METAVIR Score Research Articles

Role of Circulating Hematopoietic Fibrocytes in Chronic Hepatitis C Patients Induced Liver Fibrosis

Background: Bone marrow derived fibrocytes may play an important role in pathogenesis and resolution of liver fibrosis. These cells may offer new approaches for better understanding the pathogenesis of liver fibrosis.
 Aim of the work: To define the proportion of circulating fibrocytes with hematopoietic progenitor origin as defined by CD45 and CD34 positivity and to assess whether they are increased in patients with chronic C hepatitis in correlation to the degree of liver fibrosis.
 Subjects and Methods: Sixty HCV patients were classified according to METAVIR score into 4 stages of liver fibrosis, 15 age and sex-matched controls were included. Flowcytometric analysis for circulating levels of fibrocytes CD34+ve cells, CD45+ve cells, collagen type I+ve cells and CXCR4+ve cells was carried out using monoclonal antibodies (anti-CD34, CD45, collagen type I and CD184). GM-CSF, TGF-β and α-SMA were assessed using ELISA.
 Results and Conclusions: A significant increase in the circulating levels of GM-CSF, TGF- β and α-SMA, with a significant increase in the percentage of cells express CXCR4and in the co expression of CD34, CD45 and collagen type I positive cells in different groups of patients compared to control group, denoting the presence of an increased proportion of circulating fibrocytes in peripheral blood of these patients. The percentage of fibrocytes that positively expression CD34, CD45, collagen type I and CXCR4, were increased in step wise fashion in conjunction with worsening severity of liver disease.
 Liver fibrosis is associated with increased levels of circulating TGF-β1 and lipopolysaccharide, activation of myofibroblasts, and extensive deposition of extracellular matrix, mostly collagen Type I. TGF-β and LPS play a critical role in fibrogenesis and trigger fibrocyte recruitment to the injured liver promoting their differentiation into collagen type I producing myofibroblast, supporting that fibrocytes may become a novel target for anti fibrotic therapy.

Оценка некоторых предикторов прогрессирования фиброза печени у больных гепатитом С после успешной элиминации вируса

Objective. To determine environmental and genetic factors associated with fibrosis progression after virus elimination as a result of direct-acting antiviral therapy by follow-up dispensary observation of patients with chronic hepatitis C with moderate and severe liver fibrosis (F2–F3 according to the METAVIR scoring system). Patients and methods. This study included 301 patients (166 men and 135 women) aged 20-64 years with chronic hepatitis C virus (HCV). A sustained virologic response was achieved in all patients after therapy with direct-acting antiviral agents. Patients were followed up for an average of 38 weeks (16–64). Patients were divided into two groups in order to perform comparative evaluation: group I included 257 patients with regression of liver fibrosis and group II – 44 patients with progression of liver fibrosis. Questionnaire and clinical and laboratory data were assessed. In addition, genetic studies of 24 singlenucleotide polymorphisms of genes involved in intracellular immune signaling pathway activation, interferon synthesis, metabolic regulation and cell proliferation were performed in both groups. Results. It was revealed that validated predictors of liver fibrosis progression in patients with chronic HCV after successful virus elimination as a result of therapy with direct-acting antiviral agents are the presence of concomitant type 2 diabetes mellitus, low ALT activity and serum osteopontin levels over 80 ng/mL at the beginning of therapy. Genetic predisposition to liver fibrosis progression is mediated by carriage of the AA genotype of HNF4α rs4812829 (OR = 3.55; 95% CI 1.21–10.41; p = 0.015). Additionally, the G-allele of NAT2 rs1495741, which marks fast xenobiotic acetylation, was found to have protective properties in the dominant genetic model. Carriers of GG- and GA-genotypes had an almost 2-fold lower risk of liver fibrosis progression after antiviral therapy than AA-genotype carriers (OR = 0.49; 95% CI 0.25–0.94; p = 0.029). Conclusion. The risk after successful hepatitis C virus elimination is significantly higher in patients with concomitant type 2 diabetes mellitus. ALT activity and serum osteopontin levels at the beginning of therapy can be estimated as predictors of liver fibrosis progression among laboratory parameters. Moreover, some genetic markers in the form of single-nucleotide polymorphisms of the HNF4α and NAT2 genes were established, which can be used to predict the development of liver fibrosis in patients with hepatitis C after therapy with direct-acting antiviral agents. Key words: hepatitis C, liver fibrosis, predictors, risk factors, diabetes mellitus, osteopontin, single-nucleotide polymorphisms, HNF4α gene, NAT2 gene

Resolvin D1 attenuates CCl4 Induced Liver Fibrosis by Inhibiting Autophagy-Mediated HSC activation via AKT/mTOR Pathway.

Resolvin D1 (RvD1) was previously reported to relieve inflammation and liver damage in several liver diseases, but its potential role in liver fibrosis remains elusive. The aim of our study was to investigate the effects and underlying mechanisms of RvD1 in hepatic autophagy in liver fibrosis. In vivo, male C57BL/6 mice were intraperitoneally injected with 20% carbon tetrachloride (CCl4, 5 ml/kg) twice weekly for 6 weeks to establish liver fibrosis model. RvD1 (100 ng or 300 ng/mouse) was added daily in the last 2 weeks of the modeling period. In vitro, lipopolysaccharide (LPS)-activated LX-2 cells were co-treated with increasing concentrations (2.5–10 nM) of RvD1. The degree of liver injury was measured by detecting serum AST and ALT contents and H&E staining. Hepatic fibrosis was assessed by masson's trichrome staining and metavir scoring. The qRT-PCR, western blot, immunohistochemistry, and immunofluorescence were applied to liver tissues or LPS-activated LX-2 cells to explore the protective effects of RvD1 in liver fibrosis. Our findings reported that RvD1 significantly attenuated CCl4 induced liver injury and fibrosis by decreasing plasma AST and ALT levels, reducing collagen I and α-SMA accumulation and other pro-fibrotic genes (CTGF, TIMP-1 and Vimentin) expressions in mouse liver, restoring damaged histological architecture and improving hepatic fibrosis scores. In vitro, RvD1 also repressed the LPS induced LX-2 cells activation and proliferation. These significant improvements mainly attributed to the inhibiting effect of RvD1 on autophagy in the process of hepatic stellate cell (HSC) activation, as demonstrated by decreased ratio of LC3-II/I and elevated p62 after RvD1 treatment. In addition, using AZD5363 (an AKT inhibitor that activates autophagy) and AZD8055 (an mTOR inhibitor, another autophagy activator), we further verified that RvD1 suppressed autophagy-mediated HSC activation and alleviated CCl4 induced liver fibrosis partly through AKT/mTOR pathway. Overall, these results demonstrate that RvD1 treatment is expected to become a novel therapeutic strategy against liver fibrosis.

Digital Pathology Enables Automated and Quantitative Assessment of Inflammatory Activity in Patients with Chronic Liver Disease.

Traditional histological evaluation for grading liver disease severity is based on subjective and semi-quantitative scores. We examined the relationship between digital pathology analysis and corresponding scoring systems for the assessment of hepatic necroinflammatory activity. A prospective, multicenter study including 156 patients with chronic liver disease (74% nonalcoholic fatty liver disease-NAFLD, 26% chronic hepatitis-CH etiologies) was performed. Inflammation was graded according to the Nonalcoholic Steatohepatitis (NASH) Clinical Research Network system and METAVIR score. Whole-slide digital image analysis based on quantitative (I-score: inflammation ratio) and morphometric (C-score: proportionate area of staining intensities clusters) measurements were independently performed. Our data show that I-scores and C-scores increase with inflammation grades (p < 0.001). High correlation was seen for CH (ρ = 0.85–0.88), but only moderate for NAFLD (ρ = 0.5–0.53). I-score (p = 0.008) and C-score (p = 0.002) were higher for CH than NAFLD. Our MATLAB algorithm performed better than QuPath software for the diagnosis of low-moderate inflammation (p < 0.05). C-score AUC for classifying NASH was 0.75 (95%CI, 0.65–0.84) and for moderate/severe CH was 0.99 (95%CI, 0.97–1.00). Digital pathology measurements increased with fibrosis stages (p < 0.001). In conclusion, quantitative and morphometric metrics of inflammatory burden obtained by digital pathology correlate well with pathologists’ scores, showing a higher accuracy for the evaluation of CH than NAFLD.

Utility of transient elastography in evaluation and follow-up of Budd-Chiari syndrome patients after endovascular intervention

BackgroundLiver stiffness measurement (LSM) using transient elastography (TE) is a common noninvasive method for assessing hepatic fibrosis. The purpose of this study was to show the utility of LSM in the evaluation and short-term follow-up of Budd-Chiarri syndrome (BCS) patients as well as to investigate the relationships between LSM, hepatic venous pressure, and liver fibrosis. We conducted a prospective cohort study in which 25 patients with a confirmed diagnosis of primary BCS underwent endovascular intervention for BCS treatment. All patients had pressure gradient changes across the stenosed hepatic veins/inferior vena cava (HV/IVC) measured before and after the procedure, a transvenous liver biopsy during the procedure, and LSM (using Transient Elastography 3 days before and 3 months after the intervention as a short-term follow-up). Comparisons and correlations were made between liver stiffness, hepatic venous pressure, and fibrosis.ResultsThe HV/IVC pressure gradient measured immediately before and after the intervention had a direct proportionate correlation with the LS measured 3 days before and 3 months after the intervention (r = 0.41 & 0.44, p: 0.04 & 0.03, respectively). LSM drops significantly 3 months after intervention (from 51.62 ± 22 kPa to 28.80 ± 19.25 kPa (p: 0.001)). There is a significant improvement in LSM post-intervention among patients with moderate to severe fibrosis (METAVIR scores of ≥ F2 before intervention (p: 0.001).ConclusionsLSM in BCS patients reflects congestion rather than fibrosis stage. TE can be used for the short-term monitoring of patients with BCS following the endovascular intervention as a noninvasive tool.

Diagnosis of liver cirrhosis with two-dimensional shear wave elastography in biliary atresia before Kasai portoenterostomy.

To evaluate the application value of two-dimensional shear wave elastography (2D-SWE) for non-invasive diagnosis of liver cirrhosis (LC) in patients with biliary atresia (BA) before Kasai portoenterostomy (KP), and the cutoff value of liver stiffness measurement (LSM) for diagnosing LC. The clinical data of 51 patients with BA who were diagnosed via surgery and pathological results from May 2017 to December 2018 in the department of general surgery, Beijing Children's Hospital, Capital Medical University, were retrospectively analyzed. The liver tissue specimens obtained during KP were evaluated according to the METAVIR criteria. The LSM was obtained using the 2D-SWE technique before KP. There was a grade positive correlation between LSM and METAVIR staging, and the Spearman correlation coefficient was 0.432 (P = 0.002). The AUC for 2D-SWE diagnosing LC (METAVIR score S = 4) in patients with BA before KP was 0.843 (95% confidence interval 0.736 ~ 0.950). The best cutoff value was 16.05kPa, and the corresponding sensitivity was 75.0%, specificity was 83.7%, positive predictive value (PPV) was 46.1%, negative predictive value (NPV) was 94.7%, and the accuracy was 82.4%. 2D-SWE can be used to noninvasively diagnose LC in patients with BA before KP, and the cutoff value is 16.05kPa.

Development and multicenter validation of FIB-6: A novel, machine learning, simple bedside score to rule out liver cirrhosis and compensated advanced chronic liver disease in patients with chronic hepatitis C.

Non-invasive tests (NITs), such as Fibrosis-4 index (FIB-4) and the aspartate aminotransferase-to-platelet ratio index (APRI), developed using classical statistical methods, are increasingly used for determining liver fibrosis stages and recommended in treatment guidelines replacing the liver biopsy. Application of conventional cutoffs of FIB-4 and APRI resulted in high rates of misclassification of fibrosis stages. There is an unmet need for more accurate NITs that can overcome the limitations of FIB-4 and APRI. Machine learning with the random forest algorithm was used to develop a non-invasive index using retrospective data of 7238 patients with biopsy-proven chronic hepatitis C from two centers in Egypt; derivation dataset (n=1821) and validation set in the second center (n=5417). Receiver operator curve analysis was used to define cutoffs for different stages of fibrosis. Performance of the new score was externally validated in cohorts from two other sites in Egypt (n=560) and seven different countries (n=1317). Fibrosis stages were determined using the METAVIR score. Results were also compared with three established tools (FIB-4, APRI, and the aspartate aminotransferase-to-alanine aminotransferase ratio [AAR]). Age in addition to readily available laboratory parameters such as aspartate, and alanine aminotransferases, alkaline phosphatase, albumin (g/dl), and platelet count (/cm3 ) correlated with the biopsy-derived stage of liver fibrosis in the derivation cohort and were used to construct the model for predicting the fibrosis stage by applying the random forest algorithm, resulting in an FIB-6 index, which can be calculated easily at http://fib6.elriah.info. Application of the cutoff values derived from the derivation group on the validation groups yielded very good performance in ruling out cirrhosis (negative predictive value [NPV]=97.7%), compensated advance liver disease (NPV=90.2%), and significant fibrosis (NPV=65.7%). In the external validation groups from different countries, FIB-6 demonstrated higher sensitivity and NPV than FIB-4, APRI, and AAR. FIB-6 score is a non-invasive, simple, and accurate test for ruling out liver cirrhosis and compensated advance liver disease in patients with chronic hepatitis C and performs better than APRI, FIB-4, and AAR.

Augmented Liver Uptake of the Membrane Voltage Sensor Tetraphenylphosphonium Distinguishes Early Fibrosis in a Mouse Model.

Mitochondrial (mito-) oxidative phosphorylation (OxPhos) is a critical determinant of cellular membrane potential/voltage. Dysregulation of OxPhos is a biochemical signature of advanced liver fibrosis. However, less is known about the net voltage of the liver in fibrosis. In this study, using the radiolabeled [3H] voltage sensor, tetraphenylphosphonium (TPP), which depends on membrane potential for cellular uptake/accumulation, we determined the net voltage of the liver in a mouse model of carbon tetrachloride (CCl4)-induced hepatic fibrosis. We demonstrated that the liver uptake of 3H-TPP significantly increased at 4 weeks of CCl4-administration (6.07 ± 0.69% ID/g, p < 0.05) compared with 6 weeks (4.85 ± 1.47% ID/g) and the control (3.50 ± 0.22% ID/g). Analysis of the fibrosis, collagen synthesis, and deposition showed that the increased 3H-TPP uptake at 4 weeks corresponds to early fibrosis (F1), according to the METAVIR scoring system. Biodistribution data revealed that the 3H-TPP accumulation is significant in the fibrogenic liver but not in other tissues. Mechanistically, the augmentation of the liver uptake of 3H-TPP in early fibrosis concurred with the upregulation of mito-electron transport chain enzymes, a concomitant increase in mito-oxygen consumption, and the activation of the AMPK-signaling pathway. Collectively, our results indicate that mito-metabolic response to hepatic insult may underlie the net increase in the voltage of the liver in early fibrosis.

Reliability of Transient Elastography as a non-Invasive Technique for Detection of Fibrosis in Budd Chiari Syndrome Patients after Endovascular Intervention

Abstract Aim of the work To test the reliability of fibroscan in detection of fibrosis in patients with Budd Chiari syndrome before and after endovascular intervention (after elimination of hepatic congestion). Background transient elastography (TE) is a noninvasive methodology that has been used to monitor liver stiffness in patients with chronic viral hepatitis. One of the limitations for accurate assessment of liver fibrosis by TE is the liver congestion. Liver congestion can result from Budd Chiari syndrome (BCS).The treatment of BCS is through restoring the flow of the blood between the portal vein to the inferior vena cava, which will lead to decongestion of the liver.TE, will be tested after liver decongestion for proper detection of liver fibrosis. Patients and methods This was a prospective cohort study conducted on 25 Egyptian patients with confirmed diagnosis of primary Budd-Chiari Syndrome (BCS) in the period from June 2017 to September 2019. TE was performed three days before endovascular intervention and three months after it. Liver biopsy was taken during the intervention for assessment of METAVIR score. Comparison was done between TE assessments before and after intervention in detection of the degree of liver fibrosis in comparison to METAVIR score measured in liver biopsy. Results FVLM was the most common hypercoagulable cause in the involved patients. There was significant drop in Liver Stiffness Measurements (LSM) measured three months post-intervention indicating improvement of liver fibrosis after relieving liver congestion but still not correlated to the METAVIR scores measured in the liver biopsy. Conclusion Liver congestion has high impact on Liver stiffness measurement giving overestimation which improves significantly after decongestion of the liver by the endovascular intervention.

Role of hepatic Doppler as compared to Fibro-scan in evaluation of post virus C hepatic fibrosis

Abstract Background Chronic infection with hepatitis C virus (HCV) is a disease of global importance with a large burden of morbidity and mortality. hepatitis c virus can cause progressive liver damage which may result in liver cirrhosis and hepato-cellular carcinoma . Aim of the work The aim of our study was to examine the efficacy of hepatic Doppler indices, namely hepatic vein wave form, hepatic vein maximum velocity ,hepatic artery RI and portal vein maximum velocity and portal vein wave form for evaluating the degree of hepatic fibrosis in chronic hepatitis C (CHC) patients as compared to transient elastography. Patients and methods our study was done over period from September 2018 till April 2019 included 40 patients from eldemerdash hospital designed to include all hepatitis c positive patients with any age or sex excluding other cause of liver cirrhosis (negative hepatitis B, negative auto immune hepatitis) in absence of obesity or ascites. hepatic vein maximum velocity, hepatic artery RI, portal vein maximum velocity, TE were performed to all patients. fibrosis was assessed on semi quantitative scoring system (METAVIR score ). we examine the efficacy of each of Doppler indices in differentiating different stages of hepatic fibrosis and their diagnostic accuracy in predicting significant fibrosis and cirrhosis . Results on comparing the fibrosis &amp;non fibrosis groups by TE according to Doppler measurement we found statistically significant higher means of the portal vein maximum velocity (PVmax) for the non fibrosis group as compared to the fibrosis group (24.15±9.71 and 15.94±3.35 respectively with p value&amp;lt;0.001) also found significantly lower HAPSV/PVmax ratio for the non fibrosis as compared to fibrosis group (1.58±0.41 and 2.59±1.17 respectively with p value 0.011) finally, we found that there was significant change of the HV waveform pattern for the non fibrosis (normal triphasic ) as compared to fibrosis (monophasic or biphasic ) (x2=4.353 with p value 0.038)

Role of Ultrasound and Transient Elastography in Diagnosis of Occult Cirrhosis in Chronic Liver Disease

Background: Hepatic cirrhosis represents the most advanced stage of any chronic liver disease characterized by progressive fibrosis. Complications including decompensation, spontaneous bacterial peritonitis, variceal bleeding, and hepatocellular carcinoma, carry significant morbidity and mortality. Early identification of patients with cirrhosis is of paramount importance to initiate surveillance protocols for prevention, early diagnosis, and specific therapies for such devastating complications. Objective: To investigate prevalence of preclinical cirrhosis diagnosed by transient elastography (TE) that we defined as occult cirrhosis (OC) and to evaluate the pattern of surveillance that patients with OC received in current clinical practice. Patients and methods: Our study included 60 chronic liver disease patients recruited from National Hepatology and Tropical Medicine Research Institute (NHTMRI) outpatient clinics to assess prevalence of occult cirrhosis among patients who have not any clinical or definite sonographic signs of liver cirrhosis, the study was divided into two groups: 1) Occult cirrhosis, defined as TE ≥13kPa and F4 on METAVIR scoring system. 2) non-cirrhotic chronic liver disease (CLD) (TE<13kPa). We found that the prevalence of OC is considerable, representing 15% of all CLD patients. This proportion suggests up to 1 in 7 individuals with CLD may have their OC missed. Results: Concerning the etiology of the chronic liver disease in our study, among the occult cirrhosis group there were 8 patients diagnosed as hepatitis C virus positive representing (89%) and one patient diagnosed as hepatitis B virus positive representing (11%) and among the non-cirrhotic group there were 42 patients diagnosed as hepatitis C virus positive representing (82%) and 4 patients diagnosed as hepatitis B virus positive representing (8%) and 5 patients diagnosed as non-alcoholic fatty liver disease (NAFLD) representing (10%). In our study among the occult cirrhosis group there were 5 male patients (55%) and 4 females (45%) with their age ranged between 33-55 years with the mean age 44 years and among the non-cirrhotic group there were 32 male patients (62.7%) and 19 female patients (37.3%) their age ranged between 19-58 years with the mean age 38.5 years. Conclusion: This study suggests that occult cirrhosis is a frequent finding. Clinicians should be aware that the absence of definite sonographic signs cannot rule out the possibility of compensated cirrhosis in its preclinical stage. Patients with occult cirrhosis can exhibit rapid development of hepatocellular carcinoma, variceal bleeding, and ascites.

Real time ultrasound tissue Elastography in post-hepatitis chronic liver disease; role, limitations and pitfalls

Abstract Assessment of liver fibrosis stage is important in determining the prognosis and treatment strategy in chronic liver diseases. To overcome the limitations of liver biopsy, great efforts have been made to develop and validate non-invasive methods for detecting liver fibrosis, including serological indicators and imaging methods. Among these noninvasive methods, ultrasound-based elastography techniques are increasingly employed to assess parenchymal stiffness. Real-time transient elastography is most accurate one of them and rapidly evolving technique that can reveal the elastic properties of tissues, and display it as real time images. This technique can avoid unnecessary invasive liver biopsy. Patients and methods This descriptive study was done on 20 chronic HCV patients as diagnosed by seropositivity for HCV antibodies and HCV RNA by PCR, Patients were recruited from police hospitals outpatient clinics to find an alternative method to assess liver fibrosis. Results showed highly significant agreement between fibroscan scores and biopsy results (p value was &amp;lt; 0.001), the identification of patients with significant fibrosis (F ≥ 2) “F2, 3, 4” We reported that the cut off level of significant fibrosis (F ≥ 2) assessed by the METAVIR scoring system was 10.2 kpa, with sensitivity 86.7 % and specificity 80 %. However, cut-off level of 13.8 kpa for the detection of cirrhosis (F4) with an area under the ROC of 0.913, and a very high sensitivity and specificity (sensitivity 90.1% and specificity 85.7%). Conclusion Transient Elastography is an easy and quick clinical non-invasive method to perform. Results are available immediately, and this technique is accurate in predicting significant fibrosis ≥f2. Hence, Transient Elastography could be useful not only to evaluate liver fibrosis as to monitor liver disease progression, but also to monitor anti-viral or antifibrotic therapy effects and to help taking decisions in daily clinical practice.

The Diagnostic Accuracy of LOGIQ S8 and E9 Shear Wave Elastography for Staging Hepatic Fibrosis, in Comparison with Transient Elastography.

Background: The aim of this study was to evaluate the usefulness of two different types of 2-dimensional shear wave elastography (2D-SWE) for predicting liver fibrosis stages in comparison to transient elastography (TE), using a histologic METAVIR scoring system as the reference method. Methods: A total of 203 patients with chronic liver disease were prospectively enrolled in the study. Two different 2D-SWEs (LOGIQ S8 and E9 systems, GE Healthcare, Chalfont St Giles, UK) were assessed for liver stiffness in patients with chronic liver diseases. Patients received 2D-SWE examinations with the S8 and E9 systems, and also underwent TE (FibroScan®, Echosens, France) tests and liver biopsies on the same day. Results: The most common etiology of chronic liver disease was non-alcoholic fatty liver disease (28.7%), followed by chronic hepatitis B (25.1%). Liver fibrosis stages consisted of F0 (22.6%), F1 (29.7%), F2 (16.9%), F3 (12.8%) and F4 (17.9%). Overall, S8 and E9 were well correlated with the histologic fibrosis stages. The optimal cut-off values for S8 and E9 to differentiate significant fibrosis (≥F2) were 6.70 kPa and 6.42 kPa, respectively, while the cut-off values for S8 and E9 in distinguishing liver cirrhosis were 9.15 kPa and 8.88 kPa, respectively. Among the 195 patients who had successful measurements in both S8 and E9, liver stiffness showed good inter-equipment correlation (ICC: 0.900, p < 0.001). Regarding diagnostic ability, upon comparison (FibroScan®), there were no significant differences between 2D-SWEs and TE for detecting every stage of liver fibrosis. Conclusion: In comparison to TE, 2D-SWE with LOGIQ S8 and E9 (GE Healthcare) are useful non-invasive tools for predicting significant fibrosis and liver cirrhosis.

S1242 Hepatocellular Carcinoma in Patients Without Cirrhosis - Characteristics, Etiologies and Trends

Introduction: Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide. Current guidelines recommend surveillance for HCC in those with cirrhosis of any etiology. However, HCC may also develop in non-cirrhotic patients. The aim of this study was to evaluate the characteristics, etiologies, trends and outcomes of non-cirrhotic HCC. Methods: All histologically confirmed HCC cases diagnosed at the Centro Hospitalar Universitário de São João from January 2010 to December 2020 were identified. Patient, tumor, liver disease characteristics and outcome data were collected. The presence of cirrhosis was assessed by histology. Results: A total of 131 patients with HCC were enrolled in the study. Among them, 53 (40.5%) were noncirrhotic individuals, the majority males (75.5%), with a median age of 67 years old (IQR 54-76). In these patients, hepatitis C virus infection (15.1%) was the most common liver disease, followed by alcohol (13.2%) and hepatitis B virus (7.5%) infection. However, in most cases (58.5%) the etiology of liver disease/HCC was not identified. Most of them (58.5%) had bridging fibrosis (Metavir score F2-3). As expected, the non-cirrhotic patients had a higher platelet count (240 vs 168 x109/L, p< 0.001) and lower bilirubin levels (2.19 vs 3.05 mg/dL, p=0.002) and prothrombin time (12.3 vs 13.5 sec, p< 0.001) than the cirrhotic patients. Alpha-fetoprotein was similar in both groups (p=0.954). Patients with non-cirrhotic HCC had larger tumors (82 vs 47 mm, p< 0.001) and were less frequently within Milan criteria (20.8% vs 55.1%, p< 0.001). Nevertheless, they had better overall survival than cirrhotic HCC patients (4.8 vs 1.4 years, p=0.036). Barcelona Clinic Liver Cancer stage 0 and A (OR 0.04, 95% CI 0.007-0.217, p< 0.001) and surgical resection (OR 0.082, 95% CI 0.018-0.362, p< 0.001) were associated with reduced risk of death in patients with noncirrhotic HCC. The etiology of underlying liver disease did not affect mortality (p=0.277). Conclusion: In our cohort, a relevant number of HCCs occurred in patients without underlying cirrhosis. The average tumor size was significantly larger in the noncirrhotic patients, possibly due to a lack of surveillance in these individuals. Although noncirrhotic individuals presented with more advanced HCC, their survival was better, probably due to normal underlying liver function, lack of portal hypertension and better performance status at presentation.Table 1.: Clinical characteristics and outcomes of cirrhotic patients admitted with Acute Kidney injury stratified by non-selective beta-blocker use

Matrix metalloproteinase-1 as a non-invasive biomarker to assess liver fibrosis in children with chronic liver disease

BackgroundAbnormal extracellular matrix (ECM) turnover is linked to liver fibrosis as it reflects an imbalance between repair and progressive substitution of the liver parenchyma by scar tissue. Matrix metalloproteinases (MMPs) are the primary enzymes involved in ECM breakdown. So, this study aims to measure the value of serum matrix metalloproteinase-1 (MMP-1) in children with chronic liver diseases (CLD) in comparison with liver biopsy and serum biomarkers. A hundred twenty children with chronic liver diseases and sixty healthy children as a control group were included in this study. Both groups were evaluated via medical history, clinical, radiological, laboratory investigations, and serum MMP-1 level was measured by ELISA. Liver biopsy was performed for studied patients only.ResultsThe mean MMP-1 was 15.2 ± 5.1 ng/ml in children with CLD, and 64.7 ± 27.4 ng/ml in the control group. MMP-1 was statistically lower in the children with CLD than controls (p < 0.001). The mean ± SD of aspartate aminotransferase to platelet ratio index (APRI) and fibrosis-4 (FIB-4) scores in all studied cases showed a significant trend of increase with progressive fibrosis stage evident with histological METAVIR scoring system, while serum MMP-1 concentration was decreased significantly with increasing the degree of fibrosis in CLD group (P 0.001). Serum MMP-1 was indirectly correlated with serum biomarkers and the degree of fibrosis in patients.ConclusionsMMP-1 is a useful non-invasive marker for detection of the stage of liver fibrosis in children with chronic liver diseases.

P-47 HISTOPATHOLOGICAL ANALYSIS OF 10-YEAR PROTOCOL LIVER ALLOGRAFT BIOPSY OF ASYMPTOMATIC PEDIATRIC RECIPIENTS FROM A SINGLE BRAZILIAN CENTER

Liver transplant (LT) is the main therapeutic procedure for irreversible liver failure. Protocol liver biopsies during post-transplant follow-up in pediatric recipients with stable graft function have been done in order to identify structural changes. A more specific score, LAFSc, to analyze fibrosis and inflammation on liver allografts has been validated recently. To describe the prevalence of histological liver abnormalities in asymptomatic pediatric patients after ten years following LT from a Brazilian single center comparing two different scoring methods. Cross-sectional study of analysis of protocol percutaneous liver biopsies performed in patients who underwent liver transplant before the age of 18 years, of both genders, asymptomatic, using Tacrolimus for immunosuppression and at least 10-years post-liver transplant. From 97 recipients, 21 met inclusion criteria and had stable liver graft function. A single experienced pathologist assessed histopathological features comparing METAVIR scoring system and LAFSc. Mean follow-up was 12.8 (+/- 2.1) years after post-LT. TAC mean daily dose during biopsy time was 4mg (+/- 1.7) with mean serum level of 4.04. METAVIR scoring system pointed 10 recipients with any degree of fibrosis and 13 with inflammation findings in protocol biopsies. However, using LAFSc scoring system, 13 patients scored for abnormal findings in any of the three zones analysed, although 8 of these had total score of mild fibrosis (1-3). Of the three zones analysed in LAFSc, the highest scores were found in portal space. Centrilobular vein zone was the most affected, documented in 11 recipients. We observed a higher prevalence of abnormal findings using the new allograft fibrosis scoring system – LAFSc. It also showed more specifically the degree and location of fibrosis within hepatic lobule compared to METAVIR. Compared to other LT reports, we observed lower rates of chronic allograft hepatitis and fibrosis at 10 years.

Serum free light chain is associated with histological activity and cirrhosis in patients with chronic hepatitis B

BackgroundThe antiviral immune response is the main cause of hepatocyte damage and inflammatory necrosis. The serum free light chain, reflecting the immune function of B-cells, is strongly associated with inflammation and disease activity. We aimed to investigate the association of serum free light chain with the progression of chronic hepatitis B. MethodsA total of 208 eligible chronic hepatitis B patients who had undergone a liver biopsy were studied. Serum free light chains of all patients were measured by turbidimetry using an immunoassay. Liver histology was assessed according to the METAVIR scoring system (which grades the stage of fibrosis on a five-point scale, F0 = no fibrosis to F4 = cirrhosis, and histological activity on a four-point scale, A0 = no activity to A3 = severe activity). The association of serum free light chains with histological activity and fibrosis progression was evaluated. ResultsThe concentration of serum free light chains in CHB patients increased gradually with histological activity and fibrosis progression. The intensity of histological activity was significantly correlated with the serum free kappa chain (r = 0.658, P < 0.001) and the serum free lambda chain (0.675, P < 0.001). The stages of fibrosis were correlated with the serum free kappa chain (r = 0.683, P < 0.001) and serum free lambda chain (0.664, P < 0.001). After adjusting for age, sex and other synergic factors, the serum free kappa chain remained a potential risk factor, but the serum free lambda chain was no longer associated with liver cirrhosis. Similar to FIB-4 and RPR, the serum free kappa chain exhibited excellent performance in the prediction of liver cirrhosis. The AUCs of serum free Kappa chain, FIB-4 and RPR were 0.873, 0.880 and 0.895, respectively, which were significantly higher than those of the AAR and APRI (0.718 and 0.746). ConclusionOur work revealed that serum free light chains were associated with histological activity and cirrhosis in chronic hepatitis B, which could play a crucial role in the immunopathogenesis of HBV-associated cirrhosis. In addition, free kappa light chain could be a useful predictor of liver cirrhosis.

Predictors of early and sustained virological response of viral hepatitis C.

Due to complex interplay between host and viral factors, pathogenesis of chronic hepatitis C (CHC) is considered a challenging issue. Infection with hepatitis C virus (HCV) is not confined only to liver but can induce disturbances in many other organs and systems. Our primary aim for this study was to evaluate biological response rates and sustained virological response (SVR) in patients diagnosed with CHC, treated with Interferon-alpha (IFN-α), Pegylated (PEG)–IFN-α2a or -α2b plus Ribavirin. The second aim of the study was the identification of predictive factors for a favorable response to antiviral therapy in patients diagnosed with CHC. We enrolled in this study 210 patients diagnosed with CHC who have accomplished all inclusion and exclusion criteria, treated with PEG–IFN plus Ribavirin. Patients’ recovery progress has been evaluated by determining: age, gender; biochemical tests: alanine aminotransferase (ALT), aspartate aminotransferase (AST); serological assays – detect anti-HCV antibody and molecular assays – detect, quantify and/or characterize hepatitis C viral load (ribonucleic acid) (HCV-RNA); liver histopathological (HP) examination. According to their response to treatment, they were classified into responders (n=145) and non-responders (n=65). Liver biopsies were histopathologically evaluated for necroinflammatory grade and fibrosis stage according to the modified Ishak and Metavir scoring systems for chronic hepatitis. Demographic, laboratory, and HP results were introduced in statistical analysis. These parameters were included in area under curve (AUC) analysis in order to estimate their degree of influence on getting early virological response (EVR) and SVR. Our study demonstrates that factors connected to treatment failure in CHC are linked to older age, high hepatitis C viral load, and impaired glucose tolerance at beginning of treatment [high fasting glucose and insulin, high homeostatic model assessment of insulin resistance (HOMA-IR) index] and also to liver histology features (high fibrosis score, liver steatosis, iron infiltration, and more or less high necroinflammatory activity). Analyzing results of our study shows that HOMA-IR index, serum insulin levels, baseline HCV-RNA, baseline mean blood glucose and HP score like Ishak fibrosis score, steatosis score and liver iron score may have a predictive value for obtaining an EVR in patients diagnosed with CHC.

Evaluation of the Hemostatic Profile in Mice with Chronic Liver Disease: Effects of Protein S and Combined Gas6 and Protein S Deficiencies

IntroductionAdvanced chronic liver disease (ACLD) is characterized by changes in the coagulation system that embrace both hypo- and hypercoagulability. In this rebalanced situation, hypercoagulability can contribute to fibrosis progression and cirrhosis complications.Common hemostasis tests fail to reflect bleeding and thrombotic tendencies in ACLD patients. Therefore, a better understanding of the underlying mechanisms is mandatory for setting-up new assays and treatments.Here, we assessed the hemostatic changes by means of routine laboratory tests and thrombin generation assay (TGA) in two murine hepatotoxin-induced ACLD models.To better evaluate hemostatic changes, wild-type mice, mice carrying a partial protein S (Pros1) deficiency -thrombotic risk factor- and with a complete deficiency in Gas6, protective factor for fibrosis progression, have been investigated.MethodsAdult C57Bl6/129 Pros1+/+ and Pros1+/- mice received 1ml/kg carbon tetrachloride (CCl4) i.p. and 0.3g/L phenobarbital (PB) in drinking water 2x/week for 11 weeks. Similarly, Pros1+/+, Pros1+/-, Gas6-/-Pros1+/+, Gas6-/-Pros1+/- mice received 200mg/kg of thioacetamide (TAA) i.p. 3x/week for 12 weeks.At the end of the treatment, body, liver and spleen were weighed. Liver sections were stained with Sirius Red to score fibrosis according to METAVIR system. Plasma was analyzed for prothombin time (PT), fibrinogen (Fb), factor V (FV), factor VIII coagulant (FVIII:c) and thrombin and antithrombin complexes (TAT). TGA was performed in presence/absence of activated protein C (APC) or thrombomodulin (TM).ResultsAfter 11 weeks of CCl4/PB treatment, only 45% mice survived and developed an advanced CLD independently of their genotype (Pros1+/+ vs Pros1+/-). In particular, 42% Pros1+/+ (n=9)and 58% Pros1+/- (n=7) mice developed a severe cirrhosis (METAVIR: F4), 48% Pros1+/+ (n=10)and 42% Pros1+/- (n=5) an advanced fibrosis stage (F3) and only 10% Pros1+/+ (n=2) a modest fibrotic stage (F2). Treated mice presented hepatomegaly with darkish discoloration and nodular surface. The liver to body weight ratio was significantly increased compared to untreated controls (Pros1+/+=9.7% and Pros1+/-= 10.8% vs Pros1+/+ untreated =5.3%, P<0.05 and P<0.001, respectively).No variations were observed on PT, Fb, FV and platelets. Differently, increased FVIII:c (Pros1+/+=340±69 and Pros1+/-=378±103 vs Pros1+/+ untreated 212±62% vs hFVIII, P<0.05 and P<0.001, respectively) and TAT (Pros1+/+=30.2 and Pros1+/-=41.8±36.7 vs Pros1+/+ untreated 8.4±2.2 ug/L) were measured independently of disease severity (F3 vs F4).TAA treatment showed a reduced mortality paralleled by the development of less severe fibrotic stages in comparison to CCl4/PB treatment. In particular, 11% Pros1+/+ (n=4)and 31% Pros1+/- (n=4) mice were found at F4, 50% Pros1+/+ (n=12)and 54% Pros1+/- (n=7) at F3 while 39% Pros1+/+ (n=10)and 15% Pros1+/- (n=3) showed a moderate fibrotic disease (F2). In line with published data, preliminary data indicate a frequency reduction of the severe/advanced fibrotic stages in Gas6-/-Pros1+/+ mice: 37% (n=3) at F3 and 63% (n=5) at F2. Interestingly, this trend was reversed in Gas6-/-Pros1+/- mice: 54% (n=6) at F3 and 46% (n=5) at F2.No variations in body, liver and spleen weight and morphology were observed. The hemostatic profile did not show relevant variations of PT, Fb, FV, FVIII and platelets. TAT was increased in Pros1+/+ and Pros1+/- mice while levels in Gas6-/-Pros1+/+ mice were comparable to the untreated group (Pros1+/+=50.2±34.8 and Pros1+/-=47.7±57.9 vs Gas6-/-Pros1+/+=10.8±5.5 Gas6-/-Pros1+/-=13.9±17.9). Preliminary data on TGA showed a lack of APC resistance in all the treated groups (nAPCsr: Pros1+/+=1.0±0.3, Pros1+/-=1.3±0.7, Gas6-/-Pros1+/+=1.3±0.3and Gas6-/-Pros1+/-=1.1) but an acquired resistance to TM in Pros1+/+ and Pros1+/- mice (nTMsr: Pros1+/+=2.1±0.4, Pros1+/-=1.9±0.5, Gas6-/-Pros1+/+=1.2±0.4and Gas6-/-Pros1+/-=1.1). Primary results suggest a positive correlation between nTMsr and the METAVIR score for Pros1+/+ mice (r=0.46, n=6).ConclusionThese data provide the first evaluation of the hemostatic profile in murine ACLD at different stages. Preliminary data suggest that TM resistance might be discriminatory for the severity of the disease. Further TGA analysis and evaluation of the thrombotic risk on treated and control mice are needed. DisclosuresNo relevant conflicts of interest to declare.

Deep learning for noninvasive liver fibrosis classification: A systematic review.

While biopsy is the gold standard for liver fibrosis staging, it poses significant risks. Noninvasive assessment of liver fibrosis is a growing field. Recently, deep learning (DL) technology has revolutionized medical image analysis. This technology has the potential to enhance noninvasive fibrosis assessment. We systematically examined the application of DL in noninvasive liver fibrosis imaging. Embase, MEDLINE, Web of Science, and IEEE Xplore databases were used to identify studies that reported on the accuracy of DL for classification of liver fibrosis on noninvasive imaging. The search keywords were "liver or hepatic," "fibrosis or cirrhosis," and "neural or DL networks." Risk of bias and applicability were evaluated using the QUADAS-2 tool. Sixteen studies were retrieved. Imaging modalities included ultrasound (n=10), computed tomography (n=3), and magnetic resonance imaging (n=3). The studies analyzed a total of 40405 radiological images from 15853 patients. All but two of the studies were retrospective. In most studies the "ground truth" reference was the METAVIR score for pathological staging (n=9.56%). The majority of the studies reported an accuracy >85% when compared to histopathology. Fourteen studies (87.5%) had a high risk of bias and concerns regarding applicability. Deep learning has the potential to play an emerging role in liver fibrosis classification. Yet, it is still limited by a relatively small number of retrospective studies. Clinicians should facilitate the use of this technology by sharing databases and standardized reports. This may optimize the noninvasive evaluation of liver fibrosis on a large scale.