Bile acids (BAs) are cholesterol-derived metabolites synthesized by hepatocytes and secreted in bile to follow an inter-organ transit to the intestine, where they facilitate fat digestion and the absorption of lipids and liposoluble vitamins. Traditionally considered mere end products of cholesterol catabolism, they are now acknowledged to play intricate roles in regulating intermediary metabolism by controlling the expression of crucial genes. Additionally, they exert a significant impact on inflammation, cytotoxicity, and carcinogenesis. Moreover, BAs have a critical impact on the crosstalk between gut microbiota and host physiology, which affects the progression of liver and gastrointestinal cancers. Clinical data and results from studies of animal models support the involvement of BAs in the development of hepatocellular carcinoma, cholangiocarcinoma, colorectal adenocarcinoma, and pancreatic cancer. On the other hand, BAs and their derivatives have been proposed as pharmacological tools in strategies to restore abnormal hepatobiliary function and target cytostatic agents to cancers of the enterohepatic circuit. In the present review, we summarize basic concepts of BA physiology and regulation, as well as new advances in this expanding field of renewed interest for cancer biology, lending further support for the key role of BAs in liver and gastrointestinal cancer.

Mitochondria are present in all mammalian cells except matured red blood cells. Mitochondria consist of several metabolic pathways for glucose, fatty acids, amino acids, and bioenergetic pathways for ATP synthesis, membrane potential, and reactive oxygen production. In the liver, hepatic mitochondria play a key role in hepatic steatosis because mitochondrial metabolism produces acetyl-CoA which is the building block for synthesis of lipids and cholesterol. Mitochondria inner membrane is impermeable of metabolites, reducing equivalents, and small molecules such as phosphate, and sulfate. Thus, mitochondrial shuttles and carriers function as the routes of influx and efflux of these metabolites and molecules across the inner membrane. The signal regulation of these shuttles and mitochondrial enzymes could play a key role in coordinating the mitochondrial metabolism to adapt the cytosolic part of metabolic pathways in liver metabolic stress. Intriguingly, the interaction of mitochondria protein SH3 domain-binding protein 5 (SAB/SH3BP5) and c-Jun N-terminal kinase (JNK) was found as a pivotal role in sustained activation of JNK and phosphorylated-JNK (P-JNK) mediated activation of lipogenic pathway in nutritional excess. Knockout or knockdown of SAB prevented or reversed the hepatic steatosis, inflammation, and fibrosis, and improved metabolic intolerance and energy expenditure. Moreover, blocking the SAB peptide prevents palmitic acid-induced P-JNK interaction with SAB and inhibition of mitochondrial bioenergetics, implying the P-JNK effect on mitochondrial metabolism. This review focuses on the flow of mitochondrial metabolites in metabolic stress conditions and the contribution of mitochondria and mitochondrial stress signals in hepatic steatosis.

Background: Hepatic lipoprotein receptor-related protein 1 (LRP-1) plays a central role in peripheral amyloid beta (A ss) clearance, but its importance in Alzheimer's disease (AD) pathology is understudied. Our previous work showed that intragastric alcohol feeding to C57BL/6 J mice reduced hepatic LRP-1 expression which correlated with significant AD-relevant brain changes. Herein, we examined the role of hepatic LRP-1 in AD pathogenesis in APP/PS1 AD mice using two approaches to modulate hepatic LRP-1, intragastric alcohol feeding to model chronic heavy drinking shown by us to reduce hepatic LRP-1, and hepato-specific LRP-1 silencing. Methods: Eight-month-old male APP/PS1 mice were fed ethanol or control diet intragastrically for 5 weeks (n = 7-11/group). Brain and liver A ss were assessed using immunoassays. Three important mechanisms of brain amyloidosis were investigated: hepatic LRP-1 (major peripheral A ss regulator), blood-brain barrier (BBB) function (vascular A ss regulator), and microglia (major brain A ss regulator) using immunoassays. Spatial LRP-1 gene expression in the periportal versus pericentral hepatic regions was confirmed using NanoString GeoMx Digital Spatial Profiler. Further, hepatic LRP-1 was silenced by injecting LRP-1 microRNA delivered by the adenoassociated virus 8 (AAV8) and the hepato-specific thyroxine-binding globulin (TBG) promoter to 4-month-old male APP/PS1 mice (n = 6). Control male APP/PS1 mice received control AAV8 (n = 6). Spatial memory and locomotion were assessed 12 weeks after LRP-1 silencing using Y-maze and open-field test, respectively, and brain and liver A ss were measured. Results: Alcohol feeding reduced plaque-associated microglia in APP/PS1 mice brains and increased aggregated A ss (p < 0.05) by ELISA and 6E10-positive A ss load by immunostaining (p < 0.05). Increased brain A ss corresponded with a significant downregulation of hepatic LRP-1 (p < 0.01) at the protein and transcript level, primarily in pericentral hepatocytes (zone 3) where alcohol-induced injury occurs. Hepato-specific LRP-1 silencing significantly increased brain A ss and locomotion hyperactivity (p < 0.05) in APP/PS1 mice.

Acetaminophen overuse is a common cause of acute liver failure (ALF). During ALF, toxins are metabolized by enzymes such as CYP2E1 and transformed into reactive species, leading to oxidative damage and liver failure. Here, we found that oral magnesium (Mg) alleviated acetaminophen-induced ALF through metabolic changes in gut microbiota that inhibit CYP2E1. The gut microbiota from Mg-supplemented humans prevented acetaminophen-induced ALF in mice. Mg exposure modulated Bifidobacterium metabolism and enriched indole-3-carboxylic acid (I3C) levels. Formate C-acetyltransferase (pflB) was identified as a key Bifidobacterium enzyme involved in I3C generation. Accordingly, a Bifidobacterium pflB knockout showed diminished I3C generation and reduced the beneficial effects of Mg. Conversely, treatment with I3C or an engineered bacteria overexpressing Bifidobacterium pflB protected against ALF. Mechanistically, I3C bound and inactivated CYP2E1, thus suppressing formation of harmful reactive intermediates and diminishing hepatocyte oxidative damage. These findings highlight how interactions between Mg and gut microbiota may help combat ALF

Cell death occurs in various circumstances, such as homeostasis, stress response, and defense, via specific pathways and mechanisms that are regulated by specific activator-induced signal transductions. Among them, Jun N-terminal kinases (JNKs) participate in various aspects, and the recent discovery of JNKs and mitochondrial protein SAB interaction in signal regulation of cell death completes our understanding of the mechanism of sustained activation of JNK (P-JNK), which leads to triggering of the machinery of cell death. This understanding will lead the investigators to discover the modulators facilitating or preventing cell death for therapeutic application in acute or chronic diseases and cancer. We discuss here the mechanism and modulators of the JNK-SAB-ROS activation loop, which is the core component of mitochondria-dependent cell death, specifically apoptosis and mitochondrial permeability transition (MPT)-driven necrosis, and which may also contribute to cell death mechanisms of ferroptosis and pyroptosis. The discussion here is based on the results and evidence discovered from liver disease models, but the JNK-SAB-ROS activation loop to sustain JNK activation is universally applicable to various disease models where mitochondria and reactive oxygen species contribute to the mechanism of disease.

Supplementary Table S1. Summary of eight pazopanib (monotherapy) clinical studies included in the discovery pharmacogenetic liver toxicity analysis Supplementary Table S2. Summary of 23 pazopanib clinical studies included in the confirmatory pharmacogenetic liver toxicity analysis Supplementary Table S3. Primary tumor types for patients in the pharmacogenetic analyses Supplementary Table S4. Association between HLA-B*57:01 carriage and ALT elevation in pazopanib-treated patients with cancer. Sensitivity analyses in the combined dataset excluding 192 patients with baseline ALT>ULN Supplementary Table S5. Assessment of patients who had concurrent ALT (>3×ULN) and total bilirubin (>2×ULN) elevations Supplementary Table S6. Association between rs1800625 (chr6:32152442) genotype and ALT elevation in pazopanib-treated patients with cancer Supplementary Table S7. Joint analysis of HLA-B*57:01 and rs1800625 (chr6:32152442) genotype association with time to ALT>3xULN in pazopanib-treated patients with cancer Supplementary Figure S1. Genome-wide association study (GWAS) of common genetic variants (minor allele frequency {greater than or equal to}5%) associated with time to first ALT>3×ULN.

AIMS We confirmed that GCLC protein rapidly decreased at the same time P-JNK increased after APAP treatment. Therefore, our aims were to determine if JNK was directly responsible for decreased causing impaired recovery of GSH and if this was an important factor in determining APAP hepatotoxicity. RESULTS Immunoprecipitation of JNK after APAP identified binding to GCLC. Expression of a site directed mutated canonical JNK docking site in GCLC was resistant to degradation and lead to rapid restoration of GSH and inhibited sustained JNK activation. The JNK-resistant GCLC markedly protected against necrosis and ALT elevation. The proteolytic loss of GCLC was abrogated by inhibition of the proteosome, ubiquitination, or calpain. INNOVATION We addressed the aims by preparing mutated Gclc resistant to JNK induced degradation. The results allowed us to identify impaired GSH recovery as an important contributor to early progression of APAP toxicity after the metabolism of APAP and initial GSH depletion had occurred. CONCLUSION Activated JNK interacts directly with GCLC and leads to proteolytic degradation of GCLC. Degradation of GCLC impairs GSH recovery after APAP allowing the continued activation of JNK. Conversely, rapid recovery of GSH inhibits the sustained activation of the MAP Kinase cascade and dampens APAP toxicity by suppressing the continued activation of JNK.

Supplementary Table S1. Summary of eight pazopanib (monotherapy) clinical studies included in the discovery pharmacogenetic liver toxicity analysis Supplementary Table S2. Summary of 23 pazopanib clinical studies included in the confirmatory pharmacogenetic liver toxicity analysis Supplementary Table S3. Primary tumor types for patients in the pharmacogenetic analyses Supplementary Table S4. Association between HLA-B*57:01 carriage and ALT elevation in pazopanib-treated patients with cancer. Sensitivity analyses in the combined dataset excluding 192 patients with baseline ALT>ULN Supplementary Table S5. Assessment of patients who had concurrent ALT (>3×ULN) and total bilirubin (>2×ULN) elevations Supplementary Table S6. Association between rs1800625 (chr6:32152442) genotype and ALT elevation in pazopanib-treated patients with cancer Supplementary Table S7. Joint analysis of HLA-B*57:01 and rs1800625 (chr6:32152442) genotype association with time to ALT>3xULN in pazopanib-treated patients with cancer Supplementary Figure S1. Genome-wide association study (GWAS) of common genetic variants (minor allele frequency {greater than or equal to}5%) associated with time to first ALT>3×ULN.

BACKGROUND & AIMS:Idiosyncratic drug-induced liver injury (DILI) with autoimmune features is a liver condition with laboratory and histological characteristics similar to those of idiopathic autoimmune hepatitis (AIH), which despite being increasingly reported, remains largely undefined. We aimed to describe in-depth the features of this entity in a large series of patients from two prospective DILI registries.METHODS:DILI cases with autoimmune features collected in the Spanish DILI Registry and the Latin American DILI Network were compared with DILI patients without autoimmune features and with an independent cohort of patients with AIH.RESULTS:Out of 1,426 patients with DILI, 33 cases with autoimmune features were identified. Female sex was more frequent in AIH patients than in the other groups (p = .001). DILI cases with autoimmune features had significantly longer time to onset (p < .001) and resolution time (p = .004) than those without autoimmune features. Interestingly, DILI patients with autoimmune features who relapsed exhibited significantly higher total bilirubin and transaminases at onset and absence of peripheral eosinophilia than those who did not relapse. The likelihood of relapse increased over time, from 17% at 6 months to 50% 4 years after biochemical normalization. Statins, nitrofurantoin and minocycline were the drugs most frequently associated with this phenotype.CONCLUSIONS:DILI with autoimmune features shows different clinical features than DILI patients lacking characteristics of autoimmunity. Higher transaminases and total bilirubin values with no eosinophilia at presentation increase the likelihood of relapse in DILI with autoimmune features. As the tendency to relapse increases over time, these patients will require long-term follow-up.

Heavy alcohol consumption is a known risk factor for various forms of dementia and the development of Alzheimer’s disease (AD). In this work, we investigated how intragastric alcohol feeding may alter the liver-to-brain axis to induce and/or promote AD pathology. Four weeks of intragastric alcohol feeding to mice, which causes significant fatty liver (steatosis) and liver injury, caused no changes in AD pathology markers in the brain [amyloid precursor protein (APP), presenilin], except for a decrease in microglial cell number in the cortex of the brain. Interestingly, the decline in microglial numbers correlated with serum alanine transaminase (ALT) levels, suggesting a potential link between liver injury and microglial loss in the brain. Intragastric alcohol feeding significantly affected two hepatic proteins important in amyloid-beta (Aβ) processing by the liver: 1) alcohol feeding downregulated lipoprotein receptor-related protein 1 (LRP1, ∼46%), the major receptor in the liver that removes Aβ from blood and peripheral organs, and 2) alcohol significantly upregulated APP (∼2-fold), a potentially important source of Aβ in the periphery and brain. The decrease in hepatic LRP1 and increase in hepatic APP likely switches the liver from being a remover or low producer of Aβ to an important source of Aβ in the periphery, which can impact the brain. The downregulation of LRP1 and upregulation of APP in the liver was observed in the first week of intragastric alcohol feeding, and also occurred in other alcohol feeding models (NIAAA binge alcohol model and intragastric alcohol feeding to rats). Modulation of hepatic LRP1 and APP does not seem alcohol-specific, as ob/ob mice with significant steatosis also had declines in LRP1 and increases in APP expression in the liver. These findings suggest that liver steatosis rather than alcohol-induced liver injury is likely responsible for regulation of hepatic LRP1 and APP. Both obesity and alcohol intake have been linked to AD and our data suggests that liver steatosis associated with these two conditions modulates hepatic LRP1 and APP to disrupt Aβ processing by the liver to promote AD.

HepatologyVolume 75, Issue 3 p. 514-517 INTRODUCTION OF THE AASLD PRESIDENT Introducing Laurie DeLeve, M.D., Ph.D., Our 2022 AASLD President Neil Kaplowitz, Corresponding Author Neil Kaplowitz kaplowit@usc.edu Research Center for Liver Disease, Division of Gastrointestinal and Liver Diseases, Department of Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA Correspondence Neil Kaplowitz, Research Center for Liver Disease, Division of Gastrointestinal and Liver Diseases, Department of Medicine, Keck School of Medicine, University of Southern California, 2011 Zonal Avenue, HMR 615, Los Angeles, CA 90033, USA. Email: kaplowit@usc.eduSearch for more papers by this author Neil Kaplowitz, Corresponding Author Neil Kaplowitz kaplowit@usc.edu Research Center for Liver Disease, Division of Gastrointestinal and Liver Diseases, Department of Medicine, Keck School of Medicine, University of Southern California, Los Angeles, California, USA Correspondence Neil Kaplowitz, Research Center for Liver Disease, Division of Gastrointestinal and Liver Diseases, Department of Medicine, Keck School of Medicine, University of Southern California, 2011 Zonal Avenue, HMR 615, Los Angeles, CA 90033, USA. Email: kaplowit@usc.eduSearch for more papers by this author First published: 06 December 2021 https://doi.org/10.1002/hep.32264Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Volume75, Issue3March 2022Pages 514-517 RelatedInformation

HepatologyVolume 73, Issue 5 p. 1634-1636 Editorial Alcohol, Fasting, and Therapeutic Dosing of Acetaminophen: A Perfect Storm William M. Lee M.D., Corresponding Author William M. Lee M.D. william.lee@utsouthwestern.edu orcid.org/0000-0002-2783-5441 Digestive and Liver Disease Division, Department of Internal Medicine, University of Texas Southwestern Medical Center at Dallas, Dallas, TX ADDRESS CORRESPONDENCE AND REPRINT REQUESTS TO: William M. Lee, M.D., F.A.C.P., F.A.A.S.L.D. Division of Digestive and Liver Diseases, UT Southwestern Medical Center at Dallas 5959 Harry Hines Blvd. Ste. 420 Dallas, TX 75390-8887 E-mail: william.lee@utsouthwestern.edu Tel.: +1-214 645 6111Search for more papers by this authorNeil Kaplowitz M.D., Neil Kaplowitz M.D. Research Center for Liver Disease, Keck School of Medicine, University of Southern California, Los Angeles, CASearch for more papers by this author William M. Lee M.D., Corresponding Author William M. Lee M.D. william.lee@utsouthwestern.edu orcid.org/0000-0002-2783-5441 Digestive and Liver Disease Division, Department of Internal Medicine, University of Texas Southwestern Medical Center at Dallas, Dallas, TX ADDRESS CORRESPONDENCE AND REPRINT REQUESTS TO: William M. Lee, M.D., F.A.C.P., F.A.A.S.L.D. Division of Digestive and Liver Diseases, UT Southwestern Medical Center at Dallas 5959 Harry Hines Blvd. Ste. 420 Dallas, TX 75390-8887 E-mail: william.lee@utsouthwestern.edu Tel.: +1-214 645 6111Search for more papers by this authorNeil Kaplowitz M.D., Neil Kaplowitz M.D. Research Center for Liver Disease, Keck School of Medicine, University of Southern California, Los Angeles, CASearch for more papers by this author First published: 08 February 2021 https://doi.org/10.1002/hep.31747Citations: 1 Potential conflict of interest: Dr. Lee consults for Affibody, Genentech, Forma, Karuna, SeaGen, Cortexyme, Pfizer, and Alynlam. He received grants from Merck, Bristol-Myers Squibb, Intercept, Novo Nordisk, Eiger, and Alexion. SEE ARTICLE ON PAGE XXXX Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat No abstract is available for this article.Citing Literature Volume73, Issue5May 2021Pages 1634-1636 RelatedInformation

Hepatic ischemia/reperfusion injury (HIRI) is a serious complication that occurs following shock and/or liver surgery. Gut microbiota and their metabolites are key upstream modulators of development of liver injury. Herein, we investigated the potential contribution of gut microbes to HIRI. Ischemia/reperfusion surgery was performed to establish a murine model of HIRI. 16S rRNA gene sequencing and metabolomics were used for microbial analysis. Transcriptomics and proteomics analysis were employed to study the host cell responses. Our results establish HIRI was significantly increased when surgery occurred in the evening (ZT12, 20:00) when compared with the morning (ZT0, 08:00); however, antibiotic pretreatment reduced this diurnal variation. The abundance of a microbial metabolite 3,4-dihydroxyphenylpropionic acid was significantly higher in ZT0 when compared with ZT12 in the gut and this compound significantly protected mice against HIRI. Furthermore, 3,4-dihydroxyphenylpropionic acid suppressed the macrophage pro-inflammatory response in vivo and in vitro. This metabolite inhibits histone deacetylase activity by reducing its phosphorylation. Histone deacetylase inhibition suppressed macrophage pro-inflammatory activation and diminished the diurnal variation of HIRI. Our findings revealed a novel protective microbial metabolite against HIRI in mice. The potential underlying mechanism was at least in part, via 3,4-dihydroxyphenylpropionic acid-dependent immune regulation and histone deacetylase (HDAC) inhibition in macrophages.

Background and Aims The hepatic mitogen‐activated protein kinase (MAPK) cascade leading to c‐Jun N‐terminal kinase (JNK) activation has been implicated in the pathogenesis of nonalcoholic fatty liver (NAFL)/NASH. In acute hepatotoxicity, we previously identified a pivotal role for mitochondrial SH3BP5 (SAB; SH3 homology associated BTK binding protein) as a target of JNK, which sustains its activation through promotion of reactive oxygen species production. Therefore, we assessed the role of hepatic SAB in experimental NASH and metabolic syndrome. Approach and Results In mice fed high‐fat, high‐calorie, high‐fructose (HFHC) diet, SAB expression progressively increased through a sustained JNK/activating transcription factor 2 (ATF2) activation loop. Inducible deletion of hepatic SAB markedly decreased sustained JNK activation and improved systemic energy expenditure at 8 weeks followed by decreased body fat at 16 weeks of HFHC diet. After 30 weeks, mice treated with control–antisense oligonucleotide (control‐ASO) developed steatohepatitis and fibrosis, which was prevented by Sab‐ASO treatment. Phosphorylated JNK (p‐JNK) and phosphorylated ATF2 (p‐ATF2) were markedly attenuated by Sab‐ASO treatment. After 52 weeks of HFHC feeding, control N‐acetylgalactosamine antisense oligonucleotide (GalNAc‐Ctl‐ASO) treated mice fed the HFHC diet exhibited progression of steatohepatitis and fibrosis, but GalNAc‐Sab‐ASO treatment from weeks 40 to 52 reversed these findings while decreasing hepatic SAB, p‐ATF2, and p‐JNK to chow‐fed levels. Conclusions Hepatic SAB expression increases in HFHC diet–fed mice. Deletion or knockdown of SAB inhibited sustained JNK activation and steatohepatitis, fibrosis, and systemic metabolic effects, suggesting that induction of hepatocyte Sab is an important driver of the interplay between the liver and the systemic metabolic consequences of overfeeding. In established NASH, hepatocyte‐targeted GalNAc‐Sab‐ASO treatment reversed steatohepatitis and fibrosis.

Background & Aims: Prospective drug-induced liver injury (DILI) registries are important sources of information on idiosyncratic DILI. We aimed to present a comprehensive analysis of 843 patients with DILI enrolled into the Spanish DILI Registry over a 20-year time period. Methods: Cases were identified, diagnosed and followed prospectively. Clinical features, drug information and outcome data were collected. Results: A total of 843 patients, with a mean age of 54 years (48% females), were enrolled up to 2018. Hepatocellular injury was associated with younger age (adjusted odds ratio [aOR] per year 0.983; 95% CI 0.974-0.991) and lower platelet count (aOR per unit 0.996; 95% CI 0.994-0.998). Anti-infectives were the most common causative drug class (40%). Liver-related mortality was more frequent in patients with hepatocellular damage aged >- 65 years (p = 0.0083) and in patients with underlying liver disease (p = 0.0221). Independent predictors of liver-related death/transplantation included nR-based hepatocellular injury, female sex, higher onset aspartate aminotransferase (AST) and bilirubin values. nR-based hepatocellular injury was not associated with 6-month overall mortality, for which comorbidity burden played a more important role. The prognostic capacity of Hy's law varied between causative agents. Empirical therapy (corticosteroids, ursodeoxycholic acid and MARS) was prescribed to 20% of patients. Drug-induced autoimmune hepatitis patients (26 cases) were mainly females (62%) with hepatocellular damage (92%), who more frequently received immunosuppressive therapy (58%). Conclusions: AST elevation at onset is a strong predictor of poor outcome and should be routinely assessed in DILI evaluation. Mortality is higher in older patients with hepatocellular damage and patients with underlying hepatic conditions. The Spanish DILI Registry is a valuable tool in the identification of causative drugs, clinical signatures and prognostic risk factors in DILI and can aid physicians in DILI characterisation and management. Lay summary: Clinical information on drug-induced liver injury (DILI) collected from enrolled patients in the Spanish DILI Registry can guide physicians in the decision-making process. We have found that older patients with hepatocellular type liver injury and patients with additional liver conditions are at a higher risk of mortality. The type of liver injury, patient sex and analytical values of aspartate aminotransferase and total bilirubin can also help predict clinical outcomes. (C) 2021 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Hazard identification regarding adverse effects on the liver is a critical step in safety evaluations of drugs and other chemicals. Current testing paradigms for hepatotoxicity rely heavily on preclinical studies in animals and human data (epidemiology and clinical trials). Mechanistic understanding of the molecular and cellular pathways that may cause or exacerbate hepatotoxicity is well advanced and holds promise for identification of hepatotoxicants. One of the challenges in translating mechanistic evidence into robust decisions about potential hepatotoxicity is the lack of a systematic approach to integrate these data to help identify liver toxicity hazards. Recently, marked improvements were achieved in the practice of hazard identification of carcinogens, female and male reproductive toxicants, and endocrine disrupting chemicals using the key characteristics approach. Here, we describe the methods by which key characteristics of human hepatotoxicants were identified and provide examples for how they could be used to systematically identify, organize, and use mechanistic data when identifying hepatotoxicants.

SAB is an outer membrane docking protein for JNK mediated impaired mitochondrial function. Deletion of Sab in hepatocytes inhibits sustained JNK activation and cell death. Current work demonstrated that increasing SAB enhanced the severity of APAP liver injury. Female mice were resistant to liver injury and exhibited markedly decreased hepatic SAB protein expression versus males. The mechanism of SAB repression involved a pathway from ER a to p53 expression which induced miR34a-5p . miR34a-5p targeted the Sab mRNA coding region, repressing SAB expression. Fulvestrant or p53 knockdown decreased miR34a-5p and increased SAB in females leading to increased injury from APAP and TNF/galactosamine. In contrast, ER a agonist increased p53 and miR34a-5p which decreased SAB expression and hepatotoxicity in males. Hepatocyte-specific deletion of miR34a also increased severity of liver injury in females, which was prevented by GalNAc-ASO knockdown of Sab . Similar to mice, premenopausal human females also expressed high hepatic p53 and low SAB levels while age-matched males expressed low p53 and high SAB levels, but there was no sex difference of SAB expression in postmenopause. In conclusion, the level of SAB expression determined the severity of JNK dependent liver injury. Females expressed low hepatic SAB protein levels due to an ER a -p53- miR34a pathway which repressed SAB expression, accounting for resistance to liver injury. ABSTRACT SAB is an outer membrane docking protein for JNK mediated impaired mitochondrial function. Deletion of Sab in hepatocytes inhibits sustained JNK activation and cell death. Current work demonstrated that increasing SAB enhanced the severity of APAP liver injury. Female mice were resistant to liver injury and exhibited markedly decreased hepatic SAB protein expression versus males. The mechanism of SAB repression involved a pathway from ER α to p53 expression which induced miR34a-5p . miR34a-5p targeted the Sab mRNA coding region, repressing SAB expression. Fulvestrant or p53 knockdown decreased miR34a-5p and increased SAB in females leading to increased injury from APAP and TNF/galactosamine. In contrast, ER α agonist increased p53 and miR34a-5p which decreased SAB expression and hepatotoxicity in males. Hepatocyte-specific deletion of miR34a also increased severity of liver injury in females, which was prevented by GalNAc-ASO knockdown of Sab . mice, human females also expressed high hepatic p53 and low SAB levels while age-matched males expressed low p53 and high SAB levels, but there was no sex difference of SAB expression in postmenopause. In conclusion, the level of SAB expression determined the severity of JNK dependent liver injury. Females expressed low hepatic SAB protein levels due to an ER α -p53-miR34a pathway which repressed SAB expression, accounting for resistance to liver injury.