present study aims to investigate high-fat diet (HFD)-induced liver injury and explore the underlying mechanism in fish. The tilapia were fed on control diet and HFD for 90 days, and then the blood and liver tissues were collected to determine biochemical parameter, gene expression and protein level. The results showed that HFD feeding signally increased the levels of plasma aminotransferases and pro-inflammatory factors after 60 days. In liver and plasma, HFD feeding significantly suppressed antioxidant ability, but enhanced lipid peroxidation formation, protein oxidation and DNA damage after 60 or 90 days. Further, the Nrf2 pathway and antioxidative function-related genes were adversely changed in liver of HFD-fed tilapia after 60 and/or 90 days. Meanwhile, HFD treatment induced apoptosis via initiating mitochondrial pathway in liver after 90 days. Furthermore, after 90 days of feeding, the expression of genes or proteins related to JNK pathway and TLRs-Myd88-NF-κB pathway was clearly upregulated in HFD treatment. Similarly, the mRNA levels of inflammatory factors including tumor necrosis factor (TNF-α), interleukin-1ß (IL-1ß), IL-6, IL-8 and IL-10 were also upregulated in liver of HFD-fed tilapia after 60 and/or 90 days. In conclusion, the current study suggested that HFD feeding impaired antioxidant defense system, induced apoptosis, enhanced inflammation and led to liver injury. The adverse influences of HFD in the liver might be due to the variation of Nrf2, JNK and TLRs-Myd88-NF-κB signaling pathways.