A nationwide survey regarding the life situations of patients with thalidomide embryopathy in Japan, 2018: First report.

BACKGROUND: Clinical studies on the effects of thalidomide-induced damage on thalidomide victims as they age have only recently started to be conducted, but no studies have examined socioeconomic differences in terms of healthcare and social status between thalidomiders and the age-matched general population in Japan. Therefore, we carried out a nationwide survey focusing on the life situations of thalidomiders. METHOD: Questionnaires were sent to 274 thalidomiders in Japan. The questionnaire items basically matched those of the Comprehensive Survey of Living Conditions (CSLC) in the general population conducted by the Japanese Government. The results were compared with those of the CSLC for individuals aged 55-59 years, which was the cohort most similar in age to the average thalidomider living in Japan. RESULTS: More thalidomiders rated their health condition as relatively bad or bad compared with the general population (20.2% vs. 13.3%, respectively). A much higher percentage of thalidomiders reported having some health or physical problem caused by a disease or injury (68.8% vs. 32.6%, respectively), and thalidomiders reported visiting medical and healthcare-related facilities more frequently. A higher proportion of thalidomiders (9.2%) were unemployed, and thalidomiders tended to feel higher levels of worry and stress, especially in terms of the future. CONCLUSIONS: The results of this nationwide survey of the life situations of thalidomiders in Japan clarified their health conditions and the related associations with socioeconomic status. These findings could be expected to help improve the provision of medical and healthcare, welfare measures, and financial support for thalidomiders in the near future.

Recent development of balloon-occluded retrograde transvenous obliteration.

Gastric varices (GVs) are a major complication of portal hypertension in patients with liver cirrhosis. The mortality rate associated with the bleeding from GVs is not low. Balloon-occluded retrograde transvenous obliteration (BRTO) was first introduced by Kanagawa et al. as a treatment for isolated GVs in 1994. It has been performed most frequently in Asia, especially in Japan. Ethanolamine oleate was the original sclerosant used in the therapy. Since the late 2000s, BRTO using sodium tetradecyl sulfate foam or polidocanol foam as a sclerosant has been performed in many countries other than Japan. Then, early in the 2010s, modified BRTO techniques including vascular plug-assisted retrograde transvenous obliteration and coil-assisted retrograde transvenous obliteration were developed as an alternative treatment for GVs. This article provides a historical overview of BRTO using various sclerosants and modified BRTO techniques, such as plug-assisted retrograde transvenous obliteration and coil-assisted retrograde transvenous obliteration.

Novel and Simple Ultrasonographic Methods for Estimating the Abdominal Visceral Fat Area.

OBJECTIVES: To evaluate the abdominal visceral fat area (VFA), we developed novel ultrasonographic (US) methods for estimating. METHODS: 100 male volunteers were recruited, and their VFA was calculated by two novel US methods, the triangle method and the ellipse method. The VFA calculated by these methods was compared with the VFA calculated by CT. RESULTS: Both the VFA calculated by the triangle method (r = 0.766, p < 0.001) and the ellipse method (r = 0.781, p < 0.001) showed a high correlation coefficient with the VFA calculated by CT. Also, the VFA calculated by our novel methods were significantly increased in subjects with one or more metabolic risk factors than in those without any risk factors. Furthermore, the correlation coefficients obtained using the two methods were enhanced by the addition of multiple regression analysis (with the triangle method, r = 0.8586, p < 0.001; with the ellipse method, r = 0.8642, p < 0.001). CONCLUSIONS: The VFA calculated by the triangle or ellipse method showed a high correlation coefficient with the VFA calculated by CT. These US methods are easy to use, they involve no radiation exposure, and the measurements can be conducted frequently. We hope that our simple methods would be widely adopted for the evaluation of VFA.

Circulating AIM as an indicator of liver damage and hepatocellular carcinoma in humans.

BACKGROUND: Hepatocellular carcinoma (HCC), the fifth most common cancer type and the third highest cause of cancer death worldwide, develops in different types of liver injuries, and is mostly associated with cirrhosis. However, non-alcoholic fatty liver disease often causes HCC with less fibrosis, and the number of patients with this disease is rapidly increasing. The high mortality rate and the pathological complexity of liver diseases and HCC require blood biomarkers that accurately reflect the state of liver damage and presence of HCC. METHODS AND FINDINGS: Here we demonstrate that a circulating protein, apoptosis inhibitor of macrophage (AIM) may meet this requirement. A large-scale analysis of healthy individuals across a wide age range revealed a mean blood AIM of 4.99 ± 1.8 µg/ml in men and 6.06 ± 2.1 µg/ml in women. AIM levels were significantly augmented in the younger generation (20s-40s), particularly in women. Interestingly, AIM levels were markedly higher in patients with advanced liver damage, regardless of disease type, and correlated significantly with multiple parameters representing liver function. In mice, AIM levels increased in response to carbon tetrachloride, confirming that the high AIM observed in humans is the result of liver damage. In addition, carbon tetrachloride caused comparable states of liver damage in AIM-deficient and wild-type mice, indicating no influence of AIM levels on liver injury progression. Intriguingly, certain combinations of AIM indexes normalized to liver marker score significantly distinguished HCC patients from non-HCC patients and thus could be applicable for HCC diagnosis. CONCLUSION: AIM potently reveals both liver damage and HCC. Thus, our results may provide the basis for novel diagnostic strategies for this widespread and fatal disease.

In vitro selection of the 3'-untranslated regions of the human liver mRNA that bind to the HCV nonstructural protein 5B.

Hepatitis C virus (HCV) nonstructural protein 5B (NS5B) has RNA-dependent RNA polymerase (RdRp) activity. Because NS5B recognizes various RNA motifs besides the HCV genome, NS5B has the potential of interacting with host RNA molecules. In this study, an RNA pool enriched with the 3'-UTR sequences was generated and mRNA molecules with high affinity binding to NS5B were selected by iterative selection. Among the high binding mRNA 3'-UTR segments, we analyzed the housekeeping ribosomal protein S4, X-linked [RPS4X] mRNA 3'-UTR and the 3'-UTR of galectin-1 (GAL-1) mRNA, which is known to be one of the genes upregulated in HCV-infected liver cells and to have a wide spectrum of biological properties. By means of IP-RT-PCR, it was demonstrated that both of the mRNA molecules bind to NS5B in the cytoplasm. Interestingly, GAL-1 and RPS4X mRNA can serve as templates for NS5B RdRp, suggesting these RNA molecules are regulated in vivo by NS5B.

Effects of long-term zinc treatment in Japanese patients with Wilson disease: efficacy, stability, and copper metabolism.

Wilson disease is an autosomal recessive disorder with copper metabolism. In Japan, the standard treatment is the administration of copper chelating agents, such as D-penicillamine and trientine. In this study, the authors used zinc acetate to treat Japanese patients with Wilson disease and investigated its efficacy. The 37 patients that comprise this study were found to have Wilson disease using clinical and biochemical tests and were administrated zinc acetate for 48 weeks. The authors followed the clinical symptoms and laboratory findings of the patients by assessing their complete blood counts, biochemical findings, as well as the results of urinalysis and special laboratory tests for copper and zinc metabolism. We also examined side effects of the treatment. Zinc acetate did not aggravate the hepatic or neurological symptoms of any of the patients. Blood biochemical analysis also did not reveal elevation of alanine aminotransferase, aspartate aminotransferase, and γ-glutamyltranspeptidase levels. Zinc treatment did not aggravate the patients' clinical signs and/or laboratory findings. However, it did improve some clinical symptoms of the Wilson disease patients. Although this agent had some side effects, none of them were severe. The authors measured spot urinary copper excretion, which gave an indication of the efficacy of treatment and of the sufficient dosage of zinc. We recommend maintaining a spot urinary copper excretion less than 0.075-µg/mg creatinine. The authors conclude that zinc acetate is an effective and safe treatment for Japanese patients with Wilson disease.

RNA-dependent RNA polymerase of hepatitis C virus binds to its coding region RNA stem-loop structure, 5BSL3.2, and its negative strand.

The hepatitis C virus NS5B RNA-dependent RNA polymerase (RdRp) is a key enzyme involved in viral replication. Interaction between NS5B RdRp and the viral RNA sequence is likely to be an important step in viral RNA replication. The C-terminal half of the NS5B-coding sequence, which contains the important cis-acting replication element, has been identified as an NS5B-binding sequence. In the present study, we confirm the specific binding of NS5B to one of the RNA stem-loop structures in the region, 5BSL3.2. In addition, we show that NS5B binds to the complementary strand of 5BSL3.2 (5BSL3.2N). The bulge structure of 5BSL3.2N was shown to be indispensable for tight binding to NS5B. In vitro RdRp activity was inhibited by 5BSL3.2N, indicating the importance of the RNA element in the polymerization by RdRp. These results suggest the involvement of the RNA stem-loop structure of the negative strand in the replication process.

Adiponectin knockout mice on high fat diet develop fibrosing steatohepatitis.

BACKGROUND AND AIM: Low levels of serum adiponectin have been reported to be associated with obesity, diabetes, and non-alcoholic steatohepatitis (NASH), as well as several malignancies. Adiponectin knockout (KO) mice have been reported to cause insulin resistance and neointimal formation of the artery. We used adiponectin KO mice fed a high fat (HF) diet, and investigated the effect of adiponectin on the progression of steatohepatitis and carcinogenesis in vivo. METHODS: Adiponectin KO mice and wild type (WT) mice were fed a HF diet or normal chow for the periods of 24 and 48 weeks. The HF diet contained 60% of calories from fat. RESULTS: The adiponectin KO mice on the HF diet showed obesity, marked elevation of serum transaminase levels, and hyperlipidemia. At 24 weeks, hepatic expression of tumor necrosis factor-alpha and procollagen alpha (I) was higher in KO mice as compared with WT mice. At 48 weeks, liver triglyceride contents in KO mice on normal chow were significantly higher than those in WT mice. Hepatocyte ballooning, spotty necrosis, and pericellular fibrosis around central veins were observed in KO mice on the HF diet. The pericellular fibrosis was more severe in KO mice on the HF diet than that in WT mice (1.62% vs 1.16%, P = 0.033). Liver adenoma and hyperplastic nodules developed in a KO mouse on the HF diet at 48 weeks (12.5%, n = 1/8), whereas no tumor was detected in WT mice (n = 10). CONCLUSIONS: Adiponectin may play a protective role in the progression of NASH in the early stages by suppressing tumor necrosis factor-alpha expression and liver fibrosis.

Adiponectin suppresses hepatic SREBP1c expression in an AdipoR1/LKB1/AMPK dependent pathway.

Adiponectin, one of the insulin-sensitizing adipokines, has been shown to activate fatty acid oxidation in liver and skeletal muscle, thus maintaining insulin sensitivity. However, the precise roles of adiponectin in fatty acid synthesis are poorly understood. Here we show that adiponectin administration acutely suppresses expression of sterol regulatory element-binding protein (SREBP) 1c, the master regulator which controls and upregulates the enzymes involved in fatty acid synthesis, in the liver of +Lepr(db)/+Lepr(db) (db/db) mouse as well as in cultured hepatocytes. We also show that adiponectin suppresses SREBP1c by AdipoR1, one of the functional receptors for adiponetin, and furthermore that suppressing either AMP-activated protein kinase (AMPK) via its upstream kinase LKB1 deletion cancels the negative effect of adiponectin on SREBP1c expression. These data show that adiponectin suppresses SREBP1c through the AdipoR1/LKB1/AMPK pathway, and suggest a possible role for adiponectin in the regulation of hepatic fatty acid synthesis.

In vitro selection of RNA aptamers that bind the RNA-dependent RNA polymerase of hepatitis C virus: a possible role of GC-rich RNA motifs in NS5B binding.

We employed SELEX (systematic evolution of ligands by exponential enrichment) and identified high affinity RNA aptamers to the hepatitis C virus NS5B RNA-dependent RNA polymerase (RdRp). GC-rich stretches were identified in many of the aptamers. Deletion of the 5'-end single-stranded GC-stretch (CGGG) of the highest binding RNA impaired the binding and the inhibitory activity of the RNA to NS5B RdRp. The majority of the mutants with a single base substitution on the CGGG motif exhibited weaker binding to NS5B. Interestingly, the CGGG motif is present on the stem structure of the NS5B coding RNA (5BSL3.2), which is considered to be an important cis-acting replication element. The 5BSL3.2 RNA showed substantial binding to NS5B, while a point mutation on the CGGG motif reduced the binding of RNA to NS5B. These results suggest a GC-stretch to be the RNA element recognized by NS5B.

Dynamic functional relay between insulin receptor substrate 1 and 2 in hepatic insulin signaling during fasting and feeding.

Insulin receptor substrate (Irs) mediates metabolic actions of insulin. Here, we show that hepatic Irs1 and Irs2 function in a distinct manner in the regulation of glucose homeostasis. The PI3K activity associated with Irs2 began to increase during fasting, reached its peak immediately after refeeding, and decreased rapidly thereafter. By contrast, the PI3K activity associated with Irs1 began to increase a few hours after refeeding and reached its peak thereafter. The data indicate that Irs2 mainly functions during fasting and immediately after refeeding, and Irs1 functions primarily after refeeding. In fact, liver-specific Irs1-knockout mice failed to exhibit insulin resistance during fasting, but showed insulin resistance after refeeding; conversely, liver-specific Irs2-knockout mice displayed insulin resistance during fasting but not after refeeding. We propose the concept of the existence of a dynamic relay between Irs1 and Irs2 in hepatic insulin signaling during fasting and feeding.

Nonlinear optical properties of polydiacetylene with donor-acceptor substitution block.

The elongation finite-field (elongation-FF) method is applied to donor/acceptor substituted polydiacetylenes (PDAs) for the estimation of substituent effects on nonlinear optical (NLO) properties. The first hyperpolarizability (beta) and the second hyperpolarizability (gamma) of PDA with separated donor and acceptor substitution blocks have much larger values than those of the other substituted PDAs. For the PDAs with donor and acceptor substitution blocks, the relationship between the NLO properties and the block period is examined. It is shown, from the local density of states, that gamma of a system with a quantum well structure has a maximum value at a certain block size. This indicates that by tuning the size of proper block it is possible to achieve the largest gamma value in block polymers. Furthermore, the through-space/bond interaction analysis is performed to examine the pi-conjugation effects on the NLO properties for particular substituted PDA. It is demonstrated by our quantitative analysis that beta is affected by electron transfers in the molecule, and the quantum well structure is critical for gamma improvement.

Co-ordinate activation of lipogenic enzymes in hepatocellular carcinoma.

Hepatocellular carcinoma is a very common neoplastic disease in countries where hepatitis viruses B and/or C are prevalent. Small hepatocellular carcinoma lesions detected by ultrasonography at an early stage are often hyperechoic because they are composed of well-differentiated cancer cells that are rich in triglyceride droplets. The triglyceride content of hepatocytes depends in part on the rate of lipogenesis. Key lipogenic enzymes, such as fatty acid synthase, are co-ordinately regulated at the transcriptional level. We therefore examined the mRNA expression of lipogenic enzymes in human hepatocellular carcinoma samples from 10 patients who had undergone surgical resection. All of the samples exhibited marked elevation of expression of mRNA for lipogenic enzymes, such as fatty acid synthase, acetyl-CoA carboxylase and ATP citrate lyase, compared with surrounding non-cancerous liver tissue. In contrast, the changes in mRNA expression of SREBP-1, a transcription factor that regulates a battery of lipogenic enzymes, did not show a consistent trend. In some cases where SREBP-1 was elevated, the main contributing isoform was SREBP-1c rather than SREBP-1a. Thus, lipogenic enzymes are markedly induced in hepatocellular carcinomas, and in some cases SREBP-1c is involved in this activation.

Peroxisome proliferator-activated receptor gamma inhibition prevents adhesion to the extracellular matrix and induces anoikis in hepatocellular carcinoma cells.

Activation of the nuclear transcription factor peroxisome proliferator-activated receptor gamma (PPARgamma) inhibits growth and survival of hepatocellular carcinoma (HCC) cell lines. To further investigate the function of PPARgamma in HCC, PPARgamma expression patterns in primary tumors were examined, and the responses of two HCC cell lines to PPARgamma activation and inhibition were compared. PPARgamma expression was increased in HCC and benign-appearing peritumoral hepatocytes compared with remote benign hepatocytes. Both compound PPARgamma inhibitors and PPARgamma small interfering RNAs prevented HCC cell lines from adhering to the extracellular matrix. Loss of adhesion was followed by caspase-dependent apoptosis (anoikis). PPARgamma inhibitors had no effect on initial beta1 integrin-mediated adhesion, or on total focal adhesion kinase levels but did reduce focal adhesion kinase phosphorylation. The PPARgamma inhibitor T0070907 was significantly more efficient at causing cancer cell death than the activators troglitazone and rosiglitazone. T0070907 caused cell death by reducing adhesion and inducing anoikis, whereas the activators had no direct effect on adhesion and caused cell death at much higher concentrations. In conclusion, PPARgamma overexpression is present in HCC. Inhibition of PPARgamma function causes HCC cell death by preventing adhesion and inducing anoikis-mediated apoptosis. PPARgamma inhibitors represent a potential novel treatment approach to HCC.

Identification of soluble NH2-terminal fragment of glypican-3 as a serological marker for early-stage hepatocellular carcinoma.

For detection of hepatocellular carcinoma (HCC) in patients with liver cirrhosis, serum alpha-fetoprotein has been widely used, but its sensitivity has not been satisfactory, especially in small, well-differentiated HCC, and complementary serum marker has been clinically required. Glypican-3 (GPC3), a heparan sulfate proteoglycan anchored to the plasma membrane, is a good candidate marker of HCC because it is an oncofetal protein overexpressed in HCC at both the mRNA and protein levels. In this study, we demonstrated that its NH(2)-terminal portion [soluble GPC3 (sGPC3)] is cleaved between Arg(358) and Ser(359) of GPC3 and that sGPC3 can be specifically detected in the sera of patients with HCC. Serum levels of sGPC3 were 4.84 +/- 8.91 ng/ml in HCC, significantly higher than the levels seen in liver cirrhosis (1.09 +/- 0.74 ng/ml; P < 0.01) and healthy controls (0.65 +/- 0.32 ng/ml; P < 0.001). In well- or moderately-differentiated HCC, sGPC3 was superior to alpha-fetoprotein in sensitivity, and a combination measurement of both markers improved overall sensitivity from 50% to 72%. These results indicate that sGPC3 is a novel serological marker essential for the early detection of HCC.

Effect of apolipoprotein E polymorphism on bile lipid composition and the formation of cholesterol gallstone.

OBJECTIVE: It remains a matter of controversy whether possession of the apolipoprotein E4 (apoE4) allele is a genetic risk factor for the formation of cholesterol gallstones. The aim of the present study was to test this hypothesis by investigating the effect of apoE4 on bile lipid composition in normal subjects and in patients with cholesterol gallstones and to evaluate the distributions of apoE alleles in these two groups. METHODS: The study population consisted of 79 patients who underwent open or laparoscopic cholecystectomy for symptomatic cholesterol gallstone disease. The control group (n = 53) included 11 patients with benign gallbladder polyps and 42 normal subjects acting as donors in living donor liver transplantation. The apoE genotypes were assessed by dot blot hybridization with digoxigenin-labeled probes. Bile lipid composition was determined by enzymatic assays and high performance liquid chromatography. RESULTS: Bile lipid composition and cholesterol saturation index (CSI) were similar in the control subjects harboring the apoE4 allele and those without apoE4 (mean CSI, 85.9% and 72.2%, respectively, p = 0.69). Likewise, in the cholesterol gallstone patients, bile lipid composition and CSI were similar in the patients with and without apoE4 allele (mean CSI, 134.9% vs 152.2%, p = 0.6). Furthermore, the prevalence of the apoE4 allele was similar in the patients with cholesterol gallstones and in the control group (8.5% vs 7.6%, p = 0.46, OR = 0.88; 95% CI = 0.64-1.22). CONCLUSIONS: The apoE4 allele is not a contributory factor to cholesterol gallstone formation, at least in the Japanese population.

Des-gamma-carboxy prothrombin in cancer and non-cancer liver tissue of patients with hepatocellular carcinoma.

Des-gamma-carboxy prothrombin (DCP), also known as protein induced by vitamin K absence or antagonist II absence (PIVKA-II), has been considered as a useful serum tumor marker for hepatocellular carcinoma (HCC). However, the underlying mechanism causing the elevation of serum DCP levels in HCC patients remains unclear. This study was undertaken to identify the relationship between serum DCP levels and the expression of DCP in cancer and surrounding non-cancer liver tissues of HCC patients. Serum and tissue samples prepared from 92 patients with a single HCC nodule were subjected to clinicopathological study by measuring serum DCP levels and performing immunohistochemical staining for tissue DCP. Serum DCP levels correlated significantly with clinicopathological factors such as hepatitis markers, tumor differentiation, vascular invasion, intrahepatic metastasis, TNM stage, tumor size, tumor recurrence, and patient survival. DCP immunohistochemical staining was positive in cancer tissues for 68 (68/92, 73.9%) patients and in non-cancer tissues surrounding tumors for 24 (24/92, 26.1%) patients. There was no apparent correlation between serum DCP values and the expression of DCP in HCC tissues; however, there was a significant correlation between serum DCP levels and the expression of DCP in non-HCC tissues (p=0.0398). In conclusion, our results suggest that the origin of elevated serum DCP may lie not only in HCC tissue but also in non-cancer tissues. The HCC lesion itself appears to influence the production of DCP in surrounding non-cancer tissues.

Treatment of unresectable hepatocellular carcinoma less than 2 centimeters by transcatheter arterial chemoembolization with autologous blood clot.

GOALS: To assess the efficacy of transcatheter arterial chemoembolization using autologous blood clot as an embolizing agent (short-TAE [S-TAE]) for the treatment of unresectable hepatocellular carcinoma less than 2 cm. STUDY: Twenty-eight consecutive patients with unresectable hepatocellular carcinoma less than 2 cm in diameter were treated by S-TAE alone. All patients had documented cirrhosis (Child class B:C = 20:8). S-TAE was performed by injecting a mixture of iodized oil and anticancer drugs followed by embolization of hepatic arteries with autologous blood clot. RESULTS: A total of 147 sessions of embolization with clots were performed. S-TAE maintained patency of hepatic arteries. The overall survival rates at 1, 3, 5, and 8 years were estimated to be 89%, 52%, 34%, and 17%, respectively, which were better compared with prior records for the gelfoam method. The survival rates for Child class B patients were significantly better than that for Child class C patients (P < 0.05). The Cox proportional hazard model also demonstrated that Child staging of cirrhosis was the sole factor significantly predicting the survival (P < 0.05). CONCLUSIONS: The long-term outcomes of S-TAE for unresectable hepatocellular carcinoma less than 2 cm are satisfactory. Prognosis of these patients was significantly dependent on clinical stages of coexisting liver cirrhosis.