Forsythoside A (FTA) is a key component found in the fruit and leaves of Forsythia suspensa, having anti-inflammatory and antioxidant properties. However, it is unclear whether FTA can have a protective effect against acute alcoholic liver injury (ALI) and how it may exert this effect. This research examined the potential protective effects of FTA against acute ALI using cell and animal models. The protective properties of FTA against acute ALI were attributed to its anti-inflammatory and antioxidant actions by detecting the markers of oxidative stress and inflammation. The underlying mechanism was explored through the utilization of Western blotting, Molecular Docking, and Microscale Thermophoresis techniques. The results showed that pretreatment with high doses of FTA had a significant protective effect on acute ALI in both cell and animal models. The pretreatment with high doses of FTA inhibited alcohol-induced oxidative stress and inflammation, raising antioxidative enzyme activity in both models. Furthermore, FTA has been shown to bind to TLR4, thereby inhibiting alcohol-induced activation of the NF-κB signaling pathway, leading to a decrease in cellular oxidative stress and inflammatory reactions. This interaction also facilitates the ubiquitination-mediated degradation of TLR4, ultimately diminishing its regulatory impact on the NF-κB signaling cascade. FTA has a significant protective effect on acute ALI. It binds to TLR4 to inhibit the activation of the NF-κB signaling pathway by alcohol, thereby reducing oxidative stress and inflammation and exerting a protective effect. The results of our study provide a theoretical basis for the development of FTA for the prevention and treatment of acute ALI.

In the early 80s, the so-called "French paradox" was proposed, that is, a correlation between wine consumption, a diet rich in fats, and low mortality from coronary disease. Conversely, it is well established that excessive alcohol consumption increases the risk of cirrhosis and cancer, but few studies have investigated the effects of moderate alcohol consumption. However, all these conclusions were derived from epidemiological population studies that may be subject to distortions due to multiple factors. Here, the effect of moderate consumption of red wine on health throughout life was examined in a murine model. Different variables were evaluated in groups of female animals that were fed a standard or a fat diet throughout their adult life and given water, or wine or alcohol diluted in water in proportions similar to what is considered moderate consumption in humans. Our results showed few differences in most of the analyzed variables (body weight, liver profile and survival rate) between the different female mouse groups. The most remarkable findings were observed in the fat-diet groups that showed more frequent and severe liver lesions and a lower average ovarian weight. Moreover, moderate and prolonged ethanol consumption significantly affected telomere length only when the diet was high in fat, whereas wine consumption showed no difference compared to water, pointing to a possible predominant role of the compounds, particularly polyphenols present in wine. On the other hand, wine-drinking mice fed a fat diet had more oocytes than those in the ethanol-drinking group. Overall, our data suggest that long-term moderate red wine consumption does not substantially influence the health of female mice.

Chronic and excessive alcohol consumption induces alcohol-related liver injury (ALI), characterized by oxidative stress (OS), disrupted lipid metabolism, and gut microbiota dysbiosis. Given the lack of effective pharmacological treatments, flavonoid-rich fruits have attracted growing attention as potential intervention strategies. This study investigated the independent and combined effects of extracts from Citri reticulatae pericarpium (CRPE) and Chaenomeles speciosa (Sweet) Nakai (CSPE), previously shown to possess hepatoprotective properties, in a mouse model of ethanol-induced chronic ALI. The flavonoid composition of CRPE and CSPE was characterized using LC-MS/MS, and their potential mechanisms of action were further elucidated through transcriptomic analysis. The results showed that CRPE and CSPE, whether administered individually or in combination, effectively alleviated alcohol-induced hepatic histological damage and inflammatory responses. Furthermore, both extracts significantly reduced OS and improved lipid metabolism. Notably, CRPE, CSPE, and their combination regulated the gut microbiota, as shown by increased abundances of beneficial bacteria such as Lactobacillus and Bifidobacterium, along with elevated levels of short-chain fatty acids (SCFAs). These findings highlight that combinations of multiple fruit extracts exhibit significant potential in alleviating ALI by modulating the gut microbiota, providing valuable insights for the development of functional foods.

Binge drinking constitutes a significant public health concern. Defined as the consumption of five or more alcoholic beverages on a single occasion, binge drinking leads to acute cognitive and motor impairments and is associated with a multitude of detrimental health consequences. Therefore, the aim of this study was to analyse globally published peer-reviewed literature on binge drinking.

A thorough search of the Scopus database was conducted to gather all the relevant research. Keywords related to binge drinking were used to locate a wide range of studies. Specific criteria were subsequently applied to narrow the results, ensuring the inclusion of only the most relevant articles. This process yielded a collection of 2,763 research papers. Finally, a software program called VOSviewer was utilized to analyse and visualize the connections between these studies.

A bibliometric analysis was performed to investigate trends in binge drinking research literature published between 1980 and 2024. The findings revealed a significant increase in publications (R²=0.916; p < 0.001), with a peak in 2018 (191 articles). The majority (89.65%, n = 2,477) were research articles, followed by review articles (4.74%, n = 131). Authors from 139 countries contributed to binge-drinking research, with the USA (n = 1,550; 56.1%) and the UK (n = 216; 7.82%) leading in the volume of publications. The National Institute on Alcohol Abuse and Alcoholism (n = 65; 2.35%) and the University of North Carolina at Chapel Hill (n = 63; 2.28%) emerged as the main institutional contributors. The National Institute on Alcohol Abuse and Alcoholism in the United States was the main funding source, supporting 599 articles (21.68%), followed by the National Institutes of Health in the United States, with 544 articles (19.69%). In particular, the post-2016 period witnessed a shift in research themes toward mechanistic investigations alongside studies on societal interventions, reflecting a growing focus on mitigating the broader social impact of binge drinking.

This study is the first comprehensive analysis of trends in binge drinking research. Over the past decade, binge drinking has increased dramatically, led by the United States, the UK, and Spain. Initially, focused on social and cultural factors, research shifted after 2016 to mechanistic and animal models, shaping future research directions and strategies.

Numerous studies have examined the pathophysiological changes induced by chronic alcohol (ethanol) consumption and the underlying mechanisms, while much less attention has been devoted to understanding the health impacts of binge drinking. Binge drinking is defined as the excessive consumption of alcohol within a single drinking episode, and is the typical consumption pattern among young people in Western countries. While most young binge drinkers are not clinically alcohol dependent, binge drinking has emerged as a significant social and public health concern. The circulating alcohol consumed during binge episodes permeates cellular membranes throughout the body, exerting profound effects on multiple organs, and signaling pathways. Regular binge drinking eventually induces hepatic steatosis (fatty liver), initiates acute inflammation, and accelerates neutrophil infiltration, de novo lipogenesis, adipocyte death/lipolysis, and the production of nonoxidative alcohol metabolites, processes that synergize to damage liver tissue and impair liver function. Metabolic abnormalities such as diabetes and obesity can also exacerbate the progression of alcohol-related liver disease among binge drinkers. Several animal models have been developed to evaluate the pathophysiological changes resulting from binge drinking; however, the pathogenesis of binge drinking is not fully understood due to differences in alcohol metabolism between animal models and humans. Thus, given the high prevalence and severe health implications of binge drinking, there is an urgent need for comprehensive experimental and clinical investigations to unravel the associated pathophysiological changes. This review summarizes recent research findings on the impact of binge drinking, specifically focusing on its contributions to alcoholic liver injury.

Alcohol-associated liver disease (ALD), primarily caused by chronic excessive alcohol consumption, is a leading cause of chronic liver disease worldwide. ALD includes alcohol-associated steatotic liver, alcohol-associated hepatitis (AH), fibrosis, cirrhosis, and can even progress to hepatocellular carcinoma (HCC). Existing research indicates that the risk factors of ALD are quite numerous. In addition to drinking patterns, factors such as aldehyde dehydrogenase 2 (ALDH2) deficiency, smoking, medication administration, high-fat diet (HFD), hepatitis virus infection, and disruption of circadian rhythms can also increase susceptibility to ALD. However, there is limited understanding regarding the exacerbation of liver injury by alcohol plus additional risk factors. This review presents rodent models of EtOH + "X," which simulate the synergistic effects of alcohol and additional risk factors in causing liver injury. These models offer a further exploration of the interactions between alcohol and additional risk factors, advancing the simulation of human ALD and providing a more reliable platform for studying disease mechanisms and exploring therapeutic interventions. We summarize the modeling methods, relevant indicators of liver injury, and focus on the targets of the synergistic effects as well as the associated mechanisms.

Gut microbiota impacts host homeostasis and diseases. Chronic plus binge ethanol consumption has been linked to increased injuries than chronic or binge ethanol intake alone. We hypothesized that distinct shapes in gut microbiota composition are induced by chronic, binge, and the association of these treatments, thereby affecting host functions and contributing to sex-based differences in alcohol use disorders. Male and female C57BL/6J mice were submitted to chronic, binge, or chronic plus binge ethanol feeding. DNA was extracted from fecal microbiota, followed by analysis of the V3-V4 region of the 16S rRNA gene and sequencing on an Illumina platform. Gut microbiome analysis was performed using QIIME v2022.2.0. Functional profiling of the gut microbiome was performed using PICRUSt2. Ethanol differentially affected the gut microbiota of female and male mice. Decreased α diversity was observed in male and female mice from the chronic plus binge and chronic groups, respectively. The genera Faecalibaculum, Lachnospiraceae, and Alistipes were identified as major potential biomarkers for gut dysbiosis induced by ethanol consumption. In addition, ethanol-induced gut dysbiosis altered several metabolic pathways. Ethanol consumption modifies the mouse gut microbiome in a drinking pattern- and sex-dependent manner, potentially leading to different susceptibility to ethanol-related diseases. Chronic plus binge ethanol intake induces a more pronounced gut dysbiosis in male mice. Conversely, chronic ethanol is linked to a greater degree of gut dysbiosis in female mice. The changed gut microbiome may be potentially targeted to prevent, mitigate, or treat alcohol use disorders.NEW & NOTEWORTHY Ethanol alters the mouse gut microbiome in a drinking pattern- and sex-dependent manner. Chronic plus binge ethanol intake induces a more severe gut dysbiosis in male mice, whereas chronic ethanol consumption appears to be a more potent inductor of gut dysbiosis in female mice. Ethanol-induced gut dysbiosis alters several pathways linked to metabolism, genetic and environmental information processing, cellular processes, organism systems, and neurological human diseases.

Alcohol-associated liver disease (ALD) is the main cause of alcohol-associated mortality. However, the mechanism of ALD development is poorly understood. Epigenetic changes are thought to play an important role in ALD. We aimed to define the epigenetic changes induced by alcohol and predict drivers of these changes.

Mice were fed high-fat diet with or without 20% of alcohol in the drinking water for 20 weeks (WDA model). scATAC-seq data set was analyzed using Signac R package. To test the role of C/EBPβ, Cebpb-floxed mice were treated with AAV8-TBG-Cre or AAV8-control.

We analyzed differentially accessible regions in livers from control and alcohol-fed mice and found that activity of C/EBPβ transcription factor was associated with alcohol-induced epigenetic changes in hepatocytes. C/EBPβ protein levels were significantly upregulated in multiple models of ALD and human ALD samples. Using hepatocyte-specific Cebpb knockout mice we found that Cebpb loss protected male mice from alcohol-induced fibrosis development. We found no protection in female mice, suggesting that this mechanism is specific to male ALD. In vitro studies suggested that the protective effect of Cebpb loss was mediated by altered hepatocyte-macrophage cross talk. Cebpb knockout in hepatocytes reduced a profibrotic and promoted a pro-resolving phenotype in macrophages, thus modulating ALD development. We further identified the mediators of the cross talk. Cebpb knockout altered the expression of several HDL protein components, increasing APOA1 and apolipoprotein M and reducing apolipoprotein E and SAA levels in male mice. HDL secreted by Cebpb knockout hepatocytes was sufficient to confer anti-inflammatory and antifibrotic changes to macrophages.

Taken together, alcohol-induced C/EBPβ activation is a key driver of ALD fibrosis in males via C/EBPβ-dependent HDL remodeling.

Alcoholic liver disease (ALD) is a commonly known liver disease mediated by prolonged alcohol consumption. Aescin is a triterpene saponin that can manage several conditions, including brain trauma, arthritis, venous congestion, stroke, and thrombophlebitis. Even so, studies illustrating the aescin role in ALD are scarce. Our study explored the potential effect of aescin in ALD in mice. In the current experiment, forty mice were utilized and sorted randomly into four groups: the control group received only vehicles, the alcohol group was given 5% alcohol in drinking water for four weeks, and the aescin-treated groups were given 5% alcohol in drinking water and aescin (10 and 20 mg/kg/day) for four weeks, then two doses of 60% alcohol (3g/kg) were given in the 29th and 30th day of the experiment. Our study revealed that aescin ameliorated alcohol-mediated liver damage, including reducing inflammatory cell infiltration and vascular dilatation. The serum concentrations of liver enzymes (ALT and AST) decreased in the aescin-treated groups. The apoptosis and oxidative stress also decreased, and the antioxidant enzyme activities were restored by aescin in alcohol-treated mice. Additionally, aescin decreased ethanol-induced inflammation by downregulating p38 MAPK and tumor necrosis factor-α (TNF-α), suggesting that aescin positively reduces alcohol-caused inflammation and oxidative stress. Consequently, aescin could ameliorate alcohol-induced hepatic damage by targeting the p38 MAPK/TNF-α signalling and could be developed as a novel health product that potentially ameliorates ALD.

The etiology of developmental defects of enamel (DDE) remains incompletely understood. Prenatal alcohol exposure has been proposed as a potential risk factor for DDE. Animal studies suggest that in utero ethanol exposure can disrupt ameloblast function, leading to enamel abnormalities. This study aims to: (1) Assess the impact of prenatal alcohol consumption on the clinical and structural properties of dental enamel in offspring; and (2) Investigate the underlying mechanisms of these alterations through histological and molecular analyses. Pregnant Wistar rats will be assigned to two groups: one exposed to ethanol and a control group with no alcohol exposure. Ethanol exposure will follow a binge drinking model, with rats receiving 3 g/kg of ethanol (30% w/v) for 3 consecutive days, followed by 4 days of rest each week. This regimen will begin one week prior to conception and continue throughout pregnancy. The incisors and molars of offspring will be evaluated on the 10th (n = 22 per group) and 28th (n = 22 per group) days of life. Visible enamel changes will be documented through photographs. Enamel volume, thickness, and density will be assessed using micro-CT imaging. Mechanical properties will be evaluated using the Knoop microhardness test, while chemical composition will be analyzed through Scanning Electron Microscopy with Energy Dispersive X-ray (SEM-EDX) and Raman spectroscopy, respectively. The area of the organic enamel matrix will be quantified in histological sections. Genes Amelx, Enam, Ambn, Mmp2, Mmp9, Mmp20, Klk4, Cldn3, Cldn16, and Cldn19 will be evaluated in ameloblasts using real-time RT-PCR and protein synthesis will be confirmed by immunohistochemistry. Gelatinolytic activity in the ameloblast layer will be assessed by in situ zymography.

In recent years, there have been significant advances in pathological research on alcoholic liver disease (ALD), with suitable animal models making a significant contribution. However, the currently established animal ALD models still have some significant drawbacks, especially the inability to induce the entire human ALD lineage, which may be related to physiological differences between animals and humans. This review comprehensively summarized the most widely used experimental models of ALD, including voluntary drinking, Lieber-DeCarli, Meadows-Cook, Tsukamoto-French, NIAAA, and the "second hit" model. "Second hit" refers to an additional factor that damages the liver. There are various "second hit" models that fall into two main categories: particular diets and drugs. These models can either simulate human drinking patterns more accurately or produce varying degrees of ALD without significantly increasing animal mortality. We introduced the established method of the original models, discussed the advantages and disadvantages of the existing models from the aspects of operability and practicality, and provided existing improvement methods, hoping to provide a reference for future researchers.

Alcohol-associated liver disease (ALD) is a significant global health concern and a leading cause of liver disease-related deaths. However, the treatment options are limited due to the lack of animal models that accurately replicate ALD pathogenesis. An ideal ALD animal model should have pathological characteristics similar to those of human ALD, with a clear pathological process and ease of drug intervention. Over the years, researchers have focused on developing ideal ALD preclinical animal models by testing various methods, such as ad libitum drinking water with ethanol, acute, single large doses of ethanol gavage, multiple alcohol gavages in a short period, the Lieber-DeCarli liquid diet feeding model, the intragastric infusion model, and the Gao-binge model. With the increasing occurrence of obesity and metabolic dysfunction-associated steatotic liver disease, a new category of metabolic and alcohol-associated liver disease (MetALD) is also emerging. Studies have investigated the combined effects of a high-fat diet combined with binge alcohol or drinking water containing ethanol to mimic MetALD. In addition to mice, other species such as rats, guinea pigs, zebrafish, and non-human primates have also been tested to establish ALD preclinical models. This review aims to summarize current animal ALD models, particularly the emerging MetALD models, with the hope of providing a valuable reference for establishing more effective animal models in ALD studies in the future.

Alcohol-associated liver disease (ALD) is a common non-communicable chronic liver disease characterized by a spectrum of conditions ranging from steatosis and alcohol-associated steatohepatitis (AH) to fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). The pathogenesis of ALD involves a complex interplay of various molecular, biochemical, genetic, epigenetic, and environmental factors. While the mechanisms are well studied, therapeutic options remain limited. Alpinetin, a natural flavonoid with antioxidant and anti-inflammatory properties, has shown potential hepatoprotective effects, though its efficacy in ALD remains unexplored. This study investigated the hepatoprotective effects of alpinetin using a Lieber-DeCarli ethanol liquid diet model of ALD in C57BL/6 mice. Mice were divided into three groups: the control group, the ethanol group, and the ethanol group treated with alpinetin. Serum activity of ALT, AST, γ-GT, and ALP was measured to assess liver function, along with antioxidative and oxidative/nitrosative stress markers in liver tissue. Pro-inflammatory cytokines and endoplasmic reticulum (ER) stress parameters in liver tissue were also evaluated. Histological assessment of disease activity was performed using the SALVE grading and staging system. Treatment with alpinetin significantly reduced serum levels of ALT, AST, γ-GT, and oxidative/nitrosative stress markers while increasing antioxidative markers. The levels of pro-inflammatory cytokines and ER stress parameters were significantly decreased. Histological analysis demonstrated reduced steatosis, hepatocyte ballooning, and inflammation. These findings suggest that alpinetin holds promise as a potential therapeutic agent for managing ALD.

Acute liver injury is characterized by massive inflammatory cell infiltration, destruction of liver structure and abnormalities in liver function. Acetylcorynoline (AC) is one of the main chemical components of Corydalis bungeana Turcz. which has been shown to have a protective effect against acute liver injury. However, Whether AC is protective against acute liver injury remains unclear. This study aimed to explore the protective mechanism of AC against acute liver injury from in vivo as well as experiments in vitro. In experimental in vivo studies, AC pretreatment reduced the serum levels of ALT and AST and inhibited the expression of inflammatory factors in the liver of LPS/D-GalN-induced mice and alcohol liver disease mice. RNA-sequencing and molecular docking were used to predict that AC exerts its anti-inflammatory effects through the Toll-like receptor signaling pathway. Using RT-qPCR and Western blotting to detect expression levels of key genes and nodal proteins of the Toll-like receptor signaling pathway, AC was found to inhibit the phosphorylation of nuclear factor-kappaB (NF-ĸB) and c-Jun amino-terminal kinase (JNK). This finding was validated in cellular experiments. In conclusion, AC exerts its anti-hepatic injury effect by suppressing inflammation through inhibition of the TLR4/JNK/NF-ĸB pathway.

Apple polyphenols (APs) have gained attention for their various bioactivities, while no studies on anti-liver fibrosis activity are reported. This study evaluated the protective effect of APs on liver fibrosis using LPS-treated activated HSC-T6 cells and alcohol-treated liver fibrosis (ALF) mice. The results indicated that APs inhibited HSC-T6 cells activation in vitro and reduced the level of serum hyaluronic acid (HA) (p < 0.05), decreased fibrogenesis marker expression (p < 0.05), thereby alleviating ALF. In addition, APs (800 mg/kg b.w.) decreased the Firmicutes/Bacteroidetes ratio (p < 0.05) in ALF mice, inhibited LPS accumulation in the liver tissue and serum (p < 0.05), and significantly inhibited the TLR4/NF-κB/TGF-β signaling in mice liver. In conclusion, APs markedly ameliorated ALF, possibly by improving gut microbiota homeostasis, decreasing the translocation of bacterial endotoxins to the blood, and suppressing the TLR4/NF-κB/TGF-β signaling pathway, indicating its potential as lead compounds for functional foods and/or drugs against ALF.

Alcohol-related liver disease (ALD) is a major cause of morbidity and mortality worldwide. It encompasses conditions such as fatty liver, alcoholic hepatitis, chronic hepatitis with liver fibrosis or cirrhosis, and hepatocellular carcinoma. Numerous recent studies have demonstrated the critical role of oxidative stress, abnormal lipid metabolism, endoplasmic reticulum stress, various forms of cell death (including apoptosis, necroptosis, and ferroptosis), intestinal microbiota dysbiosis, liver immune response, cell autophagy, and epigenetic abnormalities in the pathogenesis of ALD. Currently, abstinence, corticosteroids, and nutritional therapy are the traditional therapeutic interventions for ALD. Emerging therapies for ALD mainly include the blockade of inflammatory pathways, the promotion of liver regeneration, and the restoration of normal microbiota. Summarizing the advances in animal models of ALD will facilitate a more systematic investigation of the pathogenesis of ALD and the exploration of therapeutic targets. This review summarizes the latest insight into the pathogenesis and molecular mechanisms of ALD, as well as the pros and cons of ALD rodent models, providing a basis for further research on therapeutic strategies for ALD.

Alcoholic liver disease (ALD) is a spectrum of liver damage caused by chronic alcohol consumption. The disease progresses in stages, starting with simple fatty liver, progressing to alcoholic hepatitis and potentially leading to fibrosis and cirrhosis. The pathophysiology of ALD is complex and involves several cellular and molecular mechanisms. Recent research has highlighted the role of long non-coding RNAs (LncRNAs) as critical regulators in the development and progression of ALD. This article reviews the current understanding of LncRNAs in ALD, focusing on their functions in key pathological processes and their potential as diagnostic markers and therapeutic targets.

Hepatic proteome and gut microbiota alterations are known in alcohol-associated hepatitis (AAH). Current animal models sparsely mimic human AAH. We aimed to develop an murine model that closely resembled human AAH.

Male C57BL/6N mice were pair-fed control/incremental ethanol Lieber-DeCarli diets and thioacetamide (TAA) for 12-weeks to induce AAH. Hepatic proteome was analyzed using LC-MS/MS. Gut-bacteria was determined using 16s-rRNA sequencing.

Mice exposed to EtOH+TAA displayed higher expression of liver triglycerides (1.5-fold, p = 0.001), pro-inflammatory (IL6, 1.5-fold, p = 0.002 and TNFα, 1.7-fold, p = 0.01), fibrotic (TGF-β, 2.7-fold, p = 0.01 and Col1α1, 2-fold, p = 0.01) and oxidative markers (GSH and SOD (-1.5 fold, p = 0.004 & 0.005 respectively)) as compared to EtOH alone. Histology of EtOH+TAA liver displayed pericellular liver fibrosis, increased steatosis, and neutrophil infiltration, which resembled human AAH. In the 12wk EtOH+TAA group, Desulfobacteria, Campylobacteria, and Patescibacteria increased by 2-fold (p = 0.02). Pathway combined score (CS, log10) in EtOH+TAA treatment showed upregulated hepatic ethanol oxidation (CS=1.93), fatty acid biosynthesis (CS=2.48), necrosis (CS=1.59), collagen formation (CS=1.28) and hypoxia (CS=0.68) and downregulated fatty acid beta-oxidation (CS=2.37), PPAR signaling (CS=1.35) fatty acid degradation (CS=2.35), bile acid metabolism (CS=1.87), and oxidative phosphorylation (CS=1.50), as observed in human disease.

Using an ethanol-thioacetamide combination in mice results in a faster establishment of AAH with fibrosis than previously known models. Differential protein expression strongly correlates with pathways found altered in human AAH, thus making the model mimic human disease better than other known models., respectively. Thioacetamide (TAA) was administered to enhance liver fibrosis and mimic human AAH.

Chronic alcohol users often exhibit an increased minimum alveolar concentration (MAC) of sevoflurane, yet the specific mechanism remains unclear. It has been reported that ethanol exposure can upregulate the protein expression and enzyme activity of cytochrome P450 2E1 (CYP2E1). CYP2E1 is a key enzyme that converts 2-5% of sevoflurane into equimolar amounts of hexafluoroisopropanol (HFIP) and F-. This study aims to explore whether ethanol exposure could alter sevoflurane metabolism through CYP2E1 modulation, potentially explaining the increased MAC observed in alcohol users.

Eighty adult male Sprague-Dawley (SD) rats were randomly divided into two groups and received either 50% ethanol (dose: 3 g/kg) or 0.9% saline twice daily by gavage. After 1, 2, 3, and 4 weeks of gavage, ten rats were randomly selected from each group to undergo 1-hour anesthesia with 2.3% sevoflurane. Blood samples were collected after anesthesia to measure the concentration of free HFIP using gas chromatography. Additionally, the left lobe tissue of the liver was collected for the analysis of CYP2E1 protein expression by Western blot and CYP2E1 enzyme activity by colorimetric assay. Correlations between these parameters were analyzed using Pearson's correlation.

In the ethanol group, CYP2E1 expression, activity, and the concentration of free HFIP were significantly higher at all time points compared to the control group (P < 0.05), except for protein expression in the first week (P > 0.05). Within-group comparisons indicated no significant changes in any of the parameters for the control group (P > 0.05). In the ethanol group, there was no difference in free HFIP concentration between the first and second weeks (P > 0.05), but a significant increase was observed in the third and fourth weeks (P < 0.01); protein expression and enzyme activity significantly varied over time, especially showing a notable increase from the first to the third and fourth weeks (P < 0.05). Correlation analysis revealed strong positive correlations between free HFIP concentration and CYP2E1 activity (r = 0.7898), free HFIP concentration and CYP2E1 expression (r = 0.8418), and CYP2E1 activity and expression (r = 0.8740), all with P < 0.001.

Ethanol exposure increased both the expression and enzymatic activity of CYP2E1, consequently enhancing the metabolism of sevoflurane.

Hepatocyte organoids (HOs) have superior hepatic functions to cholangiocyte-derived organoids but suffer from shorter lifespans. To counteract this, we co-cultured pig HOs with adipose-derived mesenchymal stem cells (A-MSCs) and performed transcriptome analysis. The results revealed that A-MSCs enhanced the collagen synthesis pathways, which are crucial for maintaining the three-dimensional structure and extracellular matrix synthesis of the organoids. A-MSCs also increased the expression of liver progenitor cell markers (KRT7, SPP1, LGR5+, and TERT). To explore HOs as a liver disease model, we exposed them to alcohol to create an alcoholic liver injury (ALI) model. The co-culture of HOs with A-MSCs inhibited the apoptosis of hepatocytes and reduced lipid accumulation of HOs. Furthermore, varying ethanol concentrations (0-400 mM) and single-versus-daily exposure to HOs showed that daily exposure significantly increased the level of PLIN2, a lipid storage marker, while decreasing CYP2E1 and increasing CYP1A2 levels, suggesting that CYP1A2 may play a critical role in alcohol detoxification during short-term exposure. Moreover, daily alcohol exposure led to excessive lipid accumulation and nuclear fragmentation in HOs cultured alone. These findings indicate that HOs mimic in vivo liver regeneration, establishing them as a valuable model for studying liver diseases, such as ALI.

Probiotics can regulate gut microbiota and protect against acute alcohol-induced liver injury through the gut-liver axis. However, efficacy is strain-dependent, and their mechanism remains unclear. This study investigated the effect of lactic acid bacteria (LAB), including Lacticaseibacillus paracasei E10 (E10), Lactiplantibacillus plantarum M (M), Lacticaseibacillus rhamnosus LGG (LGG), Lacticaseibacillus paracasei JN-1 (JN-1), and Lacticaseibacillus paracasei JN-8 (JN-8), on the prevention of acute alcoholic liver injury in mice. We found that LAB pretreatment reduced serum alanine transaminase (ALT) and aspartate transaminase (AST) and reduced hepatic total cholesterol (TC) and triglyceride (TG). JN-8 pretreatment exhibited superior efficacy in improving hepatic antioxidation. LGG and JN-8 pretreatment significantly attenuated hepatic and colonic inflammation by decreasing the expression of interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α) and increasing the expression of interleukin 10 (IL-10). JN-1 and JN-8 pretreatments have better preventive effects than other LAB pretreatment on intestinal barrier dysfunction. In addition, the LAB pretreatment improved gut microbial dysbiosis and bile acid (BA) metabolic abnormality. All of the strains were confirmed to have bile salt deconjugation capacities in vitro, where M and JN-8 displayed higher activities. This study provides new insights into the prevention and mechanism of LAB strains in preventing acute alcoholic liver injury.

Alcohol-related liver disease (ALD) is a major health concern worldwide, but effective therapeutics for ALD are still lacking. Tumor necrosis factor-inducible gene 6 protein (TSG-6), a cytokine released from mesenchymal stem cells, was shown to reduce liver fibrosis and promote successful liver repair in mice with chronically damaged livers. However, the effect of TSG-6 and the mechanism underlying its activity in ALD remain poorly understood.

To investigate its function in ALD mice with fibrosis, male mice chronically fed an ethanol (EtOH)-containing diet for 9 weeks were treated with TSG-6 (EtOH + TSG-6) or PBS (EtOH + Veh) for an additional 3 weeks.

Severe hepatic injury in EtOH-treated mice was markedly decreased in TSG-6-treated mice fed EtOH. The EtOH + TSG-6 group had less fibrosis than the EtOH + Veh group. Activation of cluster of differentiation 44 (CD44) was reported to promote HSC activation. CD44 and nuclear CD44 intracellular domain (ICD), a CD44 activator which were upregulated in activated HSCs and ALD mice were significantly downregulated in TSG-6-exposed mice fed EtOH. TSG-6 interacted directly with the catalytic site of MMP14, a proteolytic enzyme that cleaves CD44, inhibited CD44 cleavage to CD44ICD, and reduced HSC activation and liver fibrosis in ALD mice. In addition, a novel peptide designed to include a region that binds to the catalytic site of MMP14 suppressed CD44 activation and attenuated alcohol-induced liver injury, including fibrosis, in mice.

These results demonstrate that TSG-6 attenuates alcohol-induced liver damage and fibrosis by blocking CD44 cleavage to CD44ICD and suggest that TSG-6 and TSG-6-mimicking peptide could be used as therapeutics for ALD with fibrosis.

We aimed to investigate the effectiveness of physical training as a protective strategy to mitigate alveolar bone damage and blood antioxidant defense caused by ethanol (EtOH) consumption in a binge-drinking pattern. Male Wistar rats aged approximately 90 days were divided into four groups: control, training, EtOH, and training + EtOH. The physical training protocol was conducted on a treadmill for four consecutive weeks, while the animals in the EtOH group were administered EtOH via orogastric gavage for three consecutive days each week, following the binge drink pattern. After the training period, blood and mandibles were collected for plasma oxidative biochemistry analysis, and the alveolar bone was subjected to physicochemical composition analysis, tissue evaluation, and microtomography evaluation. Our results showed that EtOH induced oxidative stress and physical exercise promoted the recovery of antioxidant action. Physical training minimized the damage to the mineral/matrix composition of the alveolar bone due to EtOH consumption and increased the density of osteocytes in the trained group treated with EtOH than in those exposed only to EtOH. Furthermore, physical training reduced damage to the alveolar bone caused by EtOH consumption. Our findings suggest that physical training can serve as an effective strategy to reduce systemic enzymatic oxidative response damage and alleviate alveolar bone damage resulting from alcohol consumption. Further investigations are warranted to elucidate the underlying mechanisms and explore, in addition to physical training, the potential effects of other activities with varying intensities on managing alcohol-induced bone damage.

Hepatic encephalopathy (HE) is defined as decline in neurological function during chronic liver disease (CLD). Alcohol is a major etiological factor in the pathogenesis of fibrosis/cirrhosis and has also been documented to directly impact the brain. However, the role of alcohol in the development of HE in CLD remains unclear. Here, we investigated the impact of excessive alcohol administration on neurological deterioration in rats with CLD. Starting day 7 post-BDL surgery, rats were administered alcohol twice daily (51% v/v ethanol, 3 g/kg, via gavage) for 4 weeks. Motor coordination was assessed weekly using rotarod and anxiety-like behavior was evaluated with open field and elevated plus maze at 5 weeks. Upon sacrifice, brains were collected for western blot and immunohistochemical analyses to investigate neuronal integrity and oxidative stress status. Alcohol worsened motor coordination performance and increased anxiety-like behavior in BDL rats. Impairments were associated with decreased neuronal markers of NeuN and SMI311, increased apoptotic markers of cleaved/pro-caspase-3 and Bax/Bcl2, increased necroptosis markers of pRIP3 and pMLKL, decreased total antioxidant capacity (TAC), and increased 4-hydroxynonenal (4-HNE)modified proteins in the cerebellum of BDL-alcohol rats when compared to respective controls. Immunofluorescence confirmed the colocalization of cleaved caspase-3 and pMLKL in the granular neurons of the cerebellum of BDL-alcohol rats. Excessive alcohol consumption exacerbates HE which leads to associated apoptotic and necroptotic neuronal loss in the cerebellum of BDL-alcohol rats. Additionally, higher levels of 4-HNE and decreased TAC in the cerebellum of BDL-alcohol rats suggest oxidative stress is the triggering factor of apoptotic and necroptotic neuronal loss/injury.

The objective of this study was to determine the optimal extraction conditions for total flavonoids from S. bigelovii using microwave-assisted extraction and to analyze the protective effect of total flavonoids from S. bigelovii on alcoholic liver injury in mice. The optimization of the process conditions for the microwave-assisted extraction of total flavonoids from S. bigelovii was performed using response surface methodology, and an alcohol-induced acute liver injury model in mice was used to investigate the effects of different doses of total flavonoids (100 mg/kg, 200 mg/kg, and 400 mg/kg) on the levels and activities of serum alanine aminotransferase kits (ALT), glutamic oxaloacetic transaminase kits (AST), superoxide dismutase kits (SOD), glutathione peroxidase kits (GSH-Px), and malondialdehyde (MDA). We performed hematoxylin-eosin (H&E) staining analysis on pathological sections of mouse liver tissue, and qRT-PCR technology was used to detect the expression levels of the inflammatory factors IL-1 β, IL-6, and TNF-α. The results revealed that the optimal extraction process conditions for total flavonoids in S. bigelovii were a material-to-liquid ratio of 1:30 (g/mL), an ethanol concentration of 60%, an extraction temperature of 50 °C, an ultrasound power of 250 W, and a yield of 5.71 ± 0.28 mg/g. Previous studies have demonstrated that the flavonoids of S. bigelovii can significantly inhibit the levels of ALT and AST in the serum (p < 0.001), reduce MDA levels (p < 0.001), increase the activity of the antioxidant enzymes SOD and GSH-Px (p < 0.001), and inhibit the IL-1 β, IL-6, and TNF-α gene expression levels (p < 0.001) of inflammatory factors. The total flavonoids of S. bigelovii exert a protective effect against alcoholic liver injury by reducing the levels of inflammation, oxidative stress, and lipid peroxidation caused by alcohol. The results of this study lay the foundation for the high-value utilization of S. bigelovii and provide new resources for the development of liver-protective drugs.

Chronic alcohol exposure increases liver damage such as lipid accumulation and hepatitis, resulting in hepatic cirrhosis. Chronic alcohol intake is known to disturb circadian rhythms in humans and animals. DEC1, a basic helix-loop-helix transcription factor, plays an important role in the circadian rhythm, inflammation, immune responses, and tumor progression. We have previously shown that Dec1 deficiency inhibits stresses such as periodontal inflammation and perivascular fibrosis of the heart. However, the significance of Dec1 deficiency in chronic alcohol consumption remains unclear. In the present study, we investigated whether the biological stress caused by chronic alcohol intake is inhibited in Dec1 knockout mice. We treated control and Dec1 knockout mice for three months by providing free access to 10% alcohol. The Dec1 knockout mice consumed more alcohol than control mice, however, we observed severe hepatic lipid accumulation and circadian rhythm disturbance in control mice. In contrast, Dec1 knockout mice exhibited little effect on these outcomes. We also investigated the expression of peroxisome proliferator-activated receptors (PPARs) and AMP-activated protein kinase (AMPK), which are involved in the regulation of fatty acid metabolism. Immunohistochemical analysis revealed increases of phosphorylation AMPK and PPARa but decreases PPARg in Dec1 knockout mice compared to that in control mice. This indicates a molecular basis for the inhibition of hepatic lipid accumulation in alcohol-treated Dec1 knockout mice. These results suggest a novel function of Dec1 in alcohol-induced hepatic lipid accumulation and circadian rhythm disorders.

Alcohol consumption poses a significant risk for the development of chronic illnesses, one of the leading causes of "preventable" disease and death worldwide. Harmful consumption of alcohol is thought to result in approximately 2.5-3 million deaths each year, the majority of which are caused by alcohol-related liver diseases. Hepatocellular carcinoma, cirrhosis, fibrosis, steatosis, and steatohepatitis are among the liver illnesses caused by alcohol. The mechanisms behind human diseases are often mimicked and understood through the use of animal models. Rodents are the ideal animals to study alcohol-related liver diseases. In these experimental models using rodents, the ethanol ratio, method of administration, and diet to be applied vary. Within the scope of this review, it is aimed at providing information about the experimental models used today for alcohol toxicity and the advantages and disadvantages of these models.

Although alcohol is a well-known causal factor associated with liver diseases, challenges remain in inducing liver fibrosis in experimental rodent models. These challenges include rodents' natural aversion to high concentrations of alcohol, rapid alcohol metabolism, the need for a prolonged duration of alcohol administration, and technical difficulties. Therefore, it is crucial to establish an experimental model that can replicate the features of alcoholic liver fibrosis. The objective of this study was to develop a feasible rat model of alcoholic liver fibrosis that emulates human drinking patterns and combines low-dose chemicals within a relatively short time frame. We successfully developed an 8-week rat model of alcoholic liver fibrosis that mimics chronic and heavy drinking patterns. Rats were fed with a control liquid diet, an alcohol liquid diet, or alcohol liquid diet combined with multiple binges via oral gavage. To accelerate the progression of alcoholic liver fibrosis, we introduced low-dose carbon tetrachloride (CCl4) through intraperitoneal injection. This model allows researchers to efficiently evaluate potential therapeutics in preclinical studies of alcoholic liver fibrosis within a reasonable time frame.

Chronic ethanol overconsumption promotes alcohol-associated liver disease (ALD), characterized by hepatocyte injury, inflammation, hepatic stellate cell (HSC) activation, and fibrosis. Hyaluronan (HA) concentration is greater in livers and blood from advanced ALD patients than patients with advanced non-ALD. In the liver, HSCs are the major HA producers. The relationship between ethanol, HA, and HSC activation is incompletely understood. Thus, here, we tested the hypothesis that ethanol enhances HSC activation in a HA-dependent manner.

Liver tissue microarrays (TMAs) containing steatotic livers from donors with or without a history of alcohol consumption were used to measure HA and collagen content. Mice were fed a moderate (2%, v/v) ethanol-containing diet or pair-fed control diet for 2 days, after which they were given a single carbon tetrachloride (CCl4 ) injection. To inhibit HA synthesis, we provided 4-methylumbelliferone (4MU) daily. We used LX2 cells, a human HSC cell line, to determine the impact ethanol had on LPS responses, with or without concurrent 4MU exposure.

CCl4 induced liver injury, but it did not differ between ethanol or control diet fed mice with or without 4MU treatment. Ethanol feeding enhanced CCl4 -induced hepatic HA content, which was paralleled by HA synthase (Has)2 transcript abundance; 4MU treatment normalized both. Consistently, HSC activation, assessed by measuring αSMA mRNA and protein, was induced by CCl4 exposure, enhanced by ethanol feeding, and normalized by 4MU. Hepatic transcripts, but not protein, for Ccl2 were enhanced by ethanol feeding and normalized by 4MU exposure. Finally, ethanol-exposed LX2 cells made more LPS-stimulated CCL2 mRNA and protein than cells not exposed to ethanol; 4MU prevented this.

These data show that ethanol augments HSC activation through HA synthesis and enhances hepatic profibrogenic features. Therefore, targeting HSC HA production could potentially attenuate liver disease in ALD patients.

Fatty liver diseases, including alcohol-associated liver disease (ALD) and nonalcoholic fatty liver disease nonalcoholic fatty liver disease (NAFLD), affect a large number of people worldwide and become one of the major causes of end-stage liver disease, such as liver cirrhosis and hepatocellular carcinoma (HCC). Unfortunately, there are currently no approved pharmacological treatments for ALD or NAFLD. This situation highlights the urgent need to explore new intervention targets and discover effective therapeutics for ALD and NAFLD. The lack of properly validated preclinical disease models is a major obstacle to the development of clinical therapies. ALD and NAFLD models have been in the development for decades, but there are still no models that recapitulate the full spectrum of ALD and NAFLD. Throughout this review, we summarize the current in vitro and in vivo models used for research on fatty liver diseases and discuss the advantages and limitations of these models.

The effect of europinidin on alcoholic liver damage in rats was examined in this research.

A total of 24 Wistar rats were grouped in the same way into four groups: normal control (normal), ethanol control (EtOH), europinidin low dose (10 mg/kg), and europinidin higher dose (20 mg/kg). The test group rats were orally treated with europinidin-10 and europinidin-20 for 4 weeks, whereas 5 mL/kg distilled water was administered to control rats. In addition, 1 h after the last dose of the above-mentioned oral treatment, 5 mL/kg (i.p.) EtOH was injected to induce liver injury. After 5 h of EtOH treatment, samples of blood were withdrawn for biochemical estimations.

Administration of europinidin at both doses restored all of the estimated serum, i.e., liver function tests (ALT, AST, ALP), biochemical test (Creatinine, albumin, BUN, direct bilirubin, and LDH), lipid assessment (TC and TG), endogenous antioxidants (GSH-Px, SOD, and CAT), malondialdehyde (MDA), nitric oxide (NO), cytokines (TGF-β, TNF-α, IL-1β, IL-6, IFN-γ, and IL-12), caspase-3, and nuclear factor kappa B (NF-κB) associated with the EtOH group.

The results of the investigation showed that europinidin had favorable effects in rats given EtOH and may have hepatoprotective potential property.

Prenatal alcohol exposure affects the cardiovascular health of the offspring. Epigallocatechin-3-gallate (EGCG) may be a protective agent against it, but no data are available regarding its impact on cardiac dysfunction. We investigated the presence of cardiac alterations in mice prenatally exposed to alcohol and the effect of postnatal EGCG treatment on cardiac function and related biochemical pathways. C57BL/6J pregnant mice received 1.5 g/kg/day (Mediterranean pattern), 4.5 g/kg/day (binge pattern) of ethanol, or maltodextrin until Day 19 of pregnancy. Post-delivery, treatment groups received EGCG-supplemented water. At post-natal Day 60, functional echocardiographies were performed. Heart biomarkers of apoptosis, oxidative stress, and cardiac damage were analyzed by Western blot. BNP and Hif1α increased and Nrf2 decreased in mice prenatally exposed to the Mediterranean alcohol pattern. Bcl-2 was downregulated in the binge PAE drinking pattern. Troponin I, glutathione peroxidase, and Bax increased in both ethanol exposure patterns. Prenatal alcohol exposure led to cardiac dysfunction in exposed mice, evidenced by a reduced ejection fraction, left ventricle posterior wall thickness at diastole, and Tei index. EGCG postnatal therapy restored the physiological levels of these biomarkers and improved cardiac dysfunction. These findings suggest that postnatal EGCG treatment attenuates the cardiac damage caused by prenatal alcohol exposure in the offspring.

The toxic effects of alcohol consumption on population health are significant worldwide and the synergistic toxic effects of concurrent intake of Acetaminophen and alcohol is of clinical concern. The understanding of molecular mechanisms beneath such synergism and acute toxicity may be enhanced through assessing underlying metabolomics changes. The molecular toxic activities of the model hereby, is assessed though metabolomics profile with a view to identifying metabolomics targets which could aid in the management of drug-alcohol interactions. In vivo exposure of C57/BL6 mice to APAP (70 mg/kg), single dose of ethanol (6 g/kg of 40%) and APAP after alcohol consumption was employed. Plasma samples were prepared and subjected to biphasic extraction for complete LC-MS profiling, and tandem mass MS² analysis. Among the detected ions, 174 ions had significant (VIP scores >1 and FDR <0.05) changes between groups and were selected as potential biomarkers and significant variables. The presented metabolomics approach highlighted several affected metabolic pathways, including nucleotide and amino acid metabolism; aminoacyl-tRNA biosynthesis as well as bioenergetics of TCA and Krebs cycle. The impact of APAP on the concurrent administration of alcohol showed great biological interactions in the vital ATP and amino acid producing processes. The metabolomics changes show distinct metabolites which are altered to alcohol-APAP consumption while presenting several unneglectable risks on the vitality of metabolites and cellular molecules which shall be concerned.

Heavy drinking is a primary cause of alcoholic liver injury (ALI). Pituitary tumour transforming gene 1 (PTTG1) is involved in the occurrence and development of hepatocellular carcinoma (HCC), which is a well-known inflammation-related cancer with various aetiologies, including alcohol consumption. However, the role of PTTG1 in alcohol-induced liver injury and inflammation is not clear.

Blood samples were collected from patients with acute alcohol intoxication (n = 20) and healthy controls (n = 20). PTTG1 knockout (KO) mice and PTTG1 transgenic (TG) mice were given a single gavage of alcohol (5 g/kg, 50%) to construct the alcohol-induced liver injury.

We found that serum PTTG1 levels were downregulated in acute ALI patients. In addition, acute alcohol administration significantly reduced PTTG1 levels in the serum and liver of mice. Compared to wild-type mice, PTTG1 KO mice had more serious liver injury, which was accompanied by worsened hepatic endoplasmic reticulum (ER) stress and hepatocyte pyroptosis induced by alcohol. Similarly, PTTG1 deficiency exacerbated alcohol-induced cell death in primary mouse hepatocytes and LO2 cells, by increasing hepatic ER stress and pyroptosis. Importantly, TUDCA, an ER stress inhibitor, could blocked alcohol-induced hepatic pyroptosis in PTTG1 knockdown LO2 cells. Finally, overexpression of PTTG1 substantially attenuated alcohol-induced liver injury by reducing ER stress and hepatic pyroptosis in mice.

We demonstrated that PTTG1 participates in ALI and has a protective effect against alcohol-induced hepatic ER stress and pyroptosis.

Cyclin E1 is the regulatory subunit of cyclin-dependent kinase 2 (Cdk2) and one of the central players in cell cycle progression. We recently showed its crucial role for initiation of liver fibrosis and hepatocarcinogenesis. In the present study, we investigated the role of Cyclin E1 in the development of alcohol-associated liver disease (ALD).

Mice with constitutive (E1^-/-), hepatocyte-specific (Cyclin E1^Δhepa), or intestinal-epithelial-cell-specific (Cyclin E1^ΔIEC) inactivation of Cyclin E1 and corresponding wild type littermate controls (WT) were administered either a Lieber-DeCarli ethanol diet (LDE) for 3 weeks or acute ethanol binges (6 g/kg) through oral gavage. Serum parameters of liver functionality were measured; hepatic tissues were collected for biochemical and histological analyses.

The administration of acute EtOH binge and chronic LDE diet to E1^-/- mice enhanced hepatic steatosis, worsened liver damage and triggered body weight loss. Similarly, in the acute EtOH binge model, Cyclin E1^Δhepa mice revealed a significantly worsened liver phenotype. In contrast, inactivation of Cyclin E1 only in intestinal epithelial cell (IECs)did not lead to any significant changes in comparison to WT mice after acute EtOH challenge. Remarkably, both acute and chronic EtOH administration in E1^-/- animals resulted in increased levels of ADH and decreased expression of ALDH1/2. The additional application of a pan-Cdk inhibitor (S-CR8) further promoted liver damage in EtOH-treated WT mice.

Our data point to a novel unexpected role of Cyclin E1 in hepatocytes for alcohol metabolism, which seems to be independent of the canonical Cyclin E1/Cdk2 function as a cell cycle regulator.

Alcohol consumption exacerbates liver abnormalities in animal models, but whether it increases the severity of liver disease in early diabetic or prediabetic rats is unclear. To investigate the molecular mechanisms underlying alcohol-induced liver steatosis or hepatitis, we used a prediabetic animal model. Otsuka Long-Evans Tokushima Fatty (OLETF) and Long-Evans Tokushima Fatty (LETO) rats were pair-fed with an ethanol-containing liquid diet for 6 weeks. Compared with controls, OLETF and LETO rats displayed more pronounced liver steatosis and higher plasma levels of serum glutamic oxaloacetic transaminase (SGOT) and serum glutamate pyruvate transaminase (SPGT), indicating liver injury. Ethanol-fed LETO (Pd-L-E) rats showed mild liver steatosis and no inflammation compared with ethanol-fed OLETF (Pd-O-E) rats. Although precursor and active SREBP-1 levels in the liver of ethanol-fed OLETF rats significantly increased compared with control diet-fed OLETF rats (Pd-O-C), those of Pd-L-E rats did not. Bone morphogenetic protein (BMP) and TGF-β1 balance in Pd-O-E rats was significantly altered because BMP signaling was upregulated by inducing BMP2, BMP4, BMP7, BMP9, Smad1, and Smad4, whereas TGF-β1, Smad3, and Erk were downregulated. Activation of TGF-β/Smad signaling inhibited BMP2 and BMP9 expression and increased epithelial-mesenchymal transition (EMT) marker levels (Hepcidin, Snail, and Twist) in the liver of LETO rats. Livers of ethanol-fed OLETF rats showed increased levels of vimentin, FSP-1, α-SMA, MMP1, MMP13, and collagen III compared with rats of other groups, whereas EMT marker levels did not change. Thus, BMP exerted anti- and/or pro-fibrotic effects in ethanol-fed rats. Therefore, BMP and TGF-β, two key members of the TGF-β superfamily, play important but diverse roles in liver steatosis in young LETO and OLETF rats.

Alcoholic liver disease (ALD) is the leading cause of alcohol-related deaths worldwide. Experimental ALD models are expensive and difficult to reproduce. A low-cost, reproducible ALD model was developed, and liver damage compared with the gut microbiota. The aims of this study were to develop an experimental model of ALD, through a high-fat diet, the chronic use of ethanol, and intragastric alcohol binge; and to evaluate the composition of the gut microbiota and its correlation with markers of inflammatory and liver disease progression in this model.

Adult male Wistar rats were randomized (N = 24) to one of three groups: control (standard diet and water + 0.05% saccharin), ALC4 and ALC8 (sunflower seed, 10% ethanol + 0.05% saccharin for 4 and 8 wk, respectively). On the last day, ALC4/8 received alcoholic binge (5 g/kg). Clinical, nutritional, biochemical, inflammatory, pathologic, and gut microbiota data were analyzed.

ALC4/8 animals consumed more alcohol and lipids (P < 0.01) and less total energy, liquids, solids, carbohydrates, and proteins (P < 0.01), and gained less weight (P < 0.01) than controls. ALC8 had lower Lee index scores than controls and ALC4 (P < 0.01). Aminotransferases increased and albumin diminished in ALC4/8 but not in the control group (P < 0.03 for all). Glucose and aspartate transaminase/alanine aminotransaminase ratios were higher in the ALC8 rats than in the controls (P < 0.03). Cholesterol was higher in ALC4 and lower in ALC8 compared with controls (P < 0.03). Albumin and high-density lipoprotein cholesterol levels were lower in ALC8 (P < 0.03). Hepatic concentration of triacylglycerols was higher in ALC8 than in ALC4 and controls (P < 0.05). ALC4/8 presented microvesicular grade 2 and 3 steatosis, respectively, and macrovesicular grade 1. No change in the gene expression of inflammatory markers between groups was seen. ALC4/8 had lower fecal bacterial α-diversity and relative abundance of Firmicutes (P < 0.005) and greater Bacterioidetes (P < 0.0007) and Protobacteria (P < 0.001) than controls. Gut microbiota correlated with serum and liver lipids, steatosis, albumin, and aminotransferases (P < 0.01 for all).

The model induced nutritional, biochemical, histologic, and gut microbiota changes, and appears to be useful in the study of therapeutic targets.

The common aldehyde dehydrogenase 2 (ALDH2) alcohol flushing variant known as ALDH2*2 affects ∼8% of the world's population. Even in heterozygous carriers, this missense variant leads to a severe loss of ALDH2 enzymatic activity and has been linked to an increased risk of coronary artery disease (CAD). Endothelial cell (EC) dysfunction plays a determining role in all stages of CAD pathogenesis, including early-onset CAD. However, the contribution of ALDH2*2 to EC dysfunction and its relation to CAD are not fully understood. In a large genome-wide association study (GWAS) from Biobank Japan, ALDH2*2 was found to be one of the strongest single-nucleotide polymorphisms associated with CAD. Clinical assessment of endothelial function showed that human participants carrying ALDH2*2 exhibited impaired vasodilation after light alcohol drinking. Using human induced pluripotent stem cell-derived ECs (iPSC-ECs) and CRISPR-Cas9-corrected ALDH2*2 iPSC-ECs, we modeled ALDH2*2-induced EC dysfunction in vitro, demonstrating an increase in oxidative stress and inflammatory markers and a decrease in nitric oxide (NO) production and tube formation capacity, which was further exacerbated by ethanol exposure. We subsequently found that sodium-glucose cotransporter 2 inhibitors (SGLT2i) such as empagliflozin mitigated ALDH2*2-associated EC dysfunction. Studies in ALDH2*2 knock-in mice further demonstrated that empagliflozin attenuated ALDH2*2-mediated vascular dysfunction in vivo. Mechanistically, empagliflozin inhibited Na⁺/H⁺-exchanger 1 (NHE-1) and activated AKT kinase and endothelial NO synthase (eNOS) pathways to ameliorate ALDH2*2-induced EC dysfunction. Together, our results suggest that ALDH2*2 induces EC dysfunction and that SGLT2i may potentially be used as a preventative measure against CAD for ALDH2*2 carriers.

Alcoholic liver disease (ALD) is still a global health concern. Long-term alcohol intake alters the gut microbiota diversity and metabolic activity, and causes intestinal barrier dysfunction, leading to the development of ALD. This research explored the protective effects and underlying mechanisms of red raspberry (RR) on alcohol-related disorders in mice. Male C57BL/6J mice were fed a standard diet or a standard diet supplemented with 2%, 4%, and 8% weight/weight RR. Meanwhile, mice were administered 35% (v/v) ethanol (EtOH, 10 mL per kg body weight) intragastrically once daily for six weeks, except the control group mice. The results showed that RR supplementation decreased liver injury markers (alanine and aspartate transaminases) in the serum, reduced triglyceride level in the liver and downregulated hepatic cytochrome P450 2E1 mRNA expression in mice administered EtOH. In addition, EtOH-mediated oxidative stress in the liver was attenuated by RR supplementation through decreased hepatic malondialdehyde content and increased antioxidant (glutathione, glutathione peroxidase, and catalase) levels and activities in mice exposed to EtOH. Moreover, RR supplementation reversed EtOH-induced alteration in the cecal microbial composition at the phylum, order, genus, and species levels and improved the intestinal barrier function associated with the inhibition of the NF-κB/MLCK pathway, which was accompanied by upregulation of tight junctions (zonula occludens 1, occludin, claudin-1, and claudin-4) and E-cadherin mRNA and protein expressions. Accordingly, RR supplementation resulted in a decreased level of endotoxins in the serum and attenuation of the inflammatory response in the liver, illustrated by a significant decrease in tumor necrosis factor-alpha, interleukin (IL)-1β, and IL-6 levels. Overall, RR supplementation alleviated the adverse effects of chronic alcohol intake in C57BL/6J mice and could be a potential supplement for improving ALD.

Circadian rhythms are important for organisms to adapt to the environment and maintain homeostasis. Disruptions of circadian rhythms contribute to the occurrence, progression, and exacerbation of diseases, such as cancer, psychiatric disorders, and metabolic disorders. Alcohol-induced liver disease (ALD) is one of the most prevalent liver diseases. Disruptions of the circadian clock enhance the ALD symptoms using chronic mice models or genetic manipulated mice. However, chronic models are time consuming and clock gene deletions interfere with metabolisms. Here, we report that constant light (LL) condition significantly disrupted the circadian clock in an acute ALD model, resulting in aggravated ALD phenotypes in wild type mice. Comparative transcriptome analysis revealed that the alcohol feeding affected the circadian pathway, as well as metabolic pathways. The acute alcohol feeding plus the LL condition further interfered with metabolic pathways and dysregulated canonical circadian gene expressions. These findings support the idea that disrupting the circadian clock could provide an improved ALD mouse model for further applications, such as facilitating identification of potential therapeutic targets for the prevention and treatment of ALD.Abbreviations: ALD, alcohol-induced liver disease; LD, 12 h light _ 12 h dark; LL, constant light; HF, high-fat liquid control diet; ETH, ethanol-containing diet; NIAAA, National Institute on Alcohol Abuse and Alcoholism; TTFLs, transcription-translation feedback loops; FDA, US Foods and Drug Administration; NAFLD, non-alcoholic fatty liver disease; RER, respiratory exchange rate; DEGs, differentially expressed genes; H&E, haematoxylin and eosin; ALT, alanine transaminase; AST, aspartate transaminase; TG, triglycerides.

The chronic-plus-binge model of ethanol consumption, where chronically (8-week) ethanol-fed mice are gavaged a single dose of ethanol (E8G1), is known to induce steatohepatitis in mice. However, how chronically ethanol-fed mice respond to multiple binges of ethanol remains unknown.

We extended the E8G1 model to three gavages of ethanol (E8G3) spaced 24 h apart, sacrificed each group 9 h after the final gavage, analyzed liver injury, and examined gene expression changes using microarray analyses in each group to identify mechanisms contributing to liver responses to binge ethanol.

Surprisingly, E8G3 treatment induced lower levels of liver injury, steatosis, inflammation, and fibrosis as compared to mice after E8G1 treatment. Microarray analyses identified several pathways that may contribute to the reduced liver injury after E8G3 treatment compared to E8G1 treatment. The gene encoding cytochrome P450 2B10 (Cyp2b10) was one of the top upregulated genes in the E8G1 group and was further upregulated in the E8G3 group, but only moderately induced after chronic ethanol consumption, as confirmed by RT-qPCR and western blot analyses. Genetic disruption of Cyp2b10 worsened liver injury in E8G1 and E8G3 mice with higher blood ethanol levels compared to wild-type control mice, while in vitro experiments revealed that CYP2b10 did not directly promote ethanol metabolism. Metabolomic analyses revealed significant differences in hepatic metabolites from E8G1-treated Cyp2b10 knockout and WT mice, and these metabolic alterations may contribute to the reduced liver injury in Cyp2b10 knockout mice.

Hepatic Cyp2b10 expression is highly induced after ethanol binge, and such upregulation reduces acute-on-chronic ethanol-induced liver injury via the indirect modification of ethanol metabolism.