Nonalcoholic Fatty Liver Disease Activity Research Articles

The significance of 8-hydroxy-deoxyguanosine acid in the diagnosis of nonalcoholic steatohepatitis

Objective: To evaluate the significance of serum 8-hydroxy-deoxyguanosine acid(8-OHdG) in the diagnosis of nonalcoholic steatohepatitis (NASH). Methods: Patients or healthy subjects were enrolled at the Second Hospital of Tianjin Medical University and the Second People's Hospital of Tianjin from May 2013 to December 2015. A total of 41 patients with nonalcoholic fatty liver disease were enrolled in the study, including 20 nonalcoholic simple fatty liver (NAFL) patients and 21 NASH patients whose diagnosis were proven by liver biopsy. The other 32 healthy subjects were studied as controls. Serum 8-OHdG, ALT, AST and GGT were tested. Nonalcoholic fatty liver disease activity score (NAS) and expression of 8-OHdG in liver was investigated between NAFL patients and NASH patients. The correlations between serum 8-OHdG and serum ALT, AST, GGT, and 8-OHdG in liver tissue in NASH group were investigated. In addition, the receiver operating characteristic (ROC) curve analyses for ALT and 8-OHdG levels were performed in NAFL patients and NASH patients, and the cut-off value was determined. Results: Serum 8-OHdG values in healthy controls, NAFL and NASH patients were (0.19±0.16) μg/L, (0.22±0.16) μg/L, (0.42±0.21) μg/L respectively. The serum 8-OHdG and serum ALT, GGT and 8-OHdG in liver tissue were all positively correlated in NASH group with respective correlation coefficient r values as 0.454 7, 0.382 9, and 0.497 6. AUC of 8-OHdG was 0.901 with cut-off value 0.39 μg/L. Its sensitivity was 88.3% and specificity was 81.5%, which were higher than those of ALT. Conclusion: The value of serum 8-OHdG would be used as a marker for the diagnosis of NASH.

PS-024 - YH25724, a novel long-acting GLP-1/FGF21 dual agonist lowers both non-alcoholic fatty liver disease activity score and fibrosis stage in a diet-induced obese mouse model of biopsy-confirmed non-alcoholic steatohepatitis

PS-024 - YH25724, a novel long-acting GLP-1/FGF21 dual agonist lowers both non-alcoholic fatty liver disease activity score and fibrosis stage in a diet-induced obese mouse model of biopsy-confirmed non-alcoholic steatohepatitis

Anti-steatotic and anti-fibrotic effects of the KCa3.1 channel inhibitor, Senicapoc, in non-alcoholic liver disease.

AIMTo evaluate a calcium activated potassium channel (KCa3.1) inhibitor attenuates liver disease in models of non-alcoholic fatty liver disease (NAFLD).METHODSWe have performed a series of in vitro and in vivo studies using the KCa3.1 channel inhibitor, Senicapoc. Efficacy studies of Senicapoc were conducted in toxin-, thioacetamide (TAA) and high fat diet (HFD)-induced models of liver fibrosis in rats. Efficacy and pharmacodynamic effects of Senicapoc was determined through biomarkers of apoptosis, inflammation, steatosis and fibrosis.RESULTSUpregulation of KCa3.1 expression was recorded in TAA-induced and high fat diet-induced liver disease. Treatment with Senicapoc decreased palmitic acid-driven HepG2 cell death. (P < 0.05 vs control) supporting the finding that Senicapoc reduces lipid-driven apoptosis in HepG2 cell cultures. In animals fed a HFD for 6 wk, co-treatment with Senicapoc, (1) reduced non-alcoholic fatty liver disease (NAFLD) activity score (NAS) (0-8 scale), (2) decreased steatosis and (3) decreased hepatic lipid content (Oil Red O, P < 0.05 vs vehicle). Randomization of TAA animals and HFD fed animals to Senicapoc was associated with a decrease in liver fibrosis as evidenced by hydroxyproline and Masson’s trichrome staining (P < 0.05 vs vehicle). These results demonstrated that Senicapoc mitigates both steatosis and fibrosis in liver fibrosis models.CONCLUSIONThese data suggest that Senicapoc interrupts more than one node in progressive fatty liver disease by its anti-steatotic and anti-fibrotic activities, serving as a double-edged therapeutic sword.

FRI-363 - EDP-305, a novel and highly potent farnesoid X receptor agonist, improves liver steatosis, ballooning and non-alcoholic fatty liver disease (NAFLD) activity score (NAS) in a diet-induced murine model of non-alcoholic steatohepatitis

FRI-363 - EDP-305, a novel and highly potent farnesoid X receptor agonist, improves liver steatosis, ballooning and non-alcoholic fatty liver disease (NAFLD) activity score (NAS) in a diet-induced murine model of non-alcoholic steatohepatitis

Combination curcumin and vitamin E treatment attenuates diet-induced steatosis in Hfe-/- mice.

AIMTo investigate the synergistic hepato-protective properties of curcumin and vitamin E in an Hfe-/- high calorie diet model of steatohepatitis.METHODSHfe-/- C57BL/6J mice were fed either a high calorie diet or a high calorie diet with 1 mg/g curcumin; 1.5 mg/g vitamin E; or combination of 1 mg/g curcumin + 1.5 mg/g vitamin E for 20 wk. Serum and liver tissue were collected at the completion of the experiment. Liver histology was graded by a pathologist for steatosis, inflammation and fibrosis. RNA and protein was extracted from liver tissue to examine gene and protein expression associated with fatty acid oxidation, mitochondrial biogenesis and oxidative stress pathways.RESULTSHfe-/- mice fed the high calorie diet developed steatohepatitis and pericentral fibrosis. Combination treatment with curcumin and vitamin E resulted in a greater reduction of percent steatosis than either vitamin E or curcumin therapy alone. Serum alanine aminotransferase and non-alcoholic fatty liver disease (NAFLD) activity score were decreased following combination therapy with curcumin and vitamin E compared with high calorie diet alone. No changes were observed in inflammatory or fibrosis markers following treatment. Epididymal fat pad weights were significantly reduced following combination therapy, however total body weight and liver weight were unchanged. Combination therapy increased the mRNA expression of AdipoR2, Ppar-α, Cpt1a, Nrf-1 and Tfb2m suggesting enhanced fatty acid oxidation and mitochondrial biogenesis. In addition, combination treatment resulted in increased catalase activity in Hfe-/- mice.CONCLUSIONCombination curcumin and vitamin E treatment decreases liver injury in this steatohepatitis model, indicating that combination therapy may be of value in NAFLD.

Hepatic fat quantification using the two-point Dixon method and fat color maps based on non-alcoholic fatty liver disease activity score.

The two-point Dixon method for magnetic resonance imaging (MRI) is commonly used to non-invasively measure fat deposition in the liver. The aim of the present study was to assess the usefulness of MRI-fat fraction (MRI-FF) using the two-point Dixon method based on the non-alcoholic fatty liver disease activity score. This retrospective study included 106 patients who underwent liver MRI and MR spectroscopy, and 201 patients who underwent liver MRI and histological assessment. The relationship between MRI-FF and MR spectroscopy-fat fraction was used to estimate the corrected MRI-FF for hepatic multi-peaks of fat. Then, a color FF map was generated with the corrected MRI-FF based on the non-alcoholic fatty liver disease activity score. We defined FF variability as the standard deviation of FF in regions of interest. Uniformity of hepatic fat was visually graded on a three-point scale using both gray-scale and color FF maps. Confounding effects of histology (iron, inflammation and fibrosis) on corrected MRI-FF were assessed by multiple linear regression. The linear correlations between MRI-FF and MR spectroscopy-fat fraction, and between corrected MRI-FF and histological steatosis were strong (R2 = 0.90 and R2 = 0.88, respectively). Liver fat variability significantly increased with visual fat uniformity grade using both of the maps (ρ = 0.67-0.69, both P <0.001). Hepatic iron, inflammation and fibrosis had no significant confounding effects on the corrected MRI-FF (all P >0.05). The two-point Dixon method and the gray-scale or color FF maps based on the non-alcoholic fatty liver disease activity score were useful for fat quantification in the liver of patients without severe iron deposition.

Empagliflozin (an SGLT2 inhibitor), alone or in combination with linagliptin (a DPP-4 inhibitor), prevents steatohepatitis in a novel mouse model of non-alcoholic steatohepatitis and diabetes.

BackgroundSodium-glucose co-transporter-2 (SGLT2) inhibitors are new oral antidiabetic drugs that reduce hyperglycemia by promoting urinary glucose excretion. Glycosuria produced by SGLT2 inhibitors is associated with weight loss, mainly due to reduced fat volume. We investigated the effects of empagliflozin (selective SGLT2 inhibitor) and linagliptin (DPP-4 inhibitor) on steatohepatitis and fibrosis in a mouse model of non-alcoholic steatohepatitis (NASH) with diabetes.MethodsA novel NASH model was generated by administration of streptozotocin to C57BL/6J mice at 2 days old, with a high-fat diet from 4 weeks. NASH mice aged 6 weeks were divided into four groups of 6 animals: vehicle, linagliptin (10 mg/kg), empagliflozin (10 mg/kg), and linagliptin + empagliflozin. The histological non-alcoholic fatty liver disease activity score was significantly lower in the empagliflozin and linagliptin + empagliflozin groups than in the vehicle or linagliptin groups. Hepatic expression of inflammatory genes (tumor necrosis factor-α, interleukin-6, and monocyte chemoattractant protein-1) was decreased in the empagliflozin and linagliptin + empagliflozin groups compared with the vehicle group. The collagen deposition with Sirius red staining was significantly reduced in the linagliptin + empagliflozin group compared with the linagliptin or the empagliflozin group. Immunohistochemistry showed that expression of α-smooth muscle actin, a marker of myofibroblasts (fibrosis), was reduced in the linagliptin + empagliflozin group compared with the vehicle group, as was expression of type 1 and 3 collagen mRNA. Linagliptin + empagliflozin decreased expression of mRNAs for genes related to fatty acid synthesis, but did not increase mRNAs for β-oxidation-related genes.ConclusionsWhile empagliflozin alone attenuates development of NASH showing anti-steatotic and anti-inflammatory effects, combined administration of empagliflozin and linagliptin can synergistically ameliorates NASH with stronger anti-fibrotic effects.

Association of noninvasive quantitative decline in liver fat content on MRI with histologic response in nonalcoholic steatohepatitis.

Magnetic resonance imaging-estimated proton-density-fat-fraction (MRI-PDFF) has been shown to be a noninvasive, accurate and reproducible imaging-based biomarker for assessing steatosis and treatment response in nonalcoholic steatohepatitis (NASH) clinical trials. However, there are no data on the magnitude of MRI-PDFF reduction corresponding to histologic response in the setting of a NASH clinical trial. The aim of this study was to quantitatively compare the magnitude of MRI-PDFF reduction between histologic responders versus histologic nonresponders in NASH patients. This study is a secondary analysis of the MOZART trial, which included 50 patients with biopsy-proven NASH randomized to ezetimibe 10 mg/day orally or placebo for 24 weeks. The primary aim was to perform a head-to-head comparative analysis of histologic responders [defined as a ⩾2-point reduction in the nonalcoholic fatty liver disease (NAFLD) Activity Score (NAS) without worsening fibrosis] versus nonresponders, and the corresponding quantitative change in liver fat content measured via MRI-PDFF. Of the 35 patients who underwent paired liver biopsy and MRI-PDFF assessment at the beginning and end of treatment, 10 demonstrated a histologic response. Compared with histologic nonresponders, histologic responders had a statistically significant reduction in MRI-PDFF of -4.1% ± 4.9 versus -0.6 ± 4.1 (p < 0.04) with a mean relative percent change of -29.3% ± 33.0 versus +2.0% ± 24.0 (p < 0.004), respectively. Utilizing paired MRI-PDFF and liver histology data, we demonstrate that a relative reduction of 29% in liver fat on MRI-PDFF is associated with a histologic response in NASH. After external validation by independent research groups, these results can be incorporated into designing future NASH clinical trials, especially those utilizing change in hepatic fat quantified by MRI-PDFF, as a treatment endpoint.

Long-Term Pioglitazone Treatment for Patients With Nonalcoholic Steatohepatitis and Prediabetes or Type 2 Diabetes Mellitus

The metabolic defects of nonalcoholic steatohepatitis (NASH) and prediabetes or type 2 diabetes mellitus (T2DM) seem to be specifically targeted by pioglitazone. However, information about its long-term use in this population is limited. To determine the efficacy and safety of long-term pioglitazone treatment in patients with NASH and prediabetes or T2DM. Randomized, double-blind, placebo-controlled trial. (ClinicalTrials.gov: NCT00994682). University hospital. Patients (n= 101) with prediabetes or T2DM and biopsy-proven NASH were recruited from the general population and outpatient clinics. All patients were prescribed a hypocaloric diet (500-kcal/d deficit from weight-maintaining caloric intake) and then randomly assigned to pioglitazone, 45 mg/d, or placebo for 18 months, followed by an 18-month open-label phase with pioglitazone treatment. The primary outcome was a reduction of at least 2 points in the nonalcoholic fatty liver disease activity score in 2 histologic categories without worsening of fibrosis. Secondary outcomes included other histologic outcomes, hepatic triglyceride content measured by magnetic resonance and proton spectroscopy, and metabolic parameters. Among patients randomly assigned to pioglitazone, 58% achieved the primary outcome (treatment difference, 41 percentage points [95% CI, 23 to 59 percentage points]) and 51% had resolution of NASH (treatment difference, 32 percentage points [CI, 13 to 51 percentage points]) (P< 0.001 for each). Pioglitazone treatment also was associated with improvement in individual histologic scores, including the fibrosis score (treatment difference, -0.5 [CI, -0.9 to 0.0]; P= 0.039); reduced hepatic triglyceride content from 19% to 7% (treatment difference, -7 percentage points [CI, -10 to -4 percentage points]; P< 0.001); and improved adipose tissue, hepatic, and muscle insulin sensitivity (P< 0.001 vs. placebo for all). All 18-month metabolic and histologic improvements persisted over 36 months of therapy. The overall rate of adverse events did not differ between groups, although weight gain was greater with pioglitazone (2.5 kg vs. placebo). Single-center study. Long-term pioglitazone treatment is safe and effective in patients with prediabetes or T2DM and NASH. Burroughs Wellcome Fund and American Diabetes Association.

Macrophage Activation in Pediatric Nonalcoholic Fatty Liver Disease (NAFLD) Correlates with Hepatic Progenitor Cell Response via Wnt3a Pathway.

Non-alcoholic fatty liver disease is one of the most important causes of liver-related morbidity in children. In non-alcoholic fatty liver disease, the activation of liver resident macrophage pool is a central event in the progression of liver injury. The aims of the present study were to evaluate the polarization of liver macrophages and the possible role of Wnt3a production by macrophages in hepatic progenitor cell response in the progression of pediatric non-alcoholic fatty liver disease. 32 children with biopsy-proven non-alcoholic fatty liver disease were included. 20 out of 32 patients were treated with docosahexaenoic acid for 18 months and biopsies at the baseline and after 18 months were included. Hepatic progenitor cell activation, macrophage subsets and Wnt/β-catenin pathway were evaluated by immunohistochemistry and immunofluorescence. Our results indicated that in pediatric non-alcoholic fatty liver disease, pro-inflammatory macrophages were the predominant subset. Macrophage polarization was correlated with Non-alcoholic fatty liver disease Activity Score, ductular reaction, and portal fibrosis; docosahexaenoic acid treatment determined a macrophage polarization towards an anti-inflammatory phenotype in correlation with the reduction of serum inflammatory cytokines, with increased macrophage apoptosis, and with the up-regulation of macrophage Wnt3a expression; macrophage Wnt3a expression was correlated with β-catenin phosphorylation in hepatic progenitor cells and signs of commitment towards hepatocyte fate. In conclusion, macrophage polarization seems to have a key role in the progression of pediatric non-alcoholic fatty liver disease; the modulation of macrophage polarization could drive hepatic progenitor cell response by Wnt3a production.

Metabolic Damage Presents Differently in Young and Early-Aged C57BL/6 Mice Fed a High-Fat Diet

Summary Background Obesity in old individuals is increasing at alarming rates, and this population is more vulnerable to the deleterious metabolic effects of obesity than younger individuals. However, at present, there is no ideal obesity model to evaluate the interaction of aging and obesity. Methods The development of a metabolic damage model in response to a fixed period of high-fat diet (HFD) feeding was examined in mice of different ages. Mice aged 6 weeks (young group) and 44 weeks (elderly group) were fed a standard diet or HFD for 12 weeks, and their metabolic characteristics were studied. Inflammation was determined by the serum lipopolysaccharide (LPS) level. Gut microbiota composition was analyzed from stool samples. Results After 12-week feeding, weight gain; elevated serum levels of cholesterol, triglyceride, and LPS; increased homeostasis model assessment-estimated insulin resistance (HOMA-IR); fat accumulation; nonalcoholic fatty liver disease (NAFLD) activity score (NAS); and gut microbiota changes occurred in response to HFD feeding in both young and elderly groups ( p p Conclusion Aging in an obese mouse model does not increase LPS but exacerbates NAFLD severity, dyslipidemia, insulin insensitivity, fat accumulation, and gut microbiota changes. This obesity model might help elucidate target or multiple organ alterations associated with metabolic disorder and aging.

Aerobic exercise training in the treatment of non‐alcoholic fatty liver disease related fibrosis

Physiologically relevant rodent models of non-alcoholic steatohepatitis (NASH) that resemble the human condition are limited. Exercise training and energy restriction are first-line recommendations for the treatment of NASH. Hyperphagic Otsuka Long-Evans Tokushima fatty rats fed a western diet high in fat, sucrose and cholesterol for 24weeks developed a severe NASH with fibrosis phenotype. Moderate intensity exercise training and modest energy restriction provided some improvement in the histological features of NASH that coincided with alterations in markers of hepatic stellate cell activation and extracellular matrix remodelling. The present study highlights the importance of lifestyle modification, including exercise training and energy restriction, in the regulation of advanced liver disease. The incidence of non-alcoholic steatohepatitis (NASH) is rising but the efficacy of lifestyle modifications to improve NASH-related outcomes remain unclear. We hypothesized that a western diet (WD) would induce NASH in the Otsuka Long-Evans Tokushima Fatty (OLETF) rat and that lifestyle modification would improve this condition. Eight-week-old Long-Evans Tokushima Otsuka (L) and OLETF (O) rats consumed a control diet (10%kcal fat, 3.5% sucrose) or a WD (45%kcal fat, 17% sucrose, 1% cholesterol) for 24weeks. At 20weeks of age, additional WD-fed OLETFs were randomized to sedentary (O-SED), food restriction (O-FR; ∼25% kcal reduction vs. O-SED) or exercise training (O-EX; treadmill running 20mmin(-1) with a 15% incline, 60minday(-1) , 5daysweek(-1) ) conditions for 12weeks. WD induced a NASH phenotype in OLETFs characterized by hepatic fibrosis (collagen 1α1 mRNA and hydroxyproline content), as well as elevated inflammation and non-alcoholic fatty liver disease activity scores, and hepatic stellate cell activation (α-smooth muscle actin) compared to Long-Evans Tokushima Otsuka rats. FR and EX modestly improved NASH-related fibrosis markers (FR: hydroxyproline content, P<0.01; EX: collagen 1α1 mRNA, P<0.05; both: fibrosis score, P<0.01) and inflammation (both: inflammation score; FR: interleukin-1β and tumor necrosis factor α) vs. O-SED. FR reduced hepatic stellate cell activation markers (transforming growth factor-β protein and α-smooth muscle actin mRNA), whereas EX increased the hepatic stellate cell senescence marker CCN1 (P<0.01vs. O-SED). Additionally, both FR and EX normalized extracellular matrix remodelling markers to levels similar to L-WD (P>0.05). Although neither EX nor FR led to complete resolution of the WD-induced NASH phenotype, both independently benefitted liver fibrosis via altered hepatic stellate cell activation and extracellular matrix remodelling.

Fatty acid composition in serum correlates with that in the liver and non-alcoholic fatty liver disease activity scores in mice fed a high-fat diet.

In this study, we investigated the correlation between the serum fatty acid composition and hepatic steatosis, inflammation, hepatocellular ballooning scores, and liver fatty acids composition in mice fed a high-fat diet. Livers were collected for non-alcoholic fatty liver disease score analysis. Fatty acid compositions were analysed by gas chromatography. Correlations were determined by Pearson correlation coefficient. Exposed to a high-fat diet, mice developed fatty liver disease with varying severity without fibrosis. The serum fatty acid variation became more severe with prolonged exposure to a high-fat diet. This variation also correlated significantly with the variation in livers, with the types of fatty acids corresponding to liver steatosis, inflammation, and hepatocellular ballooning scores. Results of this study lead to the following hypothesis: the extent of serum fatty acid variation may be a preliminary biomarker of fatty liver disease caused by high-fat intake.

Prevalence and risk factors of steatosis after liver transplantation and patient outcomes.

Steatosis occurs frequently after liver transplantation (LT). We aimed to determine the prevalence of steatosis in adult LT recipients, to determine the effects of significant (>33%; grades 2-3) steatosis on patient survival, and to identify risk factors for the development of significant steatosis and its effect on fibrosis progression. We retrospectively examined 2360 posttransplant biopsies of 548 LT recipients. Survival was compared between patients with significant steatosis and those with grades 0-1 steatosis. Patients with significant steatosis were compared to controls without steatosis (grade 0) for clinical and laboratory factors and fibrosis progression. Steatosis was found in 309 (56.4%) patients, including 93 (17.0%) patients with significant steatosis. Steatohepatitis (nonalcoholic fatty liver disease activity score ≥ 5) was diagnosed in 57 (10.4%) patients. The prevalence of steatosis increased from 30.3% at 1 year to 47.6% at 10 years after LT (P < 0.001). Survival times did not differ between groups (P = 0.29). On multivariate analysis of pretransplant factors and initial immunosuppression (IS), alcohol-induced cirrhosis (P < 0.001) and high body mass index (BMI; P = 0.002) were associated with the development of significant steatosis, whereas increased levels of alkaline phosphatase (P = 0.01) and mycophenolate mofetil given initially (P = 0.009) appeared to protect against significant steatosis. On multivariate analysis of posttransplant factors, high BMI (P < 0.001), serum triglycerides (P < 0.001), alcohol consumption (P = 0.005), and type 2 diabetes mellitus (P = 0.048) were associated with significant steatosis, whereas high creatinine (P = 0.02) appeared to protect against significant steatosis. Significant steatosis was not associated with a higher fibrosis stage (P = 0.62). Posttransplant steatosis affects 56.4% of LT recipients, and the prevalence increases with time after LT. Recipient factors and types of IS affect the risk for significant steatosis, which is not associated with a higher fibrosis stage or worse patient survival. Liver Transplantation 22 644-655 2016 AASLD.

Effect of age on the development of chemotherapy-associated liver injury in colorectal cancer liver metastasis

Introduction: With the lengthening of the average life span, the number of elderly individuals undergoing liver resection for colorectal liver metastasis (CRLM) after chemotherapy has increased. However, the effect of age on the development of chemotherapy-associated liver injury (CALI) in CRLM is unclear. The aim of this study was to clarify the effect of age on the development of CALI in CRLM. Method: Sixty-four patients undergoing liver resection for CRLM after oxaliplatin-based chemotherapy (OBC) were studied. We divided the patients into three groups (A group: < 65 years, B group: 65–74 years, C group: >75 years) according to age at surgery and compared the development rate of CALI between the groups. The patient underwent pathological assessments to clarify the degree of histopathological liver injury of the nontumoral liver parenchyma. Results: The A group included 37 cases, B group 17 cases and C group 10 cases. There were no significant differences among the groups regarding the number of OBC cycles and duration of OBC cessation. Sinusoidal injury was observed in 27.0%, 29.4% and 30.0% of the A, B, and C groups, respectively, which was not significantly different (P = 0.479). Steatohepatitis was observed in 35.1%, 35.3% and 40.0% of the A, B, and C groups, respectively, which was not significantly different (P = 0.958). There was no significant correlation between age and sinusoidal pathological score (r = −0.102, P = 0.423) or nonalcoholic fatty liver disease activity score (r < ≤ 0.001, P = 0.997). Conclusions: The development of CALI after OBC in CRLM does not differ according to age.

The independent predictors of non‐alcoholic steatohepatitis and its individual histological features.

The diagnosis of non-alcoholic steatohepatitis (NASH) is based on the individual histological features: steatosis, lobular inflammation and ballooning. Non-alcoholic fatty liver disease (NAFLD) activity score (NAS ≥ 5) is used in clinical trials. Fibrosis dictates long-term NAFLD prognosis. Recently, more-than-mild portal inflammation has raised interest as a marker of NAFLD severity. We assessed the independent predictors of: (I) individual histological lesions of NASH; (II) diagnosis of NASH; (III) significant (stage ≥2) and advanced (stage ≥3) fibrosis; and (IV) more-than-mild portal inflammation. Data from 118 consecutive biopsy-proven NAFLD patients observed at our institution were retrospectively analyzed. At stepwise multivariate logistic regression analyses, independent predictors were as follows. For the individual histological features of NASH: insulin resistance (IR), assessed with Homeostasis Model Assessment-IR (HOMA-IR), serum uric acid (SUA) and serum total cholesterol (TCH) for moderate-to-severe steatosis; waist circumference (waist), HOMA-IR and TCH for lobular inflammation; waist, HOMA-IR, metabolic syndrome (MS), serum alanine aminotransferase (ALT), SUA and TCH for ballooning. For NASH diagnosis: waist, HOMA-IR, MS, ALT, SUA and TCH (Brunt etal.'s classification); ALT, SUA and TCH for NAS ≥ 5. For significant and advanced fibrosis, respectively: waist, MS and ALT; age, platelets, HOMA-IR, diabetes and TCH. For more-than-mild portal inflammation: serum aspartate aminotransferase (AST), serum iron, NAS ≥ 5 and significant liver fibrosis. HOMA-IR, SUA, MS, ALT and TCH are independent predictors of NASH and its individual histological lesions, notably including fibrosis. Based on our findings, these factors should be considered major pathogenic drivers of NASH and, by inference, potential targets for treatment.

Perspectives on Treatment for Nonalcoholic Steatohepatitis

It is important to provide treatment to patients with nonalcoholic steatohepatitis (NASH) because one third of patients with the metabolic syndrome die of liver disease. Basic research studies have elucidated mechanisms of NASH pathogenesis, which could lead to therapeutic targets. Health agencies have confirmed strategies for the optimal management of NASH and approved new drugs and treatments, which urgently are needed. The US Food and Drug Administration recently endorsed end points for NASH therapy. The reversal of NASH with no evidence of progression to advanced fibrosis has been defined as the end point for phase 2b and phase 3 trials in patients with NASH and early stage fibrosis. Although a decrease in the nonalcoholic fatty liver disease activity score could serve as an end point in clinical trials, it is not clear whether patients with lower scores have a lower risk of progression to advanced fibrosis. End points for clinical trials of patients with NASH cirrhosis currently are based on model for end-stage liver disease and Child-Pugh-Turcotte scores, as well as the hepatic venous pressure gradient. Different strategies are being explored to reduce liver diseases that are linked to a sedentary lifestyle, overeating, and genetic factors. In association with insulin resistance and deregulation of the lipid metabolism (accumulation of lipotoxins that promote hepatic lipogenesis, adipose tissue lipolysis, and impaired β-oxidation), these factors could increase the risk of liver steatosis with necroinflammatory lesions and fibrosis. We review the pathogenic mechanisms of NASH and therapeutic options, as well as strategies that are being developed for the treatment of injury to the liver and other organs.

Modulation of Gut Microbiota by Berberine Improves Steatohepatitis in High-Fat Diet-Fed BALB/C Mice.

Dysbiosis of the gut microbiota underlies non-alcoholic steatohepatitis (NASH). Ingredient of Chinese herbal medicine, berberine, has been proved to regulate the gut microbiota without systemic side effects. Therefore, we explored its effects on NASH induced by high-fat diet (HFD). BALB/c mice were randomized into three groups, including: control, model, and berberine treatment. With the exception of the control group with the standard diet, the model, and the treatment groups were treated by HFD. Mice from treatment group were further subjected to berberine (200 mg/kg/d) gavage since the 5th week. At the end of the 13th week, gut bacteria, liver endotoxin receptor, and inflammation cytokines were assessed by real-time PCR. NASH and its predisposing factors were evaluated biochemically and pathologically. Compared to their decreases in the model group, berberine administration restored the relative level of Bifidobacteria (2.16 ± 0.63 vs. 0.50 ± 0.08, P < 0.01) and the ratio of Bacteroidetes/ Firmicutes (0.76 ± 0.26 vs. 0.39 ± 0.11, P < 0.01), respectively, in the treatment groups. Microbiota restoration led to significant reductions in body weight, serum levels of lipids, glucose, insulin, and homeostasis model assessment of insulin resistance. Improvements were also observed in the serum transaminase activity and nonalcoholic fatty liver disease activity score, which demonstrated the attenuation of NASH. Mechanically, expression levels of CD14, IL-1, IL-6 and TNF-α were statistically down-regulated (treatment group vs model group, P < 0.01). Berberine alleviates NASH and its predisposing factors. Normalization of gut microbiota might underlie its effect.

Efficacy and safety study of cenicriviroc for the treatment of non-alcoholic steatohepatitis in adult subjects with liver fibrosis: CENTAUR Phase 2b study design

BackgroundNon-alcoholic steatohepatitis (NASH) is often accompanied by liver fibrosis, which can progress to cirrhosis; C-C chemokine receptors type 2 and 5 (CCR2/CCR5), which mediate interactions driving inflammation and fibrosis, are promising treatment targets. Cenicriviroc (CVC), a dual-CCR2/CCR5 antagonist, has potent anti-inflammatory and antifibrotic activity in animal models; in HIV-positive subjects it reduced soluble CD14 levels, aspartate aminotransferase-to-platelet count ratio index, and non-invasive hepatic fibrosis risk scores; favorable tolerability was demonstrated in ~600 subjects. Efficacy and safety of CVC 150mg for treating NASH with liver fibrosis are being evaluated over 2years (primary endpoint at Year 1 [Y1]). DesignPhase 2b, randomized, double-blind, placebo-controlled, multinational study (CENTAUR; NCT02217475). Adults with histological evidence of NASH, non-alcoholic fatty liver disease activity score (NAS)≥4, and liver fibrosis (stages 1–3 NASH clinical research network system) enrolled. Subjects have increased risk of progression to cirrhosis due to ≥1 characteristic: type 2 diabetes; body mass index>25kg/m2 with ≥1 feature of metabolic syndrome; bridging fibrosis and/or NAS≥5. Liver biopsy evaluation at Screening, Y1, and Year 2 (Y2). ObjectivesAssess histologic improvement (≥2-point in NAS with ≥1-point improvement in >1 category) without worsening of fibrosis at Y1 (primary); evaluate complete NASH resolution without worsening of fibrosis at Y2 (key secondary). DiscussionCENTAUR is the first prospective study evaluating an oral agent exclusively enrolling subjects with NASH and liver fibrosis, with increased risk of developing cirrhosis. It will compare shorter versus longer CVC treatment and assess correlations between decreased inflammation and fibrosis.

Elafibranor, an Agonist of the Peroxisome Proliferator−Activated Receptor−α and −δ, Induces Resolution of Nonalcoholic Steatohepatitis Without Fibrosis Worsening

Elafibranor is an agonist of the peroxisome proliferator-activated receptor-α and peroxisome proliferator-activated receptor-δ. Elafibranor improves insulin sensitivity, glucose homeostasis, and lipid metabolism and reduces inflammation. We assessed the safety and efficacy of elafibranor in an international, randomized, double-blind placebo-controlled trial of patients with nonalcoholic steatohepatitis (NASH). Patients with NASH without cirrhosis were randomly assigned to groups given elafibranor 80 mg (n= 93), elafibranor 120 mg (n= 91), or placebo (n= 92) each day for 52 weeks at sites in Europe and the United States. Clinical and laboratory evaluations were performed every 2 months during this 1-year period. Liver biopsies were then collected and patients were assessed 3 months later. The primary outcome was resolution of NASH without fibrosis worsening, using protocol-defined and modified definitions. Data from the groups given the different doses of elafibranor were compared with those from the placebo group using step-down logistic regression, adjusting for baseline nonalcoholic fatty liver disease activity score. In intention-to-treat analysis, there was no significant difference between the elafibranor and placebo groups in the protocol-defined primary outcome. However, NASH resolved without fibrosis worsening in a higher proportion of patients in the 120-mg elafibranor group vs the placebo group (19% vs 12%; odds ratio= 2.31; 95% confidence interval: 1.02-5.24; P= .045), based on a post-hoc analysis for the modified definition. In post-hoc analyses of patients with nonalcoholic fatty liver disease activity score ≥4 (n= 234), elafibranor 120 mg resolved NASH in larger proportions of patients than placebo based on the protocol definition (20% vs 11%; odds ratio= 3.16; 95% confidence interval: 1.22-8.13; P= .018) and the modified definitions (19% vs 9%; odds ratio= 3.52; 95% confidence interval: 1.32-9.40; P= .013). Patients with NASH resolution after receiving elafibranor 120 mg had reduced liver fibrosis stages compared with those without NASH resolution (mean reduction of 0.65 ± 0.61 in responders for the primary outcome vs an increase of 0.10 ± 0.98 in nonresponders; P < .001). Liver enzymes, lipids, glucose profiles, and markers of systemic inflammation were significantly reduced in the elafibranor 120-mg group vs the placebo group. Elafibranor was welltolerated and did not cause weight gain or cardiac events, but did produce a mild, reversible increase in serum creatinine (effect size vs placebo: increase of 4.31 ± 1.19 μmol/L; P < .001). A post-hoc analysis of data from trial of patients with NASH showed that elafibranor (120 mg/d for 1 year) resolved NASH without fibrosis worsening, based on a modified definition, in the intention-to-treat analysis and in patients with moderate or severe NASH. However, the predefined end point was not met in the intention to treat population. Elafibranor was well tolerated and improved patients' cardiometabolic risk profile. ClinicalTrials.gov number: NCT01694849.