Hiroyasu Morikawa

Endothelial to Mesenchymal Transition via Transforming Growth Factor-β1/Smad Activation Is Associated with Portal Venous Stenosis in Idiopathic Portal Hypertension

Idiopathic portal hypertension (IPH) represents noncirrhotic portal hypertension of unknown etiology, mainly due to stenosis of peripheral portal veins. This study was performed to clarify the mechanism of portal venous stenosis in IPH from the viewpoint of the contribution of the endothelial to mesenchymal transition of the portal vein endothelium via transforming growth factor-beta1 (TGF-beta1)/Smad activation. In vitro experiments using human dermal microvascular endothelial cells demonstrated that TGF-beta1 induced myofibroblastic features in human dermal microvascular endothelial cells, including spindle cell morphology, reduction of CD34 expression, and induction of S100A4, alpha-smooth muscle actin, and COL1A1 expression, as well as the increased nuclear expression of phospho-Smad2. Bone morphogenic protein-7 preserved the endothelial phenotype of human dermal microvascular endothelial cells. Immunohistochemical analysis showed that endothelial cells of the peripheral portal veins in IPH were characterized by the decreased expression of CD34 and the enhanced nuclear expression of phospho-Smad2; these results also confirmed the expression of S100A4 and COL1A1 in the portal vein endothelium. Serum TGF-beta1 levels in patients with IPH were significantly higher than those of healthy volunteers and patients with chronic viral hepatitis/liver cirrhosis, while an elevation of serum bone morphogenic protein-7 levels was not observed. These results suggest that the endothelial to mesenchymal transition of the portal venous endothelium via TGF-beta1/Smad activation is associated with portal venous stenosis in IPH, and bone morphogenic protein-7 may therefore be a suitable therapeutic candidate for IPH.

Usefulness of transient elastography for assessment of liver fibrosis in chronic hepatitis B: Regression of liver stiffness during entecavir therapy.

Aim:  The usefulness of transient elastography remains to be validated in chronic hepatitis B, particularly as a tool for monitoring the degree of liver fibrosis during treatment.

Physical inactivity and insufficient dietary intake are associated with the frequency of sarcopenia in patients with compensated viral liver cirrhosis.

The association between sarcopenia and nutritional status is thought to be an important problem in patients with cirrhosis. In this study, we investigated whether nutritional factors were related to sarcopenia in patients with liver cirrhosis.

Add-on combination therapy with adefovir dipivoxil induces renal impairment in patients with lamivudine-refractory hepatitis B virus.

Summary.  Combination therapy with adefovir dipivoxil (ADV) and lamivudine (LAM) is recommended for patients infected with LAM‐refractory hepatitis B virus (HBV). However, the effects of such therapy on renal function and serum phosphorus levels have not been fully evaluated. Combination therapy with ADV and LAM was given to 37 patients infected with LAM‐refractory HBV, including 17 with hepatic cirrhosis. Serum HBV DNA levels decreased to below 2.6 log10 copies/mL in 23 (62%) of 37 patients at 12 months, 25 (78%) of 32 patients at 24 months, and 16 (84%) of 19 patients at 36 months. Except for one cirrhotic patient, serum alanine aminotransferase levels were below 50 IU/L in all patients during combination therapy. Serum creatinine levels increased in 14 (38%) of 37 patients, and serum phosphate levels decreased to below 2.5 mg/mL in 6 (16%) of 37 patients during combination therapy. Patients who received combination therapy for 36 months or longer had a significantly incidence of elevated serum creatinine levels. Fanconi syndrome occurred in a 57‐year‐old woman with cirrhosis after ADV was added to LAM. Combination therapy with ADV and LAM can maintain biochemical remission in patients with LAM‐refractory HBV. However, the dosing interval of ADV should be adjusted according to renal function and serum phosphate levels in patients receiving long‐term treatment.

Assessment of Liver Fibrosis with Real-Time Tissue Elastography in Chronic Viral Hepatitis

Objective: The aim of this study was to assess prospectively the accuracy of measurement of liver fibrosis with real-time tissue elastography (RTE) in patients with chronic viral hepatitis. Methods: Two hundred and forty-five patients were prospectively enrolled. Nine image features were measured from strain images, and Liver Fibrosis Index (LFI) was calculated from these features. Fibrosis stage was diagnosed from pathological specimens obtained by ultrasound-guided biopsy. LFI and serological markers were compared with pathological diagnosis, and the diagnostic performance of RTE was compared. Results: LFI in stages F0-F1, F2, F3 and F4 was 1.58, 2.03, 2.40 and 2.86, respectively, demonstrating a stepwise increase with increasing severity of liver fibrosis (p < 0.001). LFI in F2 did not significantly differ from that in F3, whereas for all other combinations of stages, there were significant differences. The area under the receiver operating characteristic curve of the LFI, platelet count, aspartate/alanine aminotransferase ratio, aspartate aminotransferase-to-platelet ratio, and FibroIndex for predicting F3 stage or higher (F0-F2 vs. F3-F4) was 0.865, 0.824, 0.708, 0.789 and 0.828, respectively. Conclusions: RTE is useful for diagnosis of liver fibrosis, regardless of stage, in patients with chronic viral hepatitis.

JSUM ultrasound elastography practice guidelines: liver

In diffuse liver disease, it is extremely important to make an accurate diagnosis of liver fibrosis prior to determining indications for therapy or predicting treatment outcome and malignant potential. Although liver biopsy has long been the gold standard in the diagnosis of liver fibrosis, it is still an invasive method. In addition, the sampling error is an intrinsic problem of liver biopsy. Non-invasive serological methods for the diagnosis of liver fibrosis can be affected by factors unrelated to the liver. Recently, after the introduction of FibroScan, it became possible to measure liver fibrosis directly and non-invasively by elastography, which has attracted attention as a non-invasive imaging diagnostic tool for liver fibrosis. In addition, real-time tissue elastography is currently being used to conduct clinical trials at many institutions. Moreover, virtual touch quantification enables the observation of liver stiffness at any location by simply observing B-mode images. Furthermore, the recently developed ShearWave elastography visualizes liver stiffness on a color map. Elastography is thought to be useful for all types of diffuse liver diseases. Because of its association with portal hypertension and liver carcinogenesis, elastography is expected to function as a novel prognostic tool for liver disease. Although various elastographic devices have been developed by multiple companies, each device has its own measurement principle, method, and outcome, creating confusion in clinical settings. Therefore, it is extremely important to understand the characteristics of each device in advance. The objective of this guideline, which describes the characteristics of each device based on the latest knowledge, is for all users to be able to make the correct diagnosis of hepatic fibrosis by ultrasound elastography.

Noninvasive laboratory tests proposed for predicting cirrhosis in patients with chronic hepatitis C are also useful in patients with non-alcoholic steatohepatitis

BackgroundSeveral noninvasive tests have been proposed to predict cirrhosis in patients with chronic hepatitis C, but not in patients with non-alcoholic steatohepatitis (NASH). We assessed whether noninvasive laboratory tests designed to predict the risk of cirrhosis in patients with chronic hepatitis C virus (HCV) infection could be used in patients with NASH.MethodsThe subjects were 50 patients with biopsy-proved NASH and 100 age- and sex-matched patients with HCV. Aspartate aminotransferase (AST)/alanine aminotransferase (ALT) ratio (AAR), age-platelet (AP) index, AST-to-platelet ratio index (APRI), cirrhosis discriminant score (CDS), and the hepatitis C antiviral long-term treatment against cirrhosis (HALT-C) model were calculated.ResultsThe areas under the receiver-operating characteristic curves of the AAR, AP index, APRI, CDS, and HALT-C model for predicting cirrhosis were respectively 0.813, 0.877, 0.786, 0.949, and 0.908 in patients with NASH and 0.555, 0.652, 0.761, 0.782, and 0.782 in patients with HCV. A CDS cutoff value of less than 5 misclassified none of the 9 patients with NASH who had cirrhosis, while a value of more than 8 misclassified none of the 41 patients with NASH without cirrhosis. With the HALT-C model, a cutoff value of less than 0.6 classified non-cirrhotic NASH, while a cutoff value of 0.97 or higher classified cirrhotic NASH. The use of CDS and HALT-C model could avoid liver biopsy for predicting cirrhosis in 60 and 48% of the patients with NASH, respectively.ConclusionsNoninvasive laboratory tests designed to predict cirrhosis in patients with HCV are also useful in patients with NASH.

A Randomized Pilot Trial of Oral Branched-Chain Amino Acids in Early Cirrhosis: Validation Using Prognostic Markers for Pre-Liver Transplant Status

Because of the chronic shortage of liver donors, hepatologists are required to prolong the liver transplant waiting period by preserving the hepatic reserve of scheduled recipients. This study examined the effectiveness of oral branched‐chain amino acids (BCAAs), using outcome markers indicating pretransplant hepatic reserve. Fifty‐six consecutive eligible patients with Child class A cirrhosis without major complications were randomly assigned to receive oral BCAA granules (12.45 g/day) for least 1 year or no BCAAs. Differences between groups in the Model for End‐Stage Liver Disease (MELD) score, Child‐Turcotte‐Pugh (CTP) score, asialoscintigraphic clearance index (CI), and complications were examined. Of 50 remaining patients, 27 received BCAAs, and 23 received no BCAAs (mean duration, 3.2 years). The mean annual changes in the MELD score, CTP score, and asialoscintigraphic CI were smaller in the BCAA group than in the control group (−0.06 ± 0.23 versus 0.10 ± 0.40, P = 0.024, 0.06 ± 0.30 versus 0.30 ± 0.48, P = 0.037, and 0.00 ± 0.02 versus 0.02 ± 0.04, P = 0.040, respectively). The mean annual changes in the serum total bilirubin and the serum albumin in the BCAA group were better preserved than those in the control group (−0.07 ± 0.20 versus 0.12 ± 0.18 mg/dL, P < 0.001, and 0.07 ± 0.13 versus −0.02 ± 0.19 g/dL, P = 0.005, respectively); other laboratory variables were not significant. The incidence of overall major cirrhotic complications was lower in the BCAA group than in the control group [14.8% (4 of 27 patients) versus 30.4% (7 of 23 patients) at 3 years, P = 0.043]; only ascites was significant individually. In conclusion, early interventional oral BCAAs might prolong the liver transplant waiting period by preserving hepatic reserve in cirrhosis. Liver Transpl 15:790–797, 2009.

Cytoglobin is expressed in hepatic stellate cells, but not in myofibroblasts, in normal and fibrotic human liver.

Cytoglobin (CYGB) is ubiquitously expressed in the cytoplasm of fibroblastic cells in many organs, including hepatic stellate cells. As yet, there is no specific marker with which to distinguish stellate cells from myofibroblasts in the human liver. To investigate whether CYGB can be utilized to distinguish hepatic stellate cells from myofibroblasts in normal and fibrotic human liver, human liver tissues damaged by infection with hepatitis C virus (HCV) and at different stages of fibrosis were obtained by liver biopsy. Immunohistochemistry was performed on histological sections of liver tissues using antibodies against CYGB, cellular retinol-binding protein-1 (CRBP-1), α-smooth muscle actin (α-SMA), thymocyte differentiation antigen 1 (Thy-1), and fibulin-2 (FBLN2). CYGB- and CRBP-1-positive cells were counted around fibrotic portal tracts in histological sections of the samples. The expression of several of the proteins listed above was examined in cultured mouse stellate cells. Quiescent stellate cells, but not portal myofibroblasts, expressed both CYGB and CRBP-1 in normal livers. In fibrotic and cirrhotic livers, stellate cells expressed both CYGB and α-SMA, whereas myofibroblasts around the portal vein expressed α-SMA, Thy-1, and FBLN2, but not CYGB. Development of the fibrotic stage was positively correlated with increases in Sirius red-stained, α-SMA-positive, and Thy-1-positive areas, whereas the number of CYGB- and CRBP-1-positive cells decreased with fibrosis development. Primary cultured mouse stellate cells expressed cytoplasmic CYGB at day 1, whereas they began to express α-SMA at the cellular margins at day 4. Thy-1 was undetectable throughout the culture period. In human liver tissues, quiescent stellate cells are CYGB positive. When activated, they also become α-SMA positive; however, they are negative for Thy-1 and FBLN2. Thus, CYGB is a useful marker with which to distinguish stellate cells from portal myofibroblasts in the damaged human liver.

Frequent detection of hepatitis B virus DNA in hepatocellular carcinoma of patients with sustained virologic response for hepatitis C virus.

Hepatocellular carcinoma (HCC) develops several years after the eradication of hepatitis C virus (HCV) by interferon therapy. Risk factors for the development of HCC are only partly understood. To elucidate the role of occult hepatitis B virus (HBV) infection in hepatocarcinogenesis in patients with sustained virologic response, the prevalences of HBV‐related makers were examined. Study group comprised 16 patients with sustained virologic response (group A) and 50 with HCV (group B). Anti‐HBc and anti‐HBs in serum were examined by enzyme‐linked immunoassay. HBV DNA in liver was examined by nested polymerase chain reaction, using primers specific for genes encoding for HBx, HBsAg, HBcAg, and HBV cccDNA. Sequence of the amplified HBV DNA for ‘a’ determinant of HBsAg was determined in HCC. Anti‐HBc was positive in 10 of 16 in group A and 25 of 50 in group B. HBV DNA in liver was detected in 12 of 16 in group A and 21 of 50 in group B (P = 0.044). In group A, HBV DNA in liver was detected frequently in patients without cirrhosis and in those with a longer period from the time of HCV eradication to the development of HCC. Mutation in ‘a’ determinant of HBsAg was found in three HCC of group A. Occult HBV infection may be one of the most important risk factors in hepatocarcinogenesis of Japanese patients with sustained virologic response. J. Med. Virol. 81:1009–1014, 2009.