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Background and Aim: Nonalcoholic fatty liver disease (NAFLD) is considered to be the liver component of metabolic syndrome. However, the impact of NAFLD on metabolic syndrome is unclear. The aim of this study was to explore the influence of NAFLD on the development of metabolic disorders. Methods: Patients with NAFLD and an age, sex, and occupation-matched control group were recruited from employees of Bao-Steel Group (Shanghai, China) who had received medical check-ups biennially between 1995 and 2002. Anthropometric and laboratory data, and incidence of metabolic disorders were assessed at baseline and at follow-up of at least 4 years. SPSS 11.5 was used for statistical analysis. Results: The study consisted of 358 patients (326 men and 32 women) and 788 matched controls (711 men and 77 women) with a similar mean age of 39.0 years and median follow-up of 6 years. At the end of follow-up, incidence of obesity (47.6% vs 19.5%), hypertension (69.6% vs 16.3%), hypertriglyceridemia (39.1% vs 16.3%), hypercholesterolemia (24.5% vs 17.3%), impaired fasting glucose (IFG) (25.1% vs 11.6%), diabetes mellitus (20.3% vs 5.2%) and multiple metabolic disorders (MMD) (56.3% vs 16.3%) were significantly higher in the fatty liver group than the control group. Interestingly, the mean alanine aminotransferase (ALT) level in patients with fatty liver significantly decreased at follow-up compared with baseline (28.56 ± 18.86 vs 31.51 ± 18.34 U/L, P

The risk factors and settings for non-alcoholic fatty liver disease (NAFLD) in Asians are reviewed comprehensively. Based particularly on large community-based studies using ultrasonography, case-control series and prospective longitudinal studies, the prevalence of NAFLD in Asia is between 12% and 24%, depending on age, gender, locality and ethnicity. Further, the prevalence in China and Japan has nearly doubled in the last 10-15 years. A detailed analysis of these data shows that NAFLD risk factors for Asians resemble those in the West for age at presentation, prevalence of type 2 diabetes mellitus (T2DM) and hyperlipidemia. The apparent differences in prevalence of central obesity and overall obesity are related to criteria used to define waist circumference and body mass index (BMI), respectively. The strongest associations are with components of the metabolic syndrome, particularly the combined presence of central obesity and obesity. Non-alcoholic fatty liver disease appears to be associated with long-standing insulin resistance and likely represents the hepatic manifestation of metabolic syndrome. Not surprisingly therefore, Asians with NAFLD are at high risk of developing diabetes and cardiovascular disease. Conversely, metabolic syndrome may precede the diagnosis of NAFLD. The increasing prevalence of obesity, coupled with T2DM, dyslipidemia, hypertension and ultimately metabolic syndrome puts more than half the world's population at risk of developing NAFLD/non-alcoholic steatohepatitis/cirrhosis in the coming decades. Public health initiatives are clearly imperative to halt or reverse the global 'diabesity' pandemic, the underlying basis of NAFLD and metabolic syndrome. In addition, a perspective of NAFLD beyond its hepatic consequences is now warranted; this needs to be considered in relation to management guidelines for affected individuals.

Background and Aim: To evaluate changes in the prevalence rates of ultrasonographic fatty liver (FL) in a specific population. Methods: An analysis of the medical records of BaoSteel Group (Shanghai, China) employees was done to evaluate the prevalence of FL in this population, in which health examinations were performed biennially between 1995 and 2002. Results: The study reviewed a database of 59 131 employees, of which 27.1% received medical check-ups four times within the study period, 26.6% three times, and 24.0% twice. The prevalence rates of obesity and metabolic disorders were high at baseline and increased significantly with time. The prevalence of ultrasonographic FL increased from 3.87% to 14.04% in the overall population, while there was an increase from 4.44% to 14.64% in men and 1.56% to 11.37% in women over the 6-year study period. Increased rates of FL were also noted, from 25.88% to 51.39%, among patients with elevated serum alanine aminotransferase (ALT) levels (>40 U/L). The highest overall prevalence rates of FL were found in individuals aged 50–60 years, with all age-associated prevalence significantly higher in males than females. Conclusion: The prevalence of FL increased rapidly over the study period with increased rates of obesity and metabolic disorders; FL is becoming a major cause of abnormal ALT levels in the specific population.

Activation of hepatic stellate cells (HSCs) depending on epithelial-to-mesenchymal transition (EMT) reflects the key event of liver fibrosis. Contrastively, mesenchymal-to-epithelial transition (MET) of HSCs facilitates the fibrosis resolution. Here we investigated the effect of Fuzheng Huayu (FZHY) recipe, a Chinese herbal decoction made ofRadix Salviae Miltiorrhizae,Semen Persicae,Cordyceps sinensis,Pollen Pini, andGynostemma pentaphyllum, on liver fibrosis concerning the balance of EMT and MET in HSCs. In contrast to the increased TGF-β1/BMP-7 ratio in activated HSCs, FZHY administration induced significant upregulation of BMP-7 and downregulation of TGF-β1at both transcription and translation levels. Restoration of TGF-β1/BMP-7 ratio inhibited the expression of p38 MAPK and phosphorylated p38 MAPK, resulting in the reversal of epithelial-to-mesenchymal transition (EMT) to mesenchymal-to-epithelial transition (MET) as characterized by the abolishment of EMT markers (α-SMA and desmin) and reoccurrence of MET marker (E-cadherin).In vivotreatment of FZHY recipe also demonstrated the statistical reduction of activated HSCs with EMT phenotype, which attenuated the carbon tetrachloride- (CCl4-) induced liver fibrosis in a dose-dependent manner. These findings may highlight a novel antifibrotic role of FZHY recipe on the basis of rebalancing EMT and MET in HSCs.

The minipig can serve as a good pharmacological model for human subjects. However, the long-term pathogenesis of high-calorie diet-induced metabolic syndromes, including NASH, has not been well described in minipigs. We examined the development of metabolic syndromes in Bama minipigs that were fed a high-fat, high-sucrose diet (HFHSD) for 23 months, by using histology and serum biochemistry and by profiling the gene expression patterns in the livers of HFHSD pigs compared to controls. The pathology findings revealed microvesicular steatosis, iron overload, arachidonic acid synthesis, lipid peroxidation, reduced antioxidant capacity, increased cellular damage and inflammation in the liver. RNA-seq analysis revealed that 164 genes were differentially expressed between the livers of the HFHSD and control groups. The pathogenesis of early-stage NASH was characterized by hyperinsulinemia and by de novo synthesis of fatty acids and nascent triglycerides, which were deposited as lipid droplets in hepatocytes. Hyperinsulinemia shifted the energy supply from glucose to ketone bodies and the high ketone body concentration induced the overexpression of cytochrome P450 2E1 (CYP2E1). The iron overload, CYP2E1 and alcohol dehydrogenase 4 overexpression promoted reactive oxygen species (ROS) production, which resulted in arachidonic and linoleic acid peroxidation and, in turn, led to malondialdehyde production and a cellular response to ROS-mediated DNA damage.

Background: Chronic Hepatitis B (CHB) infection is common in patients with Non-Alcoholic Fatty Liver Disease (NAFLD). The replication level of Hepatitis B Virus (HBV) was inversely correlated with hepatic steatosis. Toll-Like Receptor (TLR) 4-mediated innate immunity plays a pivotal role in the occurrence of NAFLD and controls HBV replication. Objectives: This study aimed to investigate whether the TLR4-mediated innate immunity stimulates the pathogenesis of CHB in patients with NAFLD and to determine whether TLR4 plays a role in inhibiting HBV replication. Materials and Methods: The HBV transgenic mice were randomized into the HBV and HBV/NAFLD groups. HepG2.2.15 cells were treated with different concentrations (0 – 200 μM) of Stearic Acid (SA) to induce steatosis. The total RNA of the liver tissue was extracted for Real-Time Polymerase Chain Reaction (RT-PCR) detection, and immunohistochemistry or western blot was conducted for further validation. The Enzyme-Linked Immunosorbent Assay (ELISA) analysis was applied to evaluate the production of Interleukin 6 (IL-6), Tumor necrosis factor α (TNF-α) and Interferon β (IFN-β). Moreover, viral dynamics were analyzed using HBV DNA and HBV-related antigens (HBsAg and HBeAg). Results: Non-alcoholic fatty liver disease was induced in HBV-transgenic mice fed with High Fat Diet (HFD) for 8 - 24 weeks. Oil red-O staining positive droplets and the content of Triglyceride (TG) were increased in HepG2.2.15 cells treated with SA. TLR4, Myeloid differentiation factor 88 (MyD88), IL-6 and TNF-α expression levels were significantly higher in the HBV/NAFLD group and the steatotic HepG2.2.15 cells than those in their respective controls. Compared to the HBV group, significant reductions in serum levels of HBsAg, HBeAg, and HBV DNA titers occurred in the HBV/NAFLD group at 24 weeks, but the IFN-β level was remarkably increased. Similar data were also obtained from the steatoric HepG2.2.15 cells. Conclusions: Saturated Fatty Acids (SFAs) served as a potential ligand for TLR4 and activated TLR4 signaling pathway, which might be involved in the pathogenesis. Thus, SFAs can accelerate the mechanism of inhibiting HBV replication in CHB with NAFLD.

Associations between thyroid function and non-alcoholic fatty liver disease (NAFLD) are unknown in chronic hepatitis B (CHB)-infected patients. Thus, the aim of the study was to investigate the prevalence of thyroid dysfunction and its relationship with NAFLD in CHB.Consecutive naive CHB infected patients that had undergone liver biopsy and serum thyroid function tests between January 2007 and December 2011 were retrospective analyzed. NAFLD was diagnosed as at least 5% biopsy-proven hepatic steatosis without significant alcohol consumption.A total of 1154 non-alcoholics with CHB were included, 270 (23.39%) patients were found to have NAFLD, most of them (88.5%) with mild steatosis. The prevalence of hyperthyroidism and hypothyroidism (including subclinical and overt) was 1.56% and 1.64%, respectively, both with similar rates in patients with and without NAFLD (1.85% vs 1.47%, 1.48% vs 1.69%, respectively, both P 0.05). The serum thyroid-stimulating hormone (TSH) level in NAFLD patients was significantly higher than that in patients without NAFLD (2.22 ± 2.13 vs 1.61 ± 1.20 mIU/L, P 0.05). After adjustment for age and gender, the elevated TSH level was associated with increased odds of having steatosis (odds ratio1.54, 95% confidence interval 1.049-2.271) instead of viral factors and hepatic inflammation and fibrosis.Thyroid dysfunction is not common in CHB-infected patients, and the prevalence of hypothyroidism in CHB individuals with or without NAFLD is similar. However, increased serum TSH concentration at the normal range is a significant predictor of hepatic steatosis in patients with CHB.

Background Nonalcoholic fatty liver disease (NAFLD) is a risk factor for hepatocellular carcinoma (HCC), but the association between a high-fat diet (HFD) and HCC is not fully understood. In this study, we investigated whether a high-saturate-fat diet affects hepatocarcinogenesis induced by administration of diethylnitrosamine (DEN). Methods Adult SD rats were randomized into the following groups: normal chow diet (NCD), HFD, NCD + DEN, and HFD + DEN. The HFD contains 2% cholesterol and 10% lard oil. In mice with DEN treatment, the carcinogen was given via gavage. Mice were sacrificed at the end of 10, 12, and 14 weeks, respectively. The effects of HFD on hepatic carcinogenesis were assessed by HCC incidence, tumor differentiation, and the number and size of tumor nodules. Western blot and immunohistochemistry for proliferating cell nuclear antigen (PCNA), enzyme-linked immunosorbent assay (ELISA) for caspase-3, and real-time PCR for TNF-α and IL-6 further uncovered the proliferative and apoptotic properties of liver. Results In contrast to the NCD group, DEN treatment (NCD + DEN group) led to hepatitis, cirrhosis, hepatic tumor, and decreased body weight. Interestingly, HFD, which induced hyperlipidemia and hepatic steatosis, attenuated DEN-related malnutrition and fibrosis progression in HFD + DEN group during 10–14 weeks. Moreover, the HFD + DEN group exhibited that the proportion of well differentiated HCC was much higher than that of NCD + DEN group. The number and average volume of HCC node were also significantly lowered in HFD + DEN group (P

High‑mobility‑group‑box chromosomal protein 1 (HMGB1) is a ubiquitous and abundant nuclear protein in eukaryotic cells. Nuclear HMGB1 serves an important role in maintaining nuclear stability under stress. However, extracellular HMGB1 exerts actions, which are distinctly different compared with these intracellular functions. HMGB1, when released extracellularly, is a potent innate signal, which initiates host defense mechanisms or tissue regeneration. HMGB1 has two DNA‑binding domains: HMG A box and B box. The HMGB1 A box exhibits an antagonistic, anti‑inflammatory effect, and is a potential therapeutic target, however, the large‑scale expression and purification of the HMGB1 A box with high efficiency remains to be reported. In the present study, a SUMO‑fusion expression system was used to express and purify high levels of functional HMGB1 A box to meet the requirements of therapeutic protein production.

Liver disease is a major cause of illness and death worldwide. In China alone, liver diseases, primarily viral hepatitis (predominantly hepatitis B virus [HBV]), nonalcoholic fatty liver disease, and alcoholic liver disease, affect approximately 300 million people. The establishment of the Expanded Program on Immunization in 1992 has resulted in a substantial decline in the number of newly HBV-infected patients; however, the number of patients with alcoholic and nonalcoholic fatty liver diseases is rising at an alarming rate. Liver cancer, one of the most deadly cancers, is the second-most common cancer in China. Approximately 383,000 people die from liver cancer every year in China, which accounts for 51% of the deaths from liver cancer worldwide. Over the past 10 years, China has made some significant efforts to shed its "leader in liver diseases" title by investing large amounts of money in funding research, vaccines, and drug development for liver diseases and by recruiting many Western-trained hepatologists and scientists. Over the last two decades, hepatologists and scientists in China have made significant improvements in liver disease prevention, diagnosis, management, and therapy. They have been very active in liver disease research, as shown by the dramatic increase in the number of publications in Hepatology. Nevertheless, many challenges remain that must be tackled collaboratively. In this review, we discuss the epidemiology and characteristics of liver diseases and liver-related research in China.

Background/Aims To determine the prevalence and risk factors of fatty liver (FL) among Shanghai adults. Methods A cross-sectional ultrasonographic survey with randomized multistage stratified cluster sampling was used. Results The study included 3175 subjects (1218 men) with a mean age of 52 years. FL was found in 661 (20.82%) subjects. After adjustment by age and sex, FL prevalence was found to be 17.29%, and the prevalences of alcoholic, suspected alcoholic and nonalcoholic FL were determined to be 0.79, 1.15 and 15.35%, respectively. Generally, age, body mass index (BMI), waist circumference, blood pressure, and the prevalences of obesity, diabetes, hypertension and dyslipidemia were all significantly higher in FL patients than in controls; In contrast, the levels of high-density-lipoprotein cholesterol (HDL-C), education and physical activity were markedly lower. Multiple regression analyses showed that only nine factors (male, educational level, waist circumference, BMI, HDL-C, triglyceride, fasting plasma glucose, diabetes and hypertension) were closely related to FL. In excessive drinkers, obesity increased the risk for FL by 4.8-fold, but excessive drinking was associated with only a 1.5-fold increased risk in obese subjects. Conclusions FL in Shanghai is highly prevalent and mainly related to multiple metabolic disorders.

To investigate whether gut microbiota metabolite sodium butyrate (NaB) is an effective substance for attenuating non-alcoholic fatty liver disease (NAFLD) and the internal mechanisms.Male C57BL/6J mice were divided into three groups, normal control were fed standard chow and model group were fed a high-fat diet (HFD) for 16 wk, the intervention group were fed HFD for 16 wk and treated with NaB for 8 wk. Gut microbiota from each group were detected at baseline and at 16 wk, liver histology were evaluated and gastrointestinal barrier indicator such as zonula occluden-1 (ZO-1) were detected by immunohistochemistry and realtime-PCR, further serum or liver endotoxin were determined by ELISA and inflammation- or metabolism-associated genes were quantified by real-time PCR.NaB corrected the HFD-induced gut microbiota imbalance in mice, while it considerably elevated the abundances of the beneficial bacteriaNaB may restore the dysbiosis of gut microbiota to attenuate steatohepatitis, which is suggested to be a potential gut microbiota modulator and therapeutic substance for NAFLD.

Hepatic stellate cell (HSC) activation is the critical event of liver fibrosis. Abnormality of miR-16 expression induces their activation. However, the action model of miR-16 remains to be elucidated because of its multiple-targeted manner. Here, we report that miR-16 restoration exerted a wide-range impact on transcriptome (2,082 differentially expressed transcripts) of activated HSCs. Integrative analysis of both targetome (1,195 targets) and transcriptome uncovered the miR-16 regulatory network based upon bio-molecular interaction databases (BIND, BioGrid, Tranfac, and KEGG), cross database searching with iterative algorithm, Dijkstra's algorithm with greedy method, etc. Eight targets in the targetome (Map2k1, Bmpr1b, Nf1, Pik3r3, Ppp2r1a, Prkca, Smad2, and Sos2) served as key regulatory network nodes that mediate miR-16 action. A set of TFs (Sp1, Jun, Crebl, Arnt, Fos, and Nf1) was recognized to be the functional layer of key nodes, which mapped the signal of miR-16 to transcriptome. In result, the comprehensive action of miR-16 abrogated transcriptomic characteristics that determined the phenotypes of activated HSCs, including active proliferation, ECM deposition, and apoptosis resistance. Therefore, a multi-layer regulatory network based upon the integration of targetome and transcriptome may underlie the global action of miR-16, which suggesting it plays an inhibitory role in HSC activation.

Recent evidence has indicated that polyunsaturated fatty acids (PUFA), such as omega-3 PUFA, have protective effects on a range of chronic inflammatory conditions, including obesity, and may play a role in the reversal of steatohepatitis. However, the effects of omega-3 PUFA on adipokine expression and hepatic lipid metabolism have not been well evaluated. Thus, the aim of our study was to investigate the effects of PUFAs on adipokines, as well as lipid and glycometabolism, in a rat model of non-alcoholic steatohepatitis (NASH).Male Sprague-Dawley rats were divided into control, model and therapy groups. The control group received a normal diet, while the model and therapy groups received a high-fat diet. On the eighth week of high-fat diet, the therapy group was treated with omega-3 PUFA (1.0 g/d) daily. At the end of 20 weeks, serum biochemistry indices were measured and adipokine levels in serum and liver samples were detected with ELISA, Western blotting and real time fluorescence quantitative PCR (qRT-PCR).The weight, biochemical parameters and adipokine levels in serum of the model group were elevated compared to the control group (P0.05). In addition, the protein and mRNA expression levels of adipokines in the liver were significantly altered compared to the control group (P0.01). The therapy group was characterized by decreased weight and biochemical indices (P0.05) compared with the model group. Supplementing high-fat diet with omega-3 PUFA decreased serum levels of leptin and resistin, while adiponectin levels were slightly elevated. In liver tissue samples, the protein and mRNA expression levels of adipokines were significantly improved (P0.01) in the therapy group compared to the model group.Omega-3 PUFA improved lipid and glycometabolism in NASH rats and regulated adipokine expression, indicating that omega-3 PUFA may have a therapeutic benefit for patients with NASH.

The controlled attenuation parameter (CAP) using transient elastography (TE) was validated in chronic hepatitis C to evaluate hepatic steatosis; however, limited data are available on chronic hepatitis B (CHB). Therefore, we assessed the accuracy and the efficacy of CAP for the detection of steatosis in CHB. Consecutive CHB patients underwent liver biopsy and liver stiffness measurements (LSM) with simultaneous CAP determination using the M probe of the TE. The area under the receiver operating characteristics curve (AUROC) was used to evaluate the performance of CAP in diagnosing steatosis compared with biopsy. A total of 340 patients were included: 60 % were male, the median age was 37 years; the body mass index (BMI) was ≥28 kg/m2 for 14 % of the subjects; and the distribution of the steatosis grade was S0 58.2 %, S1 34.2 %, S2 5.0 % and S3 2.6 %. The median (range) of CAP was 218 (100–400) dB/m, and CAP correlated with the BMI (ρ = 3.622) and steatosis grade (ρ = 29.203) according to a multivariate analysis (both P

Background Prolyl oligopeptidase (POP) is a serine endopeptidase that is widely distributed in vivo, particularly in the liver. Significant changes in functional mitochondrial proteins involved with mitochondrial oxidoreductases/transporters and nucleic acid binding proteins were observed after POP inhibition in the liver, which suggested a role of POP in regulating liver energy metabolism. Steatosis in nonalcoholic fatty liver disease (NAFLD) is associated with disturbances in lipid and energy metabolism in hepatocytes. Here, we aimed to study the effect of POP on hepatocyte steatosis. Methods The human liver cell line L02 was used to investigate the biological effects of POP. An in vitro cell model of steatosis was successfully induced with oleic acid and palmitic acid. L02 cells were also subjected to S17092 (a POP inhibitor) at different concentrations for 24 or 48 h. Ac-SDKP levels and POP activity were measured to assess the rate of inhibition of POP by S17092. The POP gene and protein expression levels were detected using real-time PCR and Western blots, respectively. Oil red O staining was performed and the triglyceride levels in the L02 cells were also measured. Cell proliferation and apoptosis were detected using CCK-8 and flow cytometry, respectively. The expression of genes involved in lipid metabolism was detected using real-time PCR. The effects of POP inhibition on LC3B II were detected by Western blot. Results Compared with the control, the POP mRNA levels increased by approximately 30%, and the POP protein levels increased by almost 60% in the steatotic L02 cells. After S17092 (0.026~130 μM) incubation for 24 or 48 h, cell proliferation was significantly decreased in the free fatty acid (FFA)-treated cells at 26–130 μM; however, S17092 did not affect the proliferation of L02 cells after 24 h of incubation with S17092 at 0.026–65 μM without FFA treatment. S17092 treatment (13 and 26 μM) also elicited no significant effect on apoptosis in normal L02 cells, but FFA treatment increased cell apoptosis, which was attenuated by S17092 incubation. S17092 treatment inhibited intracellular POP activity and decreased the AcSDKP level at the concentration of 0.026–26 μM. After treatment with FFA for 24 h, oil red O staining revealed significant lipid accumulation in the cells in the model group compared with the controls; however, lipid accumulation was suppressed after the administration of S17092 (13 and 26 μM). Accordingly, the triglyceride levels in the FFA-treated cells were approximately 5-fold greater than those of the controls and were decreased by approximately 25% and 45% after the administration of S17092 at 13 and 26 μM, respectively. The mRNA levels of FASN, PPAR-γ, and SREBP-1c were higher in the FFA-treated cells than in the normal controls, and all of these levels were significantly inhibited in the presence of S17092 at both 13 and 26 μM. S17092 treatment did not affect LC3B II in the FFA-treated cells compared with FFA treatment alone. Conclusion The expression of POP increases with hepatocyte steatosis, and POP inhibitors can significantly reduce intracellular lipid accumulation, which might be related to the inhibition of genes involved in lipid synthesis.

Garlic (Allium sativam L.) is widely used in traditional herbal remedies and alternative medicine. The potential health benefits of garlic are largely attributed to its metabolic byproducts. Extensive in vivo and in vitro studies has demonstrated that the garlic derivatives possess anti-cancer effects, but the underlying mechanisms are not completely understood. In this mini-review, we aim to summarize the reported biological effects of garlic products as anti-tumor agents, and present the possible molecular mechanisms responsible for the anti-carcinogenesis effects of garlic and its derivatives.

β-catenin, a core component of Wnt/β-catenin signaling, has been shown to be an important regulator of cellular proliferation and differentiation. Abnormal activation of Wnt/β-catenin signaling promotes tissue fibrogenesis. In the present study, the role of β-catenin during liver fibrogenesis was analyzed and the functional effects of β-catenin gene silencing in hepatic stellate cells (HSCs) using small interfering (si)RNA were investigated. The expression of β-catenin in human hepatic fibrosis tissues of different grades and normal human hepatic tissues was examined using immunohistochemistry. To inhibit the Wnt/β-catenin signaling pathway, siRNA for β-catenin was developed and transiently transfected into HSC-T6 cells using Lipofectamine 2000. β-catenin expression was evaluated by quantitative polymerase chain reaction (qPCR) and western blot analysis. The expression of collagen types I and III was evaluated by qPCR and immunofluorescent staining. Cellular proliferation and the cell cycle were analyzed using a methyl thiazolyl tetrazolium assay. Apoptosis was assessed by Annexin V staining. A higher expression level of β-catenin was identified in the patients with high-grade hepatic fibrosis in comparison with that of the normal controls. Additionally, β-catenin siRNA molecules were successfully transfected into HSCs and induced inhibition of β-catenin expression in a time-dependent manner. β-catenin siRNA treatment also inhibited synthesis of collagen types I and I in transfected HSCs. Furthermore, compared with those of the control group, siRNA-mediated knockdown of β-catenin in HSC-T6 cells inhibited cell proliferation and resulted in cell apoptosis. This study suggests a significant functional role for β-catenin in the development of liver fibrosis and demonstrates that downregulation of the Wnt/β-catenin signaling pathway inhibits HSC activation. Thus, this study provides a novel strategy for the treatment of hepatic fibrosis.

To evaluate the diagnostic accuracy of some noninvasive fibrosis models in Chinese patients with nonalcoholic fatty liver disease (NAFLD).Consecutive biopsy-proven NAFLD patients were recruited from a single center from January 2005 to December 2010. Advanced fibrosis (stage 3 and 4) was defined using Kleiner criteria. The area under the receiver operating characteristic curve (AUROC) was used to compare the diagnostic accuracy of the NAFLD fibrosis score (NFS), FIB-4 index, aspartate transaminase (AST)/platelet ratio index (APRI), AST/alanine transaminase (ALT) ratio (AAR) and body mass index (BMI)-AAR-Diabetes (BARD) score.Of the patients with NAFLD, 79.6% were males with a mean age of 37.1 years, mean BMI of 26.1 kg/m(2) and 41.4% of them had nonalcoholic steatohepatitis, and 24 (15.8%) had advanced fibrosis. The AUROC of the FIB-4 index, APRI, AAR, NFS and BARD score for advanced fibrosis were 0.756, 0.742, 0.670, 0.653 and 0.642 (P 0.05 for all), respectively. A concordant negative predictive value of approximately 90% was indicated whereas the positive predictive values were modest for all tests, and only the FIB-4 index yielded a higher positive likelihood ratio of 7.65. Using these cut-off values of tests for excluding advanced fibrosis could reduce the use of liver biopsy in 56.6-74.3% of the patients, with a minor false negative rate of 5.3-9.9%.Although slightly less accurate than liver biopsy, simple noninvasive tests can reliably exclude advanced fibrosis in Chinese NAFLD patients in our center. FIB-4 index performs better than the other tests under examination.

AIM To assess disease-specific circulating microRNAs (miRNAs) in non-alcoholic steatohepatitis (NASH) patients. METHODS A total of 111 biopsy-proven non-alcoholic fatty liver disease (NAFLD) or chronic hepatitis B (CHB) patients and healthy controls from mainland China were enrolled to measure their serum levels of miR-122, -125b, -146b, -16, -21, -192, -27b and -34a. The correlations between serum miRNAs and histological features of NAFLD were determined. The diagnostic value of miRNA in NASH and significant fibrosis was analyzed and compared with that of cytokeratin-18 (CK-18), fibrosis-4 (FIB-4), and aspartate aminotransferase to platelet ratio index (APRI), respectively. RESULTS Circulating miR-122, -16, -192 and -34a showed differential expression levels between NAFLD and CHB patients, and miR-34a had an approximately 2-fold increase in NAFLD samples compared with that of CHB samples (P < 0.01). Serum miR-122, -192 and -34a levels were correlated with steatosis (R = 0.302, 0.323 and 0.470, respectively, P < 0.05) and inflammatory activity (R = 0.445, 0.447 and 0.517, respectively, P < 0.01); only serum miR-16 levels were associated with fibrosis (R = 0.350, P < 0.05) in patients with NAFLD. The diagnostic value of miR-34a for NASH (area under the receiver operating characteristic, 0.811, 95%CI: 0.670-0.953) was superior to that of alanine aminotransferase, CK-18, FIB-4 and APRI in NAFLD, but miR-16 showed a limited performance in the diagnosis of significant fibrosis in NASH. CONCLUSION Circulating miR-34a may serve as a disease-specific noninvasive biomarker for the diagnosis of NASH.

To investigate the impact of hepatic steatosis on virologic response in chronic hepatitis B (CHB) patients treated with pegylated interferon-alpha (PEG-IFNa).Ninety-six naive patients positive for hepatitis B e antigen (HBeAg) and with biopsy-proven CHB were administered PEG-IFNa-2a or PEG-IFNa-2b for 48 weeks. Virologic response (HBeAg clearance and hepatitis B virus (HBV) DNA less than 5 log10 copies/ml) and biochemical response (alanine transaminase (ALT) normalization) were compared between patients with (n=34) and without (n=62) steatosis.The HBV DNA titer in the steatosis group was significantly lower than that of the non-steatosis group (6.961.27 vs. 7.541.28 log10 copies/ml; t=2.161, P=0.033). After 48 weeks of PEG-IFNa treatments, there was no significant difference in HBeAg seroconversion or the percentage of undetectable HBV DNA (less than 3 log10 copies/ml) between steatosis and non-steatosis patients. However, the steatosis patients presented with a significantly lower complete response rate (virologic response plus biochemical response) compared to non-steatosis patients (26.5% vs. 48.4%; x² =4.373, P=0.037). Of the 45 CHB patients with undetectable HBV DNA after 48 weeks of treatment, seven did not achieve ALT normalization. The rate of patients with non-biochemical response was significantly higher in the steatosis group than in the non-steatosis group (33.3% vs. 6.67%; P=0.032).Hepatic steatosis does not affect the virologic response, but does affect the biochemical response in CHB patients treated with PEG-IFNa for 48 weeks.

Background. Questions remain about the dose-response relationship between body mass index (BMI) and primary liver cancer (PLC) risk, possible confounding by hepatitis virus infection, and differences by gender or geographic location. We performed a meta-analysis of prospective studies to explore these issues. Methods. We searched PubMed and Embase for studies of BMI and risk of PLC through November 30, 2011. Summary relative risks with their corresponding 95% confidence intervals (CIs) were calculated using a random effects model. Results. A total of 21 prospective studies (including 17,624 PLC cases) were included in our analysis. The summary relative risk for a 5-unit increment in BMI (in kg/m2) was 1.39 (95% CI: 1.25–1.55), with high heterogeneity. These positive results were robust when stratified by sex, geographic location, ascertainment of exposure and outcome, the number of cases, duration of follow-up, sample source, and cofounders. There was evidence of a nonlinear association between BMI and PLC risk, with the most pronounced increase in risk among persons with a BMI >32 kg/m2. Patients with hepatitis C virus or cirrhosis (but not patients with hepatitis B virus) with excess weight had a higher risk of PLC development than general populations with excess weight. Conclusion. Excess weight increases PLC risk. For people with HCV infection or cirrhosis, risk increases are greater than for general population.

Non-alcoholic fatty liver disease (NAFLD) overlapping with chronic hepatitis B virus (HBV) infection is undergoing a rapid increase in China. Therefore, the establishment and character of an animal model with both NAFLD and chronic HBV infection has become an urgent task.Mice with chronic HBV genotype B infection were established with a microinjection of oocytes. Transgenic and nontransgenic mice were then randomized into groups of NAFLD + HBV, HBV, NAFLD, and control and were treated with high-fat diets or common forage. At 8, 16, and 24 weeks, characteristics of NAFLD were evaluated by physical indices, liver function tests, glycolipid metabolism, and histopathological scoring. Viral dynamics were also analyzed by HBV-DNA and HBV-related antigens.Hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) were expressed, and HBV-DNA was replicated in HBV transgenic mice at different stages in the serum and liver. Hepatic steatosis was only induced after exposure of the mice to high-fat diets, and no obvious pathological changes occurred in the HBV group from 8 to 24 weeks. Compared to mice with HBV alone, significant reductions in serum levels of HBV-DNA, HBsAg and HBeAg occurred in the NAFLD + HBV group after 24 weeks (all P 0.05). Nevertheless, the NAFLD and NAFLD + HBV groups shared comparable physical and metabolic disorders and similar steatotic, inflammatory and fibrotic characteristics in the liver.High-fat diets and transgenic operations on the HBV genotype B induced a rodent model of NAFLD overlapping with chronic HBV infection, and this model reduces the HBV viral factors but not the metabolic and histologic features.

To determine the prevalence and risk factors of functional constipation (FC) by using Rome III criteria in the local adult communities.A stratified randomized and community-based study by multi-stage cluster sampling was employed. A household survey was conducted from April to May 2010. All of the participants were interviewed face-to-face by filling out the self-administered questionnaires which based on Rome III criteria for the diagnosis of FC. Self-rating anxiety scale (SAS), self-rating depression scale (SDS) and Athens insomnia scale (AIS) were carried out to evaluate the psychological characteristics and qualities of sleep.A total of 7648 subjects fulfilled the questionnaires, with the response rate as 90.0%. 211 patients met the Rome III criteria, including 90 males and 121 females. The adjusted prevalence rates of FC were 2.5% in males, 3.3% in females and with an overall rate as 2.9%. The ratio of men to women was 1:1.32, with significant difference between males and females (P=0.043). The most common group was in the 18-29 year-olds (χ2=37.359, P=0.000). FC patients were more likely to be detected in the group with normal BMI (χ2=16.087, P=0.002), having received high education (χ2=27.604, P=0.000), being intelectuals (χ2=6.922, P=0.031) and divorced (χ2=22.000, P=0.000) than in other groups. Multivariate analysis showed that excessive intake of high-fat food was significantly associated with the presence of FC (odds ratio as 1.253, P=0.000), whereas foods with high-fiber (odds ratio as 0.854, P=0.029) might serve as protective factors. Significant differences between FC groups and control groups were found in the incidence of anxiety (with odds ratio as 2.583, P=0.000) and insomnia (odds ratio as 2.443, P=0.000).The prevalence of FC in adult communities in Shanghai Songjiang district was not higher than that in other parts of the communities. Excessive intake of high-fat food, anxiety and insomnia might be risk factors for FC and foods with high-fiber contents might serve as protective factors.

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To establish and identify an animal model of non-alcoholic fatty liver disease in chronic HBV infected mice.Transgenic mice with sustaining HBV production were established by microinjection of ocyte. Then they were randomly assigned into 4 groups (male control, male NAFLD model, female control and female NAFLD model) and treated with high fat diet (2% cholesterol, 10% lard, 88% forage) and common forage, respectively. NAFLD-related physical indexes, liver and kidney function, glucose and lipid metabolism were investigated at the time points of 8 weeks, 16 weeks and 24 weeks. Meanwhile, HBV type, serum levels of HBV DNA and HBeAg, immunohistochemical staining of hepatic HBsAg were detected. The establishment of NAFLD was evaluated by serum levels of total cholesterol (TC), triglycerides, glucose, aspartate aminotransferase (AST), alkaline phosphatase (ALP), alanine aminotransferase (ALT), gamma-glutamyltransferase (GGT), etc. Histological changes were also analyzed by HE, oil red O and Masson's trichrome staining. The status of CHB was assessed on the basis of immunohistochemistry and real-time PCR.The body and liver weights, liver index in HBV transgenic mice were significantly increased in regardless of the gender of HFD feeding, and the levels of serum ALT, AST, ALP, GGT, TBIL, TBA, TC, TG, HDL-C, LDL and FBG were higher in HFD groups as compared with the control mice. Lipid droplets, cytologic ballooning and liver steatosis could be observed in most lobules of HFD groups after 8-week administration, fatty degeneration of hepatocytes, patch necrosis, mild to moderate chronic inflammatory infiltration were also observed in some of HFD feeding, reflecting the emerge of steatohepatitis. At the time point of 24-week perisinusoidal fibrosis and local fibrosis occurred in HFD groups. Immunohistochemical staining and real-time PCR analysis of the liver tissues showed positive signal of HBsAg in all groups of mice, although no significant difference was documented.Our study suggests that animal model of NAFLD can be established in HBV transgenic mice and provide a nice animal model for further studies on NAFLD with chronic hepatitis B infection.

Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and represents the third cause of mortality among deaths from cancer (Semela & Heim, 2011; EI-Serag & Rudolph, 2007; Yuen et al, 2009; Lodato et al, 2006; Hainaut & Boyle, 2008). There are upwards of 600, 000 new cases each year in Asia and sub-Saharan Africa, where populations suffer both from a high prevalence of hepatitis B virus (HBV) infection and largely uncontrolled aflatoxin exposure in food, and more than 200, 000 HCC-related deaths annually in the People’s Republic of China alone (Kew, 2002; Wang et al, 2002). Aflatoxins may play a causative role in 5%-28% of all global HCC cases (Liu & Wu, 2010). Aflatoxin is a kind of mycotoxins produced by the mold Aspergillus flavus, which can be found in legumes, corns, soybeans, rice, milk, and cheese throughout their lives. Aflatoxin B1 (AFB1) is regarded as a class I carcinogen by the World Health Organization (Henry et al, 2001; Lopez et al, 2002). Aflatoxin B1, the most commonly occurring and potent of the aflatoxins is associated with a specific AGG to AGT transversion mutation at codon 249 of the p53 gene in human HCC, providing mechanistic support to a causal link between exposure and disease (Goldman & Shields, 2003; Sugimura, 2000; Wild & Montesano, 2009). Prospective epidemiological studies have shown a more than multiplicative interaction between HBV and aflatoxins in terms of HCC risk. In this chapter, the available evidence for the mechanism of aflatoxin with HBV-infection in HCC will be reviewed.

AIM: To explore the role of high-mobility group box 1 (HMGB1) protein during liver fibrogenesis and investigate the functional effects of HMGB1 gene silencing in hepatic stellate cells (HSCs) using siRNA. METHODS: Hepatic fibrosis in rats was induced throu-gh serial subcutaneous injections of dimethylnitrosamine, and expression of HMGB1 was detected by immunohistochemistry. HMGB1 siRNAs were developed and transiently transfected into HSC-T6 cells using Lipofectamine 2000. HMGB1 expression was evaluated by real-time polymerase chain reaction (PCR) and Western blotting analysis. Expression of α-smooth muscle actin (α-SMA) and collagen types I and III was evaluated by real-time PCR. Cell proliferation and the cell cycle were determined using the methyl thiazolyl tetrazolium method. Finally, collagen content in HSC supernatant was evaluated by an enzyme-linked immunosorbent assay. RESULTS: The results showed that HMGB1 was upregulated during liver fibrosis and that its expression was closely correlated with the deposition of collagen. siRNA molecules were successfully transfected into HSCs and induced inhibition of HMGB1 expression in a time-dependent manner. Moreover, HMGB1 siRNA treatment inhibited synthesis of α-SMA and collagen types I and III in transfected HSCs. CONCLUSION: This study suggests a significant fun-ctional role for HMGB1 in the development of liver fibrosis. It also demonstrates that downregulation of HMGB1 expression might be a potential strategy to treat liver fibrosis.

AIM: To study the HER-2/neu protein expression and gene amplification in gastric carcinoma and their rela tion. METHODS: One hundred and forty-five formalin-fixed and paraffin- embedded tumor tissue samples from Chinese gastric carcinoma patients were studied with immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) methods. Clinicopathologic data about all patients were collected. RESULTS: The levels of HER-2 3+, HER-2 2+ and HER2 1+ were measurable in 6.9%, 8.3% and 17.2% of the samples, respectively. No HER-2 was stained in 67.6% of the samples. FISH showed that HER-2 gene was amplified in 18 samples, 10 HER-2 3+ samples, 5 HER-2 2+ samples, and 3 HER-2 1+ samples with IHC staining. HER-2 status was not correlated with the sex and age of patients, and tumor size, location or differentiation, but with the depth of invasion, TNM stage, lymph node and distant metastasis as well as histopathological classification of gastric cancer ( P < 0.05). CONCLUSION: All samples with IHC as HER-2 expression should be analyzed with FISH. Detection of HER-2 gene amplification can assess the malignant biological behaviors and prognosis of gastric cancer.

To investigate the anti-fibrogenesis property of intraportal vein small interfering RNA (siRNA) injection targeting connective tissue growth factor (CTGF) in a rat model of liver fibrosis induced by carbon tetrachloride (CCl4) and its effect on hepatic stellate cell (HSC) activation.24 male rats were randomly divided into four group. rats received CCl4 by subcutaneous injections every three days for 6 consecutive weeks, and meantime they also obtained either siRNA targeting CTGF (as CTGF siRNA group), saline (as model group) or a control siRNA (as control siRNA group) by intraportal vein injection to rats liver at the same approach. Other rats received saline intraportal vein injection for 6 weeks (as normal control group). The expressions of CTGF and a-SMA protein were detected by Western blot. Hepatic histology was evaluated by HE staining and Sirius red staining. The collagen staining areas were measured quantitatively using a computer-aided manipulator with slight modifications. The number of active HSC were evaluated by immunohistochemistry.Six weeks after CCl4 injection, prominent upregulations were observed in the expressions of CTGF and a-SMA protein in saline or control siRNA-treated rats livers. In rats with CTGF siRNA treatment, the protein expressions of CTGF and a-SMA in liver decreased by 95%+/-2% and 86%+/-11% (F=21.234 and 12.473, P0.01) respectively, the number of active HSC in liver decreased by 76%+/-9% (F=9.179, P0.01) as compared to the model group. The attenuation of liver fibrosis was also observed in rats with CTGF siRNA treatment.Intraportal vein siRNA injection targeting CTGF could significantly inhibit CTGF gene expression in rats, thereby attenuate liver fibrosis by decreasing the number of active HSCs.

To determine the effect of body mass index (BMI) at baseline and its change during follow-up on the incidence of non-alcoholic fatty liver diseases (NAFLD) in apparently healthy adults.Subjects free of previous liver injury, alcohol consumption of more than 140 g/week and hepatitis B virus infection were identified from employees of Shanghai BaoSteel Group who underwent voluntary medical checkups at a 2-year interval. The analyzed population consisted of 5402 non-drinking subjects (4633 men) of age 36.5+/-9.3 years (18-65 years), who had normal livers at baseline. Among them 327 subjects (6.05%) developed fatty liver in 2 years. Those who developed NAFLD showed advanced age (especially in females), elevated BMI, high levels of serum triglyceride and total cholesterol, high prevalence rates of obesity, hyperlipidemia, hypertension and hyperglyceridemia at baseline, more weight gain and increase of serum triglycerides during the 2-year period. The incidence of NAFLD increased significantly with the changes of BMI at baseline, from 1.4% in subjects with normal weight, 6.4% in overweight, 16.8% in obese patients to 24.5% in severe obesity (Chi2 test = 389.01, P = 0.000 in trend analysis). Logistic regression analysis showed a significant interaction occurred between NAFLD age, BMI and serum triglyceride at baseline, and subtle gain of BMI and triglyceride during follow-up.There is a high incidence of NAFLD in a Chinese workplace. Obesity and related metabolic disorders at baseline, and more weight gain and increased serum triglyceride during follow-up are important predictors for the development of NAFLD.