HCV-induced Liver Fibrosis Research Articles

Exploration of a hypoxia-immune-related microenvironment gene signature and prediction model for hepatitis C-induced early-stage fibrosis

BackgroundLiver fibrosis contributes to significant morbidity and mortality in Western nations, primarily attributed to chronic hepatitis C virus (HCV) infection. Hypoxia and immune status have been reported to be significantly correlated with the progression of liver fibrosis. The current research aimed to investigate the gene signature related to the hypoxia-immune-related microenvironment and identify potential targets for liver fibrosis.MethodSequencing data obtained from GEO were employed to assess the hypoxia and immune status of the discovery set utilizing UMAP and ESTIMATE methods. The prognostic genes were screened utilizing the LASSO model. The infiltration level of 22 types of immune cells was quantified utilizing CIBERSORT, and a prognosis-predictive model was established based on the selected genes. The model was also verified using qRT-PCR with surgical resection samples and liver failure samples RNA-sequencing data.ResultsElevated hypoxia and immune status were linked to an unfavorable prognosis in HCV-induced early-stage liver fibrosis. Increased plasma and resting NK cell infiltration were identified as a risk factor for liver fibrosis progression. Additionally, CYP1A2, CBS, GSTZ1, FOXA1, WDR72 and UHMK1 were determined as hypoxia-immune-related protective genes. The combined model effectively predicted patient prognosis. Furthermore, the preliminary validation of clinical samples supported most of the conclusions drawn from this study.ConclusionThe prognosis-predictive model developed using six hypoxia-immune-related genes effectively predicts the prognosis and progression of liver fibrosis. The current study opens new avenues for the future prediction and treatment of liver fibrosis.

CD4+ T-cell recovery in HIV/hepatitis C co-infected patients following successful hepatitis C treatment.

Hepatitis C virus (HCV)/HIV co-infection has been identified as a risk for impaired CD4+ T-cell recovery, possibly mediated by HCV-induced liver fibrosis and/or immune activation. As HCV direct-acting antivirals (DAAs) may partially reverse liver fibrosis and immune activation, sustained HCV virological response (SVR) may lead to improved CD4 recovery. We explored the effect of HCV DAA-induced SVR on CD4 recovery among patients living with both HCV and HIV, including those with poor CD4 recovery on antiretroviral therapy (immunological non-responders [INRs]). Subjects aged ≥18 years living with both HIV and HCV who achieved SVR with DAA were included. Pre-DAA CD4 counts were included only after sustained HIV viral suppression and HIV viral suppression was maintained for the duration of the study. Segmented regression of interrupted time series analysis was used to evaluate changes in median CD4 count in the pre-DAA period (-36 months) versus the post-DAA period (+36 months). In total, 156 patients were included. In the full cohort, median CD4 counts increased by 15% (p = 0.002) in the 6-month period following DAA initiation, whereafter CD4 counts decreased by 2.7% per 6-month period (p = 0.004). Among the 13 INRs, there was no immediate effect on median CD4 in the first 6 months after DAA initiation, whereafter there was a sustained CD4 increase (4.1% per 6-month time interval [p = 0.02]). In total, 54% of INRs recorded a post-DAA CD4 count of >350 cells/mm3. Successful DAA therapy induced a modest immediate CD4 immunological reconstitution among this cohort of patients living with both HIV and HCV, although this effect waned with time. By contrast, among INRs, achieving HCV SVR led to slower but sustained CD4 count recovery.

Direct-acting antiviral therapy may help restore HCV-induced impaired redox balance and liver fibrosis process

Abstract Objectives The aim of this study was to investigate the changes in thiol/disulfide balance, pro-fibrotic mediators (transforming growth factor-beta [TGF-β] and periostin) and a potential biomarker for the prediction of HCV-induced HCC (3β-hydroxysterol Δ24-reductase [DHCR24]) during direct-acting antiviral (DAA) therapy in chronic hepatitis C (CHC) patients. Methods This prospective cohort study included 56 non-cirrhotic, treatment-naive CHC patients who were treated with DAAs between January and June 2020. Laboratory tests, including serum total/native thiol, TGF-β , $\text{,}$ periostin, DHCR24, total bilirubin and albumin levels were measured and disulfide levels were calculated at baseline, then at 1 month and at the end of therapy (EOT). Results Of the 56 patients, all achieved a sustained virological response after DAA therapy. There was a significant decrease in serum levels of disulfide and TGF-β, (p=0.020 and p<0.001, respectively) and a significant increase in serum levels of native thiol compared with baseline levels (p=0.010). There was no significant change in levels of total thiol, DHCR24 and periostin levels. Serum TGF-β levels were found to be positively correlated with total bilirubin levels (rs=0.470, p=0.001) and negatively with albumin levels (rs=−0.483, p<0.001). A significant moderate positive correlation was determined between baseline serum DHRC24 and disulfide levels (rs=0.356, p=0.007). Conclusions The study results suggest that the DAA therapy may help to restore the impaired thiol/disulfide balance and reduce the pro-fibrotic process in CHC patients by markedly decreasing serum levels of TGF-β, a key player in HCV-induced liver fibrosis.

The Correlation Between Serum Collagen III Levels and APRI in Liver Fibrotic HCV Patients

Abstract Introduction/Objective Noninvasive Markers (NIM) of Liver Fibrosis help evaluate the stage of fibrosis in patients with no clear indication for a liver biopsy, such as patients with chronic hepatitis C who need monitoring of the stage of fibrosis during or after treatment. The perfect NIM for hepatic fibrosis should be inexpensive, safe, simple, reliable, and well-validated in different forms of chronic liver disease. Aim of the work Find if there is a correlation between serum collagen III and APRI as noninvasive markers in patients with HCV-induced liver fibrosis. Methods/Case Report 118 patients with HCV induced liver fibrosis (60 males and 58 females) and 50 healthy comparable control subjects (27 males and 23 females) were enrolled in the study. Collagen III was identified in serum samples of HCV-induced liver fibrosis patients at 70 kDa using collagen III polyclonal antibody and western blot analysis techniques. Collagen III was purified using the electroelution technique and then quantified using the ELISA technique. AST, ALT, albumin, and total bilirubin were measured. The APRI was calculated as [AST/ (upper limits of normal)/ platelets count 109/L]×100. The AST-ALT ratio was calculated as [AST/ ALT]. Results (if a Case Study enter NA) There was a statistically significant difference between the diseased and control groups (P<0.0001) regarding collagen III and APRI. Also, significant positive correlation between serum collagen III, APRI, and ALT levels (r=0.3 and p=0.02). However, there were no correlations between serum collagen III levels and AST, Albumin, Total Bilirubin, and AST/ALT ratio (r=0.15, -0.26, 0.22, and -0.027), respectively. Conclusion Collagen III and APRI is a promising diagnostic and prognostic potential in the noninvasive assessment of liver fibrosis as their levels elevate with each other. So, we can powerfully depend on them in liver fibrosis diagnosis in chronic HCV patients.

Assessment of Collagen III as a Non-Invasive Biomarker in HCV Induced Liver Fibrotic Patients

Abstract Introduction/Objective Liver biopsy is still the gold typical for assessing hepatic fibrosis. Non-invasive serum parameters measurement leads to essential changes in diagnosis. This procedure is painful, carries a risk of bleeding and infection, and is accompanied by a high sampling error rate. New biomarkers that can accurately predict the stage of fibrosis are urgently needed. Objectives This study aimed to identify, characterize, and detect serum collagen III in patients with HCV-induced liver fibrosis and healthy subjects. Methods/Case Report The study subjects were divided into two groups; group1: included 118 patients with HCV-induced liver fibrosis (60 males and 58 females). Group2: included 50 healthy comparable control subjects (27 males and 23 females). Collagen III was identified in serum samples of group1 at 70 kDa using collagen III polyclonal antibody and western blot analysis techniques. Additionally, it was purified from serum samples using the electroelution technique from preparative slab gels. The purified antigen gives a single sharp band at 70 kDa when resolved by SDS–PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and stained with coomassie blue. The 70 kDa purified collagen III showed high reactivity towards the collagen III polyclonal antibody. We used the ELISA method to quantify the collagen III, dose-response curve for purified collagen III as a function of the concentration in serum samples of liver fibrosis patients was done. Results (if a Case Study enter NA) The collagen III was identified in 62% of patients, and 38 % were negative collagen III (false negative) in group one. For group two, 96% of total healthy volunteers were negative for collagen III, and 4% were positive for collagen III (false positive). The mean ± SD of serum collagen III concentration was (8.38±3.56) and (3.2±0.59) for groups 1 and 2, respectively, with (P<0.0001). Conclusion The non-invasive assessment of liver fibrosis by collagen III has shown promising diagnostic and prognostic potential.

The Association Between Serum Hyaluronic Acid Levels and Routine Liver Function Tests in Liver Fibrotic HCV Patients

Abstract Introduction/Objective Liver fibrosis is the excessive accumulation of extracellular matrix proteins, including collagen, that occurs in most types of chronic liver diseases. Advanced liver fibrosis results in cirrhosis, liver failure, and portal hypertension and often requires liver transplantation. Markers (NIM) of Liver Fibrosis, such as hyaluronic acid, help evaluate the stage of fibrosis in patients with liver fibrosis. This study aims to investigate whether or not there is a correlation between serum hyaluronic acid levels and liver function tests in patients who suffer from chronic HCV-induced liver fibrosis. Methods/Case Report This study was conducted on 118 (60 males and 58 females) patients recruited from Internal Medicine Dept., Mansoura University Hospitals, Egypt. The patients were diagnosed with chronic HCV-induced liver fibrosis. Also, we included 50 healthy comparable control subjects (27 males and 23 females). Routine laboratory tests such as AST, ALT, albumin, total bilirubin, and platelets count were performed. The APRI was calculated as [AST/ (upper limits of normal)/ platelets count 109/L]×100. The AST-ALT ratio was calculated as [AST/ ALT]. According to the manufacturer's instructions, the serum levels of hyaluronic acid were measured by radioimmunoassay. Results (if a Case Study enter NA) We observed a statistically significant positive correlation was found between serum hyaluronic acid levels and APRI (r=0.3, p=0.001), ALT (r=0.3, p=0.003), AST (r=0.3 and p=0.01), and total bilirubin (r=0.3 and p=0.002). Furthermore, there was a statistically significant negative correlation between serum hyaluronic acid levels and albumin (r= -0.43, p=0.0001), and platelets count (r= -0.4, p=0.0001). Conclusion The levels of hyaluronic acid in the body and the results of other liver function tests tend to rise in tandem, which suggests a significant relationship between the two. This may assist us in constructing an accurate picture of the patient's condition, which will lead to a more precise diagnosis.

Urinary metabolomics of HCV patients with severe liver fibrosis before and during the sustained virologic response achieved by direct acting antiviral treatment

Hepatitis C virus (HCV) infection induces a long-term inflammatory response and oxidative-stress in the liver microenvironment, leading to hepatic fibrosis and metabolic alterations. Direct-acting-antiviral-agents (DAAs) induce HCV-clearance, even though liver damage is only partially restored. In this context, understanding the impact of viral-eradication on liver metabolic activities could allow optimizing the metabolic care of the patient. The present prospective longitudinal study aims at characterizing the urinary metabolic profile of HCV-induced severe liver fibrosis and the metabolic changes induced by DAAs and HCV-clearance by nuclear magnetic resonance-based metabolomics. The urinary metabolic profile of 23 HCV males with severe liver fibrosis and 20 age-matched healthy-controls was analyzed by NMR-based-metabolomics before starting DAAs, at the end-of-therapy, after one and three months of follow-up. The urinary metabolic profile of patients with severe liver fibrosis was associated to pseudouridine, hypoxanthine, methylguanidine and dimethylamine, highlighting a profile related to oxidative damage, and to tyrosine and glutamine, related to a decreased breakdown of aromatic aminoacids and ammonia detoxification, respectively.1-methylnicotinamide, a catabolic intermediate of nicotinamide-adenine-dinucleotide, was significantly increased in HCV-patients and restored after HCV-clearance, probably due to the reduced hepatic inflammation. 3-hydroxy-3-methylbutyrate, an intermediate of leucine-catabolism which was permanently restored after HCV-clearance, suggested an improvement of skeletal muscle protein synthesis. Finally, 3-hydroxyisobutyrate and 2,3-dihydroxy-2-methylbutyrate, intermediates of valine-catabolism, glycine and choline increased temporarily during therapy, resulting as potential biomarkers of DAAs systemic effects.

Oxidative stress, immunological and cellular hypoxia biomarkers in hepatitis C treatment-naïve and cirrhotic patients.

IntroductionHepatitis C virus (HCV) is the main cause of chronic liver disease, with calamitous complications. Its highest rate is recorded in Egypt. This study investigated whether oxidative stress, immunological chaos and cellular hypoxia are implicated in the pathophysiology of the disease.Material and methodsThis cross-sectional study aimed to evaluate the changes in blood oxidative stress, cellular hypoxia/angiogenesis and cellular immunological biomarkers in hospital-diagnosed treatment-naïve HCV-infected Upper Egyptian chronic liver disease patients vs. healthy controls (n = 40). The consecutively included patients comprised 120 with normal serum enzymes (HCV-NE) and 130 with high serum enzymes (HCV-HE), along with 120 cirrhotic patients.ResultsOxidative stress biomarkers – malondialdehyde (MDA), total peroxides and oxidative stress index (OSI) – were significantly lower in controls vs. each of the patient groups. Cirrhotic patients presented the highest levels. However, total antioxidants (TAO) showed non-significant differences among the four groups. The cellular hypoxia/angiogenesis biomarkers – lactate, vascular endothelial cell growth factor (VEGF) and its soluble receptor 1 (sVEGFR1) – vs. controls were massively increased in patient groups. VEGF was lowest while sVEGFR1 was highest among cirrhotic patients. Immunological biomarkers, – granulocyte/monocyte-colony stimulating factor (GM-CSF) and total immunoglobulin G (IgG) – were massively increased in patient groups vs. controls. GM-CSF was lowest in HCV-HE and IgG was highest in cirrhotic patients. sVEGFR1 correlated with the progression towards cirrhosis.ConclusionsOxidative stress is implicated in the progress of HCV infection with marked induction of cellular hypoxia and dysfunctional angiogenesis, and a futile immunological reaction. sVEGFR1 level correlated with progression towards HCV-induced liver fibrosis.

LPS stimulation during HCV infection induces MMP/TIMP1 imbalance in macrophages.

Introduction. During chronic hepatitis C virus (HCV) infections, HCV antigens establish cross-tolerance of endotoxins, but additional lipopolysaccharide (LPS) stimulation effects in this condition are poorly understood.Aim. This study aims to investigate the effects of the upregulated LPS on MMP and TIMP expression during chronic hepatitis C infection.Methodology. In the present study, we analysed the effect of HCV antigens and LPS stimulation on peripheral blood mononuclear cells (PBMCs) both in vivo and in vitro. Macrophages from HCV patients were isolated and their association with endotoxin tolerance was examined. MMP/TIMP1 expression and the related signalling pathways in macrophages were analysed. The macrophage and Huh7.5 cell co-culture model was used to analyse the effects of the cross-tolerance on collagen I deposition.Results. LPS levels were found to be significantly higher in HCV patients, particularly in those with HCV-induced liver fibrosis. In addition, although LPS serum level was occasionally upregulated in the patients, it did not induce intense immune response in PBMCs due to endotoxin cross-tolerance, and this was measured according to the changes in IL-6 and TNF-α levels. However, TIMP1 expression increased significantly during stimulation, exhibiting a tolerance/resistance phenotype, which was associated with TGF-β/Erk activation in macrophages. However, MMP levels did not increase due to endotoxin tolerance, which ultimately led to MMP/TIMP imbalance and influenced the deposition of collagen I.Conclusion. Increased LPS stimulation of macrophage during HCV antigen-induced endotoxin cross-tolerance contributes to MMP/TIMP1 imbalance and collagen I deposition.

956 Kasr Alainy Fibrosis Index (KAFI) - A Novel Score Based on Micro RNAs for Diagnosis of Significant Fibrosis: A Biopsy Controlled Study

INTRODUCTION: Precise identification of HCV induced liver fibrosis is needed for tailoring therapy, monitoring fibrosis regression after sustained virologic response and management of complications. Few studies investigated the role of MicroRNAs in predicting liver fibrosis. The current study evaluated the diagnostic accuracy of circulating miRNAs; miR-122, miR-192, miR-855, miR-375, miR-224, and miR-221 in prediction of hepatic fibrosis and to validate their usefulness against other commonly used noninvasive fibrosis scores using liver biopsy as the gold standard. METHODS: Seventy patients with chronic HCV, enrolled from Theodor Bilhariz research institute were included in this study. They were subjected to routine laboratory investigations, HCV-RNA, serum miR-122, 192, 885, 375, 224 and 221 by PCR, ultrasound guided liver biopsy and calculation of the following scores: Fibrosis Index (Fi), Fibro-Alfa , Hui index , aspartate aminotransferase (AST)/alanine aminotransferase (ALT) ratio (AAR), Biotechnology Research Center (BRC) score, aspartate to platelet ratio index (APRI), Fibro-Q , Gotebörg University Cirrhosis Index) (GUCI) and FIB-4 score. RESULTS: There was no statistically significant difference in laboratory parameters except for miR-122 which was significantly lower in patients with significant and advanced fibrosis (P < 0.0001 and P = 0.007, respectively). Univariate analysis identified miR-122, bilirubin and miR-855 as independent predictors of significant fibrosis. The AUC were 0.78, 0.67and 0.63, respectively (Figure 1a,c). Multivariate analysis selected the best overall formula combining miR-122, bilirubin and miR-855 as a novel noninvasive score for predicting significant liver fibrosis. KAFI Score = 4.558 – (miR-122 × 0.089 + miR-855 × 0.051 + bilirubin × 0.366), where 4.558 is a numeric constant. The area under the ROC curve (AUC) of this score for identifying significant fibrosis was 0.83 (Figure 1d). A cut-off of 0.41 had 77% sensitivity, 75% specificity, 65% positive predictive value, 85 % negative predictive value and 76% efficiency for predicting significant liver fibrosis. The AUCs of FI, Fibro- Alfa, Hui, AAR, BRC, APRI, Fibro-Q, GUCI and FIB 4 were 0.46, 0.53, 0.54, 0.60, 0.61, 0.62, 0.62, 0.62 and 0.67; respectively for predicting significant fibrosis. KAFI score yielded AUC 0.83 for predicting significant fibrosis and 0.80 for predicting advanced fibrosis. CONCLUSION: KAFI score is better than the current scores in discriminating hepatic-fibrosis in chronic hepatitis C patients

Genetic Variants in the Promoter Region of the Macrophage Migration Inhibitory Factor are Associated with the Severity of Hepatitis C Virus-Induced Liver Fibrosis.

Two polymorphisms in the promoter region of macrophage migration inhibitory factor (MIF)—rs755622 and rs5844572—exhibit prognostic relevance in inflammatory diseases. The aim of this study was to investigate a correlation between these MIF promoter polymorphisms and the severity of hepatitis C virus (HCV)-induced liver fibrosis. Our analysis included two independent patient cohorts with HCV-induced liver fibrosis (504 and 443 patients, respectively). The genotype of the single nucleotide polymorphism (SNP) -173 G/C and the repeat number of the microsatellite polymorphism -794 CATT5–8 were determined in DNA samples and correlated with fibrosis severity. In the first cohort, homozygous carriers of the C allele in the rs755622 had lower fibrosis stages compared to heterozygous carriers or wild types (1.25 vs. 2.0 vs. 2.0; p = 0.03). Additionally, ≥7 microsatellite repeats were associated with lower fibrosis stages (<F2) (p = 0.04). Comparable tendencies were observed in the second independent cohort, where fibrosis was assessed using transient elastography. However, once cirrhosis had been established, the C/C genotype and higher microsatellite repeats correlated with impaired liver function and a higher prevalence of hepatocellular carcinoma. Our study demonstrates that specific MIF polymorphisms are associated with disease severity and complications of HCV-induced fibrosis in a stage- and context-dependent manner.

FRI-262-Genetic variants in the promotor region of the macrophage migration inhibitory factor are associated with severity of HCV-induced liver fibrosis

FRI-262-Genetic variants in the promotor region of the macrophage migration inhibitory factor are associated with severity of HCV-induced liver fibrosis

Exosomal Transmission of MicroRNA from HCV Replicating Cells Stimulates Transdifferentiation in Hepatic Stellate Cells

The mechanism by which hepatitis C virus (HCV) causes fibrosis and other chronic liver diseases remains poorly understood. Previously, we observed that HCV infection induces microRNA-192 (miR-192) expression, which in turn upregulates transforming growth factor β1 (TGF-β1) in hepatocytes. In this study, we aimed to determine the roles and mechanisms of HCV-induced miR-192 expression during chronic liver injury and fibrosis and to identify potential target of the liver disease. Noticeably, miR-192 is secreted and transmitted through exosomes from HCV-replicating hepatocytes into hepatic stellate cells (HSCs). Exosomal transferred miR-192 upregulated fibrogenic markers in HSCs through TGF-β1 upregulation, resulting in the activation and transdifferentiation of HSCs into myofibroblasts. Anti-miR-192 treatment of HCV-replicating hepatocytes efficiently reduced miR-192 levels in exosomes, downregulated miR-192 and fibrogenic marker levels in HSCs, and impeded transdifferentiation of the cells. In contrast, miR-192 mimic RNA treatment significantly increased miR-192 levels in exosomes from naive hepatocytes, increased miR-192 and fibrogenic marker expression in HSCs, and induced transdifferentiation of the cells. Notably, transdifferentiation of exosome-exposed HSCs was reversed following treatment with anti-miR-192 into the HSCs. This study revealed a novel mechanism of HCV-induced liver fibrosis and identified exosomal miR-192 as a major regulator and potential treatment target for HCV-mediated hepatic fibrosis.

Metabolic Impact of HCV Clearance Examined by Urine Metabonomics in Patients with Severe Liver Fibrosis

Background: Hepatitis C virus (HCV) infection induces a long-term inflammatory response and oxidative-stress in the liver microenvironment, leading to hepatic fibrosis and metabolic alterations. Direct-acting-antiviral-agents (DAAs) induce HCV-clearance, but liver dyshomeostasis is not restored. Understanding the impact of viral-eradication on the liver metabolic activities could allow the optimization of patient’s metabolic care. The present prospective longitudinal study aims to characterize the urinary metabolic profile of HCV-induced severe liver fibrosis and the metabolic changes induced by DAAs and HCV-clearance by Nuclear-Magnetic-Resonance-(NMR)-based-metabolomics. Methods: The urinary metabolic profile of 23 HCV males with severe liver fibrosis and 20 age-matched healthy-controls was analyzed by NMR-based-metabolomics before starting DAAs, at the end-of-therapy, after one and three months of follow-up. Finding: The urinary metabolic profile of patients with severe liver fibrosis was characterized by higher levels of four metabolites related to oxidative-stress (pseudouridine, hypoxanthine, methyl-guanidine, dimethylamine) and two amino-acids (glutamine, tyrosine) compared to healthy-controls. N-methyl-nicotinamide, a catabolic intermediate of nicotinamide-adenine-dinucleotide, and 3-hydroxy-3-methylbutyric-acid, an intermediate of leucine-catabolism, returned to control levels with viral eradication. Finally, 3-hydroxyisobutyrate and 2,3-dihydroxy-2-methyl-butyrate, intermediates of valine-catabolism, increased temporarily during therapy, resulting as potential biomarkers of DAAs systemic effects. Interpretation: The identified metabolic profiles suggest that oxidative-stress persists despite HCV-eradication in the context of severe liver fibrosis, therefore the potential benefit of an antioxidant treatment after HCV-cure could be evaluated by further studies. HCV-clearance permanently restores leucine-catabolism, suggesting an improvement of skeletal muscle protein synthesis. DAA-administration temporarily modifies valine-catabolism, therefore, in case of amino-acid-supplementation, such modifications should be considered. Funding Statement: Sapienza University of Rome (Research Project 2017). Declaration of Interests: All Authors have nothing to disclose regarding conflict of interest with respect to this manuscript. Ethics Approval Statement: The study protocol conformed to the ethical guidelines of the 1975 Declaration of Helsinki and it was approved by the Institutional Review Board. All patients provided their written informed consent to participate in the study.

MiRNA-221 and miRNA-222 are promising biomarkers for progression of liver fibrosis in HCV Egyptian patients

BackgroundCurrent methodologies used to determine the progression of hepatic fibrosis rely heavily on liver biopsy, a dangerous and invasive procedure, with semi-subjective analysis of the results of the biopsy. Thus, a new approach is immensely needed for monitoring the progression of liver fibrosis in Hepatitis C virus (HCV) patients. Aim of workThe purpose of this study was to find highly specific and sensitive miRNA biomarkers that can be used to detect different stages of liver fibrosis. MethodologyThe study consisted of 42 cases of chronic hepatitis C (CHC) with early-stage fibrosis, 45 cases of CHC with late-stage fibrosis, and 40 healthy subjects with no CHC or fibrosis as controls. Expression patterns of 5 miRNAs (miR-16, miR-146a, miR-214-5p, miR-221, and miR-222) were analyzed in each group using TaqMan real-time PCR. ResultsSerum levels of miRNA-16, miRNA-146a, miRNA-221, and miRNA-222 were all significantly up-regulated in early and late stages of liver fibrosis. miRNA-222 had the highest sensitivity and specificity values in early and late fibrosis. miRNA-221 had the second highest sensitivity and specificity with the late-stage fibrosis group. Furthermore, miRNA-221 showed significant positive correlations with both miRNA-16 and miRNA-146a in the early- and late-stage fibrosis groups, with the early stage having a stronger correlation. ConclusionsThe results indicated that miRNA-16, miRNA-146a, miRNA-221, and miRNA-222 can be used to detect the presence of liver fibrosis. The high sensitivity and specificity of miRNA-222 and miRNA-221 in late-stage fibrosis indicate promising prognostic biomarkers for HCV-induced liver fibrosis.

Role of circulatory microRNAs in the pathogenesis of hepatitis C virus.

Hepatitis C virus (HCV) is associated with one of the major health problem in world that ultimate results in the liver cirrhosis and leads to carcinoma of hepatocellular components round the world. More than 185 million people were found to be infected with HCV. MicroRNAs are small oligonucleotide RNA having 18-22 nucleotides. Circulating mi-RNAs regulate the replication of HCV and HCV-induced liver fibrosis and HCC. By comparing the expression profiles of mi-RNAs of normal individuals with HCV infected patients, aberrant changes in expression of different mi-RNAs have been observed so it can be predicted that these mi-RNAs are associated with and play a central role in the hepatitis C infection and diseases associated with it. This review demonstrates the major role of circulatory microRNAs in the HCV and HCV associated ailments.

Increased incidence of cytomegalovirus coinfection in HCV-infected patients with late liver fibrosis is associated with dysregulation of JAK-STAT pathway

Herein, we examined the association between cytomegalovirus (CMV) coinfection and the progression of liver fibrosis in hepatitis C virus (HCV) infection, and investigated the effect of CMV coinfection on JAK-STAT pathway. CMV DNAemia was detected by PCR in DNA from controls (n = 120), and HCV patients with early (F0-F1, n = 131) and late (F2-F4, n = 179) liver fibrosis. By quantitative real time PCR (qRT-PCR), we examined the profile of 8 JAK-STAT transcripts in PBMCs RNA from 90 HCV patients (39 CMV positive and 51 CMV negative), 4 CMV mono-infected patients, and 15 controls. Our results demonstrated higher incidence of CMV in F2-F4 group than in control (OR 5.479, 95% CI 3.033–9.895, p < 0.0001) or F0-F1 groups (OR 2, 95% CI 1.238–3.181, p = 0.005). qRT-PCR showed downregulation of STAT2 (p = 0.006) and IRF7 (p = 0.02) in CMV positive group compared to CMV negative one. The downregulation of STAT2 and IRF7 was mainly in CMV positive patients with late fibrosis compared to CMV negative patients (p = 0.0007 for IRF7 and p = 0.01 for STAT2). Our results are the first to report that CMV coinfection is a possible risk factor for the progression of HCV-induced liver fibrosis, and thereby CMV screening and treatment are important for HCV patients.

Vascular Endothelial Growth Factor Expression in Hepatitis C Virus-Induced Liver Fibrosis: A Potential Biomarker.

The major complication of hepatitis C virus (HCV) infection is the induction of hepatic fibrosis. In this study, we investigated the correlation between the expression level of vascular endothelial growth factor (VEGFA) at mRNA and protein levels and the progression of HCV-related liver fibrosis. One hundred twenty subjects were selected for this study: 15 controls and 105 chronic HCV patients with different fibrosis grades (44 F0-F1 and 61 F2-F4). Quantitative real-time polymerase chain reaction (qRT-PCR) was used to measure VEGFA mRNA in peripheral blood mononuclear cells, while enzyme-linked immunosorbent assay (ELISA) was used to measure the secreted VEGFA protein in serum. Both qRT-PCR and ELISA results showed that HCV patients have significantly higher VEGFA expression than that of controls (P = 0.036 and 0.043, respectively). Moreover, patients with late fibrotic stages (F2-F4) exhibited the highest levels of VEGFA mRNA and protein (P = 0.008 and 0.041, respectively) when compared with controls. An area under the receiver operating characteristic curve (AUC of the ROC) for the circulatory VEGFA protein between HCV patients with fibrosis and healthy controls was 0.92 (P = 0.043). Our data suggest that VEGFA protein is a promising noninvasively diagnostic biomarker for HCV-induced liver fibrosis.

Selective inhibitor of Wnt/\u03b2-catenin/CBP signaling ameliorates hepatitis C virus-induced liver fibrosis in mouse model

Chronic hepatitis C virus (HCV) infection is one of the major causes of serious liver diseases, including liver cirrhosis. There are no anti-fibrotic drugs with efficacy against liver cirrhosis. Wnt/β-catenin signaling has been implicated in the pathogenesis of a variety of tissue fibrosis. In the present study, we investigated the effects of a β-catenin/CBP (cyclic AMP response element binding protein) inhibitor on liver fibrosis. The anti-fibrotic activity of PRI-724, a selective inhibitor of β-catenin/CBP, was assessed in HCV GT1b transgenic mice at 18 months after HCV genome expression. PRI-724 was injected intraperitoneally or subcutaneously in these mice for 6 weeks. PRI-724 reduced liver fibrosis, which was indicated by silver stain, Sirius Red staining, and hepatic hydroxyproline levels, in HCV mice while attenuating αSMA induction. PRI-724 led to increased levels of matrix metalloproteinase (MMP)-8 mRNA in the liver, along with elevated levels of intrahepatic neutrophils and macrophages/monocytes. The induced intrahepatic neutrophils and macrophages/monocytes were identified as the source of MMP-8. In conclusion, PRI-724 ameliorated HCV-induced liver fibrosis in mice. We hypothesize that inhibition of hepatic stellate cells activation and induction of fibrolytic cells expressing MMP-8 contribute to the anti-fibrotic effects of PRI-724. PRI-724 is a drug candidate which possesses anti-fibrotic effect.

Circulating ECV-Associated miRNAs as Potential Clinical Biomarkers in Early Stage HBV and HCV Induced Liver Fibrosis.

Introduction: Chronic hepatitis B (HBV) and C (HCV) virus infection is associated with the activation of hepatic stellate cells (HSCs) toward a myofibroblastic phenotype, resulting in excessive deposition of extracellular matrix, the development of liver fibrosis, and its progression toward cirrhosis. The gold standard for the detection and staging of liver fibrosis remains the liver biopsy, which is, however, associated with some mild and severe drawbacks. Other non-invasive techniques evade these drawbacks, but lack inter-stage specificity and are unable to detect early stages of fibrosis. We investigated whether circulating vesicle-associated miRNAs can be used in the diagnosis and staging of liver fibrosis in HBV and HCV patients.Methods: Plasma samples were obtained from 14 healthy individuals and 39 early stage fibrotic patients (F0–F2) with chronic HBV or HCV infection who underwent transient elastography (Fibroscan). Extracellular vesicles were extracted from the plasma and the level of miRNA-122, -150, -192, -21, -200b, and -92a was analyzed by qRT-PCR in total plasma and circulating vesicles. Finally, these same miRNAs were also quantified in vesicles extracted from in vitro activating primary HSCs.Results: In total plasma samples, only miRNA-200b (HBV: p = 0.0384; HCV: p = 0.0069) and miRNA-122 (HBV: p < 0.0001; HCV: p = 0.0007) were significantly up-regulated during early fibrosis. In circulating vesicles, miRNA-192 (HBV: p < 0.0001; HCV: p < 0.0001), -200b (HBV: p < 0.0001; HCV: p < 0.0001), -92a (HBV: p < 0.0001; HCV: p < 0.0001), and -150 (HBV: p = 0.0016; HCV: p = 0.004) displayed a significant down-regulation in both HBV and HCV patients. MiRNA expression profiles in vesicles isolated from in vitro activating primary mouse HSCs resembled the miRNA expression profile in circulating vesicles.Conclusion: Our analysis revealed a distinct miRNA expression pattern in total plasma and its circulating vesicles. The expression profile of miRNAs in circulating vesicles of fibrotic patients suggests the potential use of these vesicle-associated miRNAs as markers for early stages of liver fibrosis.