Skill Proficiency, Efficacy, and Safety of the Transradial Approach in Transarterial Treatments for Hepatocellular Carcinoma.

Introduction Abdominal angiography procedures such as transarterial chemoembolization (TACE) are essential for hepatocellular carcinoma treatment. One method commonly used is transfemoral access (TFA). However, issues associated with this method, which include postoperative compression of the puncture site and long periods of bed rest, can affect patient satisfaction. Thus, transradial access (TRA), a minimally invasive treatment method that improves treatment quality, was developed for TACE. This retrospective, multicenter study aimed to investigate the efficacy and safety of abdominal angiography using the radial artery approach. Methods In total, 1,601 patients underwent abdominal angiography using TRA and received treatment (radial access for visceral intervention (RAVI)) at 14 institutions in Japan. The treatment time, procedure completion rate, patient satisfaction, and complications were investigated. Results The success rate of RAVI was 99.4%, and the complication rate was 1.2%. Approximately 98.2% of the patients requested the radial artery approach again. There were no significant differences in the success rate of RAVI and the incidence of complications based on the operator's years of experience or the patient's age. Some patients developed minor complications such as puncture site bleeding, hematoma, vascular pain, and vasospasm. Further, serious complications (cerebral infarction (n = 1), cerebellar infarction (n = 1), and aortic dissection (n = 1)) were observed. Conclusion Similar to the conventional TFA, RAVI helped in facilitating peritoneal angiography safely. In abdominal angiography, this method can reduce patient burden and can be widely used in the future from the perspective of clinical benefit.

Quantitative measurements of M2BPGi depend on liver fibrosis and inflammation.

BACKGROUND: The relationship between liver fibrosis and inflammation and Mac-2-binding protein glycosylation isomer (M2BPGi) in patients with chronic liver disease (CLD) other than hepatitis C remains uncertain, owing to the limitations of qualitative methods. Here, we evaluated the influence of liver fibrosis and inflammation on quantitative M2BPGi (M2BPGi-Qt) in CLD, considering each etiology. METHODS: We recruited 1373 patients with CLD. To evaluate the influence of liver fibrosis and inflammation on M2BPGi-Qt levels, we assessed M2BPGi-Qt levels at each fibrosis and activity stage within different etiologies of CLD based on pathological findings. Subsequently, we evaluated if the accuracy of fibrosis staging based on M2BPGi-Qt could be improved by considering the influence of liver inflammation. RESULTS: In patients with viral hepatitis, non-alcoholic fatty liver disease, and primary biliary cholangitis, the median M2BPGi-Qt levels increased liver fibrosis progression. Median M2BPGi-Qt levels were not associated with the degree of fibrosis in patients with autoimmune hepatitis (AIH). Median M2BPGi-Qt levels increased with the progression of liver activity in all etiologies. A significant difference was found at each stage in AIH. Considering the liver inflammation, we established an algorithm, M2BPGi-Qt, to determine the alanine aminotransferase-to-platelet ratio (MAP-R) in liver cirrhosis (LC). The area under the receiver operating characteristic curve (AUC) of MAP-R was higher than that of the M2BPGi-Qt for detecting LC (AUC MAP-R = 0.759 and M2BPGi-Qt = 0.700, p < 0.001). CONCLUSIONS: The quantitative measurement system for M2BPGi depends on liver fibrosis and inflammation, regardless of etiology. Liver inflammation complicates the interpretation of M2BPGi-Qt results when assessing the fibrosis stage.

New chemiluminescent enzyme immunoassay for quantitative measurement of Mac-2 binding protein glycosylation isomer in chronic liver disease.

BACKGROUND: This study aimed to evaluate the quantitative measurement of Mac-2 binding protein glycosylation isomer (M2BPGi) levels using the new chemiluminescent enzyme immunoassay. METHODS: The data of a total of 347 patients with hepatitis C virus (HCV) infection and 150 health volunteers from 13 locations in Japan were evaluated. The quantitative system for measuring M2BPGi-Qt levels was based on a new chemiluminescent enzyme immunoassay. We evaluated the reproducibility and quantitation range in quantitative M2BPGi-Qt measurement. We also investigated the confidence ratio of M2BPGi-Qt levels measured by the new quantitative system to M2BPGi levels measured by the current semi-quantitative system for validating the clinical utility of the new method. RESULTS: The reproducibility of M2BPGi-Qt in HCV samples with negative, positive 1+, and positive 2+ was 0.77 ± 0.02 AU/mL, 2.25 ± 0.03 AU/mL, and 6.55 ± 0.21 AU/mL, respectively, and the corresponding coefficient of variation (CV)s were 2.1%, 1.3%, and 3.2%, respectively. The range of quantification assessment resulted that all CVs showed less than 5% in investigated range. Sample stability testing found that the mean percentage difference between the pre- and post-storage values of 6 samples ranged between 96.2 and 103.9%. The correlation coefficient between M2BPGi and M2BPGi-Qt in patients with HCV and the healthy volunteers was 0.986 and 0.991, respectively. M2BPGi-Qt could be quantitatively assessed in a patient with over 20 C.O.I. CONCLUSION: Compared with qualitative methods, the M2BPGi quantitative measurement system could provide a numerical value unaffected by interpretation bias, and measurements are more precise at high M2BPGi levels.

Pancreatic ductal adenocarcinoma with acinar-to-ductal metaplasia-like cancer cells shows increased cellular proliferation.

BACKGROUND/OBJECTIVES: Acinar-to-ductal metaplasia (ADM) has been shown to contribute to the development of pancreatic ductal adenocarcinoma (PDAC) in genetically engineered mouse models, but little is known about whether acinar cell plasticity contributes to carcinogenesis in human PDAC. We aimed to assess whether cancer cells that stain positive for amylase and CK19 (ADM-like cancer cells) are present in human resected PDAC and to investigate their role in tumor progression. METHODS: We immunohistochemically investigated the presence of ADM-like cancer cells, and compared the clinical and histological parameters of PDAC patients with and without ADM-like cancer cells. RESULTS: ADM-like cancer cells were detected in 16 of 60 (26.7%) PDAC specimens. Positive staining for anterior gradient protein 2 (AGR2) was observed in 14 of 16 (87.5%) PDAC specimens with ADM-like cancer cells. On the other hand, the intensity of AGR2 expression (negative, low/moderate or high) was lower in PDAC with ADM-like cancer cells (9/7) than in PDAC without these cells (11/33) (P = 0.032). The presence of ADM-like cancer cells was significantly correlated with increased cell proliferation (P = 0.012) and tended to be associated with MUC1 expression (P = 0.067). CONCLUSIONS: These results indicated that acinar cells may act as the origin of human PDAC, and that their presence may be useful for the stratification of human PDAC to predict prognosis.

Co-occurrence of non-alcoholic steatohepatitis exacerbates psoriasis associated with decreased adiponectin expression in a murine model.

Introduction: Clinical studies have suggested a bidirectional association between non-alcoholic steatohepatitis (NASH) and psoriasis, affecting each other's development and severity. Here, we explored bidirectional causal linkages between NASH and psoriasis using a murine model. Methods: NASH was induced in mice by streptozotocin injection at 2 days of age and by high-fat diet feeding (STAM™ model). Psoriasis was induced by topical application of imiquimod (IMQ) on the ear. The severities of liver damage and psoriatic skin changes were determined using histological analysis. Gene expression in the skin tissues was evaluated using quantitative PCR analysis. Serum cytokine levels were determined using enzyme-linked immunosorbent assay. To examine the innate immune responses of normal human epidermal keratinocytes (NHEKs), the cells were treated with interleukin (IL)-17A, tumor necrosis factor (TNF)-α, and AdipoRon, an adiponectin receptor agonist. Results and Discussion: There were no differences in the degree of liver tissue damage (fat deposition, inflammation, and fibrosis) between NASH mice with and those without psoriasis. Conversely, the co-occurrence of NASH significantly augmented psoriatic skin changes, represented by epidermal hyperplasia, in psoriatic mice. Pro-inflammatory cytokines were expressed in the inflamed skin of psoriatic mice, and the expression of genes, especially Il23a, Il1b, Il36g, and Mip2, was significantly upregulated by the co-occurrence of NASH. The expression of keratinocyte activation marker genes Defb4b and Krt16 was also upregulated by the co-occurrence of NASH. The serum TNF-α and IL-17 levels were increased by the co-occurrence of NASH and psoriasis. The serum adiponectin levels decreased in NASH mice compared with that in non-NASH mice. In NHEK culture, TNF-α and IL-17A synergistically upregulated CXCL1, CXCL8, and IL1B expression. The upregulated pro-inflammatory gene expression was suppressed by AdipoRon treatment, reflecting the anti-inflammatory capacity of adiponectin. Conclusion: The co-occurrence of NASH exacerbated psoriatic skin changes associated with increased serum inflammatory cytokine levels and decreased serum adiponectin levels. Combined with in vitro findings, increased inflammatory cytokine levels and decreased adiponectin levels likely promote innate immune responses in epidermal keratinocytes in psoriatic skin lesions. Overall, therapeutic intervention for co-occurring NASH is essential to achieve a favorable prognosis of psoriasis in clinical practice.

A case of successful treatment with antiretroviral therapy for HIV in a patient with marked liver dysfunction.

Background: Human immunodeficiency virus (HIV) infection is often complicated with hepatitis virus infection. Antiretroviral therapy (ART) should be initiated with caution for patients with severe virus- or drug-induced acute hepatitis while considering factors that might interfere with the initiation of therapy. Case report: Herein, we present a case of a 67-year-old woman who presented with symptoms of severe liver injury of unknown cause. Laboratory examinations revealed HIV infection. The HIV viral load was high, and treatment with ART was considered. However, a liver biopsy could not be performed because of hyperbilirubinemia and the risk of severe hepatic damage. After assessing the risk of further liver damage, ART was safely administered despite hyperbilirubinemia. Treatment with ART could successfully reduce the viral load and bilirubin levels. Conclusion: ART treatment could be safely used for patients with HIV to reduce the viral load and bilirubin levels while avoiding the risk of liver failure.

Inhibition of pancreatic cancer-cell growth and metastasis in vivo by a pyrazole compound characterized as a cell-migration inhibitor by an in vitro chemotaxis assay.

Pancreatic cancer is recalcitrant to treatment as it is highly metastatic and rapidly progressive. While observing the behavior of human pancreatic BxPC-3 cells using an optical assay device called TAXIScan, we found that several synthetic pyrazole and pyrimidine derivatives inhibited cell migration. One such compound, 14-100, inhibited metastasis of fluorescence-labeled BxPC-3 cells, which were transplanted into the pancreas of nude mice as a subcutaneously grown cancer fragment. Surprisingly, despite its low cytotoxicity, the compound also showed an inhibitory effect on cancer cell proliferation in vivo, suggesting that the compound alters cancer cell characteristics needed to grow in situ. Single-cell RNA-sequencing revealed changes in gene expression associated with metastasis, angiogenesis, inflammation, and epithelial-mesenchymal transition. These data suggest that the compound 14-100 could be a good drug candidate against pancreatic cancer.

CD26/DPP4 as a Therapeutic Target in Nonalcoholic Steatohepatitis Associated Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is generally considered an "immune-cold" cancer since T cells are not observed abundantly in HCC tumor tissue. Combination therapy with immune checkpoint inhibitors and vascular endothelial growth factor (VEGF) inhibitors is currently recognized as a first-line systemic treatment for advanced-stage HCC. Immunologically, immune checkpoint inhibitors influence the recognition of cancer cells by T cells, and VEGF inhibitors influence the infiltration of T cells into tumors. However, no drugs that facilitate the trafficking of T cells toward tumors have been developed. Chemokines are promising agents that activate T cell trafficking. On the other hand, metabolic factors such as obesity and insulin resistance are considered risk factors for HCC development. CD26/dipeptidyl peptidase 4 (DPP4) functions as a serine protease, selectively cleaving polypeptides with a proline or alanine at the penultimate N-terminal position, such as chemokines. Recently, CD26/DPP4 has been reported to attenuate anticancer immunity via chemokine cleavage and to promote insulin resistance and inflammation in the liver and/or adipose tissue via dysregulation of macrophage M1/M2 polarization. In this review, we discuss the promotive roles of CD26/DPP4 in HCC development and progression and the potential of DPP4 inhibitors as therapeutic agents for HCC.

Iron and liver cancer: an inseparable connection.

Iron is an essential element for all organisms. Iron-containing proteins play critical roles in cellular functions. The biological importance of iron is largely attributable to its chemical properties as a transitional metal. However, an excess of 'free' reactive iron damages the macromolecular components of cells and cellular DNA through the production of harmful free radicals. On the contrary, most of the body's excess iron is stored in the liver. Not only hereditary haemochromatosis but also some liver diseases with mild-to-moderate hepatic iron accumulation, such as chronic hepatitis C, alcoholic liver disease and nonalcoholic steatohepatitis, are associated with a high risk for liver cancer development. These findings have attracted attention to the causative and promotive roles of iron in the development of liver cancer. In the last decade, accumulating evidence regarding molecules regulating iron metabolism or iron-related cell death programmes such as ferroptosis has shed light on the relationship between hepatic iron accumulation and hepatocarcinogenesis. In this review, we briefly present the current molecular understanding of iron regulation in the liver. Next, we describe the mechanisms underlying dysregulated iron metabolism depending on the aetiology of liver diseases. Finally, we discuss the causative and promotive roles of iron in cancer development.

Long-term phlebotomy successfully alleviated hepatic iron accumulation in a ferroportin disease patient with a mutation in SLC40A1: a case report.

BACKGROUND: Hereditary hemochromatosis is a heterogenous group of inherited iron-overload conditions that is characterized by increased intestinal absorption and deposition in vital organs. Hepcidin is a soluble regulator that acts to attenuate both intestinal iron absorption and iron release from reticuloendothelial macrophages through internalization of ferroportin-1, an iron exporter. Ferroportin disease is hereditary hemochromatosis which is affected by SLC40A1, a gene coding ferroportin-1, and phenotypically classified into two forms (classical and nonclassical). In nonclassical form, ferroportin mutations are responsible for a gain of function with full iron export capability but insensitivity to downregulation by hepcidin. Here, we report a case of nonclassical ferroportin disease. CASE PRESENTATION: A 46-year-old Japanese man showed elevated serum iron (284 µg/dl), ferritin (1722 ng/ml), transferrin saturation ratio (91.3%), and hepcidin-25 level (139.6 ng/ml). Magnetic resonance imaging (MRI) demonstrated a marked reduction in the signal intensity of the liver in T1- and T2-weighted images. The liver histology exhibited a large amount of iron that had accumulated predominantly in hepatocytes. We identified a heterozygous 1520A > G (p.H507R) mutation in the SLC40A1 gene. Phlebotomy (400 ml at a time) was monthly performed for 3 years in this patient. Importantly, the serum hepcidin level (1.0 ng/ml) was normal when the serum ferritin level was normal and hepatic iron accumulation was remarkably reduced after 3 years of phlebotomy. CONCLUSIONS: The present case demonstrated for the first time that there was a correlation between hepatic iron levels as measured by MRI and serum hepcidin levels through long-term phlebotomy in a patient with ferroportin disease with the p.H507R mutation of in SLC40A1.

Nanoparticle-Mediated Delivery of 2-Deoxy-D-Glucose Induces Antitumor Immunity and Cytotoxicity in Liver Tumors in Mice.

BACKGROUND & AIMS: Immune checkpoint inhibitors have shed light on the importance of antitumor immunity as a therapeutic strategy for hepatocellular carcinoma (HCC). The altered glucose metabolism known as the Warburg effect recently has gained attention as a cancer immune-resistance mechanism. Considering glycolysis inhibitors as therapeutic agents, their specific delivery to cancer cells is critical not to induce adverse effects. Thus, we investigated antitumor effects of a glycolysis inhibitor, consisting of 2-deoxy-D-glucose (2DG)-encapsulated poly(lactic-co-glycolic acid) (PLGA) nanoparticles (2DG-PLGA-NPs), against hepatocellular carcinoma in mice. METHODS: The antitumor effects of 2DG-PLGA-NPs were examined using hepatoma cell lines, xenograft tumors, and hepatocarcinogenic and syngeneic mouse models. RESULTS: The 2DG-PLGA-NPs induced cytotoxic effects and antitumor immunity through enhanced T-cell trafficking. In addition, 2DG-PLGA-NPs induced decreased lactate production and increased interferon-γ-positive T cells in liver tumors. Human CD8+ T cells cocultured with 2DG-PLGA-NP-treated Huh7 cells showed their increased interferon-γ production and glucose uptake compared with the CD8+ T cells co-cultured with PLGA-NP-treated Huh7 cells. Chemotaxis of CD8+ T cells was suppressed by lactate and enhanced by glucose. Interferon-γ enhanced CD8+ T-cell chemotaxis in both an autocrine and paracrine manner. Notably, the 2DG-PLGA-NPs augmented chemokine (CXCL9/CXCL10) production in liver tumors via interferon-γ-Janus kinase-signal transducers and activator of transcription pathway and 5' adenosine monophosphate-activated protein kinase-mediated suppression of histone H3 lysine 27 trimethylation. These 2DG-PLGA-NPs not only amplified antitumor effects induced by sorafenib or an anti-programmed death-1 antibody, but also suppressed anti-programmed death-1-resistant tumors. CONCLUSIONS: The newly developed 2DG-PLGA-NPs showed antitumor immunity and cytotoxicity in liver tumors in mice, suggesting the potential of 2DG-PLGA-NPs for future clinical applications.

Iron loss triggers mitophagy through induction of mitochondrial ferritin.

Mitochondrial quality is controlled by the selective removal of damaged mitochondria through mitophagy. Mitophagy impairment is associated with aging and many pathological conditions. An iron loss induced by iron chelator triggers mitophagy by a yet unknown mechanism. This type of mitophagy may have therapeutic potential, since iron chelators are clinically used. Here, we aimed to clarify the mechanisms by which iron loss induces mitophagy. Deferiprone, an iron chelator, treatment resulted in the increased expression of mitochondrial ferritin (FTMT) and the localization of FTMT precursor on the mitochondrial outer membrane. Specific protein 1 and its regulator hypoxia-inducible factor 1α were necessary for deferiprone-induced increase in FTMT. FTMT specifically interacted with nuclear receptor coactivator 4, an autophagic cargo receptor. Deferiprone-induced mitophagy occurred selectively for depolarized mitochondria. Additionally, deferiprone suppressed the development of hepatocellular carcinoma (HCC) in mice by inducing mitophagy. Silencing FTMT abrogated deferiprone-induced mitophagy and suppression of HCC. These results demonstrate the mechanisms by which iron loss induces mitophagy and provide a rationale for targeting mitophagic activation as a therapeutic strategy.

Dipeptidyl Peptidase 4 Inhibitors Reduce Hepatocellular Carcinoma by Activating Lymphocyte Chemotaxis in Mice.

Background & Aims: CD26, a multifunctional transmembrane glycoprotein, is expressed in various cancers and functions as dipeptidyl peptidase 4 (DPP4). We investigated whether CD26 expression is associated with hepatocellular carcinoma (HCC) progression and whether DPP4 inhibitors exert antitumor effects against HCC. Methods: CD26 expression was examined in 41 surgically resected HCC specimens. The effects of DPP4 inhibitors on HCC were examined by using HCC cell lines (Huh-7 and Li-7), xenograft tumors in nude mice, and a nonalcoholic steatohepatitis-related HCC mouse model. Results: CD26 expression in HCC specimens was associated with increased serum DPP4 activity, as well as a more advanced stage, less tumor immunity, and poorer prognosis in HCC patients. The HCC cell lines and xenograft tumors exhibited CD26 expression and DPP4 activity. The DPP4 inhibitors did not exhibit antitumor effects in vitro, but natural killer (NK) and/or T-cell tumor accumulation suppressed growth of xenograft tumor and HCC in vivo. The antitumor effects of DPP4 inhibitors were abolished by the depletion of NK cells or the neutralization of CXCR3, a chemokine receptor on NK cells. EZ-TAXIScan, an optical horizontal chemotaxis apparatus, identified enhanced NK and T-cell chemotaxis by DPP4 inhibitors ex vivo in the presence of Huh-7 cells and the chemokine CXCL10, which binds to CXCR3. The DPP4 inhibitors prevented the biologically active form of CXCL10 from being truncated by Huh-7 cell DPP4 activity. DPP4 inhibitors also suppressed tumor angiogenesis. Conclusions: These results provide a rationale for verifying whether DPP4 inhibitors clinically inhibit the progression of HCC or augment the antitumor effects of molecular-targeting drugs or immunotherapies against HCC.

Genome-wide association study identified new susceptible genetic variants in HLA class I region for hepatitis B virus-related hepatocellular carcinoma.

We have performed a genome-wide association study (GWAS) including 473 Japanese HBV (hepatitis B virus)-positive HCC (hepatocellular carcinoma) patients and 516 HBV carriers including chronic hepatitis and asymptomatic carrier individuals to identify new host genetic factors associated with HBV-derived HCC in Japanese and other East Asian populations. We identified 65 SNPs with P values < 10-4 located within the HLA class I region and three SNPs were genotyped in three independent population-based replication sets. Meta-analysis confirmed the association of the three SNPs (rs2523961: OR = 1.73, P = 7.50 × 10-12; rs1110446: OR = 1.79, P = 1.66 × 10-13; and rs3094137: OR = 1.73, P = 7.09 × 10-9). We then performed two-field HLA genotype imputation for six HLA loci using genotyping data to investigate the association between HLA alleles and HCC. HLA allele association testing revealed that HLA-A * 33:03 (OR = 1.97, P = 4.58 × 10-4) was significantly associated with disease progression to HCC. Conditioning analysis of each of the three SNPs on the HLA class I region abolished the association of HLA-A*33:03 with disease progression to HCC. However, conditioning the HLA allele could not eliminate the association of the three SNPs, suggesting that additional genetic factors may exist in the HLA class I region.

Wisteria floribunda agglutinin-positive Mac-2 binding protein predicts the development of hepatocellular carcinoma in patients with non-alcoholic fatty liver disease.

AIM: As it is not practical to perform regular screening for hepatocellular carcinoma (HCC) in all patients with non-alcoholic fatty liver disease (NAFLD), there is a need to identify NAFLD patients who are at high risk for HCC. Wisteria floribunda agglutinin-positive Mac-2 binding protein (WFA+ -M2BP) has been shown to be a surrogate marker for predicting HCC as well as a liver fibrosis marker in patients with chronic hepatitis B and C. The aim of this study was to investigate whether WFA+ -M2BP predicts HCC development in NAFLD patients. METHODS: Serum WFA+ -M2BP was retrospectively measured in 331 patients with histologically proven NAFLD, 51 of whom developed HCC. The association of WFA+ -M2BP and HCC development in NAFLD patients was investigated. RESULTS: The WFA+ -M2BP values were significantly greater in NAFLD patients with HCC than in those without HCC among patients with liver fibrosis ≥stage 3. Multivariate analysis identified WFA+ -M2BP as one of the predictive factors for HCC development (odds ratio, 1.57; 95% confidence interval, 1.083-2.265; P = 0.017). The optimal cut-off index of WFA+ -M2BP for predicting HCC was 1.255 with specificity of 78.4% and sensitivity of 70.4%. The area under the receiver operating characteristic curve value for the prediction of HCC development was 0.806. The cumulative incidence rate of HCC was significantly greater in patients with WFA+ -M2BP ≥ 1.255 (n = 61) than in those with WFA+ -M2BP < 1.255 (n = 137) among patients who were followed up for more than 2 years after the diagnosis of NAFLD. CONCLUSIONS: Wisteria floribunda agglutinin-positive Mac-2 binding protein predicts HCC development and is a useful surrogate marker for identifying NAFLD patients who are at a high risk for HCC.

Genome-Wide Association Study Identifies TLL1 Variant Associated With Development of Hepatocellular Carcinoma After Eradication of Hepatitis C Virus Infection.

BACKGROUND & AIMS: There is still a risk for hepatocellular carcinoma (HCC) development after eradication of hepatitis C virus (HCV) infection with antiviral agents. We investigated genetic factors associated with the development of HCC in patients with a sustained virologic response (SVR) to treatment for chronic HCV infection. METHODS: We obtained genomic DNA from 457 patients in Japan with a SVR to interferon-based treatment for chronic HCV infection from 2007 through 2015. We conducted a genome-wide association study (GWAS), followed by a replication analysis of 79 candidate single nucleotide polymorphisms (SNPs) in an independent set of 486 patients in Japan. The study end point was HCC diagnosis or confirmation of lack of HCC (at follow-up examinations until December 2014 in the GWAS cohort, and until January 2016 in the replication cohort). We collected clinical and laboratory data from all patients. We analyzed expression levels of candidate gene variants in human hepatic stellate cells, rats with steatohepatitis caused by a choline-deficient L-amino acid-defined diet, and a mouse model of liver injury caused by administration of carbon tetrachloride. We also analyzed expression levels in liver tissues of patients with chronic HCV infection with different stages of fibrosis or tumors vs patients without HCV infection (controls). RESULTS: We found a strong association between the SNP rs17047200, located within the intron of the tolloid like 1 gene (TLL1) on chromosome 4, and development of HCC; there was a genome-wide level of significance when the results of the GWAS and replication study were combined (odds ratio, 2.37; P = 2.66 × 10-8). Multivariate analysis showed rs17047200 AT/TT to be an independent risk factor for HCC (hazard ratio, 1.78; P = .008), along with male sex, older age, lower level of albumin, advanced stage of hepatic fibrosis, presence of diabetes, and higher post-treatment level of α-fetoprotein. Combining the rs17047200 genotype with other factors, we developed prediction models for HCC development in patients with mild or advanced hepatic fibrosis. Levels of TLL1 messenger RNA (mRNA) in human hepatic stellate cells increased with activation. Levels of Tll1 mRNA increased in liver tissues of rodents with hepatic fibrogenesis compared with controls. Levels of TLL1 mRNA increased in liver tissues of patients with progression of fibrosis. Gene expression levels of TLL1 short variants, including isoform 2, were higher in patients with rs17047200 AT/TT. CONCLUSIONS: In a GWAS, we identified the association between the SNP rs17047200, within the intron of TLL1, and development of HCC in patients who achieved an SVR to treatment for chronic HCV infection. We found levels of Tll1/TLL1 mRNA to be increased in rodent models of liver injury and liver tissues of patients with fibrosis, compared with controls. We propose that this SNP might affect splicing of TLL1 mRNA, yielding short variants with high catalytic activity that accelerates hepatic fibrogenesis and carcinogenesis. Further studies are needed to determine how rs17047200 affects TLL1 mRNA levels, splicing, and translation, as well as the prevalence of this variant among other patients with HCC. Tests for the TLL1 SNP might be used to identify patients at risk for HCC after an SVR to treatment of HCV infection.

Assessment of clinical and magnetic resonance imaging features of de novo hypervascular hepatocellular carcinoma using gadoxetic acid-enhanced magnetic resonance imaging.

AIM: To clarify the clinical and magnetic resonance imaging (MRI) features of de novo hypervascular hepatocellular carcinoma (HCC) using serial gadoxetic acid-enhanced MRI. METHODS: The institutional review board approved this retrospective study. After review of 1007 MRI examinations in 240 patients with chronic liver disease, 17 newly developed hypervascular HCCs in 16 patients detected by follow-up from initial MRI examination without hepatocellular nodules were evaluated. The clinical and MRI findings such as previous treatment history for HCC, period to hypervascular HCC onset, presence or absence of hypovascular hypointense nodules on hepatobiliary phase before hypervascularization, and intralesional fat component were recorded or evaluated. Statistical evaluations included Fisher's exact test, χ2 -test, and Mann-Whitney U-test. RESULTS: In 17 HCCs, 12 (71%) were de novo hypervascular HCC without showing hypovascular hypointense nodule on hepatobiliary phase before hypervascularization (de novo group) and 5 (29%) were hypervascularized HCC developed during multistep hepatocarcinogenesis (multistep group). The incidence of previous treatment history for HCC in the de novo group (91%) was significantly higher than that in the multistep group (20%) (P = 0.013). The duration to hypervascular HCC onset from initial examination was shorter in the de novo group (mean, 291 days) than in the multistep group (mean, 509 days) (P = 0.035). The incidence of fat-containing lesion in the de novo group (0%) was lower than that in the multistep group (40%) (P = 0.074). CONCLUSION: De novo hypervascular HCC is characterized by rapid growth, patients with previous treatment history for HCC, and lack of intralesional fat, compared to hypervascular HCC with multistep progression.

Fibroblast activation protein-α-expressing fibroblasts promote the progression of pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is characterized by an extensive desmoplastic stromal response. Fibroblast activation protein-α (FAP) is best known for its presence in stromal cancer-associated fibroblasts (CAFs). Our aim was to assess whether FAP expression was associated with the prognosis of patients with PDAC and to investigate how FAP expressing CAFs contribute to the progression of PDAC. METHODS: FAP expression was immunohistochemically assessed in 48 PDAC specimens. We also generated a fibroblastic cell line stably expressing FAP, and examined the effect of FAP-expressing fibroblasts on invasiveness and the cell cycle in MiaPaCa-2 cells (a pancreatic cancer cell line). RESULTS: Stromal FAP expression was detected in 98% (47/48) of the specimens of PDAC, with the intensity being weak in 16, moderate in 19, and strong in 12 specimens, but was not detected in the 3 control noncancerous pancreatic specimens. Patients with moderate or strong FAP expression had significantly lower cumulative survival rates than those with negative or weak FAP expression (mean survival time; 352 vs. 497 days, P = 0.006). Multivariate analysis identified moderate to strong expression of FAP as one of the factors associated with the prognosis in patients with PDAC. The intensity of stromal FAP expression was also positively correlated to the histological differentiation of PDAC (P < 0.05). FAP-expressing fibroblasts promoted the invasiveness of MiaPaCa-2 cells more intensively than fibroblasts not expressing FAP. Coculture with FAP-expressing fibroblasts significantly activated cell cycle shift in MiaPaCa-2 cells compared to coculture with fibroblasts not expressing FAP. Furthermore, coculture with FAP expressing fibroblasts inactivated retinoblastoma (Rb) protein, an inhibitor of cell cycle progression, in MiaPaCa-2 cells by promoting phosphorylation of Rb. CONCLUSIONS: The present in vitro results and the association of FAP expression with clinical outcomes provide us with a better understanding of the effect of FAP-expressing CAFs on the progression of PDAC.

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.