Hepatic Glutathione Depletion Research Articles

In vivo upstream factors of mouse hepatotoxic mechanism with sustained hepatic glutathione depletion: Acetaminophen metabolite-erythrocyte adducts and splenic macrophage-generated reactive oxygen species

Investigation of acetaminophen (APAP)-induced liver damage recently indicated the significance of phagocytic NADPH oxidase (NOX)-derived reactive oxygen species (ROS) and ferroptosis in the liver. Here, we focused on phagocytosis by iron-containing erythrocyte-devouring splenic macrophages and explored upstream factors of known APAP hepatotoxic mechanisms in vivo. Splenectomy did not alter hepatic cytochrome P450 (CYP) 2E1 activity or hepatic glutathione (GSH) content. APAP injection into splenectomized mice almost completely suppressed increases in plasma alanine aminotransferase levels and centrilobular hepatic necrosis showing the spleen to be a critical tissue in APAP-induced liver damage. Hepatic GSH was recovered to approximately 50 % content at 8 h. In non-splenectomized mice, liver damage was dramatically suppressed by a sensitive redox probe (DCFH-DA), macrophage-depleting clodronate (CL), and a NOX2 inhibitor. APAP treatment resulted in markedly stronger fluorescence intensity from DCFH-DA due to excessive ROS around splenic macrophages, which was lost upon co-treatment with a CYP inhibitor and CL. Deformed erythrocytes disappeared in mice co-treated with DCFH-DA, CL, the NOX2 inhibitor, and the CYP inhibitor. Simultaneously, these four compounds significantly improved APAP-depleted GSH levels. The CYP inhibitor also prevented the formation of APAP-cell adducts in the blood and spleen. In the spleen, CL co-treatment markedly reduced the number of adducts. Splenic ferrous iron levels were significantly elevated by APAP. Therefore, we demonstrated that splenic macrophages devoured APAP metabolite-erythrocyte adducts and subsequently splenic macrophage-related ROS caused sustained hepatic GSH depletion and excessive erythrocyte deformation around 7 h. Our data indicate in vivo upstream factors of known APAP hepatotoxic mechanisms.

Depletion of hepatic glutathione and adenosine by glucocorticoid exposure in Wistar rats is pregnancy-independent

Liver diseases have gained increasing attention due to their substantial impact on health, independently as well as in association with cardio-metabolic disorders. Studies have suggested that glutathione and adenosine assist in providing protection against oxidative stress and inflammation while glucocorticoid (GC) therapy has been associated with chronic inflammatory disorders, even in pregnancy. The implications of Glucocorticoid exposure on maternal health and fetal growth is a concern, however, the possible role of glutathione and adenosine has not been thoroughly investigated. The study therefore hypothesize that exposure to glucocorticoids leads to depletion of hepatic glutathione and adenosine levels, contributing to oxidative stress and tissue injury. Additionally, we aim to investigate whether the effects of glucocorticoids on hepatic health are pregnancy dependent in female rats. Twelve Pregnant and twelve age-matched non-pregnant rats were used for this study; an exogenous administration of glucocorticoid (Dex: 0.2 mg/kg) or vehicle (po) was administered to six pregnant and six non-pregnant rats from gestational day 14 to 19 or for a period of 6 days respectively. Data obtained showed that GC exposure led to a decrease in hepatic glucose-6-phosphate dehydrogenase, glutathione peroxidase, GSH/GSSG ratio and adenosine content in both pregnant and non-pregnant rats. In addition, increased activities of adenosine deaminase and xanthine oxidase, along with increased production of uric acid and increased levels of lactate dehydrogenase, aspartate aminotransferase, alanine transferase, alkaline phosphatase and gamma-glutamyl transferase were observed. In summary, the study indicates that GC-induced liver damage is underlined by depleted hepatic adenosine and glutathione levels as well as elevated markers of tissue inflammation and/or injury. Furthermore, the findings suggest that the effects of GC exposure on hepatic health are pregnancy independent.

Hepatocyte-specific deletion of small heterodimer partner protects mice against acetaminophen-induced hepatotoxicity via activation of Nrf2.

Acetaminophen (APAP) overdose stands as the primary cause of acute liver failure in the United States. APAP hepatotoxicity involves hepatic glutathione (GSH) depletion and mitochondrial damage. To counteract the toxicity of APAP, the nuclear factor erythroid 2 like 2 (Nrf2) activates the expression of genes responsible for drug detoxification and GSH synthesis. In this study, we present evidence that the elimination of hepatocyte small heterodimer partner, a critical transcriptional repressor for liver metabolism, results in Nrf2 activation and protects mice from APAP-induced acute liver injury. Initial investigations conducted on wildtype (WT) mice revealed a swift downregulation of Shp mRNA within the first 24 h after APAP administration. Subsequent treatment of hepatocyte-specific Shp knockout (ShpHep-/-) mice with 300 mg/kg APAP for 2 h exhibited comparable bioactivation of APAP with that observed in the WT controls. However, a significant reduction in liver injury was observed in ShpHep-/- after APAP treatment for 6 and 24 h. The decreased liver injury correlated with a faster recovery of GSH, attributable to heightened expression of Nrf2 target genes involved in APAP detoxification and GSH synthesis. Moreover, in vitro studies revealed that SHP protein interacted with NRF2 protein, inhibiting the transcription of Nrf2 target genes. These findings hold relevance for humans, as overexpression of SHP hindered APAP-induced NRF2 activation in primary human hepatocytes. In conclusion, our studies have unveiled a novel regulatory axis involving SHP and NRF2 in APAP-induced acute liver injury, emphasizing SHP as a promising therapeutic target in APAP overdose-induced hepatotoxicity.

Evaluation of drug-induced liver toxicity of trovafloxacin and levofloxacin in a human microphysiological liver model

Drug-induced liver injury induced by already approved substances is a major threat to human patients, potentially resulting in drug withdrawal and substantial loss of financial resources in the pharmaceutical industry. Trovafloxacin, a broad-spectrum fluoroquinolone, was found to have unexpected side effects of severe hepatotoxicity, which was not detected by preclinical testing. To address the limitations of current drug testing strategies mainly involving 2D cell cultures and animal testing, a three-dimensional microphysiological model of the human liver containing expandable human liver sinusoidal endothelial cells, monocyte-derived macrophages and differentiated HepaRG cells was utilized to investigate the toxicity of trovafloxacin and compared it to the structurally-related non-toxic drug levofloxacin. In the model, trovafloxacin elicited vascular and hepatocellular toxicity associated with pro-inflammatory cytokine release already at clinically relevant concentrations, whereas levofloxacin did not provoke tissue injury. Similar to in vivo, cytokine secretion was dependent on a multicellular immune response, highlighting the potential of the complex microphysiological liver model for reliably detecting drug-related cytotoxicity in preclinical testing. Moreover, hepatic glutathione depletion and mitochondrial ROS formation were elucidated as intrinsic toxicity mechanisms contributing to trovafloxacin toxicity.

The Role of the Optic Atrophy 1 (OPA1) Protein in Drug‐Induced Liver Injury

The liver is the metabolic hub, and is responsible for the myriad of processes including the nutrient homeostasis and detoxification. Mitochondria of liver are critical for these functions. The detoxification process in liver, when severe, often results in liver damage through causing oxidative stress. Mitochondria are the main source of ROS and are also vulnerable to oxidant damage. Therefore, mitochondrial dysfunction is one of the prominent causes for drug‐induced liver injury.Mitochondrial fission and fusion, the main processes of mitochondrial dynamics, determine mitochondrial shape, and are important for functional maintenance of mitochondria. However, the role of mitochondrial dynamics in drug‐induced liver injury is poorly understood. In the current study, we examined the role of the optic atrophy 1 (OPA1) protein in drug‐induced liver injury. OPA1 is associated with mitochondrial inner membrane (IM) and mediates IM fusion. OPA1 also regulates cristate structure, and is required for proper electron transport and ATP production. To investigate the OPA1’s role, we used liver‐specific OPA1‐knockout (OPA1‐LKO) mice with acetaminophen (APAP) administration as a model for drug‐induced liver injury.We generated OPA1‐LKO mice by crossing OPA1 flox mice with mice carrying the Cre recombinase under the albumin promoter. Whereas whole body KO of OPA1 causes embryonic lethality, OPA1‐LKO mice appeared healthy and showed normal growth and behavior. Although the OPA1 gene in the liver was disrupted, OPA1‐LKO mice showed approximately 30% of OPA1 remaining in the liver, presumably due to less efficient albumin promoter‐mediated Cre expression and from other cell types of the liver. Liver histology revealed that OPA1‐KO livers have disorganized hepatic cords with enlarged hepatocytes. Despite a reduced OPA1 level, mitochondria in OPA1‐KO liver show near intact cristae structure and respiration, suggesting that a low level of OPA1 would support mitochondrial function in liver.We then tested the effect of OPA1 LKO on liver function under APAP stress. In APAP overdose, excess APAP metabolite depletes GSH in hepatocytes, which causes mitochondrial oxidative stress, leading to mitochondrial permeability transition, mitochondrial dysfunction, ATP depletion, and ultimately necrotic cell death. Upon administration of excess APAP, we found that OPA1‐LKO mice were more sensitive to APAP‐induced liver injury compared with the control mice. Histological analyses showed significantly more expanded focal centrilobular necrosis with vacuolization, cell swelling, and nuclear disintegration in OPA1‐KO livers. Alanine aminotransferase levels, as a clinical chemistry parameter, were higher in OPA1‐LKO mice than in control mice with APAP overdose. Furthermore, phospho‐JNK levels were higher in OPA1‐KO livers, indicating increased initial oxidative stress. However, depletion of hepatic glutathione (GSH) contents and reduction of GSH/GSSG ratio upon APAP treatment were similar between the LKO and control liver. Together, our experimental results indicate that although liver is tolerant to a reduced level of OPA1, OPA1 depletion lowers the stress threshold and makes hepatocytes more sensitive to APAP‐induced liver injury.

A structurally engineered fatty acid, icosabutate, suppresses liver inflammation and fibrosis in NASH

Although long-chain omega-3 fatty acids (LCn-3FAs) regulate inflammatory pathways of relevance to non-alcoholic steatohepatitis (NASH), their susceptibility to peroxidation may limit their therapeutic potential. We compared the metabolism of eicosapentaenoic acid (EPA) with an engineered EPA derivative (icosabutate) in human hepatocytes invitro and their effects on hepatic glutathione metabolism, oxidised lipids, inflammation, and fibrosis in a dietary mouse model of NASH, and in patients prone to fatty liver disease. Oxidation rates and cellular partitioning of EPA and icosabutate were compared in primary human hepatocytes. Comparative effects of delayed treatment with either low- (56mg/kg) or high-dose (112mg/kg) icosabutate were compared with EPA (91mg/kg) or a glucagon-like peptide 1 receptor agonist in a choline-deficient (CD), L-amino acid-defined NASH mouse model. To assess the translational potential of these findings, effects on elevated liver enzymes and fibrosis-4 (FIB-4) score were assessed in overweight, hyperlipidaemic patients at an increased risk of NASH. In contrast to EPA, icosabutate resisted oxidation and incorporation into hepatocytes. Icosabutate also reduced inflammation and fibrosis in conjunction with a reversal of CD diet-induced changes in the hepatic lipidome. EPA had minimal effect on any parameter and even worsened fibrosis in association with depletion of hepatic glutathione. In dyslipidaemic patients at risk of NASH, icosabutate rapidly normalised elevated plasma ALT, GGT and AST and reduced FIB-4 in patients with elevated ALT and/or AST. Icosabutate does not accumulate in hepatocytes and confers beneficial effects on hepatic oxidative stress, inflammation and fibrosis in mice. In conjunction with reductions in markers of liver injury in hyperlipidaemic patients, these findings suggest that structural engineering of LCn-3FAs offers a novel approach for the treatment of NASH. Long-chain omega-3 fatty acids are involved in multiple pathways regulating hepatic inflammation and fibrosis, but their susceptibility to peroxidation and use as an energy source may limit their clinical efficacy. Herein, we show that a structurally modified omega-3 fatty acid, icosabutate, overcame these challenges and had markedly improved antifibrotic efficacy in a mouse model of non-alcoholic steatohepatitis. A hepatoprotective effect of icosabutate was also observed in patients with elevated circulating lipids, in whom it led to rapid reductions in markers of liver injury.

Effect of vinyl chloride exposure on cardiometabolic toxicity.

Vinyl chloride (VC) is an organochlorine mainly used to manufacture its polymer polyvinyl chloride, which is extensively used in the manufacturing of consumer products. Recent studies suggest that chronic low dose VC exposure affects glucose homeostasis in high fat diet-fed mice. Our data suggest that even in the absence of high fat diet, exposure to VC (0.8ppm, 6h/day, 5day/week, for 12 weeks) induces glucose intolerance (1.0g/kg, i.p.) in male C57BL/6 mice. This was accompanied with the depletion of hepatic glutathione and a modest increase in lung interstitial macrophages. VC exposure did not affect the levels of circulating immune cells, endothelial progenitor cells, platelet-immune cell aggregates, and cytokines and chemokines. The acute challenge of VC-exposed mice with LPS did not affect lung immune cell composition or plasma IL-6. To examine the effect of VC exposure on vascular inflammation and atherosclerosis, LDL receptor-KO mice on C57BL/6 background maintained on western diet were exposed to VC for 12 weeks (0.8ppm, 6h/day, 5day/week). Unlike the WT C57BL/6 mice, VC exposure did not affect glucose tolerance in the LDL receptor-KO mice. Plasma cytokines, lesion area in the aortic valve, and markers of lesional inflammation in VC-exposed LDL receptor-KO mice were comparable with the air-exposed controls. Collectively, despite impaired glucose tolerance and modest pulmonary inflammation, chronic low dose VC exposure does not affect surrogate markers of cardiovascular injury, LPS-induced acute inflammation in C57BL/6 mice, and chronic inflammation and atherosclerosis in the LDL receptor-KO mice.

Zerumbone Exhibit Protective Effect against Zearalenone Induced Toxicity via Ameliorating Inflammation and Oxidative Stress Induced Apoptosis.

Zearalenone are widely occurring food contaminants that cause hepatotoxicity. This research work aimed to investigate how zerumbone, a plant-derived dietary compound, can fight ZEA-induced hepatotoxicity. ZER is found to increase the cells’ toxin resistance. This study was performed on mice challenged with ZEA. The administration of ZER decreased the level of alkaline phosphatase and alanine aminotransferase (ALT). Simultaneously, ZER attenuated the inflammatory response via significantly reducing the levels of pro-inflammatory factors, including interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α) in serum. Pretreatment with ZER reduced the hepatic malondialdehyde (MDA) concentration, as well as the depletion of hepatic superoxide dismutase (SOD), hepatic glutathione (GSH), and hepatic catalase (CAT). Moreover, it significantly ameliorated ZEA-induced liver damage and histological hepatocyte changes. ZER also relieved ZEA-induced apoptosis by regulating the PI3K/AKT pathway and Nrf2 and HO-1 expression. Furthermore, ZER increasingly activated Bcl2 and suppressed apoptosis marker proteins. Our findings suggest that ZER exhibits the ability to prevent ZEA-induced liver injury and present the underlying molecular basis for potential applications of ZER to cure liver injuries.

Eupatilin pretreatment ameliorates hepatic ischemia-reperfusion injury in mice

Eupatilin, a pharmacologically active flavone derived from Artemisia species, is known to have antioxidant and anti-inflammatory activities. Ischemia-reperfusion injury (IRI) is a major critical event that commonly occurs after liver transplantation and resection. Furthermore, inflammatory responses to IRI exacerbate the resultant hepatic injury. In this study, we investigated whether eupatilin protects against IR-induced acute liver injury in mice. Partial (70%) hepatic IRI was induced in male C57BL/6 mice by portal triad pedicle occlusion for 45 minutes followed by reperfusion for 6 hours. Eupatilin (10 mg/kg body weight, oral) was administered 4 days before the IRI. Treatment with eupatilin significantly decreased serum alanine aminotransferase and serum aspartate aminotransferase as well as liver histological changes. Eupatilin also prevented hepatic glutathione depletion and increased malondialdehyde levels induced by IRI. Western blotting indicated that eupatilin significantly increased the levels of heat shock protein 70 and B-cell lymphoma 2 protein, attenuated inducible nitric oxide synthase, and cleaved caspase-3 levels 6 hours after IRI. The expression of the toll-like receptor 2/4, and phosphorylated nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor (p-IkB-α) was significantly decreased in the eupatilin pretreatment group. Eupatilin improved the acute hepatic IRI by reducing inflammation and apoptosis. These findings suggest that eupatilin is a promising therapeutic agent against acute IR-induced hepatic damage.

Thrombospondin‐1 Inhibition Reduces Nrf2 Activity and Increases Liver Injury during Acetaminophen‐Induced Hepatotoxicity

Introduction Acetaminophen (N-Acetyl-p-Aminophenol or APAP)-induced hepatotoxicity is the most common cause of acute liver failure in the United States and Western Europe. Toxicity from APAP results from the accumulation of the toxic metabolite NAPQI, a consequence of hepatic glutathione (GSH) depletion. Therefore, pathways regulating GSH and antioxidant pathways can play a pivotal role in APAP-induced hepatotoxicity. Thrombospondin-1 (TSP1) is a homotrimeric protein that can modulate antioxidant signaling pathways and other signaling pathways involved in cell death and inflammation. Therefore, the hypothesis of this study was that TSP1 was upregulated in the liver during APAP-induced hepatotoxicity and inhibition of TSP1 can contribute to liver injury by increasing oxidative damage and hepatic cell death. Methods C57Bl/6 mice or TSP1 null mice (TSP1-/-) were administered 300 mg/kg or 600 mg/kg of APAP and were euthanized either 2-hours or 6-hours after injection. Liver injury was assessed by measuring serum ALT and AST levels as well as liver H&E and TUNEL staining. TSP1, CYP2E1, Nrf2, and SOD1 expression were assessed by real-time PCR, immunoblotting, ELISA or immunohistochemistry analyses. Liver GSH levels were measured by colorimetric assay. The GSH synthesis enzymes GCLC, GCLM and GSR were assessed by RTPCR. All data were analyzed using Graphpad Prism software with differences between two groups compared using the Student's t-test and differences between three or more groups were determined by ANOVA. Results C57Bl/6 mice administered 300 mg/kg or 600 mg/kg had a significant increase of TSP1 mRNA and protein in their livers 6-hours after injection with this effect being dose-dependent. TSP1-/- mice had a similar degree of hepatic necrosis at 2-hours and 6-hours following APAP injection compared to C57Bl/6 mice. However, TSP1-/- mice had a significant increase of serum ALT and AST levels as well as increased TUNEL staining 6-hours following APAP administration. Nrf2 and SOD1 protein expression were significantly decreased in TSP1-/-mice but not C57Bl/6 mice 6-hours after APAP injection. This was associated with decreased GSH, GCLC, GCLM and GSR in APAP-treated TSP1-/- mice compared to C57Bl/6 mice administered APAP. Conclusions Together, these data demonstrate that elimination of TSP1 protein in APAP-treated mice reduced hepatic Nrf2 expression and GSH activity leading to worse liver injury. Treatment strategies aimed at inducing Nrf2 activity or increasing TSP1 expression may be beneficial to improve outcomes during APAP-induced liver injury.

Chrysene, a four-ring polycyclic aromatic hydrocarbon, induces hepatotoxicity in mice by activation of the aryl hydrocarbon receptor (AhR)

Polycyclic aromatic hydrocarbons (PAHs) are a group of persistent organic global environmental pollutants and cause harmful effects on human health. Here, we evaluated adverse effects of chrysene, which is a four-ring PAH and an important member of 16 priority PAHs, on the liver. Chrysene was detected in some common raw and cooked Chinese food samples. Hepatotoxicity including increased relative liver weight, hepatocyte swelling and degeneration, and elevated serum alanine aminotransferase (ALT) levels were observed in chrysene-exposed C57BL/6 mice. Glutamine treatment effectively ameliorated chrysene-induced mice liver injury by decreasing serum ALT levels. Chrysene induced mice hepatic glutathione depletion and oxidative DNA damage with increased 8-hydroxy-2′-deoxyguanosine (8-OHdG) levels. Hepatic expression levels of the aryl hydrocarbon receptor (AhR), AhR-related target genes including CYP1A1, CYP1A2 and CYP1B1, and AhR nuclear translocator (ARNT) were significantly increased in chrysene-exposed C57BL/6 mice. Chrysene induced mice hepatic mRNA levels of the nuclear factor erythroid 2–related factor 2 (Nrf2) and Nrf2-mediated phase II detoxifying and antioxidant enzymes including NQO1, UGT1A1, UGT1A6, SULT1A1, GSTm1, GSTm3, Catalase (CAT), GPx1, and SOD2. We found that chrysene had toxic effects including increased relative liver weight and elevated serum ALT levels on AhR+/+ mice but not AhR−/- mice. Chrysene significantly induced hepatic mRNA levels of CYP1A1 and CYP1A2 in AhR+/+ mice but not AhR−/- mice. To our knowledge, this study is the first to demonstrate that hepatotoxicity causes by chrysene is dependent on AhR, and Nrf2 plays an important regulation role in protection against oxidative liver injury induced by chrysene.

Noninvasive monitoring of hepatic glutathione depletion through fluorescence imaging and blood testing.

Hepatic glutathione plays a key role in regulating redox potential of the entire body, and its depletion is known to increase susceptibility to oxidative stress involved in many diseases. However, this crucial pathophysiological event can only be detected noninvasively with high-end instrumentation or invasively with surgical biopsy, limiting both preclinical research and clinical prevention of oxidative stress-related diseases. Here, we report that both in vivo fluorescence imaging and blood testing (the first-line detection in the clinics) can be used for noninvasive and consecutive monitoring of hepatic glutathione depletion at high specificity and accuracy with assistance of a body-clearable nanoprobe, of which emission and surface chemistries are selectively activated and transformed by hepatic glutathione in the liver sinusoids. These findings open a new avenue to designing exogenous blood markers that can carry information of local disease through specific nanobiochemical interactions back to the bloodstream for facile and rapid disease detection.

Rutaecarpine Protects against Acetaminophen-Induced Acute Liver Injury in Mice by Activating Antioxidant Enzymes.

Rutaecarpine, an indolopyridoquinazolinone alkaloid isolated from the unripe fruit of Evodia rutaecarpa, is used to treat hypertension, postpartum hemorrhage, dysentery, and amenorrhea as a traditional medicine in Asia. We investigated the effect of rutaecarpine on acetaminophen-induced hepatotoxicity in mice. Rutaecarpine was administered orally daily for seven consecutive days, followed by intraperitoneal injection of acetaminophen in mice on day seven to induce hepatotoxicity. Rutaecarpine pretreatment significantly decreased acetaminophen-induced serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST) activities and hepatic malondialdehyde content and prevented acetaminophen-induced hepatic glutathione depletion. Furthermore, CYP2E1 expression was decreased by rutaecarpine pretreatment in a dose-dependent manner. Rutaecarpine pretreatment inhibited acetaminophen-induced expression of inflammatory cytokines by inhibiting NF-κB activation by JNK1/2. Also, rutaecarpine pretreatment promoted Nrf2-mediated activation of the antioxidant enzymes GCLC, HO-1, and NQO1. This indicates that the protective effect of rutaecarpine during acetaminophen-induced acute liver injury is mediated by the activation of antioxidant enzymes. Therefore, rutaecarpine has a protective effect of APAP-induced liver damage.

Tetrahydroxy stilbene glycoside attenuates acetaminophen-induced hepatotoxicity by UHPLC-Q-TOF/MS-based metabolomics and multivariate data analysis.

Tetrahydroxy stilbene glycoside (TSG) is a main active compound in Polygonum multiflorum. Acetaminophen (APAP) is a well-known analgesic and antipyretic drug. It is considered to be safe within a therapeutic range, in case of acute intoxication hepatotoxicity occurs. This present study aims to observe TSG-provided alleviation on APAP-induced hepatoxicity in C57BL/6 mice. APAP performs extensive necrosis and dissolves nucleus suggesting liver damage from hepatic histopathology. Serum alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase, and alkaline phosphatase analysis and liver histological evaluation showed that TSG reduced the hepatotoxicity induced by a toxic dose of APAP. Moreover, TSG alone had no hepatotoxicity. TSG eliminated hepatic glutathione depletion and cysteine adducts formation. It also reduced the expression of interleukin-10 and lowered the production of reactive oxygen species in liver tissues. Luminex was used to detect cytokine production in different groups. Herein, we used an untargeted metabolomics approach by performing ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry on treated mice to identify metabolic disruptions under APAP and TSG. Major alterations were observed for purine metabolism, amino acid metabolism, and fatty acid metabolism. These data provide metabolic evidence and biomarkers in the liver that the ABC transporters, Glycine serine and threonine metabolism, and Choline metabolism in cancer changed the most. These targets of metabolites have the potential to improve our understanding of homeostatic. Meanwhile, these metabolites revealed that TSG can alleviate inflammation caused by APAP and promote the activity of intrinsic antioxidants. In summary, TSG can regulate lipid metabolism, promote the production of antioxidant enzymes, and decrease the inflammatory response.

Combined alcoholic and non-alcoholic steatohepatitis.

SummaryWhile metabolic syndrome and alcohol consumption are the two main causes of chronic liver disease, one of the two conditions is often predominant, with the other acting as a cofactor of morbimortality. It has been shown that obesity and alcohol act synergistically to increase the risk of fibrosis progression, hepatic carcinogenesis and mortality, while genetic polymorphisms can strongly influence disease progression. Based on common pathogenic pathways, there are several potential targets that could be used to treat both diseases; based on the prevalence and incidence of these diseases, new therapies and clinical trials are needed urgently.

Acetaminophen Hepatotoxicity.

Acetaminophen (APAP) is one of the most popular and safe pain medications worldwide. However, due to its wide availability, it is frequently implicated in intentional or unintentional overdoses where it can cause severe liver injury and even acute liver failure (ALF). In fact, APAP toxicity is responsible for 46% of all ALF cases in the United States. Early mechanistic studies in mice demonstrated the formation of a reactive metabolite, which is responsible for hepatic glutathione depletion and initiation of the toxicity. This insight led to the rapid introduction of N-acetylcysteine as a clinical antidote. However, more recently, substantial progress was made in further elucidating the detailed mechanisms of APAP-induced cell death. Mitochondrial protein adducts trigger a mitochondrial oxidant stress, which requires amplification through a MAPK cascade that ultimately results in activation of c-jun N-terminal kinase (JNK) in the cytosol and translocation of phospho-JNK to the mitochondria. The enhanced oxidant stress is responsible for the membrane permeability transition pore opening and the membrane potential breakdown. The ensuing matrix swelling causes the release of intermembrane proteins such as endonuclease G, which translocate to the nucleus and induce DNA fragmentation. These pathophysiological signaling mechanisms can be additionally modulated by removing damaged mitochondria by autophagy and replacing them by mitochondrial biogenesis. Importantly, most of the mechanisms have been confirmed in human hepatocytes and indirectly through biomarkers in plasma of APAP overdose patients. The extensive necrosis caused by APAP overdose leads to a sterile inflammatory response. Although recruitment of inflammatory cells is necessary for removal of cell debris in preparation for regeneration, these cells have the potential to aggravate the injury. This review touches on the newest insight into the intracellular mechanisms of APAP-induced cells death and the resulting inflammatory response. Furthermore, it discusses the translation of these findings to humans and the emergence of new therapeutic interventions.

Codium fragile extract Pretreatment Ameliorates Hepatic Ischemia-Reperfusion Injury in Mice.

Introduction: Ulvan is a complex of sulfated polysaccharides derived from marine green seaweeds. One of marine green algae is codium fragile (CF). Ulvan demonstrated antioxidant as well as potential anti-inflammatory properties in previous studies. Ischemia-reperfusion injury (IRI) is a major critical event that commonly occurs after liver transplantation and resection. Reactive oxygen species–mediated release of related inflammatory factors have important roles in hepatic IRI. In this study, we investigated whether CF extract protects against IRI-induced acute liver injury in mice. Method: Partial (70%) hepatic IRI was induced in male C57BL/6 mice by portal triad pedicle occlusion for 45 min followed by reperfusion for 6 h. CF extract (300 mg/kg body weight [BW], oral) was administered 5 days before the IRI. Result: Treatment with CF extract significantly decreased serum alanine aminotransferase (sALT), serum aspartate aminotransferase (sAST) and serum lactate dehydrogenase (LDH) as well as liver histological changes. CF extract also prevented hepatic glutathione (GSH) depletion, increased malondialdehyde (MDA) and HO-1 levels induced by IRI. Western blotting indicated that the expression of the ERK, C-JUN, and iNOS were significantly decreased in the CF extract treatment group after IRI. RT-PCR indicated that CF extract significantly attenuated the Toll-like receptor 2 (TLR2) protein levels after IRI. The expression of tumor necrosis factor (TNF)-α, and interleukin (IL)-1β was significantly decreased in the CF extract treatment group. Conclusion: CF improved the acute hepatic IRI by reducing oxidative damage, and inflammation. These findings suggest that CF is a promising agent against acute IR-induced hepatic damage.

Exogenous supplementation of N-acetylcysteine Can Reduce Hepatotoxicity Induced by Ascites Fluid (Cell-Free) Adsorbed Over Protein-A-Containing Staphylococcus aureus Cowan-I Without Compromising Its Antitumor Effect.

Introduction:Hepatotoxicity along with enhanced mortality has remained a major concern during the development of antitumor therapy with the use of cell-free ascites fluid adsorbed (ad-AF) over Protein-A-containing Staphylococcus aureus Cowan I (SAC). Major issue with ad-AF inoculation is the significant depletion of hepatic glutathione (GSH). Exogenous supplementation of –SH contents to the host has offered an encouraging hope to explore the possibilities to use ad-AF as a therapeutic material due to its antitumor effects. GSH and l-cysteine have shown a promise with the recovery of –SH contents as well as the recovery of phase I and phase II biotransformation enzymes. Aforementioned observations prompted us to try other –SH donors.Materials and Methods:Therefore, in this study, N-acetylcysteine (NAC) was used as an exogenous source to provide –SH contents to reduce hepatotoxicity and mortality induced by ad-AF treatment.Results:Exogenous supplementation of NAC along with ad-AF treatment to ascites tumor bearers has shown a significant protection against hepatotoxicity and mortality caused by ad-AF. NAC substitution along with ad-AF has significantly enhanced the mean survival time (MST), without altering the antitumor effect of ad-AF as evident from tumor cell counts and viability.Discussion:NAC supplementation has been successful to recover hepatic –SH contents along with the significant recovery of phase I and phase II biotransformation enzymes. Marker enzymes for liver injury have also given clear-cut indications for the recovery of tumor bearers from hepatotoxicity induced by ad-AF.Conclusion:This study has shown that exogenous supplementation of NAC protects the host from the enhanced mortality and hepatotoxicity induced by ad-AF. These observations offer a hope to develop ad-AF as one of the probable treatment strategies for ascites tumors at least at experimental levels.

Hepatoprotective Effect of Baicalein Against Acetaminophen-Induced Acute Liver Injury in Mice.

Baicalein (BAI), one of the main components of Scutellaria baicalensis Georgi, possesses numerous pharmacological properties, including anti-cancer, anti-oxidative, anti-virus and anti-bacterial activities. The purpose of this study was to evaluate the hepatoprotective effect of baicalein against acetaminophen (APAP)-exposed liver injury in mice, and elucidate the underlying hepatoprotective mechanism. Baicalein pretreatment significantly alleviated the elevation of IL-6, IL-1β and TNF-α in serum and hepatic in a dose-dependent manner. It also dose-dependently reduced the hepatic malondialdehyde (MDA) concentration, as well as the depletion of hepatic superoxide dismutase (SOD), hepatic glutathione (GSH) and hepatic catalase (CAT). Moreover, pretreatment with baicalein significantly ameliorated APAP-exposed liver damage and histological hepatocyte changes. Baicalein also relieved APAP-induced autophagy by regulating AKT/mTOR pathway, LC3B and P62 expression. Furthermore, the hepatoprotective effect of baicalein to APAP-induced liver injury involved in Jak2/Stat3 and MAPK signaling pathway. Taken together, our findings suggested that baicalein exhibits the ability to prevent liver from APAP-induced liver injury and provided an underlying molecular basis for potential applications of baicalein to cure liver injuries.

Dictamnine-induced hepatotoxicity in mice: the role of metabolic activation of furan

Cortex Dictamni is extensively used as an herbal medicine worldwide, but is believed to induce hepatotoxicity and even causes mortality in many Asian and European countries. As the most abundant furoquinoline alkaloid ingredient of Cortex Dictamni, dictamnine (DIC) can be metabolically activated by CYP3A to an epoxide metabolite, which possesses the potential to induce hepatotoxicity by covalent binding with proteins. As yet, the hepatotoxicity of DIC and the role played by metabolic activation remain unknown. Here, we found that DIC caused acute liver injury in a time- and dose-dependent manner in mice. The hepatic and urinary DIC epoxide intermediates were observed in DIC-treated mice. Ketoconazole, a CYP3A inhibitor, significantly reduced the hepatotoxicity of DIC and inhibited the formation of reactive metabolites of DIC. Moreover, treatment with 2,3-dihydro-DIC, a DIC analog synthesized by selective reduction of the furan moiety, produced no hepatotoxicity in mice, and no reactive metabolite was formed, suggesting a structural necessity of furan moiety in DIC hepatotoxicity. A time course of gradual hepatic glutathione consumption was observed in DIC-treated mice, while depletion of hepatic glutathione by L-buthionine-S,R-sulfoximine enhanced the hepatotoxicity of DIC. Collectively, this study demonstrates that DIC induces acute hepatocellular injury in mice, and that metabolic activation of furan plays a crucial role in DIC-induced hepatotoxicity.