Intrahepatic angiopoietin-2 correlates with chronic hepatitis C progression and is induced in hepatitis C virus replicon systems.

BACKGROUND & AIMS: Chronic hepatitis C (CHC) is a major cause of cirrhosis and hepatocellular carcinoma and angiogenesis is closely related to the pathogenesis and progression of different chronic liver diseases (CLD). Thus, the intrahepatic expression of angiopoietins 1 and 2 (Ang1 and Ang2), as relevant mediators of pathological angiogenesis in several CLD, was investigated. In addition, the differential influence of structural and non-structural genomic regions of HCV on the expression of angiopoietins and the possible signalling involved were studied. METHODS: Ang1 and Ang2 expression was evaluated by western blotting and enzyme-linked immunosorbent assay (ELISA) in liver homogenates of CHC patients (n=47) and uninfected subjects (n=8). Their association with disease progression (according to METAVIR classification) was assessed by Spearman's correlation. Statistical differences among the expression of angiopoietins at different CHC stages were calculated by Mann-Whitney U-test. Finally, the in vitro expression of Angiopoietins in HCV replicons (complete or non-structural subgenomic) and the main signalling pathways involved were also examined. RESULTS: Ang2 levels were significantly higher in the liver of CHC patients compared to controls and significantly correlated with inflammation and fibrosis. Accordingly, an increased expression of Ang2 was found in all HCV replicons tested. Interestingly, the inhibition of MEK and PI3K signalling pathways exerted differential effects on Ang2 expression concerning to the genomic region of HCV. CONCLUSIONS: Hepatitis C virus induces Ang2 expression in hepatocytes through different signalling routes which may lead to the disregulation of vascular homeostasis in the liver. Thus, pharmacologic intervention on Ang2 signalling might constitute an important therapeutic tool.

High-resolution hepatitis C virus subtyping using NS5B deep sequencing and phylogeny, an alternative to current methods.

Hepatitis C virus (HCV) is classified into seven major genotypes and 67 subtypes. Recent studies have shown that in HCV genotype 1-infected patients, response rates to regimens containing direct-acting antivirals (DAAs) are subtype dependent. Currently available genotyping methods have limited subtyping accuracy. We have evaluated the performance of a deep-sequencing-based HCV subtyping assay, developed for the 454/GS-Junior platform, in comparison with those of two commercial assays (Versant HCV genotype 2.0 and Abbott Real-time HCV Genotype II) and using direct NS5B sequencing as a gold standard (direct sequencing), in 114 clinical specimens previously tested by first-generation hybridization assay (82 genotype 1 and 32 with uninterpretable results). Phylogenetic analysis of deep-sequencing reads matched subtype 1 calling by population Sanger sequencing (69% 1b, 31% 1a) in 81 specimens and identified a mixed-subtype infection (1b/3a/1a) in one sample. Similarly, among the 32 previously indeterminate specimens, identical genotype and subtype results were obtained by direct and deep sequencing in all but four samples with dual infection. In contrast, both Versant HCV Genotype 2.0 and Abbott Real-time HCV Genotype II failed subtype 1 calling in 13 (16%) samples each and were unable to identify the HCV genotype and/or subtype in more than half of the non-genotype 1 samples. We concluded that deep sequencing is more efficient for HCV subtyping than currently available methods and allows qualitative identification of mixed infections and may be more helpful with respect to informing treatment strategies with new DAA-containing regimens across all HCV subtypes.

Preliminary evidence of sustained expression of angiopoietin-2 during monocyte differentiation in chronic hepatitis C.

BACKGROUND: Monocytes are essential precursors of antigen-presenting cells, such as macrophages and dendritic cells, and contribute to the pathogenesis of chronic inflammatory diseases and cancer. AIMS: As Tie2-expressing monocytes (TEMs) are increased in the peripheral blood of patients with chronic hepatitis C (CHC), we aimed to examine the expression of Tie2 and angiopoietins (Ang1 and Ang2) during monocyte differentiation and maturation in CHC. METHODS: The expression of Tie2, CD11b, CD80, CD83, CD86 and MHC-II was measured by flow cytometry in peripheral blood monocytes and monocytes-derived cells (Mo-DCs) from nine healthy subjects and eight CHC patients whose HCV infection was unresolved after combination therapy. Ang1 and Ang2 levels were measured in cellular supernatants by ELISA. RESULTS: Mo-DCs from CHC patients expressed differential patterns of maturation markers compared with controls--primarily with regard to CD80. Tie2 was downregulated during monocyte differentiation in controls and CHC patients, whereas Ang1 expression was constant. However, Ang2 levels fell significantly during the differentiation of control monocytes, in contrast with those from CHC patients in whom Ang2 expression remained stable throughout differentiation. CONCLUSIONS: Altered expression of the Ang/Tie2 system in monocytes and Mo-DCs from CHC patients might account for the inflammatory and angiogenic disorders that are related to CHC. An increased understanding of Ang/Tie2 system regulation might be helpful in designing strategies to prevent CHC progression.

A predictive model of treatment outcome in patients with chronic HCV infection using IL28B and PD-1 genotyping.

BACKGROUND & AIMS: The advent of new chronic hepatitis C virus (HCV) therapies requires characterization of patients in order to predict adequate treatment. A good candidate marker is Programmed Cell Death-1 (PD-1) which is involved in progression of HCV infection. The aim of this study was to analyse the effect of several single nucleotide polymorphisms of PD-1 gene and several previously associated factors (IL28B and KIR receptors) on treatment responses. METHODS: 407 HCV chronic infected patients treated with PEG-IFN-α and ribavirin were recruited and classified according to their response to treatment. They were genotyped for PD-1 and IL28B polymorphisms, killer immunoglobulin-like receptors (KIR) and HLA genes. A multivariate logistic regression analysis and a Chi-squared Automatic Interaction Detector (CHAID) prediction model of response included these and other clinical parameters. RESULTS: Our results showed that PD-1.3/A allele was significantly associated with sustained virological response (SVR) in a multivariate logistic regression analysis (p<0.01, OR=2.57). Additionally, IL28B C/C genotype was the most significant predictor of an SVR to treatment in all HCV genotypes (74.5%). In IL28B C/C patients, the presence of PD-1.3/A allele increased the probability of an SVR to 93.3%. Moreover, when this analysis was made only with patients infected by HCV-1, the predictive value of IL28B C/C genotype with PD-1.3/A allele was 90%. CONCLUSIONS: PD-1.3/A allele is associated with SVR to treatment and notably increases the predictive value of IL28B C/C genotype. Both markers in conjunction could be a useful tool, more relevant than HCV genotype in some cases, in clinical practice.

The molecular and pathophysiological implications of hepatitis B X antigen in chronic hepatitis B virus infection.

Hepatitis B virus is considered one of the most significant environmental carcinogens in humans. Because the mechanisms of HBV replication and the development of hepatocellular carcinoma (HCC) are partially known, HBV-associated pathogenesis remains a challenge to increase its understanding. Evidence suggests that the regulatory protein hepatitis B virus X (HBx) mediates the establishment and maintenance of the chronic carrier state. HBx is a multifunctional and potentially oncogenic protein that is conserved among mammalian hepadnaviruses; it is produced very early after infection and throughout the chronic phase. HBx exerts its effects by interacting with cellular proteins and activating various signaling pathways. HBx stimulates the transcription of genes that regulate cell growth, apoptosis, and DNA repair. It also interacts with proteasome subunits and affects mitochondrial stability. Moreover, HBx participates in processes that are associated with the progression of chronic liver disease, including angiogenesis and fibrosis. This review discusses the function of HBx in the life cycle of HBV and its contribution to the pathogenesis of HCC.

Importance of host genetic factors HLA and IL28B as predictors of response to pegylated interferon and ribavirin.

OBJECTIVES: Viral factors are considered the best predictors of response to treatment for chronic hepatitis C (CHC), but genetic factors are known to have an important role in this respect. This paper investigates the relationships among the host genetic factors HLA and IL28B, viral factors, and the outcome of combination therapy. METHODS: A multicenter retrospective cohort of 428 previously untreated CHC patients was treated with pegylated interferon/ribavirin (pegIFN/RBV) for 48 weeks. In all, 378 (88%) of these patients were genotype 1 or 4, and 50 (12%) were genotype 2 or 3. RESULTS: Multivariate logistic regression showed the rs12979860 CC genotype (adjusted odds ratio (aOR)=4.3, 95% confidence interval (95% CI): 2.6-7), the HLA-DQB1*0301 allele (aOR=2.08, 95% CI: 1.2-3.5) and age, viral genotype, and viral load levels to be significantly associated with sustained virological response (SVR). When the variable rs12979860 was eliminated, the area under the receiver operating characteristic (ROC) curve (AUC) decreased significantly (0.76 vs. 0.69; P=0.03). AUC values derived from viral factors were lower than those corresponding to host genetic factors (0.67 vs. 0.72, respectively; P=0.04). The HLA-DQB1*0301 and A*0201 alleles were associated with rs12979860 CC genotype and SVR (P<0.0001). CONCLUSIONS: The HLA-DQB1*0301 allele and IL28B genotype are factors that are associated independently with SVR. There is a synergism between the HLA-DQB1*0301 and HLA-A*0201 alleles with polymorphism rs12979860 CC, which increases the SVR rate. IL28B genotype is the best predictor of SVR.

Corticosteroids modulate angiogenic soluble factors in ulcerative colitis patients.

AIMS: The aim of this study was to compare angiogenic factors in serum levels of active ulcerative colitis (UC) patients and in healthy controls, and to analyze these angiogenic levels depending on the achievement of remission after oral corticosteroid treatment throughout treatment, and according to the Truelove-Witts activity index. METHODS: Blood samples were collected from 13 patients receiving oral corticosteroids for treatment of UC flares at three different intervals--baseline, during, and after treatment--and from 26 healthy controls. Vascular endothelial growth factor (VEGF), placental growth factor (PlGF), VEGF receptor 1 (VEGFR1), angiopoietins (Ang) 1 and 2, and its receptor Tie2 were assayed by ELISA. RESULTS: While VEGF and Ang2 levels in UC patients were higher than in healthy controls (P < 0.05), UC patients showed lower levels of Ang1 than healthy individuals (P < 0.05). In UC patients who achieved clinical remission after corticosteroid treatment, a statistically significant higher baseline serum level of PlGF was observed (22 ± 5 vs. 18 ± 2; P < 0.05). Angiogenic factor levels varied during treatment; however, they did not show a statistically significant correlation to the activity of the disease. CONCLUSIONS: VEGF, Ang1, and Ang2 levels showed statistically significant differences between UC patients and healthy controls. Although determination of PlGF serum levels before corticosteroid treatment might be helpful to anticipate the response by UC patients, no angiogenic pattern that could accurately predict "a priori" this response to corticosteroid treatment was observed. Corticosteroids altered temporarily circulating levels of VEGF, angiopoietins and Tie2. No correlation was found between systemic levels of angiogenic factors and the clinical activity of UC.

Genetic contribution of endoplasmic reticulum aminopeptidase 1 polymorphisms to liver fibrosis progression in patients with HCV infection.

The endoplasmic reticulum aminopeptidase ERAP1 regulates innate and adaptive immune responses, trimming peptides and loading onto HLA class I molecules. Coding single nucleotide polymorphisms within ERAP1 are associated with autoimmune diseases, viral infections, and cancer development. Our purpose was to analyze the influence of ERAP1 variants on fibrogenesis in hepatitis C virus (HCV)-infected patients. A range of ERAP1 polymorphisms were genotyped in 722 unrelated Caucasian patients diagnosed with chronic HCV from two Spanish cohorts. Patients were classified according to their fibrosis stage. Paraffin-embedded tissue microarrays were constructed to assess ERAP1 expression (HCV = 38; alcoholic = 20) by immunohistochemistry. A statistical algorithm was applied to derive a fibrogenesis prediction model. The ERAP1 variants rs30187/T (K528, pc < 0.001) and rs27044/G (Q730, pc < 0.001) were related with severe fibrosis. These results were validated in the two independent cohorts. Furthermore, patients with the rs30187/T allele had stronger ERAP1 protein expression than those with the rs30187/C (p < 0.05). The statistical model showed that patients with rs30187 C/T and T/T genotypes took 15.58 years (median) to develop advanced fibrosis, but this value was 32.08 years in patients carrying C/C genotype (p < 0.005). ERAP1 variants may influence the clinical course of fibrogenesis in HCV-infected patients. These polymorphisms could be exploited as constitutive new markers of fibrosis evolution. The results highlight the possibility of using modulators of ERAP1 to generate a protective immune response against chronic HCV infection. KEY MESSAGES: What is known Several ERAP1 polymorphisms are associated with autoimmune diseases and cancer. ERAP1 trims peptides to HLA class I presentation. What is new here ERAP1 polymorphisms are associated with fibrogenesis. The ERAP1 polymorphisms genotype could help us in clinical management of patients. Potential translational impact The use of modulators of ERAP1 could generate a protective response depending on SNPs.

Deep-sequencing reveals broad subtype-specific HCV resistance mutations associated with treatment failure.

A percentage of hepatitis C virus (HCV)-infected patients fail direct acting antiviral (DAA)-based treatment regimens, often because of drug resistance-associated substitutions (RAS). The aim of this study was to characterize the resistance profile of a large cohort of patients failing DAA-based treatments, and investigate the relationship between HCV subtype and failure, as an aid to optimizing management of these patients. A new, standardized HCV-RAS testing protocol based on deep sequencing was designed and applied to 220 previously subtyped samples from patients failing DAA treatment, collected in 39 Spanish hospitals. The majority had received DAA-based interferon (IFN) α-free regimens; 79% had failed sofosbuvir-containing therapy. Genomic regions encoding the nonstructural protein (NS) 3, NS5A, and NS5B (DAA target regions) were analyzed using subtype-specific primers. Viral subtype distribution was as follows: genotype (G) 1, 62.7%; G3a, 21.4%; G4d, 12.3%; G2, 1.8%; and mixed infections 1.8%. Overall, 88.6% of patients carried at least 1 RAS, and 19% carried RAS at frequencies below 20% in the mutant spectrum. There were no differences in RAS selection between treatments with and without ribavirin. Regardless of the treatment received, each HCV subtype showed specific types of RAS. Of note, no RAS were detected in the target proteins of 18.6% of patients failing treatment, and 30.4% of patients had RAS in proteins that were not targets of the inhibitors they received. HCV patients failing DAA therapy showed a high diversity of RAS. Ribavirin use did not influence the type or number of RAS at failure. The subtype-specific pattern of RAS emergence underscores the importance of accurate HCV subtyping. The frequency of "extra-target" RAS suggests the need for RAS screening in all three DAA target regions.

The hepatitis B virus X protein induces paracrine activation of human hepatic stellate cells.

UNLABELLED: Chronic hepatitis B virus (HBV) infection is a major cause of liver fibrosis, eventually leading to cirrhosis and hepatocellular carcinoma. Although the involvement of the X protein of HBV (HBx) in viral replication and tumor development has been extensively studied, little is known about its possible role in the development of fibrosis. In this work we show that expression of HBx in hepatocytes results in paracrine activation and proliferation of hepatic stellate cells (HSCs), the main producers of extracellular matrix proteins in the fibrotic liver. Both human primary HSCs and rat HSCs exposed to conditioned medium from HBx-expressing hepatocytes showed increased expression of collagen I, connective tissue growth factor, alpha smooth muscle actin, matrix metalloproteinase-2, and transforming growth factor-beta (TGF-beta), together with an enhanced proliferation rate. We found that HBx induced TGF-beta secretion in hepatocytes and that the activation of HSCs by conditioned medium from HBx-expressing hepatocytes was prevented by a neutralizing anti-TGF-beta antibody, indicating the involvement of this profibrotic factor in the process. CONCLUSION: Our results propose a direct role for HBx in the development of liver fibrosis by the paracrine activation of stellate cells and reinforce the indication of antiviral treatment in patients with advanced HBV-related chronic liver disease and persistent liver replication.

Mechanisms of angiogenesis in chronic inflammatory liver disease.

Intrahepatic hypoxia may occur during the inflammatory and fibrotic processes that characterize several chronic liver diseases of viral and autoimmune origin. As a consequence, new vascular structures are formed to provide oxygen and nutrients. Angiogenesis involves a tightly regulated network of cellular and molecular mechanisms that result in the formation of functional vessels. Of particular importance are growth factors and molecules involved in matrix remodeling and cell migration, as weel as vessel maturation-related factors. In recent years a number of studies have investigated the expression and function of many pro- and antiangiogenic molecules in chronic liver diseases and liver regeneration. This review examines the potential pathogenic role of angiogenesis in the context of viral hepatitis, autoinmmune hepatitis, primary biliary cirrhosis and hepatocellular carcinoma.

Interferon-related genetic markers of necroinflammatory activity in chronic hepatitis C.

INTRODUCTION: Chronic hepatitis C (CHC) is a major cause of liver disease worldwide which often leads to progressive liver inflammation, fibrosis, cirrhosis and hepatocellular carcinoma (HCC). CHC displays heterogeneous progression depending on a broad set of factors, some of them intrinsic to each individual such as the patient's genetic profile. This study aims to evaluate the contribution of certain genetic variants of crucial interferon alpha and lambda signaling pathways to the hepatic necroinflammatory activity (NIA) grade of CHC patients. METHODS: NIA was evaluated in 119 CHC patients by METAVIR scale and classified as low (NIA = 0-2, n = 80) or high grade (NIA = 3, n = 39). In a candidate gene approach, 64 SNPs located in 30 different genes related to interferon pathways (IL-28B, IFNAR1-2, JAK-STAT and OAS1-3, among others) were genotyped using the Illumina GoldenGate® Genotyping Assay. Statistical association was determined by logistic regression and expressed as OR and 95% CI. Those SNPs significantly associated were further adjusted by other covariates. RESULTS: Seven SNPs located in IL-28B (rs12979860), JAK1 (rs11576173 and rs1497056), TYK2 (rs280519), OAS1 (rs2057778), SOCS1 (rs33932899) and RNASEL (rs3738579) genes were significantly related to severe NIA grade (p<0.05). Regarding to clinical variables, elevated NIA was notably associated with aspartate aminotransferase (AST) serum levels >40 IU/L (p<0.05) but not with other clinical factors. Multivariate logistic regression analysis of these factors reflected that AST (>40 IU/L), TYK2 rs280519 (G allele) and RNASEL rs3738579 (G allele) were factors independently associated with elevated NIA (p<0.05). AST concentration showed a moderate AUC value (AUC = 0.63), similar to TYK2 (rs280519) and RNASEL (rs3738579) SNPs (AUC = 0.61, both) in the ROC_AUC analysis. Interestingly, the model including all significant variables reached a considerable predictive value (AUC = 0.74). CONCLUSION: The identified genetic variants in interferon signaling pathways may constitute useful prognostic markers of CHC progression. Further validation in larger cohorts of patients is needed.

Angiopoietin-2/angiopoietin-1 as non-invasive biomarker of cirrhosis in chronic hepatitis C.

AIM: To evaluate the efficacy of peripheral blood concentrations of angiopoietins (Ang) as cirrhosis biomarkers of chronic hepatitis C (CHC). METHODS: Ang1 and Ang2 serum levels were measured by enzyme-linked immunosorbent assays (ELISA) in samples from 179 cirrhotic and non-cirrhotic CHC patients, classified according to the METAVIR system. Groups were compared by non-parametric Mann-Whitney U test. Subsequently, the association of peripheral concentrations of angiopoietins with the stage of fibrosis was analyzed using Spearman correlation test. Finally, the accuracy, sensitivity and specificity of circulating angiopoietins for cirrhosis diagnosis were determined by the study of the respective area under the curve of receiver operator characteristics (AUC-ROC). RESULTS: Peripheral blood concentrations of Ang1 and Ang2 in CHC patients were significantly related to fibrosis. While Ang1 was decreased in cirrhotic subjects compared to non-cirrhotic (P < 0.0001), Ang2 was significantly increased as CHC progressed to the end stage of liver disease (P < 0.0001). Consequently, Ang2/Ang1 ratio was notably amplified and significantly correlated with fibrosis (P < 0.0001). Interestingly, the individual performance of each angiopoietin for the diagnosis of cirrhosis reached notable AUC-ROC values (above 0.7, both), but the Ang2/Ang1 ratio was much better (AUC-ROC = 0.810) and displayed outstanding values of sensitivity (71%), specificity (84%) and accuracy (82.1%) at the optimal cut-off (10.33). Furthermore, Ang2/Ang1 ratio improved the performance of many other previously described biomarkers or scores of liver cirrhosis in CHC. CONCLUSION: Ang2/Ang1 ratio might constitute a useful tool for monitoring the progression of chronic liver disease towards cirrhosis and play an important role as therapeutic target.

Inhibition of tyrosine kinase receptor Tie2 reverts HCV-induced hepatic stellate cell activation.

BACKGROUND: Hepatitis C virus (HCV) infection is a major cause of chronic liver disease (CLD) and is frequently linked to intrahepatic microvascular disorders. Activation of hepatic stellate cells (HSC) is a central event in liver damage, due to their contribution to hepatic renewal and to the development of fibrosis and hepatocarcinoma. During the progression of CLDs, HSC attempt to restore injured tissue by stimulating repair processes, such as fibrosis and angiogenesis. Because HSC express the key vascular receptor Tie2, among other angiogenic receptors and mediators, we analyzed its involvement in the development of CLD. METHODS: Tie2 expression was monitored in HSC cultures that were exposed to media from HCV-expressing cells (replicons). The effects of Tie2 blockade on HSC activation by either neutralizing antibody or specific signaling inhibitors were also examined. RESULTS: Media from HCV-replicons enhanced HSC activation and invasion and upregulated Tie2 expression. Notably, the blockade of Tie2 receptor (by a specific neutralizing antibody) or signaling (by selective AKT and MAPK inhibitors) significantly reduced alpha-smooth muscle actin (α-SMA) expression and the invasive potential of HCV-conditioned HSC. CONCLUSIONS: These findings ascribe a novel profibrogenic function to Tie2 receptor in the progression of chronic hepatitis C, highlighting the significance of its dysregulation in the evolution of CLDs and its potential as a novel therapeutic target.

Diversity of killer cell immunoglobulin-like receptor (KIR) genotypes and KIR2DL2/3 variants in HCV treatment outcome.

The aim of this study was to analyse the distribution of KIR haplotypes and the KIR2DL2/3 alleles in chronic HCV-infected patients in order to establish the influence on the response to pegylated interferon plus ribavirin classical treatment. The alleles study of previously associated KIR2DL2/3 showed that KIR2DL2*001 was more frequent in non-SVR (NSVR) (42.2% vs. 27.5%, p<0.05) and KIR2DL3*001 was associated with sustained viral response (SVR) (41.6% vs. 61.2%, p<0.005). The KIR2DL3*001-HLA-C1 association was also significant (24.5% vs. 45.7%, p<0.001). From the frequencies of KIR obtained, 35 genotypes were assigned on the basis of previous studies. The centromeric A/A genotype was more frequent in SVR (44.1% vs. 34.5%, p<0.005) and the centromeric B/B genotype was found to be significantly more frequent in NSVR (20.9% vs. 11.2%, p<0.001). The logic regression model showed the importance of KIR genes in predicting the response to combined treatment, since the positive predictive value (PPV) was improved (from 55.9% to 75.3%) when the analysis of KIR was included in addition to the IFNL3 rs12979860 polymorphism. The study of KIR receptors may be a powerful tool for predicting the combined treatment response in patients with chronic HCV infection in association with the determination of IFNL3 polymorphism.

[Effects of hepatitis B virus X protein on chronic hepatitis B pathophysiology]. / Mecanismos de acción y efectos de la proteína X (HBx) en la patogenia de la hepatitis crónica B.

The infection by hepatitis B virus often promotes chronic liver inflammation which progresses to cirrhosis and hepatocellular carcinoma in a high percentage of patients. The persistent activation of the immune system causes an incessant liver damage, which fosters a disorganized stimulation of tissue repair and remodelling phenomena. In turn, the viral protein X (HBx) is essential for virus replication and therefore for the maintenance of chronic infection. However, the important oncogenic potential of HBx seems to reside, on one hand, in its ability to integrate into cellular DNA and, additionally, in the transactivation of different cellular signaling pathways involved in cell growth regulation, apoptosis and DNA repair. HBx also interacts with proteasome subunits and notably affects mitochondrial electric potential, thus altering cellular calcium homeostasis. Finally, this review discusses the pathogenic role of HBx in the progression of chronic hepatitis B through its effects on angiogenic, fibrogenic and oncogenic processes.

Angiopoietin-2 Serum Levels Improve Noninvasive Fibrosis Staging in Chronic Hepatitis C: A Fibrogenic-Angiogenic Link.

AIMS: Accurate liver fibrosis staging is crucial for the management of chronic hepatitis C (CHC). The invasiveness and cost burden of liver biopsy have driven the search for new noninvasive biomarkers of fibrosis. Based on the link between serum angiopoietin-1 and 2 levels and CHC progression, we aimed to determine the value of these angiogenic factors as noninvasive biomarkers of liver fibrosis. METHODS: Serum levels of angiopoietin-1 and -2 were measured by ELISA in 108 CHC patients who underwent pretreatment liver biopsy. The correlation between angiopoietins and clinical and demographic variables with liver fibrosis was analyzed by univariate regression. Significant factors were then subjected to multivariate analysis, from which we constructed a novel noninvasive liver fibrosis index (AngioScore), whose performance was validated in an independent series of 71 CHC patients. The accuracy of this model was compared with other documented fibrosis algorithms by De Long test. RESULTS: Angiopoietins correlated significantly with hepatic fibrosis; however, only angiopoietin-2 was retained in the final model, which also included age, platelets, AST, INR, and GGT. The model was validated and behaved considerably better than other fibrosis indices in discriminating all, significant, moderate and severe liver fibrosis (0.886, 0.920, 0.923). Using clinically relevant cutoffs, we classified CHC patients by discarding significant fibrosis and diagnosing moderate and severe fibrosis with greater accuracy, sensitivity, and specificity. CONCLUSIONS: Our novel noninvasive liver fibrosis model, based on serum angiopoietin-2 levels, outperforms other indices and should help substantially in managing CHC and monitoring long-term follow-up prognosis.

Angiogenesis: from chronic liver inflammation to hepatocellular carcinoma.

Recently, new information relating to the potential relevance of chronic hepatic inflammation to the development and progression of hepatocellular carcinoma (HCC) has been generated. Persistent hepatocellular injury alters the homeostatic balance within the liver; deregulation of the expression of factors involved in wound healing may lead to the evolution of dysplastic lesions into transformed nodules. Progression of such nodules depends directly on the development and organization of a vascular network, which provides the nutritional and oxygen requirements to an expanding nodular mass. Angiogenic stimulation promotes intense structural and functional changes in liver architecture and physiology, in particular, it facilitates transformation of dysplasia to nodular lesions with carcinogenic potential. HCC depends on the growth and spreading of vessels throughout the tumor. Because these vascular phenomena correlate with disease progression and prognosis, therapeutic strategies are being developed that focus on precluding vascular expansion in these tumors. Accordingly, an in-depth study of factors that promote and support pathological angiogenesis in chronic hepatic diseases may provide insights into methods of preventing the development of HCC and/or stimulating the regression of established HCC.