In vivo Bruton's tyrosine kinase inhibition attenuates alcohol-associated liver disease by regulating CD84-mediated granulopoiesis.

Severe alcohol-associated hepatitis (AH) is a life-threatening form of alcohol-associated liver disease. Liver neutrophil infiltration is a hallmark of AH, yet the effects of alcohol on neutrophil functions remain elusive. Identifying therapeutic targets to reduce neutrophil-mediated liver damage is essential. Bruton's tyrosine kinase (BTK) plays an important role in neutrophil development and function; however, the role of BTK in AH is unknown. Using RNA sequencing of circulating neutrophils, we found an increase in Btk expression (P = 0.05) and phosphorylated BTK (pBTK) in patients with AH compared with healthy controls. In vitro, physiologically relevant doses of alcohol resulted in a rapid, TLR4-mediated induction of pBTK in neutrophils. In a preclinical model of AH, administration of a small-molecule BTK inhibitor (evobrutinib) or myeloid-specific Btk knockout decreased proinflammatory cytokines and attenuated neutrophil-mediated liver damage. We found that pBTK was essential for alcohol-induced bone marrow granulopoiesis and liver neutrophil infiltration. In vivo, BTK inhibition or myeloid-specific Btk knockout reduced granulopoiesis, circulating neutrophils, liver neutrophil infiltration, and liver damage in a mouse model of AH. Mechanistically, using liquid chromatography-tandem mass spectrometry, we identified CD84 as a kinase target of BTK, which is involved in granulopoiesis. In vitro, CD84 promoted alcohol-induced interleukin-1ß and tumor necrosis factor-α in primary human neutrophils, which was inhibited by CD84-blocking antibody treatment. Our findings define the role of BTK and CD84 in regulating neutrophil inflammation and granulopoiesis, with potential therapeutic implications in AH.

Fecal Carriage of Multidrug-Resistant Organisms Increases the Risk of Hepatic Encephalopathy in Cirrhotic Patients: Insights from Gut Microbiota and Metabolite Features.

Background: Impact of fecal colonization by multidrug-resistant organisms (MDROs) on changes in gut microbiota and associated metabolites, as well as its role in cirrhosis-associated outcomes, has not been thoroughly investigated. Methods: Eighty-eight cirrhotic patients and 22 healthy volunteers were prospectively enrolled with analysis conducted on plasma metabolites, fecal MDROs, and microbiota. Patients were followed for a minimum of one year. Predictive factors for cirrhosis-associated outcomes were identified using Cox proportional hazards regression models, and risk factors for fecal MDRO carriage were assessed using logistic regression model. Correlations between microbiota and metabolic profiles were evaluated through Spearman's rank test. Results: Twenty-nine (33%) cirrhotic patients exhibited MDRO carriage, with a notably higher rate of hepatic encephalopathy (HE) in MDRO carriers (20.7% vs. 3.2%, p = 0.008). Cox regression analysis identified higher serum lipopolysaccharide levels and fecal MDRO carriage as predictors for HE development. Logistic regression analysis showed that MDRO carriage is an independent risk factor for developing HE. Microbiota analysis showed a significant dissimilarity of fecal microbiota between cirrhotic patients with and without MDRO carriage (p = 0.033). Thirty-two metabolites exhibiting significantly different expression levels among healthy controls, cirrhotic patients with and without MDRO carriage were identified. Six of the metabolites showed correlation with specific bacterial taxa expression in MDRO carriers, with isoaustin showing significantly higher levels in MDRO carriers experiencing HE compared to those who did not. Conclusion: Fecal MDRO carriage is associated with altered gut microbiota, metabolite modulation, and an elevated risk of HE occurrence within a year.

Mac-2-Binding Protein Glycosylation Isomer to Albumin Ratio Predicts Bacterial Infections in Cirrhotic Patients.

INTRODUCTION: Mac-2-binding protein glycosylation isomer (M2BPGi) is a novel biomarker for liver fibrosis, but little is known about its role in cirrhosis-associated clinical outcomes. This study aimed to investigate the predictive role of M2BPGi in cirrhosis-associated complications. METHODS: One hundred and forty-nine cirrhotic patients were retrospectively enrolled. Patients were followed up for 1 year, and cirrhosis-associated clinical events were recorded. Receiver operating characteristic curve (ROC) analysis was used to establish the values of the predictive models for cirrhotic outcomes, and Cox proportional hazards regression models were used to identify predictors of clinical outcomes. RESULTS: Sixty (40.3%) patients experienced cirrhosis-associated clinical events and had higher M2BPGi levels compared to those without events (8.7 vs. 5.1 cutoff index, p < 0.001). The most common cirrhosis-associated complications were bacterial infections (24.2%). On ROC analysis, M2BPGi to albumin ratio (M2BPGi/albumin) had comparable discriminant abilities for all cirrhosis-associated events (area under the ROC curve [AUC] = 0.74) compared with M2BPGi, Child-Pugh, model for end-stage liver disease, albumin-bilirubin scores, and neutrophil-to-lymphocyte ratio and was superior to M2BPGi alone for all bacterial infectious events (AUC = 0.80). Cox regression analysis revealed that the M2BPGi/albumin, but not M2BPGi alone, independently predicted all cirrhosis-associated events (hazard ratio [HR] = 1.34, p = 0.038) and all bacterial infectious events (HR = 1.51, p = 0.011) within 1 year. However, M2BPGi/albumin did not predict other cirrhotic complications and transplant-free survival. DISCUSSION/CONCLUSION: M2BPGi/albumin might serve as a potential prognostic indicator for patients with cirrhosis, particularly for predicting bacterial infections.

Metabolomic Profiling of Different Antrodia cinnamomea Phenotypes.

Antrodia cinnamomea (AC) is a precious medicinal fungus with numerous therapeutic benefits. Based on the color appearance of its fruiting bodies, AC can be divided into red AC (RAC), yellow AC (YAC), and white AC (WAC); however, the differences in their metabolomic profiles remain unknown. This study aimed to analyze the metabolomic profiles of three different AC phenotypes and examine their relationship to the color appearance of fruiting bodies. The results showed that although RAC, YAC, and WAC appear to have a relatively similar profile of index triterpenoids, their total triterpenoid contents were significantly different. Among the annotated triterpenoids, many of them were highly present in RAC but not in YAC and WAC, and the relative contents of the four ergostanes (antcamphin F, antcamphin L, antcin B, and antcin K) and one lanostane (versisponic acid D) were found to be significantly different among AC phenotypes. The metabolomic profiles of the AC fruiting bodies demonstrated a total of 140 metabolites, and 41 of them were very different among AC phenotypes. This study indicates that red, yellow, and white AC can biosynthesize the diverse structures of triterpenoids, and RAC possesses a relatively higher contents of triterpenoids and diverse unannotated metabolites than YAC and WAC.

Myosteatosis, but not Sarcopenia, Predisposes NAFLD Subjects to Early Steatohepatitis and Fibrosis Progression.

BACKGROUND & AIMS: Sarcopenia and myosteatosis are associated with advanced nonalcoholic fatty liver disease (NAFLD). However, muscle alterations in early stage NAFLD remain unclear. METHODS: Patients with nonalcoholic fatty liver (NAFL) or early nonalcoholic steatohepatitis (NASH) without significant fibrosis were selected from a prospective biopsy-proven NAFLD cohort (N = 338). The skeletal muscle index and mean muscle attenuation (MA) were measured using abdominal fat computed tomography at the third lumbar vertebra level. Severe myosteatosis was defined as the lowest quartile of sex-stratified MA values. RESULTS: Patients with early NASH (n = 87) had lower MA (45.61 ± 6.45 vs 47.48 ± 5.85 HU; P = .028) than patients with NAFL (n = 251) but a similar skeletal muscle index. Patients with more severe lobular inflammation and hepatocellular ballooning had lower MA (P = .003 and P = .041, respectively). The severe myosteatosis prevalence was higher in early NASH than in NAFL (33.3% vs 21.1%; P = .029). Patients with severe myosteatosis were more likely to have early NASH in multivariable analysis adjusted for age, sex, and metabolic factors (odds ratio, 2.45; 95% confidence interval (CI), 1.24-4.86), which was maintained after adjustment for visceral fat amount (odds ratio, 2.44; 95% CI, 1.22-4.89). During a median 29-month follow-up, 170 patients underwent repeated transient elastography. Fibrosis progression-an increase in liver stiffness measurement >2 kPa or second liver stiffness measurement ≥7 kPa-was found in 28 and 31 individuals. Severe myosteatosis was significantly associated with fibrosis progression after adjustment for various confounders (hazard ratio, 2.49; 95% CI, 1.15-5.40 and hazard ratio, 2.09; 95% CI, 1.01-4.34 for different fibrosis progression definitions). CONCLUSIONS: Severe myosteatosis is significantly associated with early NASH and fibrosis progression in early stage NAFLD.

(Pro)renin receptor inhibition attenuated liver steatosis, inflammation, and fibrosis in mice with steatohepatitis.

The (Pro)renin receptor (PRR) is reportedly involved in hepatic lipid metabolism and hepatocyte PRR knockdown protects mice against hepatosteatosis. However, the impact of PRR inhibition on liver inflammation and fibrosis in nonalcoholic steatohepatitis (NASH) remains unclear. Herein, C57BL/6 mice were fed a normal chow diet or fast food diet (FFD) for 24 weeks. Lentivirus-mediated PRR short hairpin RNA (shRNA) or handle region peptide (HRP), a PRR blocker, was administered for PRR inhibition. Mouse primary hepatocytes were cultured with palmitic acid, prorenin, siRNA-targeted PRR, and HRP. In FFD-fed mice, PRR inhibition via lentivirus-mediated PRR knockdown or HRP significantly attenuated liver steatosis, inflammation, and fibrosis. Mechanistically, PRR knockdown or HRP decreased hepatic acetyl-CoA carboxylase (ACC) abundance and upregulated peroxisome proliferator-activated receptor-alpha (PPARα). HRP treatment also decreased hepatic PRR expression. In addition, intrahepatic oxidative stress, apoptosis and inflammatory cell recruitment were ameliorated by PRR knockdown or HRP treatment, along with suppression of proinflammatory cytokine expression. PRR inhibition downregulated the hepatic expression of profibrotic factors, as well as TGF-ß1/SMAD3 pathway. In primary mouse hepatocytes, PRR knockdown with siRNA or HRP downregulated cellular ACC and increased PPARα expression. In conclusion, our findings revealed that PRR inhibition attenuated hepatic steatosis, inflammation, and fibrosis in mice with NASH. Accordingly, targeting PRR signaling may serve as a potential treatment for NASH.

Microsatellite Instability, Epstein-Barr Virus, and Programmed Cell Death Ligand 1 as Predictive Markers for Immunotherapy in Gastric Cancer.

Immunotherapy benefits selected cases of gastric cancer (GC), but the correlation between biomarkers and prognosis is still unclear. Fifty-two patients with GC who underwent immunotherapy were enrolled from June 2016 to December 2020. Their clinical features and biomarkers-microsatellite instability-high (MSI-H), programmed cell death ligand 1 (PD-L1) combined positive score (CPS), and Epstein-Barr encoding region (EBER)-were analyzed. Eight patients had MSI-H, five patients had EBER, 29 patients had CPS ≥ 1, and 20 patients had no biomarker. The overall response rates (ORRs) of the MSI-H, EBER, PD-L1 CPS ≥ 1, and all-negative group were 75%, 60%, 44.8%, and 15%, respectively. Compared with that of the all-negative group, progression-free survival (PFS) was better in the MSI-H (p = 0.018), CPS ≥ 5 (p = 0.012), and CPS ≥ 10 (p = 0.006) groups, but not in the EBER (p = 0.2) and CPS ≥ 1 groups (p = 0.35). Ten patients had combined biomarkers, CPS ≥ 1 with either MSI-H or EBER. The ORRs were 66.7% for CPS ≥ 1 and MSI-H and 75% for CPS ≥ 1 and EBER. PFS was better in patients with combined biomarkers (p = 0.01). MSI-H, EBER, and CPS are useful biomarkers for predicting the efficacy of immunotherapy.

Mac-2 binding protein glycosylation isomer is a potential biomarker to predict portal hypertension and bacterial infection in cirrhotic patients.

OBJECTIVES: Mac-2-binding protein glycosylation isomer (M2BPGi) is a novel plasma biomarker for liver fibrosis, but less is known about its role in portal hypertension. We aimed to evaluate the association between M2BPGi and hepatic venous pressure gradient (HVPG) and to investigate its predictive value on prognosis of cirrhotic patients. METHODS: Forty-eight cirrhotic patients who underwent HVPG measurement in Taipei Veterans General hospital were retrospectively enrolled. The Spearman's correlation test was used to analyze the correlation between plasma M2BPGi levels and HVPG and other parameters. Cox proportional hazards regression models were used to identify predictors for clinical outcomes. RESULTS: Plasma M2BPGi levels were higher in cirrhotic patients than healthy subjects and significantly correlated with HVPG levels (rs = 0.45, p = 0.001). On multivariate Cox regression analysis, higher plasma M2BPGi levels [≥ 6 cut-off index (C.O.I)] did not predict mortality within five years for cirrhotic patients and the result was similar in patients without hepatocellular carcinoma. Interestingly, M2BPGi ≥ 6 C.O.I was a potential predictor of bacterial infection within five years [Hazar ratio (HR) = 4.51, p = 0.003]. However, M2BPGi failed to predict occurrence of other cirrhosis-related complications, including variceal bleeding, ascites formation, spontaneous bacterial peritonitis, hepatorenal syndrome and hepatic encephalopathy. CONCLUSION: Plasma M2BPGi levels positively correlated with HVPG and higher serum M2BPGi levels might have a potential role in predicting development of bacterial infection for cirrhotic patients with portal hypertension.

Eritoran Attenuates Hepatic Inflammation and Fibrosis in Mice with Chronic Liver Injury.

Toll-like receptor 4 (TLR4) signaling plays a key role in liver inflammation and fibrosis. The therapeutic effects of eritoran, a TLR4 antagonist, in mice with chronic liver injury remained unclear. C57BL/6 mice were fed a fast-food diet (FFD) or treated with carbon tetrachloride (CCl4) to induce chronic liver injury. Eritoran (10 mg/kg) or a vehicle was randomly intraperitoneally administered to the FFD-fed mice and the CCl4-injured mice. Primary mouse liver cells were cultured with lipopolysaccharide (LPS) or eritoran. In both FFD and CCl4 mouse models, eritoran significantly reduced serum ALT levels and decreased hepatic inflammatory cell infiltration without altering hepatic steatosis. Additionally, eritoran attenuated liver fibrosis by decreasing hepatic stellate cells (HSCs) activation and the abundance of α-smooth muscle actin and transforming growth factor-ß1. Hepatic TLR4 downstream signaling including MyD88 expression, NF-κB p65 nuclear translocation, p38 and JNK phosphorylation were successfully inhibited by eritoran. In the in vitro study, LPS-induced nuclear translocation of NF-κB in primary HSCs and Kupffer cells was significantly suppressed by eritoran. In conclusion, eritoran attenuated hepatic inflammation and fibrosis by inhibition of the TLR4 signaling pathway in mice with chronic liver injury. Eritoran may serve as a potential drug for chronic liver disease.

(Pro)renin Receptor Knockdown Attenuates Liver Fibrosis Through Inactivation of ERK/TGF-ß1/SMAD3 Pathway.

BACKGROUND & AIMS: Activation of the (pro)renin receptor (PRR) up-regulates the expression of profibrotic genes in the kidney and heart. We aimed to investigate the role of PRR in hepatic fibrogenesis. METHODS: Human hepatic PRR levels were measured in patients with or without liver fibrosis. PRR expression was analyzed in primary mouse hepatic stellate cells (HSCs). Experimental fibrosis was studied in thioacetamide (TAA)-treated or methionine choline-deficient (MCD) diet-fed C57BL/6 mice. Lentivirus-mediated PRR short hairpin RNA was used to knockdown hepatic PRR expression. Lentiviral vectors expressing PRR short hairpin RNA or complementary DNA from the α-smooth muscle actin promoter were used for myofibroblast-specific gene knockdown or overexpression. RESULTS: PRR is up-regulated in human and mouse fibrotic livers, and in activated HSCs. Hepatic PRR knockdown reduced liver fibrosis by suppressing the activation of HSCs and expression of profibrotic genes in TAA or MCD diet-injured mice without significant changes in hepatic inflammation. Renin and prorenin increased the expression of PRR and production of TGF-ß1 in human activated HSC Lieming Xu-2 cells, and knockdown of PRR inactivated Lieming Xu-2 cells with decreased production of transforming growth factor (TGF)-ß1 and Mothers against decapentaplegic homolog 3 (Smad3) phosphorylation. Myofibroblast-specific PRR knockdown also attenuated liver fibrosis in TAA or MCD diet-injured mice. Mice with both myofibroblast-specific and whole-liver PRR knockdown showed down-regulation of the hepatic extracellular signal-regulated kinase (ERK)/TGF-ß1/Smad3 pathway. Myofibroblast-specific PRR overexpression worsened TAA-induced liver fibrosis by up-regulating the ERK/TGF-ß1/Smad3 pathway. CONCLUSIONS: PRR contributes to liver fibrosis and HSC activation, and its down-regulation attenuates liver fibrosis through inactivation of the ERK/TGF-ß1/Smad3 pathway. Therefore, PRR is a promising therapeutic target for liver fibrosis.

Persistent liver inflammation in chronic hepatitis C patients with advanced fibrosis after direct-acting antivirals induced sustained virological response.

BACKGROUND: Direct-acting antivirals (DAA) improve sustained virological response (SVR) rates with normalization of liver enzymes in patients with hepatitis C. However, liver inflammation may persist despite virus eradication. We aimed to investigate the rate and risk factors for persistent elevated aminotransferase levels in patients with advanced fibrosis after DAA-induced SVR. METHODS: From January 2017 to April 2018, chronic hepatitis C patients with advanced fibrosis and SVR after DAA treatment at the Taipei Veterans General Hospital were prospectively enrolled. Persistent liver inflammation after SVR was defined as an increase in levels of alanine aminotransferase (ALT) (>40 U/L) at SVR12. RESULTS: A total of 461 patients were included (57.9% females, mean age 64 years, 69.6% genotype 1b, 46.4% cirrhosis). At SVR12, there was a decline in ALT levels (90.5 ± 80.8 U/L to 25.3 ± 26.5 U/L) from baseline levels. Persistent liver inflammation at SVR12 was detected in 45 patients (9.8%). The presence of cirrhosis, markers of impaired liver functions, history of interferon-based therapy, steatosis, and elevated ALT levels at baseline was associated with persistent liver inflammation after SVR12. Results of multivariate analysis indicated that levels of baseline serum total bilirubin (odds ratio [OR]: 2.605, 95% CI: 1.158-5.858), international normalized ratio (OR: 14.389, 95% CI: 1.754-118.049), ALT (OR: 1.006, 95% CI: 1.003-1.009), and the presence of steatosis (OR: 3.635, 95% CI: 1.716-7.698) were independent predictors of persistent liver inflammation at SVR12. CONCLUSION: Persistent liver inflammation is not uncommon in chronic hepatitis C patients with advanced fibrosis after DAA-induced SVR. It is associated with impaired baseline liver function and steatosis. Long-term follow-up is required to assess the implication of liver inflammation on disease progression.

Muscle alterations are independently associated with significant fibrosis in patients with nonalcoholic fatty liver disease.

BACKGROUND & AIM: Anthropometric data are associated with nonalcoholic fatty liver disease (NAFLD) development and progression. We investigated whether the quantity and quality of muscle and visceral fat assessed by computed tomography (CT) are associated with fibrosis severity in NAFLD. METHODS: In a prospective biopsy-confirmed NAFLD cohort of 521 patients, we measured skeletal muscle index (SMI), muscle attenuation (MA) and visceral adipose tissue index (VATI) via CT. Low skeletal muscle mass (LSMM) was defined using previously validated cut-offs. Myosteatosis and visceral adiposity were defined as the lowest and highest quartile, respectively. Significant fibrosis was defined as F2-F4 in liver histology. RESULTS: Patients with significant fibrosis had lower SMI and MA and higher VATI than those without. The significant fibrosis prevalence was significantly higher in subjects with LSMM (45.1% vs 30.8%, P = .005), myosteatosis (46.1% vs 29.7%, P = .001) and visceral adiposity (46.9% vs 29.9%, P = .001) than those without. The significant fibrosis risk increased with increasing numbers of body composition components (24.5%, 35.6%, 53.0% and 72.7% in patients with 0, 1, 2 and 3 components respectively). Multivariable analysis revealed that LSMM (OR, 1.72; 95% CI, 1.05-2.84), myosteatosis (OR, 1.65; 95% CI, 1.01-2.68) and visceral adiposity (OR, 1.75; 95% CI, 1.09-2.83) were independent predictors of significant fibrosis. Subjects with sarcopenia had a higher risk of significant fibrosis (OR, 2.17; 95% CI, 1.03-4.56). CONCLUSION: Muscle alterations and visceral adiposity assessed by CT are associated with significant fibrosis in NAFLD. LSMM and myosteatosis have additive values in prediction of significant fibrosis.

Early echocardiographic signs of diastolic dysfunction predict acute kidney injury in cirrhotic patients.

BACKGROUND: Cardiovascular dysfunction in cirrhotic patients affects survival and the development of cirrhotic complications. We aimed to evaluate potential echocardiographic parameters to predict mortality and acute kidney injury (AKI) in cirrhotic patients. METHODS: A total of 103 cirrhotic patients who underwent echocardiography between February 2009 and August 2016 in Taipei Veterans General Hospital were retrospectively enrolled. Cardiac function was evaluated using transthoracic two-dimensional echocardiography with tissue Doppler imaging. Cox hazard regression analysis was used for assessing predictors for 1-year mortality and AKI within 1 year. RESULTS: Baseline echocardiographic parameters were similar between survivors (n = 92) and nonsurvivors (n = 11). Lower serum levels of albumin, as well as higher albumin-bilirubin (ALBI) scores, Child-Pugh scores, and model for end-stage liver disease scores were observed in nonsurvivors. Cox proportional hazard regression analysis revealed Child-Pugh score as the only predictor of 1-year mortality. Baseline serum creatinine (Cr) > 1.5 mg/dL, total bilirubin > 2 mg/dL, and a higher E/e' ratio predict occurrence of AKI within 1 year. Among patients with serum Cr < 1.5 mg/dL, an increased atrial filling velocity and higher ALBI scores predict AKI occurrence within 1 year. CONCLUSION: Severity of underlying liver disease but not echocardiographic parameters predicts 1-year mortality in cirrhosis. Early echocardiographic signs of diastolic dysfunction and higher ALBI scores may predict development of AKI in cirrhotic patients with serum Cr < 1.5 mg/dL.

Obeticholic acid ameliorates hepatorenal syndrome in ascitic cirrhotic rats by down-regulating the renal 8-iso-PGF2α-activated COX-TXA2 pathway.

BACKGROUNDS/AIMS: The present study explores the potential of chronic treatment with the Foresaid X receptor (FXR) agonist obeticholic acid (OCA), which inhibits oxidative stress-related pathogenesis, in ascitic cirrhotic rats with hepatorenal syndrome (HRS) developed 6 weeks after bile duct ligation (BDL). METHODS: Systemic, splanchnic, and renal hemodynamics and pathogenic cascades were measured in ascitic BDL and sham rats receiving 2-weeks of either vehicle or OCA treatments (sham-OCA and BDL-OCA groups), and NRK-52E cells, rat kidney tubular epithelial cells. RESULTS: Chronic OCA treatment significantly normalized portal hypertension, glomerular filtration rate, urine output, renal blood flow; decreased ascites, renal vascular resistance, serum creatinine, and the release of renal tubular damage markers, including urinary neutrophil gelatinase-associated lipocalin (uNGAL) and kidney injury moleculae-1 (uKim-1) in BDL-OCA rats. In the BDL group, inhibition of the renal oxidative stress (8-iso-PGF2α)-activated cyclooxygenase-thromboxane A2 [COX-TXA2] pathway, apoptosis, and tubular injury accompanied by a decrease in hyper-responsiveness to the vasoconstrictor 8-iso-PGF2α in perfused kidneys. In vitro experiments revealed that 8-iso-PGF2α induced oxidative stress, release of reactive oxygen species, and cell apoptosis, which were reversed by concomitant incubation with the FXR agonist. CONCLUSIONS: Through the inhibition of renal 8-iso-PGF2α production and the down-regulation of the COX-TXA2 pathway, our study suggests that chronic OCA treatment can ameliorate the HRS in ascitic cirrhotic rats. Thus, OCA is an agent with antioxidative stress, antivasoconstrictive, antiapoptotic properties which benefit ascitic, cirrhotic rats with systemic, hepatic, and renal abnormalities.

SIRT1-dependent mechanisms and effects of resveratrol for amelioration of muscle wasting in NASH mice.

BACKGROUND: In non-alcoholic steatohepatitis (NASH), muscle wasting was an aggravating factor for the progression of hepatic steatosis. This study explores the potential benefits of chronic treatment with resveratrol, a strong activator of SIRT1 on the muscle wasting of NASH mice. METHODS: In vivo and in vitro study, we evaluate the SIRT1-dependent mechanisms and effects of resveratrol administration for 6 weeks with high-fat-methionine and choline deficient diet-induced NASH mice and palmitate-pretreated C2C12 myoblast cells. RESULTS: Resveratrol treatment improved grip strength and muscle mass of limbs, increased running distance and time on exercise wheels in NASH mice. There is a negative correlation between muscular SIRT1 activity and 3-nitrotyrosine levels of NASH and NASH-resv mice. The SIRT1-dependent effect of muscle wasting was associated with the suppression of oxidative stress, upregulation of antioxidants, inhibition of protein degradation, activation of autophagy, suppression of apoptotic activity, upregulation of lipolytic genes and the reduction of fatty infiltration in limb muscles of NASH mice. In vitro, resveratrol alleviated palmitate acid-induced oxidative stress, lipid deposition, autophagy dysfunction, apoptotic signals, and subsequently reduced fusion index and myotube formation of C2C12 cells. The beneficial effects of resveratrol were abolished by EX527. CONCLUSIONS: Our study suggests that chronic resveratrol treatment is a potential strategy for amelioration of hepatic steatosis and muscle wasting in NASH mouse model.

Propranolol Is Associated with Lower Risk of Incidence of Hepatocellular Carcinoma in Patients with Alcoholic Cirrhosis: A Tertiary-Center Study and Indirect Comparison with Meta-Analysis.

Alcoholic cirrhosis (AC) leads to enormous disease burden and occupies a substantial proportion in the etiology of hepatocellular carcinoma (HCC), but scarce attention has been paid to this topic. Besides, propranolol has been reported to decrease the rate of HCC in viral hepatitis. We conducted a retrospective tertiary-center cohort study to identify the HCC incidence in AC patients with or without propranolol. A total of 1,046 AC patients with hospitalization had been screened, and those with regular follow-up for three years or otherwise until the date of malignancy diagnosis without meeting exclusion criteria were enrolled; finally, 23 AC patients with propranolol and 46 AC patients without propranolol were analyzed after twofold propensity-score matching. The cumulative incidence of HCC was lower in the propranolol group (log-rank test, P = 0.046). Furthermore, we undertook the meta-analysis of annual incidence of HCC in AC patients, and 1,949 publications were screened, within which eight studies were analyzed; the pooled annual incidence was 2.41%, which was higher than the calculated annual incidence of HCC in our AC cohort with propranolol (1.45%). In conclusion, propranolol is associated with decreased risk of HCC incidence in patients with AC.

Elafibranor Inhibits Chronic Kidney Disease Progression in NASH Mice.

Identification of new pharmacological approaches to inhibit the excessive fat intake-induced steatohepatitis and chronic kidney disease (CKD) is important. High-fat diet (HFD)-induced steatohepatitis and CKD share common pathogenesis involving peroxisome proliferator-activated receptor (PPAR)-α and -δ. Elafibranor, a dual PPARα/δ agonist, can ameliorate the HFD-induced steatohepatitis. Nonetheless, the effects of HFD-induced CKD had not yet explored. This study investigated the effects of elafibranor (elaf) on the progression of HFD-induced CKD in mice. In vivo and in vitro renal effects were evaluated in HFD-elaf mice receiving 12 weeks of elafibranor (from 13th to 24th week of HFD feeding) treatment. In elafibranor-treated HFD mice, increased insulin sensitivity, reduced obesity and body fat mass, decreased severity of steatohepatitis, increased renal expression of PPARα, PPARδ, SIRT1, and autophagy (Beclin-1 and LC3-II) as well as glomerular/renal tubular barrier markers [synaptopodin (podocyte marker), zona occludin-1, and cubulin], reduced renal oxidative stress and caspase-3, and less urinary 8-isoprostanes excretion were observed. Aforementioned benefits of elafibranor were associated with low renal tubular injury and tubulointerstitial fibrosis scores, less albuminuria, low urinary albumin-to-creatinine ratio, and preserved glomerular filtration rate. Acute incubation of podocytes and HK-2 cells with elafibranor or recombinant SIRT1 reversed the HFD-sera-induced oxidative stress, autophagy dysfunction, cell apoptosis, barrier marker loss, albumin endocytosis, and reuptake reduction. Besides hepatoprotective and metabolic beneficial effects, current study showed that elafibranor inhibited the progression of HFD-induced CKD through activation of renal PPARα, PPARδ, SIRT1, autophagy, reduction of oxidative stress, and apoptosis in mice with steatohepatitis.

Human relaxin-2 attenuates hepatic steatosis and fibrosis in mice with non-alcoholic fatty liver disease.

Human relaxin-2 reduces hepatic fibrosis in mice. However, the effects of relaxin-2 on hepatic steatosis and fibrosis in animals with non-alcoholic fatty liver disease (NAFLD) remain to be elucidated. C57BL/6 mice fed a high-fat diet (HFD) or methionine-choline-deficient (MCD) diet were randomly assigned to receive recombinant human relaxin-2 (25 or 75 µg/kg/day) or vehicle for 4 weeks. In HFD-fed mice, relaxin-2 decreased systemic insulin resistance and reduced body weight, epididymal fat mass and serum leptin and insulin concentrations. In livers of HFD-fed mice, relaxin-2 attenuated steatosis and increased phosphorylation of insulin receptor substrate-1, Akt and endothelial nitric oxide synthase (eNOS), and activated genes that regulate fatty acid oxidation and suppressed acetyl-CoA carboxylase. Relaxin-2 had no direct anti-steatotic effect on primary mouse hepatocytes, but S-nitroso-N-acetylpenicillamine attenuated palmitic acid-induced steatosis and activated genes regulating fatty acid oxidation in hepatocytes. In mice fed an MCD diet, relaxin-2 attenuated steatosis, inflammation and fibrosis. Relaxin-2 increased eNOS and Akt phosphorylation and transcript levels of cytochrome P450-4a10 and decreased acetyl-CoA carboxylase in MCD-fed mouse livers. Moreover, expression levels of Kupffer cell activation, hepatic stellate cell activation and hepatocyte apoptosis were decreased in MCD diet-fed mice receiving relaxin-2. In conclusion, relaxin-2 reduces hepatic steatosis by activating intrahepatic eNOS in HFD-fed mice and further attenuates liver fibrosis in MCD diet-fed mice. Therefore, human relaxin-2 is a potential therapeutic treatment for NAFLD.

Dabigatran Reduces Liver Fibrosis in Thioacetamide-Injured Rats.

BACKGROUND: Liver fibrosis can progress to cirrhosis, hepatocellular carcinoma, or liver failure. Unfortunately, the antifibrotic agents are limited. Thrombin activates hepatic stellate cells (HSCs). Therefore, we investigated the effects of a direct thrombin inhibitor, dabigatran, on liver fibrosis. METHODS: Adult male Sprague-Dawley rats were injected intraperitoneally with thioacetamide (TAA, 200 mg/kg twice per week) for 8 or 12 weeks to induce liver fibrosis. The injured rats were assigned an oral gavage of dabigatran etexilate (30 mg/kg/day) or vehicle in the last 4 weeks of TAA administration. Rats receiving an injection of normal saline and subsequent oral gavage of dabigatran etexilate or vehicle served as controls. RESULTS: In the 8-week TAA-injured rats, dabigatran ameliorated fibrosis, fibrin deposition, and phosphorylated ERK1/2 in liver, without altering the transcript expression of thrombin receptor protease-activated receptor-1. In vitro, dabigatran inhibited thrombin-induced HSC activation. Furthermore, dabigatran reduced intrahepatic angiogenesis and portal hypertension in TAA-injured rats. Similarly, in the 12-week TAA-injured rats, a 4-week treatment with dabigatran reduced liver fibrosis and portal hypertension. CONCLUSIONS: By inhibiting thrombin action, dabigatran reduced liver fibrosis and intrahepatic angiogenesis. Dabigatran may be a promising therapeutic agent for treatment of liver fibrosis.

Correlation and prognostic accuracy between noninvasive liver fibrosismarkers and portal pressure in cirrhosis: Role of ALBI score.

BACKGROUND: The role of noninvasive liver fibrosis markers which were developed to evaluate the severity of chronic liver disease remains unclear in cirrhosis. AIMS: To evaluate the correlation between noninvasive markers and hemodynamic parameters and their prognostic performance in cirrhotic patients. METHODS: A total of 242 cirrhotic patients undergoing hemodynamic study were analyzed. The correlations between noninvasive models, including FIB-4, aspartate aminotransferase to platelet ratio index, cirrhosis discriminant score, Lok index, Goteborg University Cirrhosis Index, and albumin-bilirubin (ALBI) score and hemodynamic parameters were investigated, along with their predictive accuracy for short- and long-term survival. RESULTS: There was a significant correlation between all noninvasive markers and hepatic venous pressure gradient (HVPG), and ALBI score had the best correlation (r = 0.307, p<0.001). For the prediction of 3-month and 6-month mortality, serum sodium (sNa) levels had the highest area under curve (AUC; 0.799 and 0.818, respectively) among all parameters, and ALBI score showed the best performance (AUC = 0.691 and 0.740, respectively) compared with other 5 noninvasive models. Of 159 patients with low MELD scores (<14), high ALBI score (>-1.4) and low sNa (<135 mmol/L) predicted early mortality. In the Cox multivariate model, ALBI, MELD, HVPG and sNa were independent predictors of long-term survival. CONCLUSIONS: Among noninvasive markers, ALBI score is best correlated with HVPG and associated with short-term outcome in cirrhotic patients. A high ALBI score and low sNa identify high-risk patients with low MELD scores. High MELD, HVPG, ALBI and low sNa levels are independent predictors of survival. Independent studies are required to confirm our findings.