Kupffer phase image of Sonazoid-enhanced US is useful in predicting a hypervascularization of non-hypervascular hypointense hepatic lesions detected on Gd-EOB-DTPA-enhanced MRI: a multicenter retrospective study.

BACKGROUND: It remains unknown whether Kupffer-phase images in Sonazoid-enhanced ultrasonography (US) can be used to predict hypervascularization of borderline lesions. Therefore, we aimed to clarify whether Kupffer-phase images in Sonazoid-enhanced ultrasonography can predict subsequent hypervascularization in hypovascular borderline lesions detected on hepatobiliary-phase gadolinium-ethoxybenzyl-diethylenetriamine pentaacetic acid (Gd-EOB-DTPA)-enhanced magnetic resonance imaging. METHODS: From January 2008 to March 2012, 616 low-intensity hypovascular nodules were detected in hepatobiliary-phase images of Gd-EOB-DTPA-enhanced MRI at nine institutions. Among these, 167 nodules, which were confirmed as hypovascular by Gd-EOB-DTPA-enhanced MRI and Sonazoid-enhanced US, were evaluated in this study. Potential hypervascularization factors were selected based on their clinical significance and the results of previous reports. The Kaplan-Meier model and log-rank test were used for univariate analysis and the Cox regression model was used for multivariate analysis. RESULTS: The cumulative incidence of hypervascularization of borderline lesions was 18, 37, and 43 % at 1, 2, and 3 years, respectively. Univariate analyses showed that tumor size (p = 0.0012) and hypoperfusion on Kupffer-phase images in Sonazoid-enhanced US (p = 0.004) were associated with hypervascularization of the tumor. Multivariate analysis showed that tumor size [HR: 1.086, 95 % confidence interval = 1.027-1.148, p = 0.004] and hypo perfusion on Kupffer-phase images [HR: 3.684, 95 % confidence interval = 1.798-7.546, p = 0.0004] were significantly different. CONCLUSIONS: Kupffer-phase images in Sonazoid-enhanced US and tumor diameter can predict hypervascularization of hypointense borderline lesions detected on hepatobiliary-phase Gd-EOB-DTPA-enhanced MRI.

The impact of PNPLA3 and JAZF1 on hepatocellular carcinoma in non-viral hepatitis patients with type 2 diabetes mellitus.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is an established independent risk factor for hepatocellular carcinoma (HCC). T2DM is associated with non-alcoholic steatohepatitis (NASH), which is a major cause of non-HBV and non-HCV-related HCC; nevertheless, it has been difficult to identify those patients with T2DM who have a high risk of developing HCC. The aim of this study was to identify genetic determinants that predispose T2DM patients to HCC by genotyping T2DM susceptibility loci and PNPLA3. METHODS: We recruited 389 patients with T2DM who satisfied the following three criteria: negative for HBs-Ag and anti-HCV Ab, alcohol intake <60 g/day, and history of T2DM >10 years. These patients were divided into two groups: T2DM patients with HCC (DM-HCC, n = 59) or those without HCC (DM-non-HCC, n = 330). We genotyped 51 single-nucleotide polymorphisms (SNPs) previously reported as T2DM or NASH susceptibility loci (PNPLA3) compared between the DM-HCC and DM-non-HCC groups with regard to allele frequencies at each SNP. RESULTS: The SNP rs738409 located in PNPLA3 was the greatest risk factor associated with HCC. The frequency of the PNPLA3 G allele was significantly higher among DM-HCC individuals than DM-non-HCC individuals (OR 2.53, p = 1.05 × 10(-5)). Among individuals homozygous for the PNPLA3 G allele (n = 115), the frequency of the JAZF1 rs864745 G allele was significantly higher among DM-HCC individuals than DM-non-HCC individuals (OR 3.44, p = 0.0002). CONCLUSIONS: PNPLA3 and JAZF1 were associated with non-HBV and non-HCV-related HCC development among Japanese patients with T2DM.

Appropriate use of virtual touch quantification and FibroScan M and XL probes according to the skin capsular distance.

BACKGROUND: Appropriate utilization of different diagnostic modalities is essential for the accurate liver stiffness measurements (LSM) in patients with chronic liver diseases. The aim of this study was to evaluate the efficacy of Virtual Touch Quantification (VTQ) and the FibroScan M and XL probes in term of accurate LSM and to identify factors associated with inadequate measurements in obese and non-obese Japanese patients. METHODS: A total of 664 consecutive patients with chronic liver disease were prospectively enrolled. LSM were evaluated concurrently with VTQ and the FibroScan M and XL probes. LSM quality was categorized as inadequate (success rate <60% and/or interquartile range/median value of ≥30%) or adequate. RESULTS: No significant differences in the rate of inadequate LSM were observed among the three diagnostic modalities. In multivariate analysis, skin capsule distance (SCD) was strongly associated with inadequate rates obtained with VTQ and the M probe [odds ratio (OR) 1.28, P < 0.0001 and OR 1.20, P < 0.0001, respectively]. Inadequate LSM rates with both VTQ and the M probe increased with longer SCD, with a significant difference between subgroups at an SCD of ≥22.5 mm (VTQ 54.0%; M probe 51.1%; XL probe 25.2%; P < 0.0001). The rates of inadequate LSM rates with VTQ were significantly lower than those with the XL probe at an SCD of <17.5 mm. A total of 15 liver biopsy specimens obtained from nonalcoholic fatty liver disease patients confirmed the diagnostic accuracy and high applicability of the XL probe. CONCLUSIONS: Long SCD reduced the diagnostic performance of the FibroScan® M probe and VTQ. LSM modalities should be selected according to SCD.

Branched-chain amino acids reduce hepatic iron accumulation and oxidative stress in hepatitis C virus polyprotein-expressing mice.

BACKGROUND & AIMS: Branched-chain amino acids (BCAA) reduce the incidence of hepatocellular carcinoma (HCC) in patients with cirrhosis. However, the mechanisms that underlie these effects remain unknown. Previously, we reported that oxidative stress in male transgenic mice that expressed hepatitis C virus polyprotein (HCVTgM) caused hepatic iron accumulation by reducing hepcidin transcription, thereby leading to HCC development. This study investigated whether long-term treatment with BCAA reduced hepatic iron accumulation and oxidative stress in iron-overloaded HCVTgM and in patients with HCV-related advanced fibrosis. METHODS: Male HCVTgM were fed an excess-iron diet that comprised either casein or 3.0% BCAA, or a control diet, for 6 months. RESULTS: For HCVTgM, BCAA supplementation increased the serum hepcidin-25 levels and antioxidant status [ratio of biological antioxidant potential (BAP) relative to derivatives of reactive oxygen metabolites (dROM)], decreased the hepatic iron contents, attenuated reactive oxygen species generation, and restored mitochondrial superoxide dismutase expression and mitochondrial complex I activity in the liver compared with mice fed the control diet. After 48 weeks of BCAA supplementation in patients with HCV-related advanced fibrosis, BAP/dROM and serum hepcidin-25 increased and serum ferritin decreased compared with the pretreatment levels. CONCLUSIONS: BCAA supplementation reduced oxidative stress by restoring mitochondrial function and improved iron metabolism by increasing hepcidin-25 in both iron-overloaded HCVTgM and patients with HCV-related advanced fibrosis. These activities of BCAA may partially account for their inhibitory effects on HCC development in cirrhosis patients.

Hypovascular hepatic nodules showing hypointense on the hepatobiliary-phase image of Gd-EOB-DTPA-enhanced MRI to develop a hypervascular hepatocellular carcinoma: a nationwide retrospective study on their natural course and risk factors.

OBJECTIVE: We aimed to investigate the natural outcome of nonhypervascular lesions detected in the hepatobiliary phase of gadolinium ethoxybenzyl diethylenetriamine pentaacetic acid (Gd-EOB-DTPA)-enhanced MRI by performing a longitudinal study retrospectively enrolled in a nationwide manner. METHODS: Between February 2008 and March 2011, 224 patients with 504 nodules that were diagnosed as nonhypervascular by imaging were recruited from institutions that participated in the present study. We examined the natural outcome of nonhypervascular lesions and evaluated the risk factors. RESULTS: Of the 504 nodules, 173 (34.3%) showed hypervascular transformation. The overall cumulative incidence of hypervascular transformation was 14.9% at 12 months and 45.8% at 24 months. Multivariate analysis using the Cox regression model revealed previous treatment history for hepatocellular carcinoma (HCC; relative risk = 1.498; p = 0.036, 95% CI 1.03-2.19) and hyperintensity on T2-weighted images (relative risk = 1.724; p = 0.015, 95% CI 1.11-2.67) were identified as independent factors for hypervascular transformation. CONCLUSIONS: Patients who have a previous treatment history for HCC and with hypointense nodules showing hyperintensity on T2-weighted images need careful follow-up because of the high incidence of hypervascular transformation.

Clinical usefulness of non-protein respiratory quotient measurement in non-alcoholic fatty liver disease.

AIM: Little is known about the effects of non-alcoholic fatty liver disease (NAFLD) on energy metabolism, although this disease is associated with metabolic syndrome. We measured non-protein respiratory quotient (npRQ) using indirect calorimetry, which reflects glucose oxidation, and compared this value with histological disease severity in NAFLD patients. METHODS: Subjects were 32 patients who were diagnosed with NAFLD histopathologically. Subjects underwent body composition analysis and indirect calorimetry, and npRQ was calculated. An oral glucose tolerance test was performed, and plasma glucose area under the curve (AUC glucose) was calculated. RESULTS: There were no differences in body mass index, body fat percentage or visceral fat area among fibrosis stage groups. As fibrosis progressed, npRQ significantly decreased (stage 0, 0.895 ± 0.068; stage 1, 0.869 ± 0.067; stage 2, 0.808 ± 0.046; stage 3, 0.798 ± 0.026; P < 0.005). Glucose intolerance worsened and insulin resistance increased with fibrosis stage. npRQ was negatively correlated with AUC glucose (R = -0.6308, P < 0.001), Homeostasis Model of Assessment - Insulin Resistance (R = -0.5045, P < 0.005), fasting glucose (R = -0.4585, P < 0.01) and insulin levels (R = -0.4431, P < 0.05), suggesting that decreased npRQ may reflect impaired glucose tolerance due to insulin resistance, which was associated with fibrosis progression. Estimation of fibrosis stage using npRQ was as accurate as several previously established scoring systems using receiver-operator curve analysis. CONCLUSION: npRQ was significantly decreased in patients with advanced NAFLD. Our data suggest that measurement of npRQ is useful for the estimation of disease severity in NAFLD patients.

Focal nodular hyperplasia-like nodule with reduced expression of organic anion transporter 1B3 in alcoholic liver cirrhosis.

We report a patient with alcoholic liver cirrhosis who had a 15 mm focal nodular hyperplasia (FNH)-like nodule in the liver. This FNH-like nodule was diagnosed as hepatocellular carcinoma (HCC) mainly based on hypervascularity during the hepatic arterial phase, washout pattern during the equilibrium phase and low signal intensity during the hepatobiliary phase in gadolinium-ethoxybenzyl-diethylenetriamine pentaacetic acid (Gd-EOB-DTPA)-enhanced MRI; it was surgically resected. Its histology exhibited hepatocyte hyperplasia, fibrous septa containing unpaired small arteries accompanied by reactive bile ductules, remarkable iron deposits and sinusoidal capillarization, and was compatible with the diagnosis of an FNH-like nodule. When we analyzed the images of the present nodule retrospectively, low signal intensity on in-phase and isosignal intensity on opposed-phase T1-weighted MRI may have reflected iron deposits in the FNH-like nodule. In addition, a low signal intensity on T2-weighted MRI and no detection in diffusion-weighted MRI may help in distinguishing FNH-like nodules from HCC, since these image findings are inconsistent with typical HCC. Immunohistochemical analysis revealed a markedly reduced expression of organic anion transporter (OATP) 1B3 in this nodule, which implied decreased Gd-EOB-DTPA uptake by hepatocytes and accounted for the low signal intensity during the hepatobiliary phase on Gd-EOB-DTPA-enhanced MRI. To the best of our knowledge this is the first report in which an FNH-like nodule was assessed for OATP1B3 expression.

Type 1 interferon receptor in peripheral blood mononuclear cells may predict response to intra-arterial 5-fluorouracil + interferon therapy for advanced hepatocellular carcinoma.

BACKGROUND: Type 1 interferon alpha receptor 2 (IFNAR2) in the liver has been reported to be a predictive factor for the response to intra-arterial 5-fluorouracil (5-FU) + systemic interferon (IFN)-alpha combination therapy in patients with advanced hepatocellular carcinoma. We tested whether IFNAR2 expression in peripheral blood mononuclear cells could predict the response to 5-FU + IFN. METHODS: Predictive factors for survival and response to therapy were determined in 30 patients with advanced hepatocellular carcinoma who underwent treatment with 5-FU + IFN. IFNAR2 expression in peripheral blood mononuclear cells was measured in 11 of the 30 patients. RESULTS: With a mean number of 4.2 courses of combination therapy, one patient (3%) showed a complete response, eight (27%) showed partial responses, 13 (43%) had stable disease, and eight (27%) showed progressive disease. The median survival time of responders (complete response/partial response) was 12.7 months and that of nonresponders (stable disease/progressive disease) was 7.5 months. The one-year and two-year cumulative survival rates of responders and nonresponders were 87/69% and 40/11%, respectively (P = 0.019). Multivariate analysis identified response to therapy (P = 0.037) as the sole independent determinant of survival. The expression level of IFNAR2 in peripheral blood mononuclear cells was significantly (P = 0.012) higher in responders (6.5 ± 2.4) than in nonresponders (2.4 ± 0.6), even though no clinical factors were identified as being associated with the response to the combination therapy. CONCLUSION: IFNAR2 expression in peripheral blood mononuclear cells may predict the response to 5-FU + IFN therapy in patients with advanced hepatocellular carcinoma, although these data are preliminary.

Usefulness of Sonazoid contrast-enhanced ultrasonography for hepatocellular carcinoma: comparison with pathological diagnosis and superparamagnetic iron oxide magnetic resonance images.

OBJECTIVE: We investigated the usefulness of Sonazoid contrast-enhanced ultrasonography (Sonazoid-CEUS) in the diagnosis of hepatocellular carcinoma (HCC). The examination was performed by comparing the images during the Kupffer phase of Sonazoid-CEUS with superparamagnetic iron oxide magnetic resonance (SPIO-MRI). METHODS: The subjects were 48 HCC nodules which were histologically diagnosed (well-differentiated HCC, n=13; moderately differentiated HCC, n=30; poorly differentiated HCC, n=5). We performed Sonazoid-CEUS and SPIO-MRI on all subjects. In the Kupffer phase of Sonazoid-CEUS, the differences in the contrast agent uptake between the tumorous and non-tumorous areas were quantified as the Kupffer phase ratio and compared. In the SPIO-MRI, it was quantified as the SPIO-intensity index. We then compared these results with the histological differentiation of HCCs. RESULTS: The Kupffer phase ratio decreased as the HCCs became less differentiated (P<0.0001; Kruskal-Wallis test). The SPIO-intensity index also decreased as HCCs became less differentiated (P<0.0001). A positive correlation was found between the Kupffer phase ratio and the SPIO-MRI index (r=0.839). In the Kupffer phase of Sonazoid-CEUS, all of the moderately and poorly differentiated HCCs appeared hypoechoic and were detected as a perfusion defect, whereas the majority (9 of 13 cases, 69.2%) of the well-differentiated HCCs had an isoechoic pattern. The Kupffer phase images of Sonazoid-CEUS and SPIO-MRI matched perfectly (100%) in all of the moderately and poorly differentiated HCCs. CONCLUSION: Sonazoid-CEUS is useful for estimating histological grading of HCCs. It is a modality that could potentially replace SPIO-MRI.

Effects of a late evening snack combined with alpha-glucosidase inhibitor on liver cirrhosis.

AIM: A late evening snack (LES) is recommended for protein-energy malnutrition in patients with liver cirrhosis. However, many cases of liver cirrhosis have accompanying impaired glucose tolerance and there are concerns that LESs might aggravate glucose intolerance. In this study, we concomitantly used an alpha-glucosidase inhibitor with a LES and examined the effects on glucose tolerance. In addition, we examined whether or not there was an improvement in energy metabolism by slowing glucose absorption with the concomitant use of the alpha-glucosidase inhibitor. METHODS: The subjects were 11 patients with liver cirrhosis. From before the study, all the patients had been taking a LES supplementation with a branched-chain amino acid (BCAA)-enriched nutrient mixture. The patients were started on the concomitant use of alpha-glucosidase inhibitor (0.2 mg) taken just prior to the LES. The change of glucose tolerance and energy metabolism were examined using a 75-g oral glucose tolerance test and indirect calorimetry. RESULTS: One week and three months after the start of the concomitant use of the alpha-glucosidase inhibitor, the area under the concentration curve for plasma glucose was significantly decreased. Three months after the concomitant use, the non-protein respiratory quotient was significantly improved. There were no serious side effects during the follow-ups. CONCLUSION: The concomitant use of the alpha-glucosidase inhibitor use with LES showed the possibility of improving glucose tolerance and energy metabolism. In patients with impaired glucose tolerance, the concomitant use of an alpha-glucosidase inhibitor with LES might be a useful measure for nutritional management.

The effect of supplementation with branched-chain amino acids in patients with liver cirrhosis.

AIM: We investigated the effect of supplementation with branched-chain amino acids (BCAA) in patients with liver cirrhosis on the change of energy metabolism as well as glucose tolerance. METHODS: Thirty liver cirrhosis patients underwent nutrient supervision by a dietician for one week. They were then prescribed oral supplementation with three packs of a BCAA nutrient (Livact 4.15 g/pack; Ajinomoto Pharma, Tokyo, Japan), taken three times a day: after breakfast, dinner and before sleep. The change in energy metabolism and glucose tolerance was examined using an indirect calorimeter and 75 g oral glucose tolerance test (75 g OGTT). RESULTS: Non-protein respiratory quotient (npRQ) as well as branched-chain amino acid/tyrosine ratio (BTR) showed significant improvement, especially in patients with a creatinine height index (CHI) greater than 80. There was also a significant correlation between npRQ after one week of BCAA supplementation and the CHI. The patients with CHI greater than 80 and those with borderline pattern assessed by 75 g OGTT showed significant improvement in impaired glucose tolerance. CONCLUSION: Liver cirrhosis patients with CHI greater than 80 are the first candidates for BCAA supplementation. These patients showed improvement not only in energy metabolism and BTR, but also glucose tolerance.

[Insulin sensitizer--anti-diabetic drugs, metformin and pioglitazone that can improve insulin resistance].

Nonalcoholic steatohepatitis (NASH), which is considered the hepatic manifestation of the metabolic syndrome is an increasingly cause of chronic liver disease in Japan. NASH is finally lead to liver cirrhosis and hepatocellular carcinoma as viral hepatitis, therefore, medical treatment should be considered, when NASH occurs. Treatment of patients with metabolic syndrome has been focused on the management of associated conditions such as obesity, hyperlipidemia, hypertension and hyperinsulinemia. Insulin resistance, that could accelerate liver inflammation and fibrosis by up-regulation of TNFa seems to be most important factor in many cases of NASH. The insulin-sensitizing drugs, which were biguanides (metformin) and thiazolidinediones (pioglitazone) have been shown to correct not only insulin resistance but also steatosis and inflammation in the liver. Metformin and pioglitazone might be useful drugs against NASH, however further investigations were needed.

Cytokine-dependent anti-viral role of CD4-positive T cells in therapeutic vaccination against chronic hepatitis B viral infection.

There are several lines of evidence suggesting that specific vaccine therapy with a standard hepatitis B virus (HBV) vaccination reduces HBV replication. The aim of this study was to investigate the anti-viral mechanism of vaccine therapy in chronic hepatitis B patients. Nineteen patients were assigned to receive either vaccine therapy (n = 13) or no treatment as a control (n = 6). Vaccinated patients were analyzed for T cell proliferative responses specific for envelope antigen and cytokine production by antigen-specific T cells. ELISPOT and cytotoxicity assays also were carried out for limited blood samples. Serum HBV DNA levels decreased significantly at 3 months after completion of therapy and thereafter as compared to the baseline ones, and were significantly lower in vaccinated patients than in controls at 12 and 18 months after completion of therapy. Vaccination induced antigen-specific CD4+ T cell proliferative responses in four patients (30.8%). The production of high levels of interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) by antigen-specific T cells was found in six patients (46.0%) who showed significantly lower HBV DNA levels in serum at 6 (P = 0.04) and 18 months (P = 0.005) after completion of therapy than those without high levels of cytokine production. Vaccination did not induce antigen-specific CD8+ T cells or cytotoxic T cells. These results suggest that envelope-specific CD4+ T cells may control directly HBV replication by producing anti-viral cytokines rather than providing help for cytotoxic T cells in therapeutic vaccination against chronic HBV infection.