Cancer-associated fibroblasts (CAFs) contribute to tumor progression, and microRNAs (miRs) play an important role in regulating the tumor-promoting properties of CAFs. The objectives of this study were to clarify the specific miR expression profile in CAFs of hepatocellular carcinoma (HCC) and identify its target gene signatures. Small-RNA-sequencing data were generated from nine pairs of CAFs and para-cancer fibroblasts isolated from human HCC and para-tumor tissues, respectively. Bioinformatic analyses were performed to identify the HCC-CAF-specific miR expression profile and the target gene signatures of the deregulated miRs in CAFs. Clinical and immunological implications of the target gene signatures were evaluated in The Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA_LIHC) database using Cox regression and TIMER analysis. The expressions of hsa-miR-101-3p and hsa-miR-490-3p were significantly downregulated in HCC-CAFs. Their expression in HCC tissue gradually decreased as HCC stage progressed in the clinical staging analysis. Bioinformatic network analysis using miRWalks, miRDB, and miRTarBase databases pointed to TGFBR1 as a common target gene of hsa-miR-101-3p and hsa-miR-490-3p. TGFBR1 expression was negatively correlated with miR-101-3p and miR-490-3p expression in HCC tissues and was also decreased by ectopic miR-101-3p and miR-490-3p expression. HCC patients with TGFBR1 overexpression and downregulated hsa-miR-101-3p and hsa-miR-490-3p demonstrated a significantly poorer prognosis in TCGA_LIHC. TGFBR1 expression was positively correlated with the infiltration of myeloid-derived suppressor cells, regulatory T cells, and M2 macrophages in a TIMER analysis. In conclusion, hsa-miR-101-3p and hsa-miR-490-3p were substantially downregulated miRs in CAFs of HCC, and their common target gene was TGFBR1. The downregulation of hsa-miR-101-3p and hsa-miR-490-3p, as well as high TGFBR1 expression, was associated with poor clinical outcome in HCC patients. In addition, TGFBR1 expression was correlated with the infiltration of immunosuppressive immune cells. Full article

Cancer treatment evokes impediments to liver metabolism that culminate in fatty liver. This study determined hepatic fatty acid composition and expression of genes and mediators involved in lipid metabolism following chemotherapy treatment. Female rats bearing the Ward colon tumor were administered Irinotecan (CPT-11) +5-fluorouracil (5-FU) and maintained on a control diet or a diet containing eicosapentaenoic acid (EPA) + docosahexaenoic acid (DHA) (2.3 g/100 g fish oil). Healthy animals provided with a control diet served as a reference group. Livers were collected one week after chemotherapy. Triacylglycerol (TG), phospholipid (PL), ten lipid metabolism genes, leptin, and IL-4 were measured. Chemotherapy increased TG content and reduced EPA content in the liver. Expression of SCD1 was upregulated by chemotherapy, while dietary fish oil downregulated its expression. Dietary fish oil down-regulated expression of the fatty acid synthesis gene FASN, while restoring the long chain fatty acid converting genes FADS2 and ELOVL2, and genes involved in mitochondrial β-oxidation (CPT1α) and lipid transport (MTTP1), to values similar to reference animals. Neither leptin nor IL-4 were affected by chemotherapy or diet. Depletion of EPA is associated with pathways evoking enhanced TG accumulation in the liver. Restoring EPA through diet may pose a dietary strategy to attenuate chemotherapy-associated impediments in liver fatty acid metabolism. Full article

Microbiome alterations are emerging as one of the most important factors that influence the course of alcohol use disorder (AUD). Recent advances in bioinformatics enable more robust and accurate characterization of changes in the composition of the microbiome. In this study, our objective was to provide the most comprehensive and up-to-date evaluation of microbiome alterations associated with AUD and alcoholic liver disease (ALD). To achieve it, we have applied consistent, state of art bioinformatic workflow to raw reads from multiple 16S rRNA sequencing datasets. The study population consisted of 122 patients with AUD, 75 with ALD, 54 with non-alcoholic liver diseases, and 260 healthy controls. We have found several microbiome alterations that were consistent across multiple datasets. The most consistent changes included a significantly lower abundance of multiple butyrate-producing families, including Ruminococcaceae, Lachnospiraceae, and Oscillospiraceae in AUD compared to HC and further reduction of these families in ALD compared with AUD. Other important results include an increase in endotoxin-producing Proteobacteria in AUD, with the ALD group having the largest increase. All of these alterations can potentially contribute to increased intestinal permeability and inflammation associated with AUD and ALD. Full article

Epithelial–mesenchymal transition (EMT) has been implicated in cancer progression, invasion, and metastasis. We aimed to evaluate the correlations between clinicopathological characteristics and EMT markers in patients with hepatocellular carcinoma (HCC) who underwent surgical resection and to identify the key regulator in EMT process. Fresh-frozen HCC tissues and adjacent nontumor liver tissues from 30 patients who underwent surgical resection were provided by the Gachon University Gil Medical Center Bio Bank. Human HCC cell lines, Hep3B, SNU449, and Huh7 cells were transfected with Rac1 siRNA and exposed to hypoxic conditions. The combined EMT markers expression (down-expression of E-cadherin and overexpression of p21-activated kinases 1 (PAK1)/Snail) by Western blot in HCC tissues when compared to adjacent nontumor liver tissues was significantly associated with macrovascular invasion (p = 0.021), microvascular invasion (p = 0.001), large tumor size (p = 0.021), and advanced tumor stage (p = 0.015). Patients with combined EMT markers expression showed early recurrence and poor overall survival. In vitro studies showed that Rac1 knockdown decreased the expression of EMT markers including PAK1 and Snail in hypoxia-induced Hep3B cells and suppressed the migration and invasion of hypoxia-induced HCC cells. Rac1 may be a potential therapeutic target for inhibition of EMT process through the inhibition of PAK1 and Snail in HCC. Full article

Hepatocellular carcinoma (HCC) is a common type of liver cancer and one of the highly lethal diseases worldwide. Hypoxia plays an important role in the development and prognosis of HCC. This study aimed to construct a new hypoxia-related prognosis signature and investigate its potential ceRNA axes in HCC. RNA profiles and hypoxia genes were downloaded, respectively, from the Cancer Genome Atlas hepatocellular carcinoma database and Gene Set Enrichment Analysis website. Cox regression analyses were performed to select the prognostic genes and construct the risk model. The ENCORI database was applied to build the lncRNA-miRNA–mRNA prognosis-related network. The TIMER and CellMiner databases were employed to analyze the association of gene expression in ceRNA with immune infiltration and drug sensitivity, respectively. Finally, the co-expression analysis was carried out to construct the potential lncRNA/miRNA/mRNA regulatory axes. We obtained a prognostic signature including eight hypoxia genes (ENO2, KDELR3, PFKP, SLC2A1, PGF, PPFIA4, SAP30, and TKTL1) and further established a hypoxia-related prognostic ceRNA network including 17 lncRNAs, six miRNAs, and seven mRNAs for hepatocellular carcinoma. Then, the analysis of immune infiltration and drug sensitivity showed that gene expression in the ceRNA network was significantly correlated with the infiltration abundance of multiple immune cells, the expression level of immune checkpoints, and drug sensitivity. Finally, we identified three ceRNA regulatory axes (SNHG1/miR-101-3p/PPFIA4, SNHG1/miR-101-3p/SAP30, and SNHG1/miR-101-3p/TKTL1) associated with the progression of HCC under hypoxia. Here, we constructed a prognosis gene signature and a ceRNA network related to hypoxia for hepatocellular carcinoma. Among the ceRNA network, six highly expressed lncRNAs (AC005540.1, AC012146.1, AC073529.1, AC090772.3, AC138150.2, AL390728.6) and one highly expressed mRNA (PPFIA4) were the potential biomarkers of hepatocellular carcinoma which we firstly reported. The three predicted hypoxia-related regulatory axes may play a vital role in the progression of hepatocellular carcinoma. Full article

Clinical phenotypes of familial hypobetalipoproteinemia (FHBL) are related to a number of defective apolipoprotein B (APOB) alleles. Fatty liver disease is a typical manifestation, but serious neurological symptoms can appear. In this study, genetic analysis of the APOB gene and ophthalmological diagnostics were performed for family members with FHBL. Five relatives with FHBL, including a proband who developed neurological disorders, were examined. A sequencing analysis of the whole coding region of the APOB gene, including flanking intronic regions, was performed using the next-generation sequencing (NGS) method. Electrophysiological ophthalmological examinations were also done. In the proband and his affected relatives, NGS identified the presence of the pathogenic, rare heterozygous splicing variant c.3696+1G>T. Two known heterozygous missense variants—c.2188G>A, p.(Val730Ile) and c.8353A>C, p.(Asn2785His)—in the APOB gene were also detected. In all patients, many ophthalmologic abnormalities in electrophysiological tests were also found. The identified splicing variant c.3696+1G>T can be associated with observed autosomal, dominant FHBL with coexisting neurological symptoms, and both identified missense variants could be excluded as the main cause of observed clinical signs, according to mutation databases and the literature. Electroretinography examination is a sensitive method for the detection of early neuropathy and should therefore be recommended for the care of patients with FHBL. Full article

Nonalcoholic steatohepatitis (NASH) is becoming a public health problem worldwide. While the number of research studies on NASH progression rises every year, sometime their findings are controversial. To identify the most important and commonly described findings related to NASH progression, we used an original bioinformatics, integrative, text-mining approach that combines PubMed database querying and available gene expression omnibus dataset. We have identified a signature of 25 genes that are commonly found to be dysregulated during steatosis progression to NASH and cancer. These genes are implicated in lipid metabolism, insulin resistance, inflammation, and cancer. They are functionally connected, forming the basis necessary for steatosis progression to NASH and further progression to hepatocellular carcinoma (HCC). We also show that five of the identified genes have genome alterations present in HCC patients. The patients with these genes associated to genome alteration are associated with a poor prognosis. In conclusion, using an integrative literature- and data-mining approach, we have identified and described a canonical pathway underlying progression of NASH. Other parameters (e.g., polymorphisms) can be added to this pathway that also contribute to the progression of the disease to cancer. This work improved our understanding of the molecular basis of NASH progression and will help to develop new therapeutic approaches. Full article

Circulating microparticles (MPs) are novel potential biomarkers in cancer patients. Their role in hepatocellular carcinoma (HCC) is under intensive investigation. In this study, we tested the hypothesis that MPs expressing the antigen HepPar1 are increased in the blood of subjects with HCC and may serve as markers of early recurrence after liver resection (LR). We studied 15 patients affected by HCC undergoing LR, and used flow cytometry to assess the number of circulating HepPar1+ MPs. Ten subjects without HCC (five with liver cirrhosis and five with healthy livers) were used as controls. After LR, HCC patients underwent a follow-up to check for early recurrence, which occurred in seven cases. The number of circulating HepPar1+ MPs was significantly higher in subjects affected by HCC, compared to individuals without cancer (p < 0.01). We also found that, among HCC patients, the number of circulating HepPar1+ MPs, measured before LR, was significantly higher in those who displayed early recurrence compared to those without recurrence (p = 0.02). Of note, other types of circulating MPs, such as those derived from endothelial cells (CD144+) or those produced by the activated endothelium (CD144+/CD62+), were not associated with HCC, nor could they predict HCC recurrence. HepPar1+ MPs deserve further investigation as novel biomarkers of disease and prognosis in HCC patients. Full article

Primary biliary cholangitis (PBC) is an immune-mediated cholestatic disease. Vitamin D receptor (VDR)-dependent signaling constrains an inflammatory response by targeting the miRNA155-SOCS1 (suppressor of cytokine signaling 1) axis. The VDR-miRNA155-SOCS1 pathway was investigated in the context of the autoimmune response associated with PBC. Human liver tissues from non-cirrhotic PBC (n = 22), cirrhotic PBC (n = 22), cirrhotic primary sclerosing cholangitis (PSC, n=13), controls (n = 23), and peripheral blood mononuclear cells (PBMC) obtained from PBC (n = 16) and PSC (n = 10) patients and healthy subjects (n = 11) were used for molecular analyses. VDR mRNA and protein expressions were substantially reduced in PBC livers (51% and 59%, respectively). Correspondingly, the decrease of SOCS1 protein expression in PBC livers, after normalization to a marker of lymphocytes and forkhead family transcriptional regulator box P3 (FOXP3, marker of Treg), was observed, and this phenomenon was accompanied by enhanced miRNA155 expression. In PSC livers, protein expressions of VDR and SOCS1 were comparable to the controls. However, in PBM cells, protein expressions of VDR and SOCS1 were considerably decreased in both PBC and PSC. We demonstrated that VDR/miRNA155-modulated SOCS1 expression is decreased in PBC which may lead to insufficient negative regulation of cytokine signaling. These findings suggest that the decreased VDR signaling in PBC could be of importance in the pathogenesis of PBC. Full article

Ischemia/reperfusion injury (IRI) occurs inevitably in liver transplantations and frequently during major resections, and can lead to liver dysfunction as well as systemic disorders. High-mobility group box 1 (HMGB1) plays a pathogenic role in hepatic IRI. In the normal liver, HMGB1 is located in the nucleus of hepatocytes; after ischemia reperfusion, it translocates to the cytoplasm and it is further released to the extracellular space. Unlike the well-explored functions of nuclear and extracellular HMGB1, the role of cytoplasmic HMGB1 in hepatic IRI remains elusive. We hypothesized that cytoplasmic HMGB1 interacts with binding proteins involved in the hepatocellular response to IRI. In this study, binding proteins of cytoplasmic HMGB1 during hepatic IRI were identified. Liver tissues from rats with warm ischemia reperfusion (WI/R) injury and from normal rats were subjected to cytoplasmic protein extraction. Co-immunoprecipitation using these protein extracts was performed to enrich HMGB1-protein complexes. To separate and identify the immunoprecipitated proteins in eluates, 2-dimensional electrophoresis and subsequent mass spectrometry detection were performed. Two of the identified proteins were verified using Western blotting: betaine–homocysteine S-methyltransferase 1 (BHMT) and cystathionine γ-lyase (CTH). Therefore, our results revealed the binding of HMGB1 to BHMT and CTH in cytoplasm during hepatic WI/R. This finding may help to better understand the cellular response to IRI in the liver and to identify novel molecular targets for reducing ischemic injury. Full article

Autophagy, an evolutionary conserved multifaceted lysosome-mediated bulk degradation system, plays a vital role in liver pathologies including hepatocellular carcinoma (HCC). Post-translational modifications (PTMs) and genetic variations in autophagy components have emerged as significant determinants of autophagy related proteins. Identification of a comprehensive spectrum of genetic variations and PTMs of autophagy related proteins and their impact at molecular level will greatly expand our understanding of autophagy based regulation. In this study, we attempted to identify high risk missense mutations that are highly damaging to the structure as well as function of autophagy related proteins including LC3A, LC3B, BECN1 and SCD1. Number of putative structural and functional residues, including several sites that undergo PTMs were also identified. In total, 16 high-risk SNPs in LC3A, 18 in LC3B, 40 in BECN1 and 43 in SCD1 were prioritized. Out of these, 2 in LC3A (K49A, K51A), 1 in LC3B (S92C), 6 in BECN1 (S113R, R292C, R292H, Y338C, S346Y, Y352H) and 6 in SCD1 (Y41C, Y55D, R131W, R135Q, R135W, Y151C) coincide with potential PTM sites. Our integrated analysis found LC3B Y113C, BECN1 I403T, SCD1 R126S and SCD1 Y218C as highly deleterious HCC-associated mutations. This study is the first extensive in silico mutational analysis of the LC3A, LC3B, BECN1 and SCD1 proteins. We hope that the observed results will be a valuable resource for in-depth mechanistic insight into future investigations of pathological missense SNPs using an integrated computational platform. Full article

Hepatocellular carcinoma (HCC) is currently the third most common cause of cancer-related death in the Asia-Pacific region. Our previous work showed that knockdown of CD98 significantly inhibits malignant HCC cell phenotypes in vitro and in vivo. The level of CD98 in the membrane is tightly regulated to mediate complex processes associated with cell–cell communication and intracellular signaling. In addition, the intracellular domain of CD98 (CD98-ICD) seems to be of vital importance for recycling CD98 to the membrane after it is endocytosed. The intracellular and transmembrane domains of CD98 associate with β-integrins (primarily β1 but also β3), and this association is essential for CD98 mediation of integrin-like signaling and complements dominant suppression of β1-integrin. We speculated that isolated CD98-ICD would similarly suppress β1-integrin activation and inhibit the malignant behaviors of cancer cells. In particular, the exact role of CD98-ICD has not been studied independently in HCC. In this study, we found that ectopic expression of CD98-ICD inhibited the malignant phenotypes of HCC cells, and the mechanism possibly involves β1-integrin suppression. Moreover, the expression levels of CD98, β1-integrin-A (the activated form of β1-integrin) and Ki-67 were significantly increased in HCC tissues relative to those of normal liver tissues. Therefore, our preliminary study indicates that ectopic CD98-ICD has an inhibitory role in the malignant development of HCC, and shows that CD98-ICD acts as a dominant negative mutant of CD98 that attenuates β1-integrin activation. CD98-ICD may emerge as a promising candidate for antitumor treatment. Full article

Current evidence suggests that gut dysbiosis drives obesity and non-alcoholic fatty liver disease (NAFLD) pathogenesis. Toll-like receptor 2 (TLR2) and TLR6 specifically recognize components of Gram-positive bacteria. Despite the potential implications of TLR2 in NAFLD pathogenesis, the role of TLR6 has not been addressed. Our aim is to study a potential role of TLR6 in obesity-related NAFLD. Forty morbidly obese patients undergoing bariatric surgery were prospectively studied. Cell surface expression of TLR2 and TLR6 was assessed on peripheral blood mononuclear cells (PBMCs) by flow cytometry. Freshly isolated monocytes were cultured with specific TLR2/TLR6 agonists and intracellular production of cytokines was determined by flow-cytometry. In liver biopsies, the expression of TLR2 and TLR6 was analyzed by immunohistochemistry and cytokine gene expression using RT-qPCR. TLR6 expression in PBMCs from non-alcoholic steatohepatitis (NASH) patients was significantly higher when compared to those from simple steatosis. The production of pro-inflammatory cytokines in response to TLR2/TLR6 stimulation was also significantly higher in patients with lobular inflammation. Hepatocyte expression of TLR6 but not that of TLR2 was increased in NAFLD patients compared to normal liver histology. Deregulated expression and activity of peripheral TLR6 in morbidly obese patients can mirror the liver inflammatory events that are well known drivers of obesity-related NASH pathogenesis. Moreover, TLR6 is also significantly overexpressed in the hepatocytes of NAFLD patients compared to their normal counterparts. Thus, deregulated TLR6 expression may potentiate TLR2-mediated liver inflammation in NAFLD pathogenesis, and also serve as a potential peripheral biomarker of obesity-related NASH. Full article

Forkhead box P1 (FOXP1) belongs to a family of winged-helix transcription factors that are involved in the processes of cellular proliferation, differentiation, metabolism, and longevity. FOXP1 can affect cell proliferation and migratory ability in hepatocellular carcinoma (HCC) in vitro. However, little is known about the mechanism of FOXP1 in the proliferation of HCC cells. This study aimed to further explore the function of FOXP1 on the proliferation of HCC cells as well as the relevant mechanism involved. Western blot analysis, tumor xenograft models, and flow cytometry analysis were performed to elucidate the function of FOXP1 in the regulation of cell proliferation in human HCC. We observed that silencing FOXP1 significantly suppressed the growth ability of HCC cells both in vitro and in vivo. In addition, knockdown of FOXP1 induced G1/S phase arrest, and the expression of total and phosphorylated Rb (active type) as well as the levels of E2F1 were markedly decreased at 24 h; however, other proteins, including cyclin-dependent kinase (CDK) 4 and 6 and cyclin D1 did not show noticeable changes. In conclusion, downregulation of FOXP1 inhibits cell proliferation in hepatocellular carcinoma by inducing G1/S phase cell cycle arrest, and the decrease in phosphorylated Rb is the main contributor to this G1/S phase arrest. Full article

Interleukin enhancer binding factor 2 (ILF2), a transcription factor, regulates cell growth by inhibiting the stabilization of mRNA. Currently, its role has gained recognition as a factor in the tumorigenic process. However, until now, little has been known about the detailed role ILF2 plays in hepatocellular carcinoma (HCC). In this study, we investigated the expression levels of ILF2 in HCC tissue with Western blot and immunohistochemical assays. To examine the effect of ILF2 on liver cancer cell growth and apoptosis, small interfering RNAs (siRNAs) targeting ILF2 were recombined to create lentiviral overexpression vectors. Our results showed higher expression levels of ILF2 mRNA and ILF2 protein in HCC tissue compared with matched peritumoral tissue. Expression of ILF2 may regulate cell growth and apoptosis in liver cancer cells via regulation of B-cell lymphoma 2 (Bcl-2), Bcl-2 related ovarian killer (Bok), Bcl-2-associated X protein (BAX), and cellular inhibitor of apoptosis 1 (cIAP1). Moreover, we inoculated nude mice with liver cancer cells to investigate the effect of ILF2 on tumorigenesis in vivo. As expected, a rapid growth was observed in cancer cells inoculated with a lentiviral vector coding Flag-ILF2 (Lenti-ILF2) compared with the control cells. Hence, these results promote a better understanding of ILF2’s potential role as a therapeutic target in HCC. Full article

The chemokine-like receptor 1 (CMKLR1) ligands resolvin E1 and chemerin are known to modulate inflammatory response. The progression of non-alcoholic fatty liver disease (NAFLD) to non-alcoholic steatohepatitis (NASH) is associated with inflammation. Here it was analyzed whether hepatic CMKLR1 expression is related to histological features of NASH. Therefore, CMKLR1 mRNA was quantified in liver tissue of 33 patients without NAFLD, 47 patients with borderline NASH and 38 patients with NASH. Hepatic CMKLR1 mRNA was not associated with gender and body mass index (BMI) in the controls and the whole study group. CMKLR1 expression was similar in controls and in patients with borderline NASH and NASH. In male patients weak positive correlations with inflammation, fibrosis and NASH score were identified. In females CMKLR1 was not associated with features of NAFLD. Liver CMKLR1 mRNA tended to be higher in type 2 diabetes patients of both genders and in hypercholesterolemic women. In summary, this study shows that hepatic CMKLR1 mRNA is weakly associated with features of NASH in male patients only. Full article

Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-activated nuclear receptor that regulates cellular lipid and glucose metabolism and also plays an inhibitory role in various cancers. However, the role of PPARγ in hepatocellular carcinoma (HCC) remains controversial. This study aimed to investigate the prognostic value of PPARγ in HCC and its role in inhibiting tumor progression, namely, HCC cell growth, migration, and angiogenesis. Immunohistochemical PPARγ staining was examined in 83 HCC specimens to investigate the clinicopathological correlations between PPARγ expression and various parameters. The functional role of PPARγ was determined via PPARγ overexpression and knockdown in HCC cells. Patients with low HCC tissue PPARγ expression were significantly younger (p = 0.006), and exhibited more tumor numbers (p = 0.038), more macroscopic vascular invasion (MVI) (p = 0.008), and more advanced TNM (size of primary tumor, number of regional lymph nodes, and distant metastasis) stages at diagnosis (p = 0.013) than patients with high HCC tissue PPARγ expression. PPARγ knockdown increased HCC cell growth, migration, and angiogenesis, while PPARγ overexpression reduced HCC cell growth, migration, and angiogenesis. These results suggest that low PPARγ expression is an independent predictor of more MVI in HCC patients. PPARγ contributes to the suppression of HCC cell growth, migration, and angiogenesis. Therefore, PPARγ may be a therapeutic target in HCC patients. Full article

In chronic liver diseases, regardless of their etiology, the development of fibrosis is the first step toward the progression to cirrhosis, portal hypertension, and hepatocellular carcinoma. Hepatic stellate cells (HSCs) are the main profibrogenic cells that promote the pathogenesis of liver fibrosis, and so it is important to identify the molecules that regulate HSCs activation and liver fibrosis. Niemann-Pick type C2 (NPC2) protein plays an important role in the regulation of intracellular cholesterol homeostasis by directly binding with free cholesterol. However, the roles of NPC2 in HSCs activation and liver fibrosis have not been explored in detail. Since a high-cholesterol diet exacerbates liver fibrosis progression in both rodents and humans, we propose that the expression of NPC2 affects free cholesterol metabolism and regulates HSCs activation. In this study, we found that NPC2 is decreased in both thioacetamide- and carbon tetrachloride-induced liver fibrosis tissues. In addition, NPC2 is expressed in quiescent HSCs, but its activation status is down-regulated. Knockdown of NPC2 in HSC-T6 cells resulted in marked increases in transforming growth factor-β1 (TGF-β1)-induced collagen type 1 α1 (Col1a1), α-smooth muscle actin (α-SMA) expression, and Smad2 phosphorylation. In contrast, NPC2 overexpression decreased TGF-β1-induced HSCs activation. We further demonstrated that NPC2 deficiency significantly increased the accumulation of free cholesterol in HSCs, increasing Col1a1 and α-SMA expression and activating Smad2, and leading to sensitization of HSCs to TGF-β1 activation. In contrast, overexpression of NPC2 decreased U18666A-induced free cholesterol accumulation and inhibited the subsequent HSCs activation. In conclusion, our study has demonstrated that NPC2 plays an important role in HSCs activation by regulating the accumulation of free cholesterol. NPC2 overexpression may thus represent a new treatment strategy for liver fibrosis. Full article

Toll-like receptor 4 (TLR4)/nuclear factor kappa B (NF-κB) signaling plays a dominant role in the pathogenesis of liver ischemia-reperfusion (IR) injury. Dexmedetomidine (Dex) protects the liver against IR injury via α₂-adrenoceptor activation, but the contribution of TLR4 signaling remains unknown. The authors aimed to examine whether pretreatment with Dex produces hepatic protection and investigate the influence of Dex on TLR4/NF-κB signaling. Dex was given via intraperitoneal injection 30 min prior to orthotopic autologous liver transplantation (OALT) in rats, and three α₂-adrenoceptor antagonists including atipamezole (a nonselective α₂ receptor blocker), ARC-239 (a specific α_2B/C blocker) and BRL-44408 (a specific α_2A blocker) were injected intraperitoneally 10 min before Dex administration. Histopathologic evaluation of the liver and the measurement of serum alanine aminotransferase activity, TLR4/NF-κB expression in the liver, and pro-inflammatory factors (serum tumor necrosis factor-α, interleukin-1β and hepatic myeloperoxidase) concentrations were performed 8 h after OALT. Dex ameliorated liver injury after OALT probably by suppressing the TLR4/NF-κB pathway and decreasing inflammatory mediator levels. The protective effects of Dex were reversed by atipamezole and BRL-44408, but not by ARC-239, suggesting that these effects were mediated in part by the α_2A subtype. In conclusion, Dex attenuates liver injury partly via the α_2A-adrenoceptor subtype, and the mechanism is due to the suppression of the TLR4/NF-κB pathway. Full article

Hepatitis C virus (HCV) substantially affects lipid metabolism, and remodeling of sphingolipids appears to be essential for HCV persistence in vitro. The aim of the current study is the evaluation of serum sphingolipid variations during acute HCV infection. We enrolled prospectively 60 consecutive patients with acute HCV infection, most of them already infected with human immunodeficiency virus (HIV), and serum was collected at the time of diagnosis and longitudinally over a six-month period until initiation of antiviral therapy or confirmed spontaneous clearance. Quantification of serum sphingolipids was performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Spontaneous clearance was observed in 11 out of 60 patients (18.3%), a sustained viral response (SVR) in 43 out of 45 patients (95.5%) receiving an antiviral treatment after follow-up, whereas persistence of HCV occurred in six out of 60 patients (10%). C24-ceramide (C24-Cer)-levels increased at follow-up in patients with spontaneous HCV eradication (p < 0.01), as compared to baseline. Sphingosine and sphinganine values were significantly upregulated in patients unable to clear HCV over time compared to patients with spontaneous clearance of HCV infection on follow-up (p = 0.013 and 0.006, respectively). In summary, the persistence of HCV after acute infection induces a downregulation of C24Cer and a simultaneous elevation of serum sphingosine and sphinganine concentrations. Full article

Transmembrane 4 superfamily member 1 (TM4SF1) is a member of tetraspanin family, which mediates signal transduction events regulating cell development, activation, growth and motility. Our previous studies showed that TM4SF1 is highly expressed in liver cancer. HepG2 cells were transfected with TM4SFl siRNA and TM4SF1-expressing plasmids and their biological functions were analyzed in vitro and in vivo. HepG2 cells overexpressing TM4SF1 showed reduced apoptosis and increased cell migration in vitro and enhanced tumor growth and metastasis in vivo, whereas siRNA-mediated silencing of TM4SF1 had the opposite effect. TM4SF1 exerts its effect by regulating a few apoptosis- and migration-related genes including caspase-3, caspase-9, MMP-2, MMP-9 and VEGF. These results indicate that TM4SF1 is associated with liver tumor growth and progression, suggesting that TM4SF1 may be a potential target for treatment of liver cancer in future. Full article

Background: Heptocelluar carcinoma (HCC) is insensitive to chemotherapy due to limited bioavailability and acquired drug resistance. Smad3 plays dual roles by inhibiting cell growth initially and promoting the progression of advanced tumors in HCC. However, the role of smad3 in chemosensitivity of HCC remains elusive. Methods: The role of smad3 in chemosensitivity of HCC was measured by cell viability, apoptosis, plate colony formation assays and xenograft tumor models. Non-smad signaling was detected by Western blotting to search for the underlying mechanisms. Results: Smad3 enhanced the chemosensitivity of HCC cells to cisplatin. Smad3 upregulated p21^Waf1/Cip1 and downregulated c-myc and bcl2 with the treatment of cisplatin. Moreover, overexpression of smad3 repressed the phosphorylation of AKT, and vice versa. Inhibition of PI3K/AKT pathway by LY294002 restored chemosensitivity of smad3-deficiency cells to cisplatin in HCC. Conclusion: Smad3 sensitizes HCC cells to the effects of cisplatin by repressing phosphorylation of AKT and combination of inhibitor of AKT pathway and conventional chemotherapy may be a potential way to solve drug resistance in HCC. Full article

Previous studies have found higher levels of serum malondialdehyde (MDA) in hepatocellular carcinoma (HCC) patients compared to healthy controls and higher MDA concentrations in tumoral tissue of HCC patients than in non-tumoral tissue. However, the association between pre-transplant serum levels of MDA and survival in HCC patients after liver transplantation (LT) has not been described, and the aim of the present study was to determine whether such an association exists. In this observational study we measured serum MDA levels in 127 patients before LT. We found higher pre-LT serum MDA levels in 15 non-surviving than in 112 surviving patients one year after LT (p = 0.02). Exact binary logistic regression analysis revealed that pre-LT serum levels of MDA over 3.37 nmol/mL were associated with mortality after one year of LT (Odds ratio = 5.38; 95% confidence interval (CI) = from 1.580 to infinite; p = 0.007) adjusting for age of the deceased donor. The main finding of our study was that there is an association between serum MDA levels before LT for HCC and 1-year survival after LT. Full article

Liver progenitor cells (LPCs) are activated in chronic liver damage and may contribute to liver fibrosis. Our previous investigation reported that LPCs produced connective tissue growth factor (CTGF/CCN2), an inducer of liver fibrosis, yet the regulatory mechanism of the production of CTGF/CCN2 in LPCs remains elusive. In this study, we report that Activin A is an inducer of CTGF/CCN2 in LPCs. Here we show that expression of both Activin A and CTGF/CCN2 were upregulated in the cirrhotic liver, and the expression of Activin A positively correlates with that of CTGF/CCN2 in liver tissues. We go on to show that Activin A induced de novo synthesis of CTGF/CCN2 in LPC cell lines LE/6 and WB-F344. Furthermore, Activin A contributed to autonomous production of CTGF/CCN2 in liver progenitor cells (LPCs) via activation of the Smad signaling pathway. Smad2, 3 and 4 were all required for this induction. Collectively, these results provide evidence for the fibrotic role of LPCs in the liver and suggest that the Activin A-Smad-CTGF/CCN2 signaling in LPCs may be a therapeutic target of liver fibrosis. Full article

Hepatocellular carcinoma (HCC) is one of the most commonly diagnosed cancers worldwide. Chemoprevention of HCC can be achieved through the use of natural or synthetic compounds that reverse, suppress or prevent the development of cancer progression. In this study, we investigated the antiproliferative effects and the mechanism of action of two compounds, 2,3,4′-trimethoxy-2′-hydroxy-chalcone (CH1) and 3′-bromo-3,4-dimethoxy-chalcone (CH2), over human hepatoma cells (HepG2 and Huh-7) and cultured mouse hepatocytes (HepM). Cytotoxic effects were observed over the HepG2 and Huh-7, and no effects were observed over the HepM. For HepG2 cells, treated separately with each chalcone, typical apoptotic laddering and nuclear condensation were observed. Additionally, the caspases and Bcl-2 family proteins activation by using Western blotting and immunocytochemistry were studied. Caspase-8 was not activated, but caspase-3 and -9 were both activated by chalcones in HepG2 cells. Chalcones also induced reactive oxygen species (ROS) accumulation after 4, 8 and 24 h of treatment in HepG2 cells. These results suggest that apoptosis in HepG2 was induced through: (i) a caspase-dependent intrinsic pathway; and (ii) by alterations in the cellular levels of Bcl-2 family proteins, and also, that the chalcone moiety could be a potent candidate as novel anticancer agents acting on human hepatomas. Full article

Ghrelin is a stomach-derived growth hormone secretagogue that promotes various physiological effects, including energy metabolism and amelioration of inflammation. The purpose of this study was to investigate the protective mechanism of ghrelin against liver fibrosis. Liver fibrosis was induced in C57BL/6 mice by intraperitoneal injection of CCl₄ (2.0 mL/kg of 10% CCl₄ v/v solution in peanut oil) two times per week for eight weeks. Ghrelin (10 μg/kg) was intraperitoneally injected two times per week for eight weeks. A second murine liver fibrosis model was induced by bile duct ligation (BDL) and concurrent ghrelin administration for four weeks. Hematoxylin eosin (H&E), and Masson’s trichrome were used to detect pathological changes to liver tissue. Western blotting was used to detect protein levels of transforming growth factor (TGF)-β1, phosphorylated Smad3 (p-Smad3), I-collage, α-smooth muscle actin (α-SMA), matrix metalloproteinases (MMPs) 2, tissue inhibitor of matrix metalloproteinases (TIMPs) 1, phosphorylated NF-κB (p-NF-κB), and microtubule-associated protein light chain 3 (LC3). In addition, qRT-PCR was used to detect mRNA levels of TGF-β1, I-collage, α-SMA, MMP2, TIMP1 and LC3, while levels of TGF-β1, p-Smad3, I-collage, α-SMA, and LC3 were detected immunohistochemically. Levels of aspartate aminotransferase and alanine aminotransferase were significantly decreased by ghrelin treatment. Ghrelin administration also significantly reduced the extent of pathological changes in both murine liver fibrosis models. Expression levels of I-collage and α-SMA in both models were clearly reduced by ghrelin administration. Furthermore, ghrelin treatment decreased protein expression of TGF-β1 and p-Smad3. The protein levels of NF-κB and LC3 were increased in the CCl₄- and BDL-treatment groups but were significantly reduced following ghrelin treatment. In addition, ghrelin inhibited extracellular matrix formation by decreasing NF-κB expression and maintaining the balance between MMP2 and TIMP1. Our results demonstrated that ghrelin attenuates liver fibrosis via inhibition of the TGF-β1/Smad3 and NF-κB signaling pathways, as well as autophagy suppression. Full article

The study of bone morphogenetic proteins (BMPs) role in tumorigenic processes, and specifically in the liver, has gathered importance in the last few years. Previous studies have shown that BMP9 is overexpressed in about 40% of hepatocellular carcinoma (HCC) patients. In vitro data have also shown evidence that BMP9 has a pro-tumorigenic action, not only by inducing epithelial to mesenchymal transition (EMT) and migration, but also by promoting proliferation and survival in liver cancer cells. However, the precise mechanisms driving these effects have not yet been established. In the present work, we deepened our studies into the intracellular mechanisms implicated in the BMP9 proliferative and pro-survival effect on liver tumor cells. In HepG2 cells, BMP9 induces both Smad and non-Smad signaling cascades, specifically PI3K/AKT and p38MAPK. However, only the p38MAPK pathway contributes to the BMP9 growth-promoting effect on these cells. Using genetic and pharmacological approaches, we demonstrate that p38MAPK activation, although dispensable for the BMP9 proliferative activity, is required for the BMP9 protective effect on serum withdrawal-induced apoptosis. These findings contribute to a better understanding of the signaling pathways involved in the BMP9 pro-tumorigenic role in liver tumor cells. Full article

Proteomic analyses have already been used in a number of hepatological studies and provide important information. However, few reports have focused on changes in the cytoplasmic proteome. The present study therefore aimed to evaluate changes in cytoplasmic proteome of rats in response to alcoholic hepatotoxicity. Rats were fed a Liber-DeCarli liquid diet containing ethanol for four weeks. Cytoplasmic proteins except mitochondrial proteins from the livers of these animals were investigated using two-dimensional gel electrophoresis and mass spectrometry. Alcohol induced a decrease in body weight gain and an increase in alanine transaminase (ALT), cholesterol, and phospholipid levels. Histopathological observations revealed hepatic damage characterized by necrosis and fatty change in alcohol-treated group at week 2, which continues until week 4. Our proteomic analysis revealed that 25 proteins were differentially expressed in the ethanol-fed group. Of these, 12 cytoplasmic proteins are being reported for the first time. Taken together, our results provide further insights into the disease mechanism and therapeutic information of alcoholic liver disease. Full article

Hepatocyte-specific Phosphatase and tensin homolog (Pten)-knockout (KO) mice exhibit hepatic lesions analogous to non-alcoholic steatohepatitis (NASH). 1,8-cineole is a monoterpene oxide and it has several biological effects including hepatoprotective effects. In this study we revealed that 1,8-cineole ameliorates NASH of Pten KO mice. Pten KO mice were assigned to a control group without any medication or to a 1,8-cineole group injected with 50 mg/kg i.p. twice per week for eight weeks. At eight weeks, livers from each group were processed to measure triglyceride (TG) content, gene expression analysis, western blot analysis, and histological examination including Oil red O staining. 1,8-cineole ameliorated hepatic steatosis in Pten KO mice, revealed by TG content and Oil red O staining. Moreover, 1,8-cineole downregulated collagen 1a1 expression and improved liver fibrosis. Thus, 1,8-cineole has potential as a candidate to treat NASH by inactivating the Akt/PI3-kinase pathway. Full article

Invasion and metastasis of hepatocellular carcinoma (HCC) is a major cause for lethal liver cancer. Signaling pathways associated with cancer progression are frequently reconfigured by aberrant phosphorylation of key proteins. To capture the key phosphorylation events in HCC metastasis, we established a methodology by an off-line high-pH HPLC separation strategy combined with multi-step IMAC and LC–MS/MS to study the phosphoproteome of a metastatic HCC cell line, MHCC97-H (high metastasis). In total, 6593 phosphopeptides with 6420 phosphorylation sites (p-sites) of 2930 phosphoproteins were identified. Statistical analysis of gene ontology (GO) categories for the identified phosphoproteins showed that several of the biological processes, such as transcriptional regulation, mRNA processing and RNA splicing, were over-represented. Further analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations demonstrated that phosphoproteins in multiple pathways, such as spliceosome, the insulin signaling pathway and the cell cycle, were significantly enriched. In particular, we compared our dataset with a previously published phosphoproteome in a normal liver sample, and the results revealed that a number of proteins in the spliceosome pathway, such as U2 small nuclear RNA Auxiliary Factor 2 (U2AF2), Eukaryotic Initiation Factor 4A-III (EIF4A3), Cell Division Cycle 5-Like (CDC5L) and Survival Motor Neuron Domain Containing 1 (SMNDC1), were exclusively identified as phosphoproteins only in the MHCC97-H cell line. These results indicated that the phosphorylation of spliceosome proteins may participate in the metastasis of HCC by regulating mRNA processing and RNA splicing. Full article

The cAMP-regulated phosphoprotein 19 (ARPP-19) plays a key role in cell mitotic G2/M transition. Expression of ARPP-19 was increased in human hepatocellular carcinoma (HCC) compared to adjacent non-tumorous liver tissues in 36 paired liver samples, and the level of ARPP-19 in HCC tissues was positively correlated with the tumor size. To determine the interrelationship between ARPP-19 expression and HCC, we silenced ARPP-19 expression in the human hepatocarcinoma HepG2 and SMMC-7721 cells using lentivirus encoding ARPP-19 siRNA. HepG2 and SMMC-7721 cells with ARPP-19 knockdown displayed lowered cell growth rate, retarded colony formation and increased arrest at the G2/M phase transition. Silencing ARPP-19 in HCC cells resulted in decreased protein levels of phospho-(Ser) CDKs substrates and increased levels of inactivated cyclin division cycle 2 (Cdc2). Therefore, ARPP-19 may play a role in HCC pathogenesis through regulating cell proliferation. Full article

Lipid accumulation in the human liver seems to be a crucial mechanism in the pathogenesis and the progression of non-alcoholic fatty liver disease (NAFLD). We aimed to evaluate gene expression of different fatty acid (FA) metabolism-related genes in morbidly obese (MO) women with NAFLD. Liver expression of key genes related to de novo FA synthesis (LXRα, SREBP1c, ACC1, FAS), FA uptake and transport (PPARγ, CD36, FABP4), FA oxidation (PPARα), and inflammation (IL6, TNFα, CRP, PPARδ) were assessed by RT-qPCR in 127 MO women with normal liver histology (NL, n = 13), simple steatosis (SS, n = 47) and non-alcoholic steatohepatitis (NASH, n = 67). Liver FAS mRNA expression was significantly higher in MO NAFLD women with both SS and NASH compared to those with NL (p = 0.003, p = 0.010, respectively). Hepatic IL6 and TNFα mRNA expression was higher in NASH than in SS subjects (p = 0.033, p = 0.050, respectively). Interestingly, LXRα, ACC1 and FAS expression had an inverse relation with the grade of steatosis. These results were confirmed by western blot analysis. In conclusion, our results indicate that lipogenesis seems to be downregulated in advanced stages of SS, suggesting that, in this type of extreme obesity, the deregulation of the lipogenic pathway might be associated with the severity of steatosis. Full article

Non-alcoholic fatty liver disease (NAFLD) is a spectrum of metabolic abnormalities ranging from simple triglyceride accumulation in the hepatocytes to hepatic steatosis with inflammation, ballooning and fibrosis. It has been demonstrated that the pathogenesis of NAFLD involves increased oxidative stress, with consumption of the major cellular antioxidant, glutathione (GSH). Liver has a fundamental role in sulfur compound metabolism, although the data reported on plasma thiols status in NAFLD are conflicting. We recruited 63 NAFLD patients, and we analyzed all plasma thiols, such as homocysteine (Hcy), cysteine (Cys), cysteinylglycine (CysGly) and GSH, by high-performance liquid chromatography (HPLC) with fluorescence detection. Hcy, Cys and CysGly plasma levels increased in NAFLD patients (p < 0.0001); whereas GSH levels were decreased in NAFLD patients when compared to controls (p < 0.0001). On the contrary, patients with steatohepatitis exhibited lower levels of Hcy and Cys than subjects without. Furthermore, a positive correlation was found between Hcy and Cys and the presence of fibrosis in children with NAFLD. Taken together, these data demonstrated a defective hepatic sulfur metabolism in children with NAFLD, and that high levels of Hcy and Cys probably correlates with a pattern of more severe histological liver damage, due to mechanisms that require further studies. Full article

We showed previously that pretreatment of butyrate, which is an endogenous histone deacetylase (HDAC) inhibitor normally fermented from undigested fiber by intestinal microflora, seriously alleviated ischemia reperfusion (I/R)-induced liver injury by inhibiting the nuclear factor κB (NF-κB) pathway. The goal of this study was to investigate the effect of butyrate administrated at the onset of ischemia for HDAC inhibition in hepatic I/R injury. Sprague Dawley rats were subjected to warm ischemia for 60 min followed by 6 and 24 h of reperfusion. Butyrate was administrated at the onset of ischemia. Liver injury was evaluated by serum levels of aminotransferase, inflammatory factors, and histopathology. The levels of acetylated histone H3 and expression of heat shock protein (Hsp) 70 were measured by Western blot. After reperfusion, the levels of acetylated histone H3 significantly decreased. Butyrate treatment markedly prevented the reduction of acetylated histone H3 and upregulated the expression of Hsp70, thereby reducing liver injury. Our study demonstrated that I/R resulted in marked reduction of histone acetylation; butyrate exerted a great hepatoprotective effect through HDAC inhibition and Hsp70 induction. Full article

Objective: The present study aims to identify the differently expressed microRNA (miRNA) molecules and target genes of miRNA in the immune tolerance (IT) and immune activation (IA) stages of chronic hepatitis B (CHB). Methods: miRNA expression profiles of peripheral blood mononuclear cells (PBMCs) at the IT and IA stages of CHB were screened using miRNA microarrays and authenticated using a quantitative real-time polymerase chain reaction (RT-PCR). Gene ontology (GO) and the Kyoto encyclopedia of genes and genomes (KEGG) were used to analyze the significant functions and pathways of possible target genes of miRNAs. Assays of the gain and loss of function of the miRNAs were performed to verify the target genes in THP-1 cell lines. The luciferase reporter test was used on 293T cells as direct targets. Results: Significantly upregulated miR-548 and miR-4804 were observed in the miRNA microarrays and confirmed by RT-PCR in PBMCs at the IT and IA stages of CHB. GO and KEGG analysis revealed that MiR-548 and miR-4804 could be involved in numerous signaling pathways and protein binding activity. IFNγR1 was predicted as a target gene and validated as the direct gene of MiR-548. Significant negative correlation was found between the miR-548ah and mRNA levels of IFN-γR1 in CHB patients. Conclusions: The abnormal expression profiles of miRNA in PBMCs could be closely associated with immune activation of chronic HBV infection. miR-548, by targeting IFN-γR1, may represent a mechanism that can facilitate viral pathogenesis and help determine new therapeutic molecular targets. Full article

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with poor prognosis due to resistance to conventional chemotherapy and limited efficacy of radiotherapy. Previous studies have noted the induction of endoplasmic reticulum stress or apurinic endonuclease 1 (APE1) expression in many tumors. Therefore, the aim of this study was to investigate the relationship between endoplasmic reticulum (ER stress) and APE1 in hepatocellular carcinoma. Here we investigate the expression of APE1 during ER stress in HepG2 and Huh-7 cell lines. Tunicamycin or brefeldin A, two ER stress inducers, increased APE1 and GRP78, an ER stress marker, expression in HepG2 and Huh-7 cells. Induction of APE1 expression was observed through transcription level in response to ER stress. APE1 nuclear localization during ER stress was determined using immunofluorescence assays in HepG2 cells. Furthermore, expression of Hepatitis B virus pre-S2∆ large mutant surface protein (pre-S2∆), an ER stress-induced protein, also increased GRP78 and APE1 expression in the normal hepatocyte NeHepLxHT cell line. Similarly, tumor samples showed higher expression of APE1 in ER stress-correlated liver cancer tissue in vivo. Our results demonstrate that ER stress and HBV pre-S2∆ increased APE1 expression, which may play an important role in resistance to chemotherapeutic agents or tumor development. Therefore, these data provide an important chemotherapeutic strategy in ER stress and HBV pre-S2∆-associated tumors. Full article

Obesity-related metabolic abnormalities, including adipokine imbalance and chronic inflammation, are involved in liver carcinogenesis. Chemerin, a novel adipokine, plays a critical role in adipogenesis, energy metabolism, and inflammation. We evaluated the impact of serum chemerin levels on liver functional reserves in hepatocellular carcinoma (HCC) patients and on the recurrence and prognosis of HCC. This study included 44 patients with any stage of HCC who underwent curative treatment at Gifu Municipal Hospital (Gifu, Japan) between 2006 and 2007. Recurrence-free survival and overall survival were estimated using the Kaplan-Meier method. Serum albumin levels (Pearson’s correlation coefficient; r = 0.3110, p = 0.0399), platelet counts (r = 0.4159, p = 0.0050), and prothrombin times (r = 0.3775, p = 0.0115) were significantly correlated with serum chemerin levels in patients with HCC, and they were inversely correlated with Child-Pugh scores (r = −0.3732, p = 0.0126), serum alanine aminotransferase levels (r = −0.3864, p = 0.0105), and total bilirubin levels (r = −0.4023, p = 0.0068). Among these variables, a multiple comparison test identified that platelet counts and total bilirubin levels were associated with serum chemerin levels (p < 0.0083). No significant correlation was found between serum chemerin levels and recurrence-free survival (p = 0.3691) or overall survival (p = 0.7916). In HCC patients, serum chemerin concentrations were correlated with liver functional reserves and platelet counts, but not with recurrence or prognosis. Full article

Background: Wilson disease (WD) is characterized by hepatic copper accumulation with progressive liver damage to cirrhosis. This study aimed to characterize the toxic milk mouse from The Jackson Laboratory (Bar Harbor, ME, USA) (tx-j) mouse model of WD according to changes over time in hepatic copper concentrations, methionine metabolism, global DNA methylation, and gene expression from gestational day 17 (fetal) to adulthood (28 weeks). Methods: Included liver histology and relevant biochemical analyses including hepatic copper quantification, S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) liver levels, qPCR for transcript levels of genes relevant to methionine metabolism and liver damage, and DNA dot blot for global DNA methylation. Results: Hepatic copper was lower in tx-j fetuses but higher in weanling (three weeks) and adult tx-j mice compared to controls. S-adenosylhomocysteinase transcript levels were significantly lower at all time points, except at three weeks, correlating negatively with copper levels and with consequent changes in the SAM:SAH methylation ratio and global DNA methylation. Conclusion: Compared to controls, methionine metabolism including S-adenosylhomocysteinase gene expression is persistently different in the tx-j mice with consequent alterations in global DNA methylation in more advanced stages of liver disease. The inhibitory effect of copper accumulation on S-adenosylhomocysteinase expression is associated with progressively abnormal methionine metabolism and decreased methylation capacity and DNA global methylation. Full article

The interleukin 28B (IL28B) rs12979860 polymorphism is associated with treatment outcome in hepatitis C virus (HCV) genotype 1 and 4 patients. Its association with the histological features of chronic hepatitis C and disease severity needs further clarifications. To assess the correlation between IL28B genotype, HCV genotype and liver biopsy findings in untreated patients. Materials and Methods: Pre-treatment liver biopsies from 335 HCV Caucasian patients (59% males, age 50 years) enrolled in the MIST study were staged for fibrosis and inflammation according to the METAVIR and the Ishak scoring systems; steatosis was dichotomized as <5% or ≥5%. IL28B was typed by Taqman Single Nucleotide Polymorphism (SNP) genotyping assay. HCV genotype was 1 in 151 (45%), 2 in 99 (30%), 3 in 50 (15%) and 4 in 35 (10%) patients. IL28B genotype was CC in 117 (34%), CT in 166 (49%) and TT in 52 (15%). At univariate analysis, the IL28B CC genotype was associated with severe portal inflammation in HCV-1 patients (CC vs. CT/TT: 86% vs. 63%, p = 0.005), severe lobular inflammation in HCV-2 patients (CC vs. CT/TT: 44% vs. 23%, p = 0.03), and less fatty infiltration in HCV-1 patients (CC vs. CT/TT: 72% vs. 51%, p = 0.02). Despite the lack of any association between IL28B and fibrosis stage, in HCV-3 patients IL28B CC correlated with METAVIR F3-F4 (CC vs. CT/TT: 74% vs. 26%, p = 0.05). At multivariate analysis, the genotype CC remained associated with severe portal inflammation in HCV-1, only (Odds Ratio (OR): 95% Confidence Interval (CI): 3.24 (1.23–8.51)). IL28B genotype is associated with the histological features of chronic hepatitis C in a HCV genotype dependent manner, with CC genotype being independently associated with severe portal inflammation. Full article

Mesenchymal stem cells from human bone marrow (hMSC) have the potential to differentiate into hepatocyte-like cells in vitro and continue to maintain important hepatocyte functions in vivo after transplantation into host mouse livers. Here, hMSC were differentiated into hepatocyte-like cells in vitro (hMSC-HC) and transplanted into livers of immunodeficient Pfp/Rag2^−/− mice treated with a sublethal dose of acetaminophen (APAP) to induce acute liver injury. APAP induced a time- and dose-dependent damage of perivenous areas of the liver lobule. Serum levels of aspartate aminotransferase (AST) increased to similar levels irrespective of hMSC-HC transplantation. Yet, hMSC-HC resided in the damaged perivenous areas of the liver lobules short-term preventing apoptosis and thus progress of organ destruction. Disturbance of metabolic protein expression was lower in the livers receiving hMSC-HC. Seven weeks after APAP treatment, hepatic injury had completely recovered in groups both with and without hMSC-HC. Clusters of transplanted cells appeared predominantly in the periportal portion of the liver lobule and secreted human albumin featuring a prominent quality of differentiated hepatocytes. Thus, hMSC-HC attenuated the inflammatory response and supported liver regeneration after acute injury induced by acetaminophen. They hence may serve as a novel source of hepatocyte-like cells suitable for cell therapy of acute liver diseases. Full article

Aldo-keto reductase family 1, member B10 (AKR1B10), a cancer-related oxidoreductase, is expressed in well-differentiated hepatocellular carcinomas (HCCs). However, AKR1B10 levels are minimal in normal liver tissues (NLs), similar to the 70-kilodalton heat shock protein (HSP70) and glypican-3. Moreover, the role of AKR1B10 in chronic hepatitis or cirrhosis, which are considered preneoplastic conditions for HCC, has not been fully elucidated. The aim of this study was to evaluate the expression of AKR1B10, HSP70, and glypican-3 in 61 HCC tissue samples compared to corresponding non-tumorous liver tissues (NTs), comprising 42 chronic hepatitis and 19 cirrhosis cases to clarify the significance of molecular changes at the preneoplastic stages of HCC. Immunohistochemical analysis demonstrated that the median expression levels of AKR1B10 were higher in HCCs than in NTs (p < 0.001) and higher in NTs than NLs (p < 0.001) with 54.8%, 2.1%, and 0.3% expression in HCCs, NTs, and NLs, respectively. HSP70 and glypican-3 were expressed in HCCs, but minimally in NTs and NLs with no significant difference between expression in NTs and NLs. Furthermore, a multivariate analysis identified an association between hepatic steatosis and AKR1B10 expression in NTs (p = 0.020). Of the three protein expressed in well-differentiated HCCs, only AKR1B10 was upregulated in preneoplastic conditions, and a steatosis-related factor might influence its expression. Full article

Non-alcoholic steatohepatitis (NASH) represents a risk factor for the development of hepatocellular carcinoma (HCC) and is characterized by quantitative and qualitative changes in hepatic lipids. Since elongation of fatty acids from C16 to C18 has recently been reported to promote both hepatic lipid accumulation and inflammation we aimed to investigate whether a frequently used mouse NASH model reflects this clinically relevant feature and whether C16 to C18 elongation can be observed in HCC development. Feeding mice a methionine and choline deficient diet to model NASH not only increased total hepatic fatty acids and cholesterol, but also distinctly elevated the C18/C16 ratio, which was not changed in a model of simple steatosis (ob/ob mice). Depletion of Kupffer cells abrogated both quantitative and qualitative methionine-and-choline deficient (MCD)-induced alterations in hepatic lipids. Interestingly, mimicking inflammatory events in early hepatocarcinogenesis by diethylnitrosamine-induced carcinogenesis (48 h) increased hepatic lipids and the C18/C16 ratio. Analyses of human liver samples from patients with NASH or NASH-related HCC showed an elevated expression of the elongase ELOVL6, which is responsible for the elongation of C16 fatty acids. Taken together, our findings suggest a detrimental role of an altered fatty acid pattern in the progression of NASH-related liver disease. Full article

Many investigations have revealed that a low recurrence rate of hepatocellular carcinoma (HCC) is associated with high serum albumin levels in patients; therefore, high levels of serum albumin are a major indicator of a favorable prognosis. However, the mechanism inhibiting the proliferation of HCC has not yet been elucidated, so we investigated the effect of serum albumin on HCC cell proliferation. Hep3B was cultured in MEM with no serum or containing 5 g/dL human albumin. As control samples, Prionex was added to generate the same osmotic pressure as albumin. After 24-h incubation, the expressions of α-fetoprotein (AFP), p53, p21, and p57 were evaluated with real-time PCR using total RNA extracted from the liver. Protein expressions and the phosphorylation of Rb (retinoblastoma) were determined by Western blot analysis using total protein extracted from the liver. For flow cytometric analysis of the cell cycle, FACS analysis was performed. The percentages of cell cycle distribution were evaluated by PI staining, and all samples were analyzed employing FACScalibur (BD) with appropriate software (ModFit LT; BD). The cell proliferation assay was performed by counting cells with using a Scepter handy automated cell counter (Millipore). The mRNA levels of AFP relative to Alb(−): Alb(−), Alb(+), and Prionex, were 1, 0.7 ± 0.2 (p < 0.001 for Alb(−)), and 1 ± 0.3, respectively. The mRNA levels of p21 were 1, 1.58 ± 0.4 (p = 0.007 for Alb(−) and p = 0.004 for Prionex), and 0.8 ± 0.2, respectively. The mRNA levels of p57 were 1, 4.4 ± 1.4 (p = 0.002 for Alb(−) and Prionex), and 1.0 ± 0.1, respectively. The protein expression levels of Rb were similar in all culture media. The phosphorylation of P807/811 and P780 of Rb protein was reduced in Alb(+). More cells in the G0/G1 phase and fewer cells in S and G2/M phases were obtained in Alb(+) than in Alb(−) (G0/G1: 60.9%, 67.7%, 61.5%; G2/M: 16.5%, 13.1%, 15.6%; S: 22.6%, 19.2%, 23.0%, Alb(−), Alb(+), Prionex, respectively). The same results were obtained in HepG2. Cell proliferation was inhibited in 5 g/dL albumin medium in both HepG2 cells and Hep3B cells in 24 h culture by counting cell numbers. The presence of albumin in serum reduces the phosphorylation of Rb proteins and enhances the expression of p21 and p57, following an increase in the G0/G1 cell population, and suppresses cell proliferation. These results suggest that albumin itself suppresses the proliferation of hepatocellular carcinoma. Full article

Long non-coding RNA HOTAIR exerts regulatory functions in various biological processes in cancer cells, such as proliferation, apoptosis, mobility, and invasion. We previously found that HOX transcript antisense RNA (HOTAIR) is a negative prognostic factor and exhibits oncogenic activity in hepatocellular carcinoma (HCC). In this study, we aimed to investigate the role and molecular mechanism of HOTAIR in promoting HCC cell migration and invasion. Firstly, we profiled its gene expression pattern by microarray analysis of HOTAIR loss in Bel-7402 HCC cell line. The results showed that 129 genes were significantly down-regulated, while 167 genes were significantly up-regulated (fold change >2, p < 0.05). Bioinformatics analysis indicated that RNA binding proteins were involved in this biological process. HOTAIR suppression using RNAi strategy with HepG2 and Bel-7402 cells increased the mRNA and protein expression levels of RNA binding motif protein 38 (RBM38). Moreover, the expression levels of RBM38 in HCC specimens were significantly lower than paired adjacent noncancerous tissues. In addition, knockdown of HOTAIR resulted in a decrease of cell migration and invasion, which could be specifically rescued by down-regulation of RBM38. Taken together, HOTAIR could promote migration and invasion of HCC cells by inhibiting RBM38, which indicated critical roles of HOTAIR and RBM38 in HCC progression. Full article

The aim of this study was to determine the effect and mechanism of tamoxifen (TAM)-induced steatosis in vitro. HepG 2 (Human hepatocellular liver carcinoma cell line) cells were treated with different concentrations of TAM for 72 h. Steatosis of hepatocytes was determined after Oil Red O staining and measurement of triglyceride (TG) concentration. The expressions of genes in the TG homeostasis pathway, including sterol regulatory element-binding protein-1c (SREBP-1c), peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), stearoyl-CoA desaturase (SCD), carnitine palmitoyltransferase 1 (CPT1) and microsomal triglyceride transfer protein (MTP), were examined using quantitative real-time PCR and Western blot analysis. Cell proliferation was examined using the cell counting kit-8 (CCK-8) assay. We found that hepatocytes treated with TAM had: (1) induced hepatocyte steatosis and increased hepatocyte TG; (2) upregulation of SREBP-1c, FAS, ACC, SCD and MTP mRNA expressions (300%, 600%, 70%, 130% and 160%, respectively); (3) corresponding upregulation of protein expression; and (4) no difference in HepG 2 cell proliferation. Our results suggest that TAM can induce hepatocyte steatosis in vitro and that the enhancement of fatty acid synthesis through the upregulations of SREBP-1c and its downstream target genes (FAS, ACC and SCD) may be the key mechanism of TAM-induced hepatocyte steatosis. Full article

Golgi protein 73 (GP73), which is up-regulated in hepatocellular carcinoma (HCC), has recently been identified as a novel serum marker for HCC diagnosis. Several reports also noted the increased levels of GP73 expression in chronic liver disease in patients with acute hepatitis of various etiologies, chronic Hepatitis C virus (HCV) infection and alcoholic liver disease. The molecular mechanisms of GP73 expression in HCV related liver disease still need to be determined. In this study, we aimed to evaluate the effect of HCV infection on GP73 expression. GP73 was highly expressed in Huh7, Hep3B, 293T and HUVEC cells, and was low-expressed in HepG2 cells. HCV infection led to down-regulation of GP73 in Huh7 and HepG2/CD81 cells at the early stage of infection. CXCL10 decreased GP73 expression in Huh7 and HepG2 cells. Up-regulation of GP73 was noted in hepatocytes with cytopathic effect at advanced stage of HCV infection, and further research is needed to determine the unknown factors affecting GP73 expression. In conclusion, our study provided additional evidence for the roles of GP73 in liver disease. Full article

Smad3 is a key mediator of the transforming growth factor (TGF)-β1 signaling pathway that plays central role in inflammation and fibrosis. In present study, we evaluated the effect of Smad3 deficiency in Smad3^−/− mice with carbon tetrachloride (CCl₄)-induced liver fibrosis. The animals were received CCl₄ or olive oil three times a week for 4 weeks. Histopathological analyses were performed to evaluate the fibrosis development in the mice. Alteration of protein expression controlled by Smad3 was examined using a proteomic analysis. CCl₄-induced liver fibrosis was rarely detected in Smad3^−/− mice compared to Smad3^+/+. Proteomic analysis revealed that proteins related to antioxidant activities such as senescence marker protein-30 (SMP30), selenium-binding proteins (SP56) and glutathione S-transferases (GSTs) were up-regulated in Smad3^−/− mice. Western blot analysis confirmed that SMP30 protein expression was increased in Smad3^−/− mice. And SMP30 levels were decreased in CCl₄-treated Smad3^+/+ and Smad3^−/− mice. These results indicate that Smad3 deficiency influences the proteins level related to antioxidant activities during early liver fibrosis. Thus, we suggest that Smad3 deteriorate hepatic injury by inhibitor of antioxidant proteins as well as mediator of TGF-β1 signaling. Full article

The chronic intake of fructose has been linked to insulin resistance, obesity, dyslipidemia and nonalcoholic fatty liver disease (NAFLD), which in turn, may progress to nonalcoholic steatohepatitis (NASH). We aimed to evaluate the magnitude of the effects of the chronic consumption of high-fructose (HFr) and high fat (HF) alone or combined. Four groups of male mice were fed different diets for 16 weeks: standard chow (9% fat: SC), HF diet (42% fat), HFr diet (34% fructose) and HF/HFr diet (42% fat, 34% fructose). The food intake was not different among the groups, and the body mass was not greater in the HFr group than in the SC group. The homeostasis model assessment for insulin resistance (HOMA-IR), as well as plasmatic total cholesterol and triglycerides were greater in the groups HF, HFr, and HF/HFr group than in the SC group. We observed in the groups HF, HFr and HF/HFr, compared to the group SC, nonalcoholic fatty liver disease (NAFLD) with a predominance of lipogenesis mediated by SREBP-1c and PPAR-γ, and a reduction of the oxidation mediated by PPAR-α. We also observed an increase in gluconeogenesis mediated by the GLUT-2 and the PEPCK. Importantly, we identified areas of necroinflammation indicating a transition from NAFLD to nonalcoholic steatohepatitis in the HFr and HF/HFr groups. This study is relevant in demonstrating that fructose consumption, even in the absence of obesity, causes serious and deleterious changes in the liver with the presence of the dyslipidemia, insulin resistance (IR), and NAFLD with areas of necroinflammation. These conditions are associated with a poor prognosis. Full article

This study aimed to investigate the associations of the serial changes of serum levels of various growth factors with liver regeneration after hepatectomy in healthy liver donors. Sixteen healthy liver donors who underwent conventional liver resection were included. Serum levels of various growth factors before hepatectomy and on postoperative day (POD) 1, 3, 5 and 7 were measured. Liver volume data calculated by multi-detector computed tomography using workstation. The ratio of remnant liver volume on POD 0 to liver volume before the operation was 51% ± 20%. The ratio of liver volume on POD 14 to liver volume on POD 0 were inversely correlated with remnant liver volume on POD 0 (r = −0.91). The ratio of liver volume on POD 14 to liver volume on POD 0 were significantly correlated with serum hepatocyte growth factor (HGF) levels on POD 1 (r = 0.54), serum leptin levels on POD 1 (r = 0.54), and serum macrophage colony-stimulating factor (M-CSF) levels on POD 5 (r = 0.76) and POD 7 (r = 0.80). These results suggest that early-phase elevation of serum levels of HGF, leptin and M-CSF may be associated with the acceleration of liver regeneration after hepatectomy in humans. Full article

The hepatitis B virus (HBV) counts as a major global health problem, as it presents a significant causative factor for liver-related morbidity and mortality. The development of hepatocellular carcinomas (HCC) as a characteristic of a persistent, chronic infection could be caused, among others, by the pleiotropic function of the viral regulatory protein HBx. The latter is known to modulate an onset of cellular and viral signaling processes with emerging influence in liver pathogenesis. However, the flexible and multifunctional nature of HBx impedes the fundamental understanding of related mechanisms and the development of associated diseases, and has even led to partial controversial results in the past. Based on the cellular distribution of HBx—nuclear-, cytoplasmic- or mitochondria-associated—this review encompasses the current knowledge and previous investigations of HBx in context of cellular signaling pathways and HBV-associated pathogenesis. In addition, particular focus is set on the clinical relevance and potential novel therapeutic applications in the context of HBx. Full article

Thirty-five years ago, precision-cut liver slices (PCLS) were described as a promising tool and were expected to become the standard in vitro model to study liver disease as they tick off all characteristics of a good in vitro model. In contrast to most in vitro models, PCLS retain the complex 3D liver structures found in vivo, including cell–cell and cell–matrix interactions, and therefore should constitute the most reliable tool to model and to investigate pathways underlying chronic liver disease in vitro. Nevertheless, the biggest disadvantage of the model is the initiation of a procedure-induced fibrotic response. In this review, we describe the parameters and potential of PCLS cultures and discuss whether the initially described limitations and pitfalls have been overcome. We summarize the latest advances in PCLS research and critically evaluate PCLS use and progress since its invention in 1985. Full article

Hepatocellular carcinoma (HCC) is a prime public health concern that accounts for most of the primary liver malignancies in humans. The most common etiological factor of HCC is hepatitis B virus (HBV). Despite recent advances in treatment strategies, there has been little success in improving the survival of HCC patients. To develop a novel therapeutic approach, evaluation of a working hypothesis based on different viewpoints might be important. Long interspersed element 1 (L1) retrotransposons have been suggested to play a role in HCC. However, the molecular machineries that can modulate L1 biology in HBV-related HCC have not been well-evaluated. Here, we summarize the profiles of expression and/or activation status of L1-related genes in HBV-related HCC, and HBV- and HCC-related genes that may impact L1-mediated tumorigenesis. L1 restriction factors appear to be suppressed by HBV infection. Since some of the L1 restriction factors also limit HBV, these factors may be exhausted in HBV-infected cells, which causes de-suppression of L1. Several HBV- and HCC-related genes that interact with L1 can affect oncogenic processes. Thus, L1 may be a novel prime therapeutic target for HBV-related HCC. Studies in this area will provide insights into HCC and other types of cancers. Full article

Industrialized society-caused dysregular human behaviors and activities such as overworking, excessive dietary intake, and sleep deprivation lead to perturbations in the metabolism and the development of metabolic syndrome. Non-alcoholic fatty liver disease (NAFLD), the most common chronic liver disease worldwide, affects around 30% and 25% of people in Western and Asian countries, respectively, which leads to numerous medical costs annually. Insulin resistance is the major hallmark of NAFLD and is crucial in the pathogenesis and for the progression from NAFLD to non-alcoholic steatohepatitis (NASH). Excessive dietary intake of saturated fats and carbohydrate-enriched foods contributes to both insulin resistance and NAFLD. Once NAFLD is established, insulin resistance can promote the progression to the more severe state of liver endangerment like NASH. Here, we review current and potential studies for understanding the complexity between insulin-regulated glycolytic and lipogenic homeostasis and the underlying causes of NAFLD. We discuss how disruption of the insulin signal is associated with various metabolic disorders of glucoses and lipids that constitute both the metabolic syndrome and NAFLD. Full article

Lifestyle-related disorders, such as the metabolic syndrome, have become a primary risk factor for the development of liver pathologies that can progress from hepatic steatosis, hepatic insulin resistance, steatohepatitis, fibrosis and cirrhosis, to the most severe condition of hepatocellular carcinoma (HCC). While the prevalence of liver pathologies is steadily increasing in modern societies, there are currently no approved drugs other than chemotherapeutic intervention in late stage HCC. Hence, there is a pressing need to identify and investigate causative molecular pathways that can yield new therapeutic avenues. The transcription factor p53 is well established as a tumor suppressor and has recently been described as a central metabolic player both in physiological and pathological settings. Given that liver is a dynamic tissue with direct exposition to ingested nutrients, hepatic p53, by integrating cellular stress response, metabolism and cell cycle regulation, has emerged as an important regulator of liver homeostasis and dysfunction. The underlying evidence is reviewed herein, with a focus on clinical data and animal studies that highlight a direct influence of p53 activity on different stages of liver diseases. Based on current literature showing that activation of p53 signaling can either attenuate or fuel liver disease, we herein discuss the hypothesis that, while hyper-activation or loss of function can cause disease, moderate induction of hepatic p53 within physiological margins could be beneficial in the prevention and treatment of liver pathologies. Hence, stimuli that lead to a moderate and temporary p53 activation could present new therapeutic approaches through several entry points in the cascade from hepatic steatosis to HCC. Full article

Whole-transcriptome analyses have revealed that a large proportion of the human genome is transcribed in non-protein-coding transcripts, designated as long non-coding RNAs (lncRNAs). Rather than being “transcriptional noise”, increasing evidence indicates that lncRNAs are key players in the regulation of many biological processes, including transcription, post-translational modification and inhibition and chromatin remodeling. Indeed, lncRNAs are widely dysregulated in human cancers, including hepatocellular carcinoma (HCC). Functional studies are beginning to provide insights into the role of oncogenic and tumor suppressive lncRNAs in the regulation of cell proliferation and motility, as well as oncogenic and metastatic potential in HCC. A better understanding of the molecular mechanisms and the complex network of interactions in which lncRNAs are involved could reveal novel diagnostic and prognostic biomarkers. Crucially, it may provide novel therapeutic opportunities to add to the currently limited number of therapeutic options for HCC patients. In this review, we summarize the current status of the field, with a focus on the best characterized dysregulated lncRNAs in HCC. Full article

The thyroid hormones (THs) 3,3′,5,5′-tetraiodo-l-thyronine (T4) and 3,5,3′-triiodo-l-thyronine (T3) influence many metabolic pathways. The major physiological function of THs is to sustain basal energy expenditure, by acting primarily on carbohydrate and lipid catabolism. Beyond the mobilization and degradation of lipids, at the hepatic level THs stimulate the de novo fatty acid synthesis (de novo lipogenesis, DNL), through both the modulation of gene expression and the rapid activation of cell signalling pathways. 3,5-Diiodo-l-thyronine (T2), previously considered only a T3 catabolite, has been shown to mimic some of T3 effects on lipid catabolism. However, T2 action is more rapid than that of T3, and seems to be independent of protein synthesis. An inhibitory effect on DNL has been documented for T2. Here, we give an overview of the mechanisms of THs action on liver fatty acid metabolism, focusing on the different effects exerted by T2 and T3 on the regulation of the DNL. The inhibitory action on DNL exerted by T2 makes this compound a potential and attractive drug for the treatment of some metabolic diseases and cancer. Full article

Autoimmune hepatitis (AIH) is characterized by a progressive destruction of the liver parenchyma and a chronic fibrosis. The current treatment of autoimmune hepatitis is still largely dependent on the administration of corticosteroids and cytostatic drugs. For a long time the development of novel therapeutic strategies has been hampered by a lack of understanding the basic immunopathogenic mechanisms of AIH and the absence of valid animal models. However, in the past decade, knowledge from clinical observations in AIH patients and the development of innovative animal models have led to a situation where critical factors driving the disease have been identified and alternative treatments are being evaluated. Here we will review the insight on the immunopathogenesis of AIH as gained from clinical observation and from animal models. Full article

Primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC) represent the major clinical entities of chronic cholestatic liver diseases. Both disorders are characterized by portal inflammation and slowly progress to obliterative fibrosis and eventually liver cirrhosis. Although immune-pathogenic mechanisms have been implicated in the pathogenesis of PBC and PSC, neither disorder is considered to be a classical autoimmune disease, as PSC and PBC patients do not respond to immune-suppressants. Furthermore, the decreased bile flow resulting from the immune-mediated tissue assault and the subsequent accumulation of toxic bile products in PBC and PSC not only perpetuates biliary epithelial damage, but also alters the composition of the intestinal and biliary microbiota and its mutual interactions with the host. Consistent with the close association of PSC and inflammatory bowel disease (IBD), the polyclonal hyper IgM response in PBC and (auto-)antibodies which cross-react to microbial antigens in both diseases, an expansion of individual microbes leads to shifts in the composition of the intestinal or biliary microbiota and a subsequent altered integrity of epithelial layers, promoting microbial translocation. These changes have been implicated in the pathogenesis of both devastating disorders. Thus, we will discuss here these recent findings in the context of novel and alternative therapeutic options. Full article

The prevalence of metabolic disorders, such as type 2 diabetes (T2D), obesity, and non-alcoholic fatty liver disease (NAFLD), which are common risk factors for cardiovascular disease (CVD), has dramatically increased worldwide over the last decades. Although dietary habit is the main etiologic factor, there is an imperfect correlation between dietary habits and the development of metabolic disease. Recently, research has focused on the role of the microbiome in the development of these disorders. Indeed, gut microbiota is implicated in many metabolic functions and an altered gut microbiota is reported in metabolic disorders. Here we provide evidence linking gut microbiota and metabolic diseases, focusing on the pathogenetic mechanisms underlying this association. Full article

Chronic hepatitis B virus (HBV) infection is one of the most associated factors in hepatocarcinogenesis. HBV is able to integrate into the host genome and encode the multi-functional hepatitis B virus x protein (HBx). Although the mechanism between HBx and carcinogenesis is still elusive, recent studies have shown that HBx was able to influence various signaling pathways, as well as epigenetic and genetic processes. This review will examine and summarize recent literature about HBx’s role in these various processes. Full article

The epidermal growth factor receptor (EGFR) is a transmembrane receptor tyrosine kinase that is activated by several ligands leading to the activation of diverse signaling pathways controlling mainly proliferation, differentiation, and survival. The EGFR signaling axis has been shown to play a key role during liver regeneration following acute and chronic liver damage, as well as in cirrhosis and hepatocellular carcinoma (HCC) highlighting the importance of the EGFR in the development of liver diseases. Despite the frequent overexpression of EGFR in human HCC, clinical studies with EGFR inhibitors have so far shown only modest results. Interestingly, a recent study has shown that in human HCC and in mouse HCC models the EGFR is upregulated in liver macrophages where it plays a tumor-promoting function. Thus, the role of EGFR in liver diseases appears to be more complex than what anticipated. Further studies are needed to improve the molecular understanding of the cell-specific signaling pathways that control disease development and progression to be able to develop better therapies targeting major components of the EGFR signaling network in selected cell types. In this review, we compiled the current knowledge of EGFR signaling in different models of liver damage and diseases, mainly derived from the analysis of HCC cell lines and genetically engineered mouse models (GEMMs). Full article

Chronic liver dysfunction or injury is a serious health problem worldwide. Chronic liver disease involves a wide range of liver pathologies that include fatty liver, hepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. The efficiency of current synthetic agents in treating chronic liver disease is not satisfactory and they have undesirable side effects. Thereby, numerous medicinal herbs and phytochemicals have been investigated as complementary and alternative treatments for chronic liver diseases. Since some herbal products have already been used for the management of liver diseases in some countries or regions, a systematic review on these herbal medicines for chronic liver disease is urgently needed. Herein, we conducted a review describing the potential role, pharmacological studies and molecular mechanisms of several commonly used medicinal herbs and phytochemicals for chronic liver diseases treatment. Their potential toxicity and side effects were also discussed. Several herbal formulae and their biological effects in chronic liver disease treatment as well as the underlying molecular mechanisms are also summarized in this paper. This review article is a comprehensive and systematic analysis of our current knowledge of the conventional medicinal herbs and phytochemicals in treating chronic liver diseases and on the potential pitfalls which need to be addressed in future study. Full article

A complex antioxidant system has been developed in mammals to relieve oxidative stress. However, excessive reactive species derived from oxygen and nitrogen may still lead to oxidative damage to tissue and organs. Oxidative stress has been considered as a conjoint pathological mechanism, and it contributes to initiation and progression of liver injury. A lot of risk factors, including alcohol, drugs, environmental pollutants and irradiation, may induce oxidative stress in liver, which in turn results in severe liver diseases, such as alcoholic liver disease and non-alcoholic steatohepatitis. Application of antioxidants signifies a rational curative strategy to prevent and cure liver diseases involving oxidative stress. Although conclusions drawn from clinical studies remain uncertain, animal studies have revealed the promising in vivo therapeutic effect of antioxidants on liver diseases. Natural antioxidants contained in edible or medicinal plants often possess strong antioxidant and free radical scavenging abilities as well as anti-inflammatory action, which are also supposed to be the basis of other bioactivities and health benefits. In this review, PubMed was extensively searched for literature research. The keywords for searching oxidative stress were free radicals, reactive oxygen, nitrogen species, anti-oxidative therapy, Chinese medicines, natural products, antioxidants and liver diseases. The literature, including ours, with studies on oxidative stress and anti-oxidative therapy in liver diseases were the focus. Various factors that cause oxidative stress in liver and effects of antioxidants in the prevention and treatment of liver diseases were summarized, questioned, and discussed. Full article

The story of high mobility group protein B1 (HMGB1) in cancer is complicated and the function of HMGB1 in different cancers is uncertain. This review aims to retrieve literature regarding HMGB1 from English electronic resources, analyze and summarize the role of the HMGB1 signaling pathway in hepatocellular carcinoma (HCC), and provide useful information for carcinogenesis and progression of HCC. Results showed that HMGB1 could induce cell proliferation, differentiation, cell death, angiogenesis, metastasis, inflammation, and enhance immunofunction in in vitro and in vivo HCC models. HMGB1 and its downstream receptors RAGE, TLRs and TREM-1 may be potential anticancer targets. In conclusion, HMGB1 plays an important role in oncogenesis and represents a novel therapeutic target, which deserves further study. Full article

Molecular dissection of hepatocellular adenomas has brought forward a diversity of well-defined entities. Their distinction is important for routine practice, since prognosis is tightly related to the individual subgroup. Very recent activity has generated new details on the molecular background of hepatocellular adenoma, which this article aims to integrate into the current concepts of taxonomy. Full article

Liver fibrosis, the main characteristic of chronic liver diseases, is strongly associated with the activation of hepatic stellate cells (HSCs), which are responsible for extracellular matrix production. As such, investigating the effective regulators controlling HSC activation provides important clues for developing therapeutics to inhibit liver fibrosis. Thymosin beta 4 (Tβ4), a major actin-sequestering protein, is known to be involved in various cellular responses. A growing body of evidence suggests that Tβ4 has a potential role in the pathogenesis of liver fibrosis and that it is especially associated with the activation of HSCs. However, it remains unclear whether Tβ4 promotes or suppresses the activation of HSCs. Herein, we review the potential role of Tβ4 in liver fibrosis by describing the effects of exogenous and endogenous Tβ4, and we discuss the possible signaling pathway regulated by Tβ4. Exogenous Tβ4 reduces liver fibrosis by inhibiting the proliferation and migration of HSCs. Tβ4 is expressed endogenously in the activated HSCs, but this endogenous Tβ4 displays opposite effects in HSC activation, either as an activator or an inhibitor. Although the role of Tβ4 has not been established, it is apparent that Tβ4 influences HSC activation, suggesting that Tβ4 is a potential therapeutic target for treating liver diseases. Full article

Primary biliary cirrhosis (PBC) is a chronic progressive autoimmune cholestatic liver disease characterized by highly specific antimitochondrial antibodies (AMAs) and the specific immune-mediated injury of small intrahepatic bile ducts. Unique apoptotic feature of biliary epithelial cells (BECs) may contribute to apotope presentation to the immune system, causing unique tissue damage in PBC. Perpetuation of inflammation may result in senescence of BECs, contributing to irreversible loss of bile duct. In addition to the classic liver manifestations, focal inflammation and tissue damage are also seen in salivary glands and urinary tract in a significant proportion of PBC patients. These findings provide potent support to the idea that molecular mimicry may be involved in the breakdown of autoimmune tolerance and mucosal immunity may lead to a systematic epithelitis in PBC patients. Thus, PBC is considered a generalized epithelitis in clinical practice. Full article

Wilson’s disease (WD), also known as hepatolenticular degeneration, is an autosomal recessive inherited disorder resulting from abnormal copper metabolism. Reduced copper excretion causes an excessive deposition of the copper in many organs such as the liver, central nervous system (CNS), cornea, kidney, joints, and cardiac muscle where the physiological functions of the affected organs are impaired. The underlying molecular mechanisms for WD have been extensively studied. It is now believed that a defect in P-type adenosine triphosphatase (ATP7B), the gene encoding the copper transporting P-type ATPase, is responsible for hepatic copper accumulation. Deposited copper in the liver produces toxic effects via modulating several molecular pathways. WD can be a lethal disease if left untreated. A better understanding of the molecular mechanisms causing the aberrant copper deposition and organ damage is the key to developing effective management approaches. Full article

Non-alcoholic fatty liver disease (NAFLD) is characterized by excessive fat accumulation in the liver. It ranges from simple steatosis to its more aggressive form, non-alcoholic steatohepatitis (NASH), which may develop into hepatic fibrosis, cirrhosis, or hepatocellular carcinoma (HCC) if it persists for a long time. However, the exact pathogenesis of NAFLD and the related metabolic disorders remain unclear. Epigenetic changes are stable alterations that take place at the transcriptional level without altering the underlying DNA sequence. DNA methylation, histone modifications and microRNA are among the most common forms of epigenetic modification. Epigenetic alterations are involved in the regulation of hepatic lipid metabolism, insulin resistance, mitochondrial damage, oxidative stress response, and the release of inflammatory cytokines, all of which have been implicated in the development and progression of NAFLD. This review summarizes the current advances in the potential epigenetic mechanism of NAFLD. Elucidation of epigenetic factors may facilitate the identification of early diagnositic biomarkers and development of therapeutic strategies for NAFLD. Full article

Pediatric hepatoblastoma (HB) is commonly treated by neoadjuvant chemotherapy and surgical tumor resection according to international multicenter trial protocols. Complete tumor resection is essential and survival rates up to 95% have now been achieved in those tumors classified as standard-risk HB. Drug resistance and occurrence of metastases remain the major challenges in the treatment of HB, especially in high-risk tumors. These conditions urgently require the development of alternative therapeutic strategies. One of those alternatives is the modulation of apoptosis in HB cells. HBs regularly overexpress anti-apoptotic proteins of the Bcl-family in comparison to healthy liver tissue. This fact may contribute to the development of chemoresistance of HB cells. Synthetic small inhibitory molecules with BH3-mimetic effects, such as ABT-737 and obatoclax, enhance the susceptibility of tumor cells to different cytotoxic drugs and thereby affect initiator proteins of the apoptosis cascade via the intrinsic pathway. Besides additive effects on HB cell viability when used in combination with cytotoxic drugs, BH3-mimetics also play a role in preventing metastasation by reducing adhesion and inhibiting cell migration abilities. Presumably, including additive BH3-mimetic drugs into existing therapeutic regimens in HB patients might allow dose reduction of established cytotoxic drugs and thereby associated immanent side effects, while maintaining the antitumor activity. Furthermore, reduction of tumor growth and inhibition of tumor cell dissemination may facilitate complete surgical tumor resection, which is mandatory in this tumor type resulting in improved survival rates in high-risk HB. Currently, there are phase I and phase II clinical trials in several cancer entities using this potential target. This paper reviews the available literature regarding the use of BH3-mimetic drugs as single agents or in combination with chemotherapy in various malignancies and focuses on results in HB cells. Full article

Proteins regulate many cellular functions and analyzing the presence and abundance of proteins in biological samples are central focuses in proteomics. The discovery and validation of biomarkers, pathways, and drug targets for various diseases can be accomplished using mass spectrometry-based proteomics. However, with mass-limited samples like tumor biopsies, it can be challenging to obtain sufficient amounts of proteins to generate high-quality mass spectrometric data. Techniques developed for macroscale quantities recover sufficient amounts of protein from milligram quantities of starting material, but sample losses become crippling with these techniques when only microgram amounts of material are available. To combat this challenge, proteomicists have developed micro-scale techniques that are compatible with decreased sample size (100 μg or lower) and still enable excellent proteome coverage. Extraction, contaminant removal, protein quantitation, and sample handling techniques for the microgram protein range are reviewed here, with an emphasis on liquid chromatography and bottom-up mass spectrometry-compatible techniques. Also, a range of biological specimens, including mammalian tissues and model cell culture systems, are discussed. Full article

Autoimmune hepatitis (AIH) is an immune-mediated disorder that affects the liver parenchyma. Diagnosis usually occurs at the later stages of the disease, complicating efforts towards understanding the causes of disease development. While animal models are useful for studying the etiology of autoimmune disorders, most of the existing animal models of AIH do not recapitulate the chronic course of the human condition. In addition, approaches to mimic AIH-associated liver inflammation have instead led to liver tolerance, consistent with the high tolerogenic capacity of the liver. Recently, we described a new mouse model that exhibited spontaneous and chronic liver inflammation that recapitulated the known histopathological and immunological parameters of AIH. The approach involved liver-extrinsic genetic engineering that interfered with the induction of T-cell tolerance in the thymus, the very process thought to inhibit AIH induction by liver-specific expression of exogenous antigens. The mutation led to depletion of specialized thymic epithelial cells that present self-antigens and eliminate autoreactive T-cells before they exit the thymus. Based on our findings, which are summarized below, we believe that this mouse model represents a relevant experimental tool towards elucidating the cellular and molecular aspects of AIH development and developing novel therapeutic strategies for treating this disease. Full article

Thioredoxin domain-containing 5 (TXNDC5) is a member of the protein disulfide isomerase family, acting as a chaperone of endoplasmic reticulum under not fully characterized conditions As a result, TXNDC5 interacts with many cell proteins, contributing to their proper folding and correct formation of disulfide bonds through its thioredoxin domains. Moreover, it can also work as an electron transfer reaction, recovering the functional isoform of other protein disulfide isomerases, replacing reduced glutathione in its role. Finally, it also acts as a cellular adapter, interacting with the N-terminal domain of adiponectin receptor. As can be inferred from all these functions, TXNDC5 plays an important role in cell physiology; therefore, dysregulation of its expression is associated with oxidative stress, cell ageing and a large range of pathologies such as arthritis, cancer, diabetes, neurodegenerative diseases, vitiligo and virus infections. Its implication in all these important diseases has made TXNDC5 a susceptible biomarker or even a potential pharmacological target. Full article

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease in industrialized countries. The increasing prevalence of NAFLD mirrors the outbreak of obesity in western countries, highlighting the connection between these two conditions. Nevertheless, there is currently no specific pharmacotherapy for its treatment. Accepted management begins with weight loss and exercise. Moreover, exercise can provide metabolic benefits independently of weight loss. It is known how long-term aerobic training produces improvements in hepatic triglycerides, visceral adipose tissue and free fatty acids, even if there is no weight reduction. A recent study from Boström et al. unravels a potential molecular mechanism that may explain how exercise, independently of weight loss, can potentially improve metabolic parameters through a new messenger system (irisin) linking muscle and fat tissue. Irisin has been proposed to act as a hormone on subcutaneous white fat cells increasing energy expenditure by means of a program of brown-fat-like development. Moreover, it was also shown that irisin plasma concentration was higher in people who exercise, suggesting a molecular mechanism by which exercise may improve metabolism. The present systematic review is based on the possibility that irisin might represent a hypothetical connection between NAFLD pathogenesis and disease progression. Full article

Liver fibrosis is a common phenomenon that is associated with several pathologies and is characterized by excessive extracellular matrix deposition that leads to progressive liver dysfunction. Bone morphogenetic protein 9 (BMP9) is the most recently discovered member of the BMP family. BMP9 bound with high affinity to activin receptor-like kinase 1 (ALK1) and endoglin in non-parenchymal liver cells. In addition, BMP9 activated Smad1/Smad5/Smad8 and induced the expression of the target genes inhibitor of differentiation 1 (Id1), hepcidin, Snail and the co-receptor endoglin in liver cells. Although the role of BMP9 in liver fibrosis is currently poorly understood, the presence of BMP9-activated proteins and its target genes have been reported to be associated with liver fibrosis development. This review summarizes the indirect connection between BMP9 and liver fibrosis, with a focus on the BMP9 signaling pathway members ALK1, endoglin, Id1, hepcidin and Snail. The observations on the role of BMP9 in regulating liver fibrosis may help in understanding the pathology mechanisms of liver disease. Furthermore, BMP9 could be served as a potent biomarker and the target of potential therapeutic drugs to treat hepatocytes fibrosis. Full article

Due to improved medical care, life expectancy in patients with cystic fibrosis (CF) has veritably improved over the last decades. Importantly, cystic fibrosis related liver disease (CFLD) has become one of the leading causes of morbidity and mortality in CF patients. However, CFLD might be largely underdiagnosed and diagnostic criteria need to be refined. The underlying pathomechanisms are largely unknown, and treatment strategies with proven efficacy are lacking. This review focuses on current invasive and non-invasive diagnostic standards, the current knowledge on the pathophysiology of CFLD, treatment strategies, and possible future developments. Full article

Hepatocellular carcinoma (HCC) is one of the most common cancers with a dismal outcome. The complicated molecular pathogenesis of HCC caused by tumor heterogeneity makes it difficult to identify druggable targets useful for treating HCC patients. One approach that has a potential for the improvement of patient prognosis is the identification of cancer driver genes that play a critical role in the development of HCC. Recent technological advances of high-throughput methods, such as gene expression profiles, DNA copy number alterations and somatic mutations, have expanded our understanding of the comprehensive genetic profiles of HCC. Integrative analysis of these omics profiles enables us to classify the molecular subgroups of HCC patients. As each subgroup classified according to genetic profiles has different clinical features, such as recurrence rate and prognosis, the tumor subclassification tools are useful in clinical practice. Furthermore, a global genetic analysis, including genome-wide RNAi functional screening, makes it possible to identify cancer vulnerable genes. Identification of common cancer driver genes in HCC leads to the development of an effective molecular target therapy. Full article

Hepatic fibrosis is a wound-healing response to various chronic stimuli, including viral hepatitis B or C infection. Activated myofibroblasts, predominantly derived from the hepatic stellate cells (HSCs), regulate the balance between matrix metalloproteinases and their tissue inhibitors to maintain extracellular matrix homeostasis. Transforming growth factor-β and platelet-derived growth factor are classic profibrogenic signals that activate HSC proliferation. In addition, proinflammatory cytokines and chemokines coordinate macrophages, T cells, NK/NKT cells, and liver sinusoidal endothelial cells in complex fibrogenic and regression processes. In addition, fibrogenesis involves angiogenesis, metabolic reprogramming, autophagy, microRNA, and epigenetic regulations. Hepatic inflammation is the driving force behind liver fibrosis; however, host single nucleotide polymorphisms and viral factors, including the genotype, viral load, viral mutation, and viral proteins, have been associated with fibrosis progression. Eliminating the underlying etiology is the most crucial antifibrotic therapy. Growing evidence has indicated that persistent viral suppression with antiviral therapy can result in fibrosis regression, reduced liver disease progression, decreased hepatocellular carcinoma, and improved chances of survival. Preclinical studies and clinical trials are currently examining several investigational agents that target key fibrogenic pathways; the results are promising and shed light on this debilitating illness. Full article

Non-alcoholic fatty liver disease (NAFLD) which includes steatosis and steatohepatitis, in particular non-alcoholic steatohepatitis (NASH), is a rising health problem world-wide and should be separated from alcoholic steatohepatitis (ASH). NAFLD is regarded as hepatic manifestation of the metabolic syndrome (MetSy), being tightly linked to obesity and type 2 diabetes mellitus (T2DM). Development of steatosis, liver fibrosis and cirrhosis often progresses towards hepatocellular carcinogenesis and frequently results in the indication for liver transplantation, underlining the clinical significance of this disease complex. Work on different murine models and several human patients studies led to the identification of different molecular key players as well as epigenetic factors like miRNAs and SNPs, which have a promoting or protecting function in AFLD/ASH or NAFLD/NASH. To which extent they might be translated into human biology and pathogenesis is still questionable and needs further investigation regarding diagnostic parameters, drug development and a better understanding of the genetic impact. In this review we give an overview about the currently available knowledge and recent findings regarding the development and progression of this disease. Full article

Hepatocellular carcinoma (HCC) is a prevalent primary liver cancer that is derived from hepatocytes and is characterised by high mortality rate and poor prognosis. While HCC is driven by cumulative changes in the hepatocyte genome, it is increasingly recognised that the liver microenvironment plays a pivotal role in HCC propensity, progression and treatment response. The microenvironmental stimuli that have been recognised as being involved in HCC pathogenesis are diverse and include intrahepatic cell subpopulations, such as immune and stellate cells, pathogens, such as hepatitis viruses, and non-cellular factors, such as abnormal extracellular matrix (ECM) and tissue hypoxia. Recently, a number of novel environmental influences have been shown to have an equally dramatic, but previously unrecognized, role in HCC progression. Novel aspects, including diet, gastrointestinal tract (GIT) microflora and circulating microvesicles, are now being recognized as increasingly important in HCC pathogenesis. This review will outline aspects of the HCC microenvironment, including the potential role of GIT microflora and microvesicles, in providing new insights into tumourigenesis and identifying potential novel targets in the treatment of HCC. Full article

In chronic liver disease leading to fibrosis, hepatic stellate cells (HSC) differentiate into myofibroblasts. Myofibroblastic HSC have taken center stage during liver fibrogenesis, due to their remarkable synthesis of extracellular matrix proteins, their secretion of profibrogenic mediators and their contribution to hypertension, due to elevated contractility. MicroRNAs (miRNAs) are small, noncoding RNA molecules of 19–24 nucleotides in length. By either RNA interference or inhibition of translational initiation and elongation, each miRNA is able to inhibit the gene expression of a wide panel of targeted transcripts. Recently, it was shown that altered miRNA patterns after chronic liver disease highly affect the progression of fibrosis by their potential to target the expression of extracellular matrix proteins and the synthesis of mediators of profibrogenic pathways. Here, we underline the role of miRNAs in the interplay of the profibrogenic cell communication pathways upon myofibroblastic differentiation of hepatic stellate cells in the chronically injured liver. Full article

Chronic hepatitis C virus (HCV) infection is an important cause of morbidity and mortality in people coinfected with human immunodeficiency virus (HIV). Several studies have shown that HIV infection promotes accelerated HCV hepatic fibrosis progression, even with HIV replication under full antiretroviral control. The pathogenesis of accelerated hepatic fibrosis among HIV/HCV coinfected individuals is complex and multifactorial. The most relevant mechanisms involved include direct viral effects, immune/cytokine dysregulation, altered levels of matrix metalloproteinases and fibrosis biomarkers, increased oxidative stress and hepatocyte apoptosis, HIV-associated gut depletion of CD4 cells, and microbial translocation. In addition, metabolic alterations, heavy alcohol use, as well drug use, may have a potential role in liver disease progression. Understanding the pathophysiology and regulation of liver fibrosis in HIV/HCV co-infection may lead to the development of therapeutic strategies for the management of all patients with ongoing liver disease. In this review, we therefore discuss the evidence and potential molecular mechanisms involved in the accelerated liver fibrosis seen in patients coinfected with HIV and HCV. Full article

Nonalcoholic fatty liver disease (NAFLD) affects about 30% of the general population in the United States and includes a spectrum of disease that includes simple steatosis, non-alcoholic steatohepatitis (NASH), fibrosis and cirrhosis. Significant insight has been gained into our understanding of the pathogenesis of NALFD; however the key metabolic aberrations underlying lipid accumulation in hepatocytes and the progression of NAFLD remain to be elucidated. Accumulating and emerging evidence indicate that hepatic mitochondria play a critical role in the development and pathogenesis of steatosis and NAFLD. Here, we review studies that document a link between the pathogenesis of NAFLD and hepatic mitochondrial dysfunction with particular focus on new insights into the role of impaired fatty acid oxidation, the transcription factor peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), and sirtuins in development and progression of NAFLD. Full article

Kupffer cells reside within the liver sinusoid and serve as gatekeepers. They produce pro- and anti-inflammatory cytokines and other biologically important molecules upon the engagement of pattern recognition receptors such as Toll-like receptors. Kupffer cell-ablated mice established by in vivo treatment with clodronate liposomes have revealed many important features of Kupffer cells. In this paper, we review the importance of Kupffer cells in murine acute liver injuries and focus on the following two models: lipopolysaccharide (LPS)-induced liver injury, which is induced by priming with Propionibacterium acnes and subsequent challenge with LPS, and hypercoagulability-mediated acute liver failure such as that in concanavalin A (Con A)-induced hepatitis. Kupffer cells are required for LPS sensitization induced by P. acnes and are a major cellular source of interleukin-18, which induces acute liver injury following LPS challenge. Kupffer cells contribute to Con A-induced acute liver failure by initiating pathogenic, intrasinusoidal thrombosis in collaboration with sinusoidal endothelial cells. The mechanisms underlying these models may shed light on human liver injuries induced by various etiologies such as viral infection and/or abnormal metabolism. Full article

Hepatic diseases, like viral hepatitis, autoimmune hepatitis, hereditary hemochromatosis, non-alcoholic fatty liver disease (NAFLD) and Wilson’s disease, play an important role in the development of liver cirrhosis and, hence, hepatocellular carcinoma. In this review, the current treatment options and the molecular mechanisms of action of the drugs are summarized. Unfortunately, the treatment options for most of these hepatic diseases are limited. Since hepatitis B (HBV) and C (HCV) infections are the most common causes of liver cirrhosis and hepatocellular carcinoma, they are the focus of the development of new drugs. The current treatment of choice for HBV/HCV infection is an interferon-based combination therapy with oral antiviral drugs, like nucleos(t)ide analogues, which is associated with improving the therapeutic success and also preventing the development of resistances. Currently, two new protease inhibitors for HCV treatment are expected (deleobuvir, faldaprevir) and together with the promising drug, daclatasvir (NS5A-inhibitor, currently in clinical trials), adequate therapy is to be expected in due course (circumventing the requirement of interferon with its side-effects), while in contrast, efficient HBV therapeutics are still lacking. In this respect, entry inhibitors, like Myrcludex B, the lead substance of the first entry inhibitor for HBV/HDV (hepatitis D) infection, provide immense potential. The pharmacokinetics and the mechanism of action of Myrcludex B are described in detail. Full article

Non-alcoholic steatohepatitis (NASH) is a severe form of non-alcoholic fatty liver disease (NAFLD), in which most patients exhibit non-progressive, non-alcoholic fatty liver (NAFL) attributable to simple steatosis. Multiple hits, including genetic differences, fat accumulation, insulin resistance and intestinal microbiota changes, account for the progression of NASH. NAFLD is strongly associated with obesity, which induces adipokine secretion, endoplasmic reticulum (ER) and oxidative stress at the cellular level, which in turn induces hepatic steatosis, inflammation and fibrosis. Among these factors, gut microbiota are acknowledged as having an important role in initiating this multifactorial disease. Oxidative stress is considered to be a key contributor in the progression from NAFL to NASH. Macrophage infiltration is apparent in NAFL and NASH, while T-cell infiltration is apparent in NASH. Although several clinical trials have shown that antioxidative therapy with vitamin E can effectively control hepatitis pathology in the short term, the long-term effects remain obscure and have often proved to be ineffective in many other diseases. Several long-term antioxidant protocols have failed to reduce mortality. New treatment modalities that incorporate current understanding of NAFLD molecular pathogenesis must be considered. Full article

Bone morphogenetic proteins (BMP-2 to BMP-15) belong to the Transforming Growth Factor (TGF)-β superfamily and, besides their well-documented roles during embryogenesis and bone formation, some of them have recently been described to be involved in the pathogenesis of different organs, including the liver. The role of BMPs in liver damage responses including hepatocellular carcinoma (HCC) development has only begun to be addressed and strong evidence supports the concept of a pro-tumorigenic role of BMP signaling in HCC cells. BMP-9 (also termed Growth and Differentiation Factor (GDF)-2) represents the most recently discovered member of the BMP family. We have previously demonstrated that in HCC patient samples BMP-9 expression was positively associated with the tumor seize (“T stage”) and that it enhanced cell migration and induced epithelial to mesenchymal transition (EMT) in HCC cells in vitro. In another study we recently found that BMP-9 promotes growth in HCC cells, but not in non-transformed hepatocytes. Published as well as unpublished results obtained with primary hepatocytes support the concept of a dual function of BMP-9 in the liver: while in primary, non-malignant cells BMP-9 stabilizes the epithelial phenotype and inhibits proliferation, in HCC cells it induces cell growth and the acquisition of a migratory phenotype. In this review article we summarize current knowledge about BMPs in liver diseases, with special focus on the role of BMP-9 in HCC development and progression, that may provide new clues for a better understanding of the contribution of BMP-signaling to chronic liver diseases. Full article

The hepatitis C virus (HCV) is a global health problem affecting more than 170 million people. A chronic HCV infection is associated with liver fibrosis, liver cirrhosis and hepatocellular carcinoma. To enable viral persistence, HCV has developed mechanisms to modulate both innate and adaptive immunity. The recruitment of antiviral immune cells in the liver is mainly dependent on the release of specific chemokines. Thus, the modulation of their expression could represent an efficient viral escape mechanism to hamper specific immune cell migration to the liver during the acute phase of the infection. HCV-mediated changes in hepatic immune cell chemotaxis during the chronic phase of the infection are significantly affecting antiviral immunity and tissue damage and thus influence survival of both the host and the virus. This review summarizes our current understanding of the HCV-mediated modulation of chemokine expression and of its impact on the development of liver disease. A profound knowledge of the strategies used by HCV to interfere with the host’s immune response and the pro-fibrotic and pro-carcinogenic activities of HCV is essential to be able to design effective immunotherapies against HCV and HCV-mediated liver diseases. Full article

Chronic hepatitis B virus (HBV) infection, affecting approximately 240 million people worldwide, is a major public health problem that elevates the risk of developing liver cirrhosis and hepatocellular carcinoma. Given that current anti-HBV drugs are limited to interferon-based regimens and nucleos(t)ide analogs, the development of new anti-HBV agents is urgently needed. The viral entry process is generally an attractive target implicated in antiviral strategies. Using primary cells from humans and Tupaia belangeri, as well as HepaRG cells, important determinants of viral entry have been achieved. Recently, sodium taurocholate cotransporting polypeptide (NTCP) was identified as an HBV entry receptor and enabled the establishment of a susceptible cell line that can efficiently support HBV infection. This finding will allow a deeper understanding of the requirements for efficient HBV infection, including the elucidation of the molecular entry mechanism. In addition, pharmacological studies suggest that NTCP is able to serve as a therapeutic target. This article summarizes our current knowledge on the mechanisms of HBV entry and the role of NTCP in this process. Full article