Retrospective evaluation of leflunomide as an adjunctive therapy in dogs with non-associative immune-mediated thrombocytopenia: 20 cases (2008-2021).

OBJECTIVE: To describe leflunomide as an adjunctive therapy in the treatment of non-associative immune-mediated thrombocytopenia. MATERIALS AND METHODS: A retrospective study of dogs with a diagnosis of non-associative immune-mediated thrombocytopenia treated with leflunomide March 2008 to September 2021 was conducted. Data collected included signalment, clinical signs, physical examination findings and diagnostic testing performed. Medications administered, duration of hospital stay, time to platelet concentration >150×109/L and adverse events during leflunomide therapy were recorded. Relapses within a year of diagnosis were reported. RESULTS: A total of 20 client-owned dogs met inclusion criteria. Nineteen of 20 dogs (95%) achieved a platelet concentration >150×109/L with leflunomide and prednisone combination therapy and four dogs (21.1%) relapsed during treatment or shortly after treatment. Adverse effects included diarrhoea (n=5), mild lymphopenia (n=9) and mild intermittent anaemia (n=1). A single dog developed hepatotoxicity presumed to be secondary to leflunomide therapy that resolved after drug discontinuation. One dog was treated for aspiration pneumonia during treatment. Two dogs were euthanased while receiving leflunomide. CLINICAL SIGNIFICANCE: Length of hospitalisation, time to platelet recovery, treatment response and relapse rate were comparable with alternative treatment protocols. Most adverse effects did not require leflunomide dose adjustment; however, two dogs died while undergoing leflunomide treatment and there is compelling evidence that one of these dogs experienced fatal infection secondary to immune-suppression. Hepatotoxicity remains a known complication of leflunomide treatment and serial biochemistry testing is recommended.

Evaluation of commercial doses of a feed additive and silymarin on broiler performance with and without CCl4-induced liver damage.

Improving productive performance is a daily challenge in the poultry industry. Developing cost-effective additives and strategies that improve performance in antibiotic-free poultry production is critical to maintaining productivity and efficiency. This study evaluates the influence of a commercially available phytogenic feed additive (CA-PFA, that comprises silymarin, betaine and curcumin extracts as main ingredients) and silymarin on commercial broilers' productive performance and liver function with and without carbon tetrachloride (CCl4)-induced liver damage. The experiment was conducted in a completely randomized design, with six treatments, eight replicates, and eight birds per replicate in 18 one-day-old male broilers (Cobb Vantress 500) each; under a 3 × 2 factorial arrangement (3 diets x 2 levels of CCl4, 0 and 1 mL/kg body weight orally). The experimental treatments included 3 diets, commercially recommended doses of CA-PFA (500 mg/kg of feed; this dose provides 70 mg/kg of silymarin, besides the other active ingredients included in the formulation), silymarin (250 mg/kg of feed, containing 28% of active ingredient; this dose provides 70 mg/kg of silymarin as active ingredient) and an additive-free basal diet as a control. A standard commercial silymarin was used as a reference due to its well-known and extensively studied hepatoprotective properties that can mitigate the negative effects of CCl4 in the liver. The data were analyzed as a 2-way ANOVA, and the means showing significant (P ≤ 0.05) differences were then compared using the Post-Hoc Tukey HSD test. No interaction was detected between factors. Exposure to CCl4 had a noticeable detrimental effect on alertness, productive performance, and liver function of broilers without a significant increase in mortality. Including CA-PFA in the diet improved productive performance compared to the basal diet from day 21 to the end of the trial, on day 42. While no influence in feed intake was detected for any treatment, CA-PFA improved body weight gain (BWG) and feed conversion ratio (FCR) significantly (P < 0.05) from day 21 to the end of the trial in healthy and CCl4-exposed birds. The results show that CA-PFA supplementation improves performance parameters in broilers with and without CCl4-induced liver damage, when compared to a basal diet and the addition of a standard commercial silymarin product.

Hepatoprotective Effect of Vitamin B12 in Acetaminophen Induce Hepatotoxicity in Male Rats.

Acetaminophen is a pharmaceutical synthesized non-opioid analgesic that belongs to the "aniline analgesics" class of medicine. Because it lacks a significant anti-inflammatory effect, it is not classified as a non-steroidal anti-inflammatory therapeutic medication (NSAID). As an over-the-counter pain reliever and antipyretic, Acetaminophen is the active metabolite of phenacetin and acetanilide, but it is less toxic than either precursor. According to some medical studies, Acetaminophen toxicity can be treated with vitamin B12. Acetaminophen-poisoned Male Wister rats were the subject model of the current study, which examines the effects of vitamin B12 on their hepatic health. There were three groups of animals: Acetaminophen treated animals (750 ml/kg), vitamin B12-treated animals (0.63 g/kg), and a control group that received distilled water (750 ml/kg). All animals were given oral medication for seven days. On the seventh day, the animal was sacrificed. Plasma levels of Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Glutathione (GSH), total antioxidant capacity (TAC), Caspase3, Malondialdehyde (MDA), Interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-) were measured in the cardiac blood samples. Vitamin B12 lowers liver enzyme levels in the blood, increases overall antioxidant levels, and compensates for tissue glutathione deficiency while lowering serum elevations. TNF-α and interleukin-6 levels are also reduced by caspase3. Acetaminophen-induced hepatic necrosis and inflammatory cell infiltration were both considerably reduced by vitamin B12 supplementation. According to this study, vitamin B12 was found to have a protective effect against acetaminophen-induced hepatotoxicity.

Susceptibility to adriamycin-induced hepatotoxicity in mice depends on PRKDC polymorphism.

Adriamycin (ADR) is an effective chemotherapy drug for various cancers but has serious side effects. ADR-induced liver damage is a common problem during therapy, but the underlying mechanism remains to be fully understood. In contrast, ADR-induced glomerular damage is well studied in rodents, and sensitivity to ADR-induced nephropathy is because of the R2140C polymorphism of Prkdc gene. To investigate whether strain differences or sensitivity to ADR-induced liver damage are related to Prkdc polymorphism, this study compared the sensitivity to ADR-induced liver damage among C57BL/6J (B6J), B6-PrkdcR2140C, and BALB/c mice. Although B6J exhibits resistance to ADR-induced liver injury, BALB/c and B6-PrkdcR2140C are more susceptible to liver injury, which is exacerbated by the presence of R2140C mutation in PRKDC.

Beneficial effects of quercetin on vincristine-induced liver injury in rats: Modulating the levels of Nrf2/HO-1, NF-kB/STAT3, and SIRT1/PGC-1α.

Our experimental objective was to investigate the hepatotoxic effect of vincristine (VCR) administration in rats and determined whether combined therapy with Quercetin (Quer) ensured protection. Five groups with seven rats each were used for this purpose, and experimental groups were formulated as follows: Control group; Quer group; VCR group; VCR plus Quer 25 group; VCR plus Quer 50 group. The results showed that VCR significantly increased the activity of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) enzymes. Besides, VCR caused considerable increases in the malondialdehyde (MDA) contents, along with significant decreases in reduced glutathione levels, superoxide dismutase, catalase, and glutathione peroxidase enzyme activities in the rat livers. Quer treatment in VCR toxicity markedly decreased the activity of ALT, AST, ALP enzymes, and MDA contents and enhanced the activities of antioxidant enzymes. The results also showed that VCR significantly increased the levels of NF-kB, STAT3, and the expression of caspase 3, Bax, and MAP LC3 and decreased the expression of Bcl2 and levels of Nrf2, HO-1, SIRT1, and PGC-1α. Compared to the VCR group, Quer treatment exhibited significantly lower levels of NF-kB, STAT3, and the expression of caspase 3, Bax, and MAP LC3, and higher levels of Nrf2, HO-1, SIRT1, and PGC-1α. In conclusion, our study demonstrated that Quer could alleviate the harmful effects of VCR via activation of NRf2/HO-1 and SIRT1/PGC-1α pathways, and via attenuation of oxidative stress, apoptosis, autophagy, and NF-kB/STAT3 pathways.

Modified rougan decoction alleviates lipopolysaccharide-enrofloxacin-induced hepatotoxicity via activating the Nrf2/ARE pathway in chicken.

Liver injury plays a heavy burden on the chicken industry. Although modified rougan decoction is a prescription for the treatment of liver disease based on the classical prescription of rougan decoction (containing peony and licorice). However, the effect and mechanism of modified rougan decoction on the liver remain unclear. In this study, the effects of the water extracts (MRGD) and the alcohol precipitates of water extracts (MRGDE) against lipopolysaccharide-enrofloxacin (LPS-ENR)-induced hepatotoxicity were discussed in vivo and in vitro. The isolated hepatocytes and 128 one-day-old Hyline chickens were considered research objects. The indices of liver injury and oxidative stress were evaluated by hematoxylin and eosin (H&E) stained and the assay kits, and the nuclear erythroid 2-related factor 2 (Nrf2)/antioxidant responsive element (ARE) pathway was detected by the RT-PCR, western blot, and immunofluorescence tests. All data were analyzed using the IBM SPSS 20.0 software. In vivo, the structural integrity of the liver was maintained, AST, ALT, and MDA levels were decreased, and antioxidant enzymes were increased, confirming that the oxidative stress was reduced and liver injury was alleviated. Correspondingly, MRGD and MRGDE were observed to improve cell viability and decrease lactate dehydrogenase (LDH) in vitro, and the cell oxidative damage was reduced. In addition, the nuclear translocation of Nrf2 was improved significantly, and the mRNA and protein expression levels of the related genes were upregulated. In conclusion, MRGD and MRGDE can exert a protective effect against LPS-ENR-induced hepatotoxicity by activating the Nrf2/ARE pathway, which might be a potential therapeutic prescription for preventing or treating liver injury. Notably, no significant difference was found between the 2 extracts, suggesting that a depth extraction method did not always improve the efficacy of natural medicine. Our results provided new insights into finding effective hepatoprotective medicine.

Selenium regulates Nrf2 signaling to prevent hepatotoxicity induced by hexavalent chromium in broilers.

Hexavalent chromium (Cr(Ⅵ)) is considered to be a common environmental pollutant, which widely exists in industrial effluents and wastes and then potentially noxious effects to the health of the poultry. Studies have reported that selenium (Se), which is one of the essential trace elements of the poultry and participates in the oxidative metabolism, can alleviate Cr(Ⅵ)-induced organ damage by inhibiting oxidative stress, but its specific molecular mechanism remains unclear. Herein, animal models of Cr(Ⅵ)- and Se-exposure were constructed using broilers to investigate the antagonistic mechanism of Se to Cr(Ⅵ)-induced hepatotoxicity. In this experiment, the four groups of broiler models were used as the research objects: control, Se, Se plus Cr, and Cr groups. Histopathology and ultrastructure liver changes were observed. Liver-somatic index, serum biochemistry, oxidative stress, Nrf2 pathway related factors, and autophagy-related genes were also determined. Overall, Se was found to ameliorate the disorganized structure, hepatic insufficiency, and oxidative damage caused by Cr(Ⅵ) exposure. Electron microscopy analysis further showed that the number of autophagosomes was obviously decreased after Se treatment compared to Cr group. Furthermore, gene and protein expression analyses illustrated that the levels of Nrf2, glutathione peroxidase 1 (GPx-1), NAD(P)H: quinone oxidoreductase 1 (NQO1), and mechanistic target of rapamycin (mTOR) in the Se&Cr group was upregulated, along with decreased expression of Beclin 1, ATG5 and LC3 compared to the Cr group. These suggest that Se can repair the oxidative lesion and autophagy induced by Cr(Ⅵ) exposure in broiler livers by upregulating the Nrf2 signaling pathway.

Prevalence of hepatotoxicity and myelosuppression with alternate day use of azathioprine and glucocorticoids for treatment of dermatological conditions in dogs.

BACKGROUND: The use of azathioprine (AZA) in dogs is limited by the potential for hepatotoxicity and myelosuppression. HYPOTHESIS/OBJECTIVES: To determine the prevalence of AZA-associated hepatotoxicity in dogs with dermatological conditions receiving alternate-day AZA. The hypothesis was that dogs receiving AZA every other day (EOD) would have a lower prevalence of hepatotoxicity compared to published data for dogs receiving daily AZA. A secondary aim was to determine the prevalence of AZA-associated myelosuppression over the same time period and population. ANIMALS: Forty-one client-owned dogs with dermatological conditions treated with AZA EOD and glucocorticoids with clinical and haematological follow-up available for a minimum of two months of AZA therapy. METHODS: Retrospective analysis of data from April 1994 to July 2020. Hepatotoxicity was defined as elevation of alanine aminotransferase (ALT) at least twofold above the reference range. RESULTS: Azathioprine-associated hepatotoxicity was observed in two of 41 dogs (4.9%), with onset at 18 and 40 days, respectively. One dog receiving AZA at 1.9 mg/kg EOD had a fourfold increase in ALT. The other dog (AZA dose 2.3 mg/kg EOD) had a 30-fold increase in ALT. Azathioprine was not associated with thrombocytopenia, anaemia or neutropenia in any dogs. Lymphopenia developed in one dog (2.4%) with onset at 105 days. CONCLUSIONS AND CLINICAL RELEVANCE: Alternate-day AZA administration with tapering glucocorticoids was well-tolerated in dogs with dermatological conditions.

Hepatotoxic mechanism of diclofenac sodium on broiler chicken revealed by iTRAQ-based proteomics analysis.

BACKGROUND: At the therapeutic doses, diclofenac sodium (DFS) has few toxic side effects on mammals. On the other hand, DFS exhibits potent toxicity against birds and the mechanisms remain ambiguous. OBJECTIVES: This paper was designed to probe the toxicity of DFS exposure on the hepatic proteome of broiler chickens. METHODS: Twenty 30-day-old broiler chickens were randomized evenly into two groups (n = 10). DFS was administered orally at 10 mg/kg body weight in group A, while the chickens in group B were perfused with saline as a control. Histopathological observations, serum biochemical examinations, and quantitative real-time polymerase chain reaction were performed to assess the liver injury induced by DFS. Proteomics analysis of the liver samples was conducted using isobaric tags for relative and absolute quantification (iTRAQ) technology. RESULTS: Ultimately, 201 differentially expressed proteins (DEPs) were obtained, of which 47 were up regulated, and 154 were down regulated. The Gene Ontology classification and Kyoto Encyclopedia of Genes and Genomes pathway analysis were conducted to screen target DEPs associated with DFS hepatotoxicity. The regulatory relationships between DEPs and signaling pathways were embodied via a protein-protein interaction network. The results showed that the DEPs enriched in multiple pathways, which might be related to the hepatotoxicity of DFS, were "protein processing in endoplasmic reticulum," "retinol metabolism," and "glycine, serine, and threonine metabolism." CONCLUSIONS: The hepatotoxicity of DFS on broiler chickens might be achieved by inducing the apoptosis of hepatocytes and affecting the metabolism of retinol and purine. The present study could provide molecular insights into the hepatotoxicity of DFS on broiler chickens.

Protective effects of methanolic leaf extracts of Monanthotaxis caffra against aflatoxin B1-induced hepatotoxicity in rats.

Aflatoxins are potent hepatotoxic and carcinogenic secondary metabolites produced by toxigenic fungi. The present study investigated the protective effect of methanolic leaf extracts of Monanthotaxis caffra (MLEMC) against aflatoxin B1-induced toxicity in male Sprague-Dawley rats. The rats were randomly divided into 6 groups of 8 animals each. Five groups were administered orally for seven days with three different concentrations of MLEMC (100 mg/kg, 200 mg/kg and 300 mg/kg), curcumin (10 mg/kg) or vehicle (25% propylene glycol). The following day, these groups were administered 1 mg/kg b.w. of aflatoxin B1 (AFB1). The experiment was terminated three days after administration of AFB1. Group 6 represented untreated healthy control. Serum aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, lactate dehydrogenase, creatinine and liver histopathology were evaluated. Methanolic leaf extracts of M. caffra decreased the levels of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase and creatinine in the sera of rats as compared with the AFB1 intoxicated group. Co-administration of MLEMC improved the histological characteristics of the hepatocytes in contrast to the AFB1 treated group, which had mild to severe hepatocellular injuries including bile duct proliferation, bile duct hyperplasia, lymphoplasmacytic infiltrate and fibrosis. Extracts of M. caffra were beneficial in mitigating the hepatotoxic effects of AFB1 in rats by reducing the levels of liver enzymes and preventing hepatic injury.

Cadmium and molybdenum co-induce pyroptosis and apoptosis via the PTEN/PI3K/AKT axis in the livers of Shaoxing ducks (Anas platyrhynchos).

Cadmium (Cd) and excessive molybdenum (Mo) have adverse impacts on animals. However, the hepatotoxicity co-induced by Cd and Mo in ducks has not been fully elucidated. In order to explore the impacts of Cd and Mo co-exposure on pyroptosis and apoptosis by the PTEN/PI3K/AKT pathway in the livers of ducks, 40 healthy 7-day-old Shaoxing ducks (Anas platyrhynchos) were randomly assigned into 4 groups, and Cd or/and Mo were added to the basic diet per kilogram (kg): control group (0 mg Mo and 0 mg Cd), Mo group (100 mg Mo), Cd group (4 mg Cd), and Mo + Cd group (100 mg Mo and 4 mg Cd), with 16 weeks feed management. Results signified that Cd or/and Mo caused trace element imbalance, liver function and histomorphological abnormalities in the duck liver, and activated the PTEN/PI3K/AKT pathway through increasing PTEN mRNA and protein levels, reducing PI3K, AKT mRNA and p-AKT/AKT protein levels, which triggered pyroptosis and apoptosis via increasing Caspase-1, NLRP3, NEK7, ASC, GSDME, GSDMA, IL-1ß and IL-18 mRNA levels, Caspase-1 p20, NLRP3, ASC and GSDMD protein levels, and IL-1ß and IL-18 contents, and increasing Bak-1, Bax, Cyt C and Caspase-3 mRNA levels and cleaved Caspase-3/Caspase-3 protein level, and downregulating Bcl-2 mRNA level and the ratio of Bcl-2 to Bax, respectively. Overall, the results illustrate that pyroptosis and apoptosis induced by Cd or/and Mo may be associated with activating the PTEN/PI3K/AKT pathway in the livers of ducks. There may be a synergy between these two elements.

The difference of chows affects mouse physiological conditions.

Acetaminophen-induced liver injury in mice is a model system of human acetaminophen overdose and oxidative stress in vivo. The system is technically established, and we usually obtain severe liver damage in the treated mice; however, it is possible that the degree of liver damage is affected by the type of chow fed to mice. Thus, in this experiment, we investigated the effect of different chows on mice by comparing acetaminophen-induced liver damage, liver antioxidant level, and serum amino-acid concentrations. The results showed that differences in chows, even standard ones, affected mouse physiological conditions, with the response to oxidative stress greatly affected.

Therapeutic effects of thymoquinone in doxorubicin-induced hepatotoxicity via oxidative stress, inflammation and apoptosis.

Cancer is a lethal disease that is characterized by uncontrolled cell division and proliferation, and it results in death in many organisms. Doxorubicin (DOX) is a therapeutic agent used for treatment of many cancer types, but it induces serious hepatotoxicity. In this study, we aimed to determine possible hepato-therapeutic effects of thymoquinone (THQ) on DOX-induced hepatotoxicity in rats. Rats were divided into five groups (n = 8): Control, THQ (10 mg/kg/day/i.p for 14 days), Olive Oil (equal volume with THQ for 14 days), DOX (single dose, 15 mg/kg/i.p on 7th day) and DOX + THQ (10 mg/kg/day/i.p and DOX 15 mg/kg/i.p on 7th day). At the end of the experiment, liver tissues were extracted and evaluated histopathologically. eNOS, iNOS and Cas-3 immunostaining were performed to determine the expression levels. TUNEL method was used to determine apoptotic index. Furthermore, liver tissue total antioxidant status (TAS), total oxidant status (TOS), TNF-α and TGF-ß levels were measured by ELISA assay. The DOX group showed histopathological deterioration compared to Control group. Moreover, apoptotic index, eNOS, iNOS and Cas-3 expressions increased in DOX group. While TAS level of the DOX group decreased, TOS level increased. TNF-α and TGF-ß levels increased in DOX group. However, there was improvement in DOX + THQ group compared to DOX group. Moreover, apoptotic cell number, eNOS, iNOS and Cas-3 expressions decreased in DOX + THQ group compared to DOX group. We concluded that thymoquinone can be used as a phytotherapeutic for reducing DOX-induced liver damage.

Chitosan nanoparticles alleviated endocrine disruption, oxidative damage, and genotoxicity of Bisphenol-A- intoxicated female African catfish.

Bisphenol-A (BPA) is widely used in production of plastic products. It can reach the ecosystems affecting aquatic organisms most likely fishes. The purpose of this study was to study the toxic effects of BPA on the biochemical variables and oxidative stress in female African catfish, Clarias gariepinus and to estimate the protective role of chitosan nanoparticles (CSNPs) against BPA toxicity. Five groups in triplicates of fish were divided as follows: group I was control, group II was treated with CSNPs (0.66 ml/L), group III was exposed to BPA (1.43 µg/L), group IV was treated with BPA (1.43 µg/L) plus CSNPs (0.33 ml/L), and group V was treated with BPA (1.43 µg/L) plus CSNPs (0.66 ml/L) for 30 days. Blood and liver tissue samples were collected at the end of experiment for the biochemical and oxidative stress biomarkers analyses. Results exhibited that serum Follicle Stimulating Hormone (FSH) and 17-ß Estradiol (E2) were significantly decreased in female catfish. While, serum Testosterone (T.) and Luteinizing Hormone (LH) were increased after exposure to BPA. Marked increment in superoxide dismutase (SOD) and malondialdehyde (MDA) levels of hepatic tissue of catfish exposed to BPA. Furthermore, significant reduction in hepatic catalase (CAT), glutathione peroxidase (GSH-px), total antioxidant capacity (TAC), reduced glutathione (GSH), and glutathione S-transferase (GST) levels were decreased significantly in BPA-exposed catfish compared to the control group. However, administration of female C. gariepinus with the low and high doses (0.33 ml/L and 0.66 ml/L) of CNPs restored the biochemical parameters to be close to the normal values of the control group and also, reduced oxidative stress induced by BPA toxicity. This improvement was evident in fish administrated with the high CSNPs dose (0.66 ml/L) compared to catfish exposed to BPA in group (III). Furthermore, the percentage of hepatic DNA damage was detected in group III exposed to BPA alone. However, it was declined after co- administration with both the low and high doses of CSNPs. The study has revealed that treatment with CSNPs has antagonistic functions against the toxicity of BPA in female African catfish.

Hepatogenous photosensitisation in cows grazing turnips (Brassica rapa) in South Africa.

Holstein cows on a farm in the Humansdorp district, Eastern Cape province, South Africa, developed reddened, painful teat skin 3 days after grazing a mixed forage crop dominated by bulb turnip (Brassica rapa, Barkant cultivar). The crop was grazed 45 days after planting and 10% of the herd developed symptoms. More characteristic non-pigmented skin lesions started manifesting 1-2 days after the appearance of the teat lesions. Affected cows had elevated serum activities of gamma-glutamyl transferase, glutamate dehydrogenase and aspartate aminotransferase. These blood chemistry findings confirmed a secondary (hepatogenous) photosensitivity. As a result of the severity of the teat and skin lesions, seven cows were slaughtered and tissue samples from five of them were collected for histopathological examination. Liver lesions in cows that were culled 3 or more weeks after the onset of the outbreak showed oedematous concentric fibrosis around medium-sized bile ducts and inflammatory infiltrates in portal tracts. Characteristic lesions associated with other known hepatobiliary toxicities were not found. No new cases were reported 5 days after the cattle were removed from the turnips. The sudden introduction of the cows, without any period of transitioning or adaptation to grazing turnips, as well as the short latent period, clinical signs of photosensitisation, blood chemistry and histopathology, confirmed a diagnosis of Brassica-associated liver disease, a condition seen in New Zealand but not previously described in South Africa. Brassica forage crops are potentially toxic under certain conditions and farmers must be aware of these risks.

Can the exposure system adopted influence the results of the atrazine toxicity in hepatic tissue of fish?

The widespread use of atrazine, a herbicide used to control weeds, has contributed to the increased contamination of aquatic environments. To assess the toxicological effects of a xenobiotic on a nontarget organism in the laboratory, different models of toxicological exposure systems have been widely used. Therefore, the aim of this study was to evaluate and compare the action of sublethal concentrations of atrazine on the hepatic histology of Oreochromis niloticus, considering two models of exposure: static (where atrazine was only added once) and semi-static (where atrazine was periodically renewed). Fish were exposed to a concentration of 2 ppm atrazine for 15 days, which was verified by high-performance liquid chromatography. The livers were stained with hematoxylin and eosin and histopathological data were collected. In addition, they were submitted to immunohistochemistry for inducible nitric oxide synthase (iNOS). A maximum variation of 45% (static) and 12.5% (semi-static) was observed between the observed and nominal atrazine concentration. Nuclear and cytoplasmic changes were observed in both experimental models. Hepatocytes from the livers of the static system showed a degenerative appearance, while in the semi-static system, intense cytoplasmic vacuolization and necrosis were observed. iNOS positive cells were identified only in macrophages in the hepatocytes of fish in the semi-static system. These results directly showed how the choice of exposure system can influence the results of toxicological tests. However, future analysis investigating the by-products and nitrogen products should be carried out since the histopathological findings revealed the possibility of these compounds serving as secondary contamination routes.

Assessment of Hepatotoxicity Induced by Aluminum Oxide Nanoparticles in Oreochromis niloticus Using Integrated Biomarkers: Exposure and Recovery.

The hepatotoxic impacts of 2, 4, and 8 mg/L of Al2O3 nanoparticles (31.4 ± 4.8 nm) were evaluated in Oreochromis niloticus after 7 days of exposure and 15 days of recovery periods. The biochemical analysis of aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase in plasma showed significant increases in both 4 and 8 mg/L Al2O3 NPs exposed groups. The antioxidant biomarkers showed concentration-dependent elevations in catalase, superoxide dismutase, glutathione peroxidase activities, and thiobarbituric acid reactive substances levels. Glutathione reduced contents showed significant reductions in both 4 and 8 mg/L Al2O3 nanoparticles exposed groups. Several hepatic histopathological alterations were recorded ranging from adaptive responses (e.g. melanomacrophages aggregation) to permanent damage (e.g. necrosis). The recovery period using toxicant-free water led to an obvious reduction in the Al content in liver, liver and antioxidant enzymes in addition to regressive histopathological alterations based on the frequency of alterations occurrence and the extent of affected areas.

Involvement of neutrophils in rat livers by low-dose thioacetamide administration.

The administration with high dose (close to LD50) of thioacetamide (TAA), a hepatotoxicant used widely to induce experimental liver lesions, develops hepatocellular necrosis and subsequent inflammation (mainly M1-/M2-macrophages without neutrophil infiltration) in rats. We analyzed rat livers treated with a low dose TAA (50 mg/kg/body weight) at 6, 12, 18, 24 and 48 hr. The lesions in the affected centrilobular areas consisted of slight hepatocyte degeneration at 12 hr, and inflammatory cell infiltration at 18 and 24 hr; the lesions recovered until 48 hr. Translocation of intranuclei to cytoplasm of HMGB1, a representative molecule of damage-associated molecular patterns, was seen in some hepatocytes mainly at 6, 12, and 18 hr. As an interesting finding, at 12 hr, myeloperoxidase-positive neutrophil infiltration was observed in the affected centrilobular area. Additionally, CD68 M1-/CD163 M2-macrophages increased consistently at 12 to 48 hr. CXCL1, a chemokine for induction of neutrophils, began to increase at 6 hr and gradually increased at 12, 18 and 24 hr, apparently corresponding to the appearance of neutrophils. Collectively, the present findings at the low dose TAA indicated that along with M1-/M2-macrophages, neutrophils were characteristically seen, which might be elicited by cytoplasmic translocation of HMGB1 from nuclei. These finding would be useful for evaluation of hepatotoxicity at the early stages.

Chronic facial eczema in sheep: description of gross and histological changes in the liver and association with serum gamma-glutamyltransferase activity at the time of sporidesmin intoxication.

AIMS: To determine the gross and histological changes developing in the liver of sheep 8 months after a single period of exposure to sporidesmin and to examine associations between the severity of gross and histological changes to the liver and the activity of gamma-glutamyltransferase (GGT) measured in serum in the sheep at the time of intoxication. METHODS: A group of 50 Romney ewes grazing a mixed ryegrass/white clover pasture were accidentally exposed to sporidesmin for up to 5 weeks. Seventeen sheep showed photosensitisation and four were subject to euthanasia. The remaining sheep were moved to safer pasture and a blood sample collected and analysed for serum GGT activity. The sheep were slaughtered 8 months later. Livers were classified into grossly normal, moderately affected, or severely affected and histology performed to assess portal fibrosis, biliary hyperplasia, portal inflammation, and hepatocellular necrosis. RESULTS: Serum GGT activity ranged from 59 to 1571 IU/L (reference range 32-70 IU/L). Thirteen of the 46 sheep developed clinical signs of facial eczema. However, at slaughter all except four sheep had grossly detectable changes to the shape of the liver including atrophy of the left lobe and the lateral part of the right lobe. Hypertrophy was typically limited to the medial part of the right lobe. In severely affected sheep the liver hypertrophy formed a nodular bulging mass. Changes in the liver shape were classified as severe in 25 and moderate in 17 sheep. Severely affected livers contained significantly more fibrosis than moderately affected livers (p = 0.001, Cliff's delta (d) = 0.68). While there was significantly greater fibrosis and biliary hyperplasia in the left than right lobes, histological changes were present throughout all samples taken of affected livers. Serum GGT activity taken during acute intoxication were correlated to subsequent fibrosis and biliary hyperplasia. CONCLUSIONS: Hepatic fibrosis develops in sheep after a single episode of sporidesmin intoxication, even in sheep with only mildly elevated GGT activity at the time of intoxication. Furthermore, the severity of the subsequent hepatic fibrosis was predicted by the degree of elevation of serum GGT activity during intoxication. CLINICAL RELEVANCE: More research is required to determine how the presence and severity of hepatic fibrosis affect animal production. However, if hepatic fibrosis does decrease production, the consistent development of fibrosis after sporidesmin ingestion reinforces the importance of avoiding exposure of livestock to sporidesmin. ABBREVIATIONS: GGT: Gamma-glutamyltransferase; d: Cliff's delta.