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Background & Aims: Non-invasive scores have been proposed to identify patients with fibrotic, metabolic dysfunction-associated steatohepatitis (MASH), who are at the highest risk of progression to complications of cirrhosis and may benefit from pharmacologic treatments. However, data in patients with type 2 diabetes (T2DM) are lacking. The aim of this multicenter prospective study was to perform a head-to-head comparison of FAST (FibroScan-aspartate aminotransferase [AST]), MAST (MRI-AST), MEFIB (magnetic resonance elastography [MRE] plus FIB-4), and FNI (fibrotic NASH index) for detecting fibrotic MASH in patients with T2DM., Methods: A total of 330 outpatients with T2DM and biopsy-proven metabolic dysfunction-associated steatotic liver disease (MASLD) from the QUID-NASH study (NCT03634098), who underwent FibroScan, MRI-proton density fat fraction and MRE at the time of liver biopsy were studied. The main outcome was fibrotic MASH, defined as NAS ≥4 (with at least one point for each parameter) and fibrosis stage ≥2 (centrally reviewed)., Results: All data for score comparisons were available for 245 patients (median age 59 years, 65% male, median BMI 31 kg/m 2 ; fibrotic MASH in 39%). FAST and MAST had similar accuracy (AUROCs 0.81 vs. 0.79, p = 0.41) but outperformed FNI (0.74; p = 0.01) and MEFIB (0.68; p <0.0001). When using original cut-offs, MAST outperformed FAST, MEFIB and FNI when comparing the percentage of correctly classified patients, in whom liver biopsy would be avoided (69% vs. 48%, 46%, 39%, respectively; p <0.001). When using cut-offs specific to our population, FAST outperformed FNI and MAST (56% vs. 40%, and 38%, respectively; p <0.001)., Conclusion: Our findings show that FAST, MAST, MEFIB and FNI are accurate non-invasive tools to identify patients with T2DM and fibrotic MASH in secondary/tertiary diabetes clinics. Cut-offs adapted to the T2DM population should be considered., Impact and Implications: Among patients with type 2 diabetes (T2DM), identifying those with metabolic dysfunction-associated steatohepatitis and significant fibrosis, who are the most at risk of developing clinical liver-related outcomes and who may benefit from pharmacologic treatments, is an unmet need. In this prospective multicenter study, we compared four non-invasive scores, three based on imaging (MRI or ultrasound technologies) and one on laboratory blood tests, for this purpose, using original and study-specific cut-offs. Our findings show that FAST, MAST, MEFIB and FNI are accurate non-invasive tools to identify patients with T2DM and fibrotic MASH in secondary/tertiary diabetes clinics. Cut-offs adapted to the T2DM population should be considered., Trial Registration Number: NCT03634098., (Copyright © 2024 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background and Aims: Nonalcoholic Steatohepatitis (NASH) is a major cause of end-stage liver diseases such as cirrhosis and hepatocellular carcinoma resulting ultimately in increased liver-related mortality. Fibrosis is the main driver of mortality in NASH. Procollagen C-Proteinase Enhancer-1 (PCPE-1) plays a key role in procollagen maturation and collagen fibril formation. To assess its role in liver fibrosis and NASH progression, knock-out mice were evaluated in a dietary NASH model., Methods: Global constitutive Pcolce-/- and WT male mice were fed with a Choline Deficient Amino acid defined High Fat Diet (CDA HFD) for 8 weeks. Liver triglycerides, steatosis, inflammation and fibrosis were assessed at histological, biochemical and gene expression levels. In addition, human liver samples from control and NASH patients were used to evaluate the expression of PCPE-1 at both mRNA and protein levels., Results: Pcolce gene deficiency prevented diet-induced liver enlargement but not liver dysfunction. Furthermore, liver triglycerides, steatosis and inflammation were not modified in Pcolce-/- male mice compared to WT under CDA HFD. However, a significant decrease in liver fibrosis was observed in Pcolce-/- mice compared to WT under NASH diet, associated with a decrease in total and insoluble collagen content without any significant modifications in the expression of genes involved in fibrosis and extracellular matrix remodeling. Finally, PCPE-1 protein expression was increased in cirrhotic liver samples from both NASH and Hepatitis C patients., Conclusions: Pcolce deficiency limits fibrosis but not NASH progression in CDA HFD fed mice., Competing Interests: The authors have declared that no competing interests exist.

Background and Aims: Porto-sinusoidal vascular liver disease (PSVD) is a rare cause of portal hypertension. PSVD is still often misdiagnosed as cirrhosis, emphasizing the need to improve PSVD diagnosis strategies. Data on liver stiffness measurement using transient elastography (TE-LSM) in PSVD are limited. The aim of this study was to evaluate the accuracy of TE-LSM to discriminate PSVD from cirrhosis in patients with signs of portal hypertension., Approach and Results: Retrospective multicenter study comparing TE-LSM in patients with PSVD, according to Vascular Liver Disease Interest Group criteria, with patients with compensated biopsy-proven cirrhosis associated with alcohol (n = 117), HCV infection (n = 110), or NAFLD (n = 46). All patients had at least one sign of portal hypertension among gastroesophageal varices, splenomegaly, portosystemic collaterals, history of ascites, or platelet count < 150 × 10 9 /L. The 77 patients with PSVD included in the test cohort had lower median TE-LSM (7.9 kPa) than the patients with alcohol-associated, HCV-related, and NAFLD-related cirrhosis (33.8, 18.2, and 33.6 kPa, respectively; P < 0.001). When compared with cirrhosis, a cutoff value of 10 kPa had a specificity of 97% for the diagnosis of PSVD with a 85% positive predictive value. A cutoff value of 20 kPa had a sensitivity of 94% for ruling out PSVD with a 97% negative predictive value. Of the patients, 94% were well-classified. Even better results were obtained in a validation cohort including 78 patients with PSVD., Conclusions: This study including a total of 155 patients with PSVD and 273 patients with cirrhosis demonstrates that TE-LSM < 10 kPa strongly suggests PSVD in patients with signs of portal hypertension. Conversely, when TE-LSM is >20 kPa, PSVD is highly unlikely., (© 2020 by the American Association for the Study of Liver Diseases.)

Background & Aims: We estimated the accuracy of FibroScan vibration-controlled transient elastography controlled attenuation parameter (CAP) and liver stiffness measurement (LSMs) in assessing steatosis and fibrosis in patients with suspected nonalcoholic liver disease (NAFLD)., Methods: We collected data from 450 consecutive adults who underwent liver biopsy analysis for suspected NAFLD at 7 centers in the United Kingdom from March 2014 through January 2017. FibroScan examinations with M or XL probe were completed within the 2 weeks of the biopsy analysis (404 had a valid examination). The biopsies were scored by 2 blinded expert pathologists according to nonalcoholic steatohepatitis clinical research network criteria. Diagnostic accuracy was estimated using the area under the receiver operating characteristic curves (AUROCs) for the categories of steatosis and fibrosis. We assessed effects of disease prevalence on positive and negative predictive values. For LSM, the effects of histological parameters and probe type were appraised using multivariable analysis., Results: Using biopsy analysis as the reference standard, we found that CAP identified patients with steatosis with an AUROC of 0.87 (95% confidence interval [CI] 0.82-0.92) for S≥S1, 0.77 (95% CI 0.71-0.82) for S≥S2, and 0.70 (95% CI 0.64-0.75) for S=S3. Youden cutoff values for S≥S1, S≥S2, and S≥S3 were 302 dB/m, 331 dB/m, and 337 dB/m, respectively. LSM identified patients with fibrosis with AUROCs of 0.77 (95% CI 0.72-0.82) for F≥F2, 0.80 (95% CI 0.75-0.84) for F≥F3, and 0.89 (95% CI 0.84-0.93) for F=F4. Youden cutoff values for F≥F2, F≥F3, and F=F4 were 8.2 kPa, 9.7 kPa, and 13.6 kPa, respectively. Applying the optimal cutoff values, determined from this cohort, to populations of lower fibrosis prevalence increased negative predictive values and reduced positive predictive values. Multivariable analysis found that the only parameter that significantly affected LSMs was fibrosis stage (P<10 -16 ); we found no association with steatosis or probe type., Conclusions: In a prospective analysis of patients with NAFLD, we found CAP and LSM by FibroScan to assess liver steatosis and fibrosis, respectively, with AUROC values ranging from 0.70 to 0.89. Probe type and steatosis did not affect LSM., Study Registration: ClinicalTrials.gov Identifier: NCT01985009., (Copyright © 2019 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background: Hepatopulmonary syndrome (HPS) is characterized by an arterial oxygenation defect, defined by an increased alveolar-arterial oxygen gradient, induced by pulmonary vascular dilatations in the context of liver disease. The pathogenesis of HPS is poorly understood. Morphologic changes associated with HPS are unknown. This study aimed at describing imaging and pathology changes associated with HPS., Methods: We performed a case-control study in candidates for transplant with suspicion of cirrhosis. Each patient with HPS (Pao 2  ≤ 70 mm Hg) was matched to three control subjects for age, cause, and liver disease severity. Pretransplant thoracic and abdominal imaging and explanted livers were reviewed., Results: CT scans and Doppler ultrasounds from 21 patients with HPS were compared with those from 63 control subjects. HPS was associated with a two- to threefold higher prevalence of obstructed intrahepatic portal branches, of slowed or hepatofugal portal blood flow, and of large abdominal portosystemic shunts. Hepatic artery diameter was also larger in patients with HPS. Explanted livers from 19 patients with HPS were compared with those from 57 control subjects. HPS was associated with a fourfold higher prevalence of portal venule thrombosis and a ninefold higher prevalence of extensive vascular proliferation within fibrous septa. Obstruction of centrilobular venules, sinusoidal dilatation, and liver parenchymal extinction were also more common in patients with HPS., Conclusions: HPS is associated with intrahepatic vascular changes and with features suggesting severe portal hypertension. These results raise the hypothesis that intrahepatic vascular changes precipitate the development of HPS, opening new therapeutic perspectives for HPS., (Copyright © 2018 American College of Chest Physicians. Published by Elsevier Inc. All rights reserved.)

Microvesicles (MVs) are extracellular vesicles released by cells following activation or apoptosis. Some MV subpopulations augment with cirrhosis severity and contribute to portal hypertension. This study aimed at determining if plasma MV levels can estimate the presence of hepatic venous pressure gradient (HVPG) ≥10 mm Hg and predict mortality in patients with advanced chronic liver disease. All patients with severe fibrosis or cirrhosis undergoing liver catheterization between 2013 and 2015 at two centers were prospectively included. We measured circulating levels of annexin V + , platelet, leukocyte, endothelial, and hepatocyte MVs. The test cohort included 139 patients. Hepatocyte MV levels were 4.0-fold and 2.2-fold higher in patients with Child-Pugh C than in those with Child-Pugh A or B liver disease, respectively. Levels of other MV subpopulations were not influenced by liver disease severity. Hepatocyte MV levels correlated with HVPG but could not identify patients with HVPG ≥10 mm Hg. Hepatocyte MV level >65 U/L predicted 6-month mortality independently of Child-Pugh score and of Model for End-Stage Liver Disease (MELD). Patients with hepatocyte MV levels >65 U/L and MELD >15 had a higher 6-month mortality than other patients (23% versus 3%; P = 0.001). These findings were confirmed in a validation cohort including 103 patients., Conclusion: Circulating MV levels cannot identify patients with HVPG ≥10 mm Hg; by contrast, hepatocyte MV levels strongly improve prediction of 6-month mortality in patients with advanced chronic liver disease; therapies associated with decreased levels of circulating hepatocyte MV might be attractive strategies in patients with severe cirrhosis. (Hepatology 2018)., (© 2018 by the American Association for the Study of Liver Diseases.)

Liver fibrosis is the common response to chronic liver injury, and leads to cirrhosis and its complications. Persistent inflammation is a driving force of liver fibrosis progression. Mucosal-associated invariant T (MAIT) cells are non-conventional T cells that display altered functions during chronic inflammatory diseases. Here, we show that circulating MAIT cells are reduced in patients with alcoholic or non-alcoholic fatty liver disease-related cirrhosis while they accumulate in liver fibrotic septa. Using two models of chronic liver injury, we demonstrate that MAIT cell-enriched mice show increased liver fibrosis and accumulation of hepatic fibrogenic cells, whereas MAIT cell-deficient mice are resistant. Co-culture experiments indicate that MAIT cells enhance the proinflammatory properties of monocyte-derived macrophages, and promote mitogenic and proinflammatory functions of fibrogenic cells, via distinct mechanisms. Our results highlight the profibrogenic functions of MAIT cells and suggest that targeting MAIT cells may constitute an attractive antifibrogenic strategy during chronic liver injury.

Background & Aims: Intestinal failure-associated liver disease is rare in adults and risk factors are unclear. The aim of this study was to determine risk factors of liver fibrosis in adults receiving home parenteral nutrition for intestinal failure and its impact on survival., Methods: We retrospectively analysed patients with irreversible intestinal failure who underwent a liver biopsy between 2000 and 2013. Significant liver fibrosis was defined as ≥F2 according to NASH-CRN score., Results: Thirty-two patients (46 years [29-60]) underwent liver biopsy 55 months (9-201) after beginning parenteral nutrition. Twenty-six patients (81%) had a short bowel (gut < 200 cm), including 12 (37%) with an ultra-short bowel (gut < 20 cm). Eighteen patients (56%) had liver fibrosis (4 F2, 10 F3, 4 F4), associated with steatohepatitis (72%) and/or cholestasis (17%). Factors associated with occurrence of liver fibrosis included ultra-short bowel (83% vs 13% at 60 months; P < .001), alcohol consumption (73% vs 33% at 60 months; P < .001) and diabetes (80% vs 34% at 60 months; P = .01). Home parenteral nutrition composition, quantity, or duration, episodes of sepsis, abandoned bowel segment were not associated with fibrosis. Ultra-short bowel [risk ratio 12.4, P < .001] and alcohol consumption [risk ratio 7.4, P = .009] independently predicted the development of liver fibrosis on multivariate analysis. After a median follow-up of 118 months (72-155), survival was poorer in patients who developed liver fibrosis than in those without (59% vs 92% at 120 months; P = .02)., Conclusion: An ultra-short bowel and alcohol consumption are independent risk factors for liver fibrosis in adults requiring HPN., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Purpose To determine the relationship of liver fibrosis, inflammation, and steatosis with the magnetic resonance (MR) viscoelastic and diffusion parameters in patients with chronic liver disease and to compare the diagnostic accuracy of the imaging parameters in staging liver fibrosis. Materials and Methods Consecutive patients with chronic liver disease scheduled for liver biopsy were prospectively recruited from November 2010 to October 2012 for this institutional review board-approved study after they provided written informed consent. Sixty-eight patients underwent three-dimensional MR elastography and intravoxel incoherent motion diffusion-weighted MR imaging with a 1.5-T MR system. Fibrosis, inflammation, and steatosis were assessed with the METAVIR and steatosis, activity, and fibrosis (or SAF) scoring systems. Spearman correlation and multiple regression analyses were performed to determine the relationship between liver fibrosis, inflammation, steatosis, and alanine aminotransferase (ALT) levels and viscoelastic and diffusion parameters. The accuracy of three-dimensional MR elastography and diffusion-weighted MR imaging in the determination of fibrosis stage was assessed with Obuchowski measures. Results At multiple regression analysis, fibrosis was the only variable associated with viscoelastic parameters (β = 0.6, P < .001, R 2 = 0.33 for shear modulus; β = 0.6, P < .001, R 2 = 0.32 for elasticity). Fibrosis had a weaker independent association with the apparent diffusion coefficient (β = -0.3, P = .02, R 2 = 0.33) than did steatosis (β = -0.5, P < .001, R 2 = 0.33). Steatosis was the only factor independently associated with the pure diffusion coefficient (β = -0.4, P = .002, R 2 = 0.22). Inflammation and ALT level were not associated with the viscoelastic or diffusion parameters. The diagnostic accuracy of fibrosis staging was significantly higher when measuring the shear modulus rather than the apparent diffusion coefficient (Obuchowski measures, 0.82 ± 0.04 vs 0.30 ± 0.06; P < .001). Conclusion Fibrosis is independently associated with the MR viscoelastic parameters and is less associated with the diffusion parameters than is steatosis. These results and those of diagnostic accuracy suggest that MR elastography should be preferred over diffusion-weighted MR imaging in the staging of liver fibrosis. © RSNA, 2016.

Hepatic fibrosis arises from inflammation in the liver initiated by resident macrophage activation and massive leukocyte accumulation. Hepatic macrophages hold a central position in maintaining homeostasis in the liver and in the pathogenesis of acute and chronic liver injury linked to fibrogenesis. Interferon regulatory factor 5 (IRF5) has recently emerged as an important proinflammatory transcription factor involved in macrophage activation under acute and chronic inflammation. Here, we revealed that IRF5 is significantly induced in liver macrophages from human subjects developing liver fibrosis from nonalcoholic fatty liver disease or hepatitis C virus infection. Furthermore, IRF5 expression positively correlated with clinical markers of liver damage, such as plasma transaminase and bilirubin levels. Interestingly, mice lacking IRF5 in myeloid cells (MKO) were protected from hepatic fibrosis induced by metabolic or toxic stresses. Transcriptional reprogramming of macrophages lacking IRF5 was characterized by immunosuppressive and antiapoptotic properties. Consequently, IRF5 MKO mice respond to hepatocellular stress by promoting hepatocyte survival, leading to complete protection from hepatic fibrogenesis. Our findings reveal a regulatory network, governed by IRF5, that mediates hepatocyte death and liver fibrosis in mice and humans. Therefore, modulating IRF5 function may be an attractive approach to experimental therapeutics in fibroinflammatory liver disease.

Disorders of glucose metabolism, namely glucose intolerance and diabetes, are frequent in patients with chronic liver diseases. In patients with cirrhosis, diabetes can be either a classical type 2 diabetes mellitus or the so-called hepatogenous diabetes, i.e. a consequence of liver insufficiency and portal hypertension. This review article provides an overview of the possible pathophysiological mechanisms explaining diabetes in patients with cirrhosis. Cirrhosis is associated with portosystemic shunts as well as reduced hepatic mass, which can both impair insulin clearance by the liver, contributing to peripheral insulin resistance through insulin receptors down-regulation. Moreover, cirrhosis is associated with increased levels of advanced-glycation-end products and hypoxia-inducible-factors, which may play a role in the development of diabetes. This review also focuses on the clinical implications of diabetes in patients with cirrhosis. First, diabetes is an independent factor for poor prognosis in patients with cirrhosis. Specifically, diabetes is associated with the occurrence of major complications of cirrhosis, including ascites and renal dysfunction, hepatic encephalopathy and bacterial infections. Diabetes is also associated with an increased risk of hepatocellular carcinoma in patients with chronic liver diseases. Last, the management of patients with concurrent diabetes and liver disease is also addressed. Recent findings suggest a beneficial impact of metformin in patients with chronic liver diseases. Insulin is often required in patients with advanced cirrhosis. However, the favourable impact of controlling diabetes in patients with cirrhosis has not been demonstrated yet., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Tumor recurrence occurs in almost 10% after liver transplantation (LT) for early stage hepatocellular carcinoma (HCC). Evidence-based recommendation regarding these techniques and their schedule remains unclear. The aim was to analyze the imaging characteristics of HCC recurrence after LT according to the initial primary tumor characteristics to provide relevant evidence regarding their role., Methods: Among 336 cirrhotic patients undergoing LT between 2000 and 2011, 25 (7.4%) experienced HCC recurrence. Their pre-LT characteristics, long-term outcome and imaging characteristics of recurrences were retrospectively analyzed., Results: All patients were men and median age at the time of recurrence was 54 (41-64) years. Before LT, 13 (52%) patients had HCC outside the Milan criteria. Median time to HCC recurrence was 13.8 months (1-75) after LT and 8 patients (32%) experienced recurrence longer than 24 months after LT. The most frequently involved organs were the lungs (n = 13, 52%) and the bones (n = 9, 36%). Recurrent HCC involved more than 1 organ in 11 patients (44%). Recurrences were limited to the liver in 1 patient (4%), exclusively extrahepatic in 18 patients (72%), and were both intra and extrahepatic in 6 patients (24%). Delay of recurrence was shorter in the presence of microvascular invasion (17 vs 28 months), but this was not significant (P = 0.208). Poorly differentiated tumors showed a trend toward decreased recurrence-free survival (8 vs 28 months, P = 0.075)., Conclusions: Both pattern and delay of HCC recurrence support performing regular whole-body imaging initially every 6 to 12 months and continuing beyond 2 years after LT for HCC, especially in patients with pejorative pathological features of the primary tumor that could influence HCC recurrence pattern.

Purpose: To compare the value of enhancement and pharmacokinetic parameters measured at dynamic gadoxetate-enhanced magnetic resonance (MR) imaging in determining hepatic organic anion transporter expression in control rats and rats with advanced liver fibrosis., Materials and Methods: Institutional animal review board approval was received before the study began. Advanced liver fibrosis was created in rats by means of carbon tetrachloride injections over an 8-week period. In 17 rats with liver fibrosis and eight control rats, dynamic gadoxetate-enhanced MR images of the liver were obtained during 1 hour after injection of 0.025 mmol gadoxetate per kilogram of body weight. Enhancement parameters (maximum enhancement [Emax], time to peak [Tmax], and elimination half-life) were measured on enhancement-versus-time curves, and pharmacokinetic parameters (hepatic extraction fraction [HEF] and mean residence time [MRT]) were obtained by means of deconvolution analysis of the concentration-versus-time curves in the liver and the portal vein. The parameters were correlated at simple and multiple regression analysis with the expression of the hepatic anion uptake transporter organic anion-transporting polypeptide 1A1 (Oatp1a1), the hepatobiliary transporter multidrug resistance-associated protein 2 (Mrp2), and the backflux transporter Mrp4, as determined with reverse transcription polymerase chain reaction., Results: In rats with advanced liver fibrosis, the Emax, Tmax, HEF, and MRT decreased significantly relative to those in control rats, whereas the elimination half-life increased significantly. The enhancement and pharmacokinetic parameters correlated significantly with the expression of the transporters at simple regression analysis. At multiple regression analysis, HEF was the only parameter that was significantly associated with the expression of Oatp1a1 and Mrp2 (P < .001, r = 0.74 and P < .001, r = 0.70, respectively)., Conclusion: The pharmacokinetic parameter HEF at dynamic gadoxetate-enhanced MR imaging is independently correlated with hepatic organic anion transporter expression., (© RSNA, 2014.)

Cirrhosis is a lesion at risk of hepatocellular carcinoma (HCC). Identifying mechanisms associated with the transition from cirrhosis to HCC and characterizing biomarkers of cirrhosis at high risk of developing into cancer are crucial for improving early diagnosis and prognosis of HCC. We used MALDI imaging to compare mass spectra obtained from tissue sections of cirrhosis without HCC, cirrhosis with HCC, and HCC, and a top-down proteomics approach to characterize differential biomarkers. We identified a truncated form of monomeric ubiquitin lacking the two C-terminal glycine residues, Ubi(1-74), the level of which increased progressively, from cirrhosis without HCC to cirrhosis with HCC to HCC. We showed that kallikrein-related peptidase 6 (KLK6) catalysed the production of Ubi(1-74) from monomeric ubiquitin. Furthermore, we demonstrated that KLK6 was induced de novo in cirrhosis and increased in HCC in parallel with accumulation of Ubi(1-74). We investigated in vitro the possible consequences of Ubi(1-74) accumulation and demonstrated that Ubi(1-74) interferes with the normal ubiquitination machinery in what is likely to be a kinetic process. Our data suggest that de novo KLK6 expression during early liver carcinogenesis may induce production of Ubi(1-74) by post-translational modification of ubiquitin. Given the deleterious effect of Ubi(1-74) on protein ubiquitination and the major role of ubiquitin machinery in maintenance of cell homeostasis, Ubi(1-74) might severely impact a number of critical cellular functions during transition from cirrhosis to cancer. Ubi(1-74) and KLK6 may serve as markers of cancer risk in patients with cirrhosis., (Copyright © 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Background and Aims: There is evidence for an association between thrombosis in the hepatic microcirculation and liver fibrosis. The aim of this study was to evaluate the role of daily low-dose aspirin (75 or 100mg, given for prevention of hepatic artery thrombosis) in fibrosis progression to ≥ F2 fibrosis score in liver-transplant recipients with recurrent hepatitis C virus (HCV)., Methods: All HCV-positive patients who had undergone liver transplantation (LT) between 2000 and 2010 were included. Exclusion criteria were negative HCV RNA, previous LT or death within a year of LT. Liver fibrosis was assessed by histological evaluation. Data were censored at the date of the last histological evaluation before starting anti HCV therapy. Progression to fibrosis F ≥ 2 was analyzed with a multistate model with time-dependent covariables., Results: One hundred and eighty-eight patients were included. In univariate analysis, older recipient and donor age, male donor gender, activity score ≥ A2 after LT, number of steroid boluses and aspirin intake (HR: 0.75 [0.57-0.97]; P=0.03) influenced the risk of progression to fibrosis ≥ F2. In multivariate analysis, adjusted on site, older donor age, male donor gender, activity score ≥ A2 and number of steroids boluses, remained independent predictors of fibrosis progression, while younger recipient age and aspirin intake (HR: 0.65 [0.47-0.91]; P=0.01) were associated with a slower fibrosis progression., Conclusion: Low-dose aspirin treatment might be associated with a lower risk of liver fibrosis progression in patients with HCV recurrence after LT., (Copyright © 2014 Elsevier Masson SAS. All rights reserved.)

Objective: To assess in a high-resolution model of thin liver rat slices which viscoelastic parameter at three-dimensional multifrequency MR elastography has the best diagnostic performance for quantifying liver fibrosis., Materials and Methods: The study was approved by the ethics committee for animal care of our institution. Eight normal rats and 42 rats with carbon tetrachloride induced liver fibrosis were used in the study. The rats were sacrificed, their livers were resected and three-dimensional MR elastography of 5 ± 2 mm liver slices was performed at 7T with mechanical frequencies of 500, 600 and 700 Hz. The complex shear, storage and loss moduli, and the coefficient of the frequency power law were calculated. At histopathology, fibrosis and inflammation were assessed with METAVIR score, fibrosis was further quantified with morphometry. The diagnostic value of the viscoelastic parameters for assessing fibrosis severity was evaluated with simple and multiple linear regressions, receiver operating characteristic analysis and Obuchowski measures., Results: At simple regression, the shear, storage and loss moduli were associated with the severity of fibrosis. At multiple regression, the storage modulus at 600 Hz was the only parameter associated with fibrosis severity (r = 0.86, p<0.0001). This parameter had an Obuchowski measure of 0.89+/-0.03. This measure was significantly larger than that of the loss modulus (0.78+/-0.04, p = 0.028), but not than that of the complex shear modulus (0.88+/-0.03, p = 0.84)., Conclusion: Our high resolution, three-dimensional multifrequency MR elastography study of thin liver slices shows that the storage modulus is the viscoelastic parameter that has the best association with the severity of liver fibrosis. However, its diagnostic performance does not differ significantly from that of the complex shear modulus.

Objectives: Combining noninvasive tests increases diagnostic accuracy for staging liver fibrosis in hepatitis C virus (HCV)-infected patients, but this strategy remains to be validated in HIV/HCV coinfection. We compared the performances of transient elastography (TE), Fibrotest (FT), the aspartate aminotransferase-to-platelet ratio index (APRI) and two algorithms combining TE and FT (Castera) or APRI and FT (SAFE) in HIV/HCV coinfection., Methods: One hundred and sixteen HIV/HCV-coinfected patients (64% male; median age 44 years) enrolled in two French multicentre studies (the HEPAVIH cohort and FIBROSTIC) for whom TE, FT and APRI data were available were included in the study. Diagnostic accuracies for significant fibrosis (METAVIR F ≥ 2) and cirrhosis (F4) were evaluated by measuring the area under the receiver-operating characteristic curve (AUROC) and calculating percentages of correctly classified (CC) patients, taking liver biopsy as a reference., Results: For F ≥ 2, both TE and FT (AUROC = 0.87 and 0.85, respectively) had a better diagnostic performance than APRI (AUROC = 0.71; P < 0.005). Although the percentage of CC patients was significantly higher with Castera's algorithm than with SAFE (61.2% vs. 31.9%, respectively; P < 0.0001), this percentage was lower than that for TE (80.2%; P < 0.0001) or FT (73.3%; P < 0.0001) taken separately. For F4, TE (AUROC = 0.92) had a better performance than FT (AUROC = 0.78; P = 0.005) or APRI (AUROC = 0.73; P = 0.025). Although the percentage of CC patients was significantly higher with the SAFE algorithm than with Castera's (76.7% vs. 68.1%, respectively; P < 0.050), it was still lower than that for TE (85.3%; P < 0.033)., Conclusions: In HIV/HCV-coinfected patients, TE and FT have a similar diagnostic accuracy for significant fibrosis, whereas for cirrhosis TE has the best accuracy. The use of the SAFE and Castera algorithms does not seem to improve diagnostic performance., (© 2013 British HIV Association.)

Background & Aims: Transient elastography (TE) is a validated non-invasive tool to evaluate hepatic fibrosis in patients with hepatitis C virus (HCV) infection. Whether TE may sense changes of liver fibrosis following therapeutic HCV eradication has never been evaluated., Methods: 37 HCV cirrhotics with paired pre- and post-sustained virological response (SVR) liver biopsies (LB) underwent TE at the time of post-SVR LB. Liver fibrosis was staged with the METAVIR scoring system and the area of fibrosis (%) was assessed morphometrically., Results: Thirty-three patients had valid TE measurements after 61 (48-104) months from an SVR, and 20 (61%) of them had cirrhosis regression. On post-SVR LB, the median area of fibrosis was 2.3%, being significantly reduced from baseline (p<0.0001). Median TE value was 9.8 kPa being lower in regressed vs. not regressed patients (9.1 kPa vs. 12.9 kPa, p=0.01). TE was <12 kPa in 5 (38%) F4 patients and in 19 (95%) ≤F3 patients (p=0.0007). The diagnostic accuracy of TE for diagnosing F4 after treatment was 61% sensitivity, 95% specificity, 12.3 LR+, 0.4 LR-, and AUROC 0.77. A significant correlation was found between TE and both fibrosis stage (r=0.56; p=0.001) and morphometry (r=0.56, p=0.001) as well as between fibrosis stage and area of fibrosis (r=0.72, p=0001)., Conclusions: Following therapeutic eradication of HCV, the predictive power of the viremic cut-off of 12 kPa was low as a consequence of liver remodelling and fibrosis reabsorption. LB still remains the only reliable approach to stage liver fibrosis following an SVR., (Copyright © 2013 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background and Aims: The role of insulin resistance (IR) in chronic hepatitis C genotype 4 (CHC-4) patients is still under assessment. The aims of this study are to assess the prevalence and predictors of IR and its influence along with clinical, metabolic, virological, and histological factors on the severity of liver fibrosis in 100 Egyptian patients with CHC-4., Patients and Methods: In 100 untreated patients with CHC-4, IR was assessed using the Homeostasis Model Assessment and defined greater than 3. By logistic regression (LR), independent factors associated with IR and significant fibrosis (SF=fibrosis, Metavir score≥F2) were assessed in nondiabetic and noncirrhotic patients., Results: One hundred patients were included; 54% were men and 46% were women. The mean age of the patients was 40.46±9.41 years. Of the total patients, 55% were overweight and 28% were obese. Metabolic syndrome was observed in 26% of patients; five of them were known to be diabetic. All patients were genotype 4. Most of our patients had mild viremia (<2 00 000 IU/ml), whereas only 16% had higher viral load (>2 00 000 IU/ml). There was no correlation between IR and hepatitis C virus viremia (r=-0.069; P=0.492). Necroinflammation was moderate-severe (A2-A3) in 25% of patients. SF (F2-F4) was found in 46% of patients and 11% had cirrhosis (F4). Most of our patients, 54%, had moderate steatosis and 21% had severe steatosis. IR was present in 46% of patients; 39 (42.9%) were nondiabetic, which is correlated significantly with BMI (r=0.395; P<0.01). IR was found to increase significantly with the fibrosis stage (P=0.001), insignificant fibrosis, 18.5%, SF (F2-F4), 71.4%, and cirrhosis (F4), 100%. By LR, IR was independently and significantly associated with age more than 40 years, obesity (BMI>30 kg/m), SF, and severe steatosis (>30%). IR was also significantly associated with metabolic syndrome. SF was present in 46 patients (46%). It was associated with IR, moderate-severe necroinflammation, and severe steatosis. By LR, in noncirrhotic patients, SF was associated with age more than 40 years, obesity (BMI>30 kg/m), moderate/severe liver inflammation, and severe steatosis., Conclusion: In CHC-4 patients, IR is highly prevalent and independently associated with age, obesity, SF, and severe steatosis. Management of IR might significantly improve the prognosis of CHC-4 patients.

Unlabelled: Nonalcoholic fatty liver disease (NAFLD) is highly prevalent and being overweight is a significant risk factor. The aim was to build an algorithm along with a scoring system for histopathologic classification of liver lesions that covers the entire spectrum of lesions in morbidly obese patients. A cohort of 679 obese patients undergoing liver biopsy at the time of bariatric surgery was studied. An algorithm for segregating lesions into normal liver, NAFLD, or nonalcoholic steatohepatitis (NASH) was built based on semiquantitative evaluation of steatosis, hepatocellular ballooning, and lobular inflammation. For each case, the SAF score was created including the semiquantitative scoring of steatosis (S), activity (A), and fibrosis (F). Based on the algorithm, 230 obese patients (34%) were categorized as NASH, 291 (43%) as NAFLD without NASH, and 158 (23%) as not NAFLD. The activity score (ballooning + lobular inflammation) enabled discriminating NASH because all patients with NASH had A ≥ 2, whereas no patients with A < 2 had NASH. This score was closely correlated with both alanine aminotransferase (ALT) and aspartate aminotransferase (AST) (P < 0.0001, analysis of variance [ANOVA]). Comparison of transaminase levels between patients with normal liver and pure steatosis did not reveal significant differences, thus lending support to the proposal not to include steatosis in the activity score but to report it separately in the SAF score. In the validation series, the interobserver agreement for the diagnosis of NASH was excellent (κ = 0.80) between liver pathologists. There was no discrepancy between the initial diagnosis and the diagnosis proposed using the algorithm., Conclusion: We propose a simple but robust algorithm for categorizing liver lesions in NAFLD patients. Because liver lesions in obese patients may display a continuous spectrum of histologic lesions, we suggest describing liver lesions using the SAF score., (Copyright © 2012 American Association for the Study of Liver Diseases.)

Unlabelled: Although annular fibrosis is the hallmark of cirrhosis, other microscopic changes that affect liver function such as sinusoid capillarization or loss of metabolic zonation are common. A sustained virological response (SVR) may halt fibrosis deposition in hepatitis C virus (HCV)-infected patients, but its impact on the other cirrhosis-associated lesions is unknown. The aim of this study was to assess the impact of an SVR on cirrhosis-related histopathological features. Paired pre- and posttreatment liver biopsies from 38 HCV patients with cirrhosis with an SVR were analyzed. Fibrosis was staged using the METAVIR scoring system, and the area of fibrosis was measured using morphometry. Ductular proliferation, metabolic zonation, sinusoid capillarization, and hepatic stellate cell activation were assessed by anti-cytokeratin-7, anti-glutamine synthetase (GS), anti-cytochrome P4502E1 (CYP2E1), anti-CD34, and anti α-smooth muscle actin (αSMA). After 61 months from an SVR, cirrhosis regression was observed in 61%, and the collagen content decreased in 89%. Although periportal and lobular necroinflammation vanished, portal inflammation persisted in 66%. Ductular proliferation decreased in 92%. Before treatment, metabolic zonation was lost, as shown by GS and CYP2E1, in 71% and 88%, respectively, with normalization in 79% and 73%, after an SVR. Conversely, no changes in sinusoidal capillarization were observed after treatment, as assessed by CD34 (P = 0.41) and αSMA (P = 0.95). Finally, no differences in all the immunohistochemical scores emerged whether or not cirrhosis persisted., Conclusion: Cirrhosis regression and decreased fibrosis are frequently observed among HCV patients with cirrhosis with an SVR. Despite ductular proliferation vanishing and lobular zonation restoration, portal inflammation and sinusoidal capillarization may not regress after viral eradication., (Copyright © 2012 American Association for the Study of Liver Diseases.)

Background & Aims: Circulating membrane-shed microparticles (MPs) participate in regulation of vascular tone. We investigated the cellular origins of MPs in plasma from patients with cirrhosis and assessed the contribution of MPs to arterial vasodilation, a mechanism that contributes to portal hypertension., Methods: We analyzed MPs from blood samples of 91 patients with cirrhosis and 30 healthy individuals (controls) using flow cytometry; their effects on the vascular response to vasoconstrictors were examined in vitro and in vivo., Results: Circulating levels of leuko-endothelial (CD31(+)/41(-)), pan-leukocyte (CD11a(+)), lymphocyte (CD4(+)), and erythrocyte (CD235a(+)) MPs were higher in patients with cirrhosis than in controls. Plasma of patients with cirrhosis contained hepatocyte-derived MPs (cytokeratin-18(+)), whereas plasma from controls did not. The severity of cirrhosis and systemic inflammation were major determinants of the levels of leuko-endothelial and hepatocyte MPs. MPs from patients with advanced cirrhosis significantly impaired contraction of vessels in response to phenylephrine, whereas MPs from healthy controls or from patients of Child-Pugh class A did not. This effect depended on cyclooxygenase type 1 and required phosphatidylserine on the surface of MPs. Intravenous injection of MPs from patients with cirrhosis into BALB/C mice decreased mean arterial blood pressure., Conclusions: Cirrhosis is associated with increases in circulating subpopulations of MPs, likely resulting from systemic inflammation and liver cell damage. The overall pool of circulating MPs from patients with advanced cirrhosis impairs vasoconstrictor responses and decreases blood pressure, contributing to the arterial vasodilation associated with portal hypertension., (Copyright © 2012 AGA Institute. Published by Elsevier Inc. All rights reserved.)

"Cirrhosis" is a morphologic term that has been used for almost 200 years to denote the end stage of a variety of chronic liver diseases. The term implies a condition with adverse prognosis due to the well-known complications of portal hypertension, hepatocellular carcinoma, and liver failure. However, recent advances in the diagnosis and treatment of chronic liver diseases have changed the natural history of cirrhosis significantly. This consensus document by the International Liver Pathology Study Group challenges the usefulness of the word cirrhosis in modern medicine and suggests that this is an appropriate time to consider discontinuing the use of this term. The role of pathologists should evolve to the diagnosis of advanced stage of chronic liver disease, with emphasis on etiology, grade of activity, features suggestive of progression or regression, presence of other diseases, and risk factors for malignancy, within the perspective of an integrated clinicopathologic assessment.

MALDI imaging mass spectrometry (MALDI IMS) is a powerful tool for comprehending the spectrum of peptides/proteins expressed in tissue sections. The aim of the present study was to investigate, using MALDI IMS, the proteome of hepatocellular carcinomas (HCC) and to compare it with peritumoral cirrhosis so as to characterize new biomarkers of HCC. Frozen liver tissues corresponding to HCC and background cirrhosis (n = 30) were selected and subjected to MALDI IMS. We found a set of proteins/peptides with a differential intensity level that most accurately delineated cancer from adjacent cirrhotic tissue. Using a support vector machine algorithm, we generated a classification model in the train set that enabled segmenting images from the independent validation set and that in most cases matched histologic analysis. The most discriminating peak (m/z 8565) more intense in HCC was characterized as the monomeric ubiquitin. An immunohistochemical study in a large series of HCC/cirrhosis sampled on tissue microarray supported that ubiquitin was overexpressed in HCC. We demonstrated also that this increase was not related to an upregulation of ubiquitin gene transcription in HCC, thus suggesting a post-transcriptional mechanism. This approach might provide a new tool for diagnosis of difficult HCC cases and an opportunity for identifying candidate biomarkers.

During liver fibrogenesis, quiescent hepatic stellate cells switch their phenotype toward a myofibroblastic-like pattern with a gain in motility. Here, we show that SCG10 (superior cervical ganglia 10) mRNA expression, a microtubule-destabilizing protein that favors cell growth and motility in neurons, both increases and correlates with the stage of fibrosis in patients with chronic hepatitis C. We also show the de novo expression of SCG10 mRNA in two rat models of liver fibrosis. We demonstrate that activated hepatic stellate cells appear to be the major cellular sources of SCG10 in the liver. Tracking of the SCG10 pathway in hepatic stellate cells shows that SCG10 initially accumulates in the perinuclear Golgi area then migrates in small vesicle-like structures along individual microtubules. Moreover, SCG10 vesicles cluster at the distal ends of microtubules in areas where tubules are spread and decompacted, suggesting their preferential association with destabilized and dynamic microtubules. Inhibition of SCG10 expression by gene-specific short interfering RNA in primary rat hepatic stellate cells is associated with a significant reduction in microtubule-dependent cellular functions, such as proliferation and migration. In conclusion, the de novo expression of SCG10 by hepatic stellate cells may play a major role in cellular mechanisms associated with HSC activation, namely cell motility and division, through interference with microtubules. SCG10 may represent a potential molecular target for anti-fibrosis therapies.

Purpose: To prospectively assess the utility of perfusion computed tomography (CT) for differentiating minimal from intermediate fibrosis in treatment-naïve patients with chronic hepatitis C virus (HCV) infection., Materials and Methods: This study was approved by the Institutional Review Board, and informed consent was obtained. Fifty-two patients with treatment-naïve HCV infection underwent perfusion CT and percutaneous liver biopsy on the same day. Portal vein, arterial, and total liver perfusion; mean transit time; and distribution volumes for the right and left liver lobes were measured. Liver samples were scored for fibrosis, and fibrosis area was determined. Differences in quantitative perfusion parameters between patients with minimal fibrosis (score of F1) and those with intermediate fibrosis (score of F2 or F3) were tested., Results: In patients with intermediate fibrosis (F2 and F3) compared with those with minimal fibrosis (F1), the portal venous perfusion (87 mL min(-1) 100 mL(-1) +/- 27 [standard deviation] vs 138 mL min(-1) 100 mL(-1) +/- 112, P = .042) and total liver perfusion (107 mL min(-1) 100 mL(-1) +/- 31 vs 169 mL min(-1) 100 mL(-1) +/- 137, P = .02) were significantly decreased, and the mean transit time was significantly increased (16 seconds +/- 4 vs 13 seconds +/- 5, P = .025). At multivariate analysis, only the mean transit time was an independent factor (odds ratio, 1.18; 95% confidence interval: 1.02, 1.37; P = .030). Receiver operating characteristic curve analysis showed that a mean transit time threshold of 13.4 seconds allowed discrimination between minimal and intermediate fibrosis with a sensitivity of 71% and a specificity of 65%., Conclusion: The results of this study show that perfusion changes occur early during fibrosis in chronic HCV infection and can be detected with perfusion CT. Perfusion CT may help to discriminate minimal from intermediate fibrosis. Mean transit time appears to be the most promising perfusion parameter for differentiating between fibrosis stages, although the large amount of overlap in the measured parameters limits the clinical utility of this test at present.

Background/objective: Little is known on the morphological changes in the kidneys of cirrhotic patients with abnormal urinalysis and/or high serum creatinine levels. This retrospective, one-point-in-time study aimed to report the results of the analysis of renal biopsy specimens obtained in patients with cirrhosis., Methods: We retrieved information on 65 patients who underwent transvenous renal biopsy for proteinuria >0.5 g/day and/or microscopic haematuria and/or unexplained renal impairment (defined by serum creatinine levels >1.5 mg/dl)., Results: Fifty-one per cent of the patients had proteinuria >0.5 g/day, 58% had haematuria and 83% had renal impairment. Renal biopsy disclosed injury to glomeruli in 77% of the patients, to vessels in 69% and to the tubulointerstitium system in 94% (chronic in 77%; acute in 75%). Fibrous endarteritis was the most common renal vascular lesion. Injuries to different structures were frequently combined. Isolated glomerular alterations were found in only two patients. Acute tubular necrosis was significantly more common in patients with fibrous endarteritis than in those without. Among 18 patients with renal impairment, proteinuria <0.5 mg/day and no haematuria, 10 had glomerular lesions, 13 had chronic tubulointerstitial lesions and 12 acute tubulointerstitial lesions., Conclusion: In patients with cirrhosis, various types of renal injuries are frequently combined. Chronic lesions (vascular or tubulointerstitial) may influence the outcome, in particular in patients who subsequently undergo liver transplantation and receive anticalcineurins. Renal vascular lesions may increase the risk of acute tubular necrosis. In patients with renal impairment, the absence of significant proteinuria and haematuria do not rule out the presence of renal lesions.

Unlabelled: Metabolic syndrome (MS) is a newly identified risk factor in chronic liver disease (CLD) and hepatocellular carcinoma (HCC). The aim of this study was to analyze the pathological characteristics of HCC and nontumoral liver in patients with MS as the only risk factor for liver disease in comparison with those that developed in the course of other CLDs in order to provide further insight into the physiopathology of HCC associated with MS. HCC patients with features of MS as the only risk factor for liver diseases (MS group, n = 31) were compared to HCC patients with overt causes of CLD (CLD group, n = 81) or without causes of CLD (cryptogenic group, n = 16) who underwent surgical resection during the same period of time. Among the patients of the MS group, there were 30 males and 1 female. In comparison with the patients with HCC of the CLD group, the patients with MS were older (mean age: 67+/- 7 versus 59 +/- 14 years, P < 0.01), and the background liver was significantly more often free of significant fibrosis (F0-F2: 65% in the MS group versus 26% in the CLD group, P < 0.001). In addition, HCCs associated with MS were more often well differentiated (65% versus 28%, P < 0.001). Five HCCs, all from the MS group, developed on a preexisting liver cell adenoma, with three of them showing typical histological features of telangiectatic adenoma., Conclusion: This study shows that HCCs in patients with features of MS as the only risk factor for liver disease have distinct morphological characteristics and mainly occur in the absence of significant fibrosis in the background liver. In addition, some of them arise through malignant transformation of a preexisting liver cell adenoma.

Background & Aims: Our study was designed to test the association between insulin resistance (IR) and hepatitis C virus (HCV) genotypes, serum HCV RNA level and liver fibrosis stage in a large prospective cohort of chronic hepatitis C (CHC) patients., Methods: Six hundred consecutive patients (CHC, n = 500; chronic hepatitis B (CHB), n = 100) were evaluated on the day of liver biopsy. IR (Homeostasis Model for Assessment of Insulin Resistance) and all components of the metabolic syndrome were assessed. By logistic regression, independent factors associated with IR and those associated with significant fibrosis were assessed in nondiabetic and noncirrhotic CHC, respectively. Parameters of IR were compared between hepatitis B and 240 CHC matched by epidemiologic, metabolic, and histologic features., Results: IR was present in 32.4% of the 462 nondiabetic CHC and associated with the metabolic syndrome, genotypes 1 and 4, significant fibrosis, and severe steatosis. IR was diagnosed in 15% of 145 CHC without metabolic syndrome or significant fibrosis, and associated with genotypes 1 and 4, high serum HCV RNA level, and moderate-severe necroinflammation. Significant fibrosis was present in 51.1% of the 454 noncirrhotic CHC patients and associated with male sex, age >40 years, IR, moderate-severe necroinflammation, and severe steatosis. IR was less frequent in CHB than in matched CHC (5% vs 35%, respectively, P < .001)., Conclusions: IR is a specific feature of CHC, associated with genotypes 1 and 4 and high serum HCV RNA level. Significant fibrosis is associated with IR independent from steatosis.

Background: FibroTest has been validated for the diagnosis of liver fibrosis in patients with chronic hepatitis C., Aim: To compare FibroTest with a new proteome-based model for the prediction of advanced liver fibrosis., Methods: Sera from 191 consecutive patients with simultaneous liver biopsy and FibroTest on fresh sera were used for retrospective mass spectrometry analysis. A new fibrosis index was constructed combining proteomic peaks, selected on differential expression according to fibrosis stages in logistic regression analyses. The main end point was the diagnosis of advanced fibrosis on liver biopsy., Results: Eight out of 1000 peaks were selected for the construction of the proteomic index. The area under the receiver operating curve (AUROC) of the proteomic index was 0.88 (95% CI: 0.82-0.92), significantly greater than the FibroTest AUROC of 0.81 (95% CI: 0.74-0.86; P = 0.04); the AUROC of the proteomic and FibroTest combination was 0.88 (95% CI: 0.83-0.92). Seven of the eight selected peaks were highly associated with the FibroTest score, with different patterns of association with the five components of FibroTest., Conclusions: A proteomic index combining eight peaks had an excellent accuracy value for the diagnosis of advanced fibrosis in patients with chronic hepatitis C. However, despite a statistical significance, the small improvement delivered by proteomics impairs clinical applications because of its cost and its variability compared with the well validated FibroTest.

Background/aims: We evaluated the test performance profile (TPP) of blood tests of liver fibrosis., Methods: Three hundred and fifty-six patients with C chronic hepatitis were included in two centers. Metavir staging of liver specimens by two independent pathologists and the following tests were evaluated: Fibrotest (FT), APRI, FibroMeter (FM), and Hepascore (HS)., Results: Metavir stages were: F0: 4%, F1: 55%, F2: 26%, F3: 11%, and F4: 4%. The AUROCs were not significantly different, respectively, FT, FM, APRI, HS: >or=F2: 0.79, 0.78, 0.76, >or=0.76; F3: 0.81, 0.85, 0.81, 0.81; and F4: 0.86, 0.94, 0.92, 0.89. The TPP relies on the paired comparison of blood-test misclassification based on liver specimen, e.g. FT vs FM, respectively: F0+1: 18 vs 28% (p=0.0003), >or=F2: 43 vs 31% (p=0.004). There was no center effect., Conclusions: In those populations, the four blood tests had a similar performance for significant fibrosis (F>or=2), lying in the lower range of published results which is attributable to a low >or=F2 prevalence, and for >or=F3 and F4. However, FM and FT had performance profiles significantly different as a function of fibrosis stages or diagnostic target (fibrosis cut-off). This has to be considered during the interpretation process. Moreover, the performance should be reported with different diagnostic targets.

Cirrhosis is a heterogeneous tissue composed of polyclonal regenerative and monoclonal neoplastic, potentially malignant nodules from which hepatocellular carcinoma (HCC) might develop. The aim of this study was to investigate proteomic profile changes associated with clonal expansion of cirrhotic nodules and malignant transformation of monoclonal nodules. Seventy-one cirrhotic nodules from 10 female patients with six HCC were dissected from liver surgical specimen by laser capture microdissection. Clonal status of each nodule was assessed by the study of X-chromosome inactivation pattern using the human androgen receptor. Protein profiles were determined by surface-enhanced laser desorption ionisation-time-of-flight technology using Q10 arrays (Cyphergen ProteinChip). Molecular weight of differentially expressed protein peaks was assessed. An average of 50 protein peaks was obtained for each nodule's profile. Comparison of protein profiles in polyclonal (n=45) and monoclonal cirrhotic nodules (n=26) identified three differentially expressed protein peaks (10,092, 54,025 and 62,133 Da). All were upregulated in monoclonal nodules. Twelve peaks were differentially expressed between monoclonal nodules and HCC with nine proteins upregulated in cancer samples. This study confirms that proteome analysis can be achieved from a limited number of microdissected cells, and provides further insight into the process of clonal expansion and malignant transformation of cirrhotic nodules.

Background and Aims: Splanchnic vein thrombosis is a significant source of complications in candidates for liver transplantation. The aims of this study were: (a) to determine the prevalence of and risk factors for splanchnic vein thrombosis in cirrhotic patients awaiting transplantation and (b) to assess the usefulness of anticoagulation., Methods: A total of 251 cirrhotic patients listed for transplantation were analysed. All underwent systematic screening for thrombosis with Doppler ultrasonography. During the second period of the study, all patients with thrombosis received anticoagulation up to transplantation while during the first period none had received anticoagulation., Results: The incidence of splanchnic vein thrombosis at evaluation was 8.4%. Seventeen additional patients (7.4%) developed de novo thrombosis after evaluation. Independent risk factors for thrombosis were low platelet count (77.4 (36.3) v 111.6 (69.2) 10(9)/l; p = 0.001), a past history of variceal bleeding (47.4% v 29.1%; p = 0.003), and a prolonged interval from listing to transplantation (8.5 (6.8) v 4.8 (4.4) months; p = 0.002). The proportion of partial or complete recanalisation was significantly higher in those who received (8/19) than in those who did not receive (0/10, p = 0.002) anticoagulation. Survival was significantly lower in those who had complete portal vein thrombosis at the time of surgery (p = 0.04)., Conclusion: These results support a systematic screening for splanchnic vein thrombosis in patients awaiting transplantation. They suggest that in these patients, anticoagulation is safe and has a significant impact on recanalisation as well as prevention of extension of thrombosis.

Surface-enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF MS) is a proteomic technique that enables the profiling of proteins present in any biological material studied. We used this approach to identify new biomarkers of hepatocellular carcinoma (HCC) in the sera of patients with cirrhosis. Sera from 82 patients with cirrhosis, either without (n = 38) or with (n = 44) HCC, were analyzed by SELDI-TOF MS, and the results of the two groups were compared. The most efficient protein peaks leading to discrimination of patients with HCC were selected (receiver operative characteristic curves). The highest-scoring peak combination was established in a first group of serum samples (multinomial regression) and was tested in an independent group. The protein corresponding to the highest discrimination was purified and characterized further. The intensity of 30 protein peaks significantly differed between cirrhotic patients with and without HCC. An algorithm including the six highest-scoring peaks allowed correct classification (presence or absence of HCC) of 92.5% of patients in the test sample set and 90% in the validation sample set. The highest discriminating peak (8900 Da) was purified further and was characterized as the C-terminal part of the V10 fragment of vitronectin. An in vitro study suggested that the increase of the 8900-Da fragment in the serum of patients with HCC may proceed from the cleavage of native vitronectin with metalloproteases, a family of enzymes whose activity is enhanced in HCC. In conclusion, global protein profiling is an efficient approach that enabled us to identify a catalytic fragment ofvitronectin as a new serum marker of HCC in patients with chronic liver diseases.

Female sex is a protective factor for the progression of fibrosis in patients with chronic hepatitis C virus (HCV) infection. Experimental data suggest that estrogens may have an antifibrotic effect. The objective of this study was to evaluate the influence of past pregnancies, oral contraceptives, menopause, and hormone replacement therapy (HRT) on liver fibrosis progression in HCV-infected women. Four hundred seventy-two HCV-infected women received a survey regarding prior pregnancies, menopause, and the use of oral contraceptives and HRT. The impact of these variables on liver fibrosis and its progression were evaluated using multivariate analyses considering all putative confounding factors. Two hundred one women completed the survey (43% response rate), 157 of whom had an estimated date of HCV infection (96 postmenopausal women, 96 women with previous pregnancies, and 105 women with past use of oral contraceptives). Through multivariate analyses, the estimated rate of fibrosis progression was higher in postmenopausal (P < .05) and nulliparous (P = .02) women and was associated with greater histological activity (P < .001). Prior use of oral contraceptives had no significant influence. Among postmenopausal women, the estimated rate of fibrosis progression (+/-SE) was lower in women who received HRT compared with untreated patients (0.099 +/- 0.016 vs. 0.133 +/- 0.006 METAVIR units/yr; P = .02) and was similar to that of premenopausal women (0.093 +/- 0.012 METAVIR units/yr; P value not significant). In conclusion, menopause appears to be associated with accelerated liver fibrosis progression in HCV-infected women, an effect that may be prevented by HRT. Pregnancies may have a beneficial impact on the long-term progression of liver fibrosis.

Background: Although hepatitis C virus (HCV) recurrence is almost universal after orthotopic liver transplantation (OLT), the impact of viral infection on liver graft is highly variable and difficult to predict. Because of the possible relationship between replicative senescence (RS) and the accelerated development of liver fibrosis, we aimed to assess the potential role of RS in the severity of HCV-related chronic hepatitis recurrence after OLT., Methods: One hundred three liver biopsies from 56 patients receiving transplants for HCV-related cirrhosis were studied, including 30 revascularization biopsies and 52 and 21 biopsies performed during and beyond the first year of OLT, respectively. The presence of senescent cells in liver grafts was assessed by the senescence-associated beta-galactosidase (SA-beta-Gal) staining method. Chronic hepatitis was defined by fibrosis stage and necrotico-inflammatory activity grade using the METAVIR score., Results: A total of 34 of the 103 (33%) frozen liver biopsies displayed SA-beta-Gal-positive cells, including 6 (20%) of the revascularization biopsies, 14 (34%) of the biopsies performed within the first year, and 10 (46%) of the biopsies performed beyond 1 year of follow-up. The presence of senescent cells in revascularization biopsies was significantly associated with the degree of ischemic necrosis at time of OLT (P = 0.01) and hepatitis C recurrence in the first year after OLT (P = 0.05). Furthermore, the presence of RS in the biopsy performed within the first year was associated with further development of fibrosis (P = 0.05)., Conclusions: These data show that RS has a significant impact upon the course of liver transplantation, especially in the long-term progression of fibrosis observed in HCV-infected patients.

Liver transplantation offers good results in patients with small hepatocellular carcinoma. However, 3 to 15% of patients still have recurrence, suggesting that factors other than the size and number of nodules are implicated. The aim of our study was to identify predictive factors of recurrence in patients with small hepatocellular carcinoma. Seventy consecutive patients fulfilling Milano criteria and who were transplanted for hepatocellular carcinoma were studied. Forty-six patients had pretransplantation adjuvant local therapy. The size and number of tumors, the clinical and biological characteristics of the patients were recorded before liver transplantation, and histological analysis was performed on the explanted liver. Overall survival rates at 1 and 3 years were 81% and 66%, respectively. Recurrence-free survival rates at 1 and 3 years were 80% and 65%, respectively. Seven patients had tumor recurrence with 1- and 3-year recurrence rates of 5% and 10%, respectively. Satellite nodules on the explanted liver were the only statistically significant predictor of recurrence (P=.0003). None of the patients who did not have satellite nodules had recurrence. There was a significant correlation between satellite nodules and microvascular invasion. Patients with pretransplantation adjuvant therapy had significantly more tumor necrosis, but did not have less satellite nodules. In conclusion, microscopic satellite nodules are a significant predictive factor of tumor recurrence in patients transplanted for small hepatocellular carcinoma.

Fibrosis is a common endpoint of clinical trials in chronic hepatitis C, and liver biopsy remains the gold standard for fibrosis evaluation. However, variability in the distribution of fibrosis within the liver is a potential limitation. Our aim was to assess the heterogeneity of liver fibrosis and its influence on the accuracy of assessment of fibrosis with liver biopsy. Surgical samples of livers from patients with chronic hepatitis C were studied. Measurement of fibrosis was performed on the whole section by using both image analysis and METAVIR score (reference value). From the digitized image of the whole section, virtual biopsy specimens of increasing length were produced. Fibrosis was assessed independently on each individual virtual biopsy specimen. Results were compared with the reference value according to the length of the biopsy specimen. By using image analysis, the coefficient of variation of fibrosis measurement with 15-mm long biopsy specimens was 55%; and for biopsy specimens of 25-mm length it was 45%. By using the METAVIR scoring system, 65% of biopsies 15 mm in length were categorized correctly according to the reference value. This increased to 75% for a 25-mm liver biopsy specimen without any substantial benefit for longer biopsy specimens. Sampling variability of fibrosis is a significant limitation in the assessment of fibrosis with liver biopsy. In conclusion, this study suggests that a length of at least 25 mm is necessary to evaluate fibrosis accurately with a semiquantitative score. Sampling variability becomes a major limitation when using more accurate methods such as automated image analysis.

Cirrhosis is considered to be the precursor of most hepatocellular carcinomas. To gain insight into the early molecular mechanisms of liver carcinogenesis, this study compared, using real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), the expression levels of 31 selected genes in normal livers, cirrhotic nodules, and hepatocellular carcinomas. Since cirrhosis is composed of a mixture of polyclonal and monoclonal nodules, gene expression levels were also compared according to the clonal status of the cirrhotic nodules. The expression of eight of the 31 genes studied was significantly increased (NEGF2, ANGPT1, ARF, KRT19, SFN, CLDN4, MMP7, and ETV4) in cirrhotic nodules compared with normal liver, while only one was decreased (LYVE1). The same trend of variation was observed in cirrhosis and hepatocellular carcinomas for all of these genes except KRT19. When gene expression variation was compared according to the clonal status of cirrhotic nodules, only the LYVE1 expression level was significantly different. The LYVE1 gene expression level decreased progressively from polyclonal cirrhotic nodules to monoclonal cirrhotic nodules (polyclonal nodules 0.39 +/- 0.25; monoclonal nodules 0.20 +/- 0.14; p < 0.05) and to hepatocellular carcinoma (0.07 +/- 0.1). In conclusion, this study highlights the fact that among genes strongly dysregulated in hepatocellular carcinoma, some are already abnormally expressed in cirrhosis. The decrease in the expression level of one of these genes, LYVE1, was associated with monoclonality in cirrhotic nodules., (Copyright 2003 John Wiley & Sons, Ltd.)

Liver fibrosis is the hallmark of every chronic liver disease. It is also the major factor of morbidity and mortality due to the development of cirrhosis and its complications including hepatocellular carcinoma. But even at the beginning of the process of liver fibrosis and due to the strategic position of the extracellular matrix at the interface between blood flow and epithelial compartment, any quantitative or qualitative modification of extracellular matrix will rapidly affect structure and function of the liver. The development of several animal models of liver fibrosis as well as isolation and cultivation of hepatic stellate cells, the major fibrogenic cell type in the liver, led to the gathering of recent knowledge on the mechanism of liver fibrosis. Activation of hepatic stellate cells is a key event in this process and many details on this finely tuned mechanism are now available. In addition to these experimental data, experience from chronic hepatitis C now allows the development of new concepts and perspectives such as liver fibrosis regression and antifibrotic therapies., (Copyright 2003 John Wiley & Sons, Ltd.)

MANAGING THE RESPONSIBLE AGENT: Hepatic fibrosis with its end-point, cirrhosis, are the principle complications responsible for morbidity and mortality in chronic liver diseases. It is therefore important to address the question of whether these lesions can disappear, once installed in the liver. Regression can only occur when the agent responsible for the fibrosis (virus, alcohol, poison, iron, autoantibodies, etc) is eradicated or controlled. THE FORMS OF REGRESSION: Once the agent controlled, regression of fibrosis can either be spontaneous, a rare situation, although some bona fide cases of fibrosis or even cirrhosis reversion have been reported in the literature, or assisted by specific therapy. It is therefore necessary to take into consideration the development of new treatments based on enhanced knowledge of the mechanisms of fibrosis. THE ACTIVITY AND EFFICACY OF TREATMENTS: These treatments target one of the three following mechanisms: the blockade of hepatic stellate cell activation, enzymatic digestion of fibrous tissue and stimulation of liver cell regeneration. Although these treatments have shown efficacy on experimental models of fibrosis, to date, there are no published results formally confirming the efficacy and safety of these treatments in man.

Dystroglycan is a membrane component of the dystrophin-glycoprotein transmembrane complex. Its expression is required for the spatial organization of laminin on the cell surface and for basement membrane assembly. In view of the constitution of a perisinusoidal basement membrane during liver fibrosis, we studied dystroglycan expression in liver fibrosis. Dystroglycan mRNA and protein expression were investigated by immunofluorescence, Western blot, and quantitative RT-PCR (TaqMan) in normal human and rat liver and in isolated rat hepatic stellate cells. On Western blot, a 43-kd band corresponding to beta-dystroglycan was observed in protein extracted from normal and fibrotic human and rat livers. The specific 43-kd protein was also detected in lysates from rat hepatic stellate cells but not from hepatocytes. By immunofluorescence, patchy deposits of beta-dystroglycan were detected on membrane of hepatic stellate cells in culture. On Western blot and quantitative RT-PCR, an up-regulation of beta-dystroglycan was shown during spontaneous activation of hepatic stellate cells in culture. Direct evidence for the role of dystroglycan in laminin-hepatic stellate cell interaction was shown because specific antibody directed against alpha-dystroglycan inhibited partially hepatic stellate cell adhesion on laminin-coated plates. This mechanism was calcium dependent because EDTA inhibited cell/laminin adhesion, an effect reversed by addition of Ca(2+). This study shows that dystroglycan is expressed on the membrane of hepatic stellate cells and is up-regulated during liver fibrosis. Dystroglycan interaction with laminin should be implicated in the concentration of pericellular laminin and in the constitution of a perisinusoidal basement membrane during liver fibrosis.

Nonalcoholic steatohepatitis (NASH) may progress to liver fibrosis and cirrhosis. Mechanisms directly involved in the development of fibrosis have been poorly investigated. Because connective tissue growth factor (CTGF) is an intermediate key molecule involved in the pathogenesis of fibrosing chronic liver diseases and is potentially induced by hyperglycemia, the aims of this study were to (1) study the expression of CTGF in vivo both in human liver biopsy specimens of patients with NASH and in an experimental model of obesity and type II diabetes (Zucker rats); and (2) analyze the effects of hyperglycemia and insulin in vitro on hepatic stellate cells. In vivo, CTGF overexpression was observed in the liver tissue of all of the 16 patients with NASH. CTGF immunostaining was mild in 7 cases (44%) and moderate or strong in 9 cases (56%). Staining was mainly detected in the liver extracellular matrix in parallel with the amount of liver fibrosis. Liver from fa/fa rats also showed CTGF overexpression by comparison with Fa/fa rats both at the messenger RNA (mRNA) level (3-fold increase) and protein level. In vitro, both CTGF mRNA and protein were significantly increased when hepatic stellate cells were incubated with either glucose or insulin. A slight increase in type I procollagen mRNA level was also observed in hepatic stellate cells incubated with glucose. In conclusion, this study suggests that hyperglycemia and insulin are key-factors in the progression of fibrosis in patients with NASH through the up-regulation of CTGF.

Most hepatocellular carcinomas (HCC) arise from malignant transformation of regenerative cirrhotic nodules. Because HCC has a very poor prognosis, detection of these premalignant lesions may improve the management of patients with cirrhosis. In this regard, clonal analysis of liver micronodules should be of particular interest in order to differentiate polyclonal regenerative micronodules from monoclonal neoplastic potentially malignant micronodules. To address this issue, 112 micronodules from 15 cases of explanted liver cirrhosis were carefully microdissected from paraffin-embedded tissue using a laser capture microscopy system. Clonal analysis was performed by analyzing X-chromosome inactivation, as indicated by the methylation status of the human androgen receptor gene (HUMARA). For each microdissected micronodule, a large set of pathological features was evaluated and correlated with their clonal status. Clonal analysis showed that 57 micronodules (51%) were monoclonal and 55 (49%) were polyclonal. Prevalence of monoclonal nodules ranged from 25% to 71% according to cases. In all cases, mono- and polyclonal nodules were randomly distributed in the cirrhotic liver. Although the clonal status was not significantly affected by the presence or absence of macronodules in the adjacent liver, size of monoclonal micronodules was significantly larger than size of polyclonal micronodules (mean size of the monoclonal nodules: 3 + 0.1 mm vs mean size of the polyclonal nodules: 2.5 +/- 0.1 mm, p = 0.007). Among the elementary pathological features evaluated, only the presence of iron overload was correlated with a monoclonal status (p = 0.04). In conclusion, clonal analysis of liver cirrhosis shows that 51% of micronodules are monoclonal lesions, supporting the notion that liver cirrhosis is a multineoplastic lesion. Because monoclonality is a marker of neoplasia, cirrhosis with accumulation of monoclonal nodules may be carefully followed, and monoclonal nodules should be screened for additional markers to assess their biological behavior.

Connective tissue growth factor (CTGF) stimulates in vitro fibroblast proliferation and extracellular matrix synthesis. The aim of this study was to assess the role of CTGF in liver fibrogenesis. CTGF expression was investigated both at the protein and mRNA level in biopsies of chronic liver diseases, in experimental models of liver fibrosis, and in hepatic stellate cells in culture. CTGF immunostaining was observed in most human liver biopsies with significant fibrosis. An increase of CTGF immunostaining was associated with a higher score of fibrosis both in the group of chronic hepatitis C (chi(2) = 9.3; P <.01) and in the non-hepatitis C group (chi(2) = 7.2; P <.02). In situ hybridization showed CTGF mRNA expression in spindle cells in both the fibrous septa and sinusoidal lining. In experimental models of liver fibrosis, CTGF accumulated in parallel with the development of septal fibrosis and cirrhosis. Quantification of CTGF mRNA by a real-time reverse-transcription polymerase chain reaction (RT-PCR) assay showed a significant increase of CTGF mRNA in both CCl(4)-induced and bile duct-ligated rat models of liver fibrosis. Expression of CTGF protein and mRNA was definitively assigned to hepatic stellate cells, because CTGF was detected by Western blot both in lysate and supernatant of a hepatic stellate cell line derived from rats. These cells also displayed CTGF protein and mRNA as shown by immunohistochemistry and in situ hybridization. In conclusion, this study shows that CTGF is strongly expressed during liver fibrogenesis, and hepatic stellate cells seem to be the major cellular sources of CTGF in the liver.

Several arguments suggest that most hepatocellular carcinomas (HCCs) occurring in human cirrhotic livers arise from large hepatocellular nodules or macronodules. Except for nodules with obvious features of HCC, there exist no consistent criteria enabling the differentiation between benign regenerative and neoplastic, potentially malignant macronodules. Surrogate markers able to accurately discriminate those lesions that will evolve toward a HCC are required. In this study, we investigated the clonality of 26 macronodules isolated from eight cases of explanted cirrhotic livers in women by analyzing X-chromosome inactivation, as indicated by the methylation status of the human androgen receptor gene (HUMARA). For each macronodule, a large set of pathological features was evaluated and used to classify the macronodules into four groups: entirely benign-looking nodule (type 1), low-grade dysplastic nodule (type 2), high-grade dysplastic nodule (type 3), and HCC (type 4). Clonal analysis showed that 14 macronodules (54%) were monoclonal and 12 (46%) were polyclonal. Monoclonality was detected in 5 of 11 (45%) nodules from groups of entirely benign-looking and low-grade dysplastic nodules (types 1 and 2) and in 9 of 15 (60%) nodules from the group of high-grade dysplastic nodule and HCC (types 3 and 4). Neither the etiology of cirrhosis nor the size or histological classification of macronodules was correlated with the clonal status. In conclusion, clonal analysis of macronodules enables the differentiation between mono- and polyclonal macronodules in cirrhosis. Because monoclonal macronodules are prone to evolve to HCC, the determination of the clonal status of a macronodule could provide additional information for evaluating the prognosis of these lesions.

Aims: To assess the occurrence of lipid peroxidation in chronic hepatitis C and to evaluate its relation to pathological features and liver iron concentrations., Methods: Liver biopsy samples of 43 patients with untreated chronic hepatitis C were studied by immunohistochemistry using specific antibodies directed against two major aldehyde metabolites of lipid peroxidation, malondialdehyde (MDA), and 4-hydroxynonenal (HNE)., Results: MDA and HNE adducts (aldehydes covalently linked to another molecule) were detected in the liver samples in 77% and 30% of cases, respectively. MDA adducts were detected both in the extracellular matrix and sinusoidal cells localised in areas of periportal and lobular necrosis. HNE adducts appeared in the cytoplasm of only a few hepatocytes. Comparison of the semiquantitative assessment of adducts (MDA and HNE indexes) with the grading and the staging of chronic hepatitis showed that the MDA index was correlated with fibrosis score (p < 0.001) and the grade of activity (p < 0.01). There was also a tendency to correlation with liver iron concentration (p = 0.09). No correlation was observed between the HNE index and pathological features or liver iron concentration., Conclusion: Lipid peroxidation products are detectable in the liver of chronic hepatitis C patients. The presence of MDA adducts in areas of active fibrogenesis and the correlation between the MDA index and fibrosis score suggest a role for lipid peroxidation in liver fibrosis.

Aims: To assess which pathological features are associated with a sensitive marker of liver fibrogenesis and thus of the potential development of fibrosis in hepatitis C., Methods: The degree of liver fibrogenesis was evaluated by quantification of type I collagen mRNA and transforming growth factor (TGF) beta 1 mRNA (a major profibrogenic cytokine) in liver biopsy specimens from 28 patients with chronic hepatitis C and five controls, using a quantitative reverse transcription polymerase chain reaction (RT-PCR) assay. Results of mRNA quantification were correlated with histological lesions scored semiquantitatively in the same specimens., Results: Type I collagen mRNA was more strongly expressed in patients than in controls and correlated with the degree of fibrosis, but not with any of the necro-inflammatory lesions (portal inflammation, piecemeal necrosis, and lobular necrosis). TGF beta 1 mRNA concentration was higher in patients than in controls and correlated with histological grade of activity and lobular necrosis. This result was confirmed by in situ hybridisation experiments which showed that TGF beta 1 mRNA was mainly expressed in areas of focal lobular necrosis in chronic hepatitis C., Conclusion: This study shows that fibrosis, rather than necro-inflammatory lesions or activity scores, is associated with fibrogenesis and thus with potential aggravation of the fibrous deposit in chronic hepatitis C. Lobular necrosis is an important predictor of prognosis in chronic hepatitis C, as shown by its strong association with TGF beta 1 mRNA expression.