Suguru Ogura

The expression and function of Toll-like receptors 3 and 9 in human colon carcinoma

Toll-like receptors (TLRs) are pattern-recognition receptors that are important in immune signaling. TLR recognition of various viral components including double-stranded RNA (TLR3) and unmethylated CpG-DNA (TLR9) plays a crucial role in cell survival. However, TLR expression and function in colon carcinoma cells are not well clarified. We investigated the expression of TLR3 and TLR9 in colon carcinoma cells using immunohistochemical methods. The function of TLR3 and TLR9 signaling in carcinoma cell lines was studied by direct cell stimulation with, or by cell transfection of, polyinosinic-polycytidylic acid (Poly I:C), a synthetic form of dsRNA, and by cell stimulation with CpG-oligodeoxynucleotides (ODNs), respectively. Positive TLR3 and TLR9 immunohistochemical staining was observed in 91 and 86% of human hepatocellular carcinoma (HCC) tissues, respectively. Cell surface stimulation of TLR3 with Poly I:C did not affect cell viability but it did activate NF-κB activity. By contrast, stimulation of intracellular TLRs with transfected Poly I:C significantly induced apoptosis. Cell surface stimulation of TLR9 with CpG-ODNs promoted cell proliferation, and, furthermore, these CpG-ODN TLR9 agonists reduced the cytotoxicity of the anticancer drug adriamycin. Cell surface expression of TLR3 and TLR9 in colon carcinoma cells plays an important role in cell survival. In addition, the proapoptotic activity of intracellularly expressed TLR3 may provide the possibility of using TLR3 agonists as novel clinical cytotoxic agents against colon carcinoma cells.

Des-γ-carboxy prothrombin ratio measured by P-11 and P-16 antibodies is a novel biomarker for hepatocellular carcinoma

Serum tumor markers, including α‐fetoprotein (AFP) and des‐γ‐carboxy prothrombin (DCP), are currently used in the diagnosis of hepatocellular carcinoma (HCC). There is, however, an aberrant increase in serum DCP in patients with obstructive jaundice, vitamin K deficiency or who are taking warfarin, resulting from a problem with the current methodology for measurement of this marker. This study aimed to elucidate the utility of a new biomarker, NX‐PVKA, for early diagnosis of HCC. A total of 96 patients were included in the HCC group. The control group included 138 liver cirrhosis (LC) patients without HCC. Serum concentrations of conventional DCP, AFP, AFP‐L3 and NX‐PVKA were measured. The NX‐PVKA ratio was calculated by dividing DCP by NX‐PVKA. In patients not taking warfarin, the area under the curve values of DCP, NX‐PVKA ratio, AFP and AFP‐L3 were 0.715, 0.690, 0.737 and 0.654, respectively, confirming the clinical utility of these markers in detecting HCC. In cases with DCP > 35 mAU/mL in particular, a significant increase in the NX‐PVKA ratio was observed in patients with HCC. In those cases, the cut‐off value for the NX‐PVKA ratio that was optimized by the receiver operating characteristic (ROC) curve was 1.15. In addition, the sensitivity and specificity for diagnosing HCC were 69.2% and 75.9%, respectively. Patients with HCC had higher NX‐PVKA ratios compared to patients with LC taking warfarin (P = 0.063). These results suggest that, when used in combination with DCP, the NX‐PVKA ratio is a promising novel marker for the detection of HCC.

Human β-defensin-3 inhibits migration of colon cancer cells via downregulation of metastasis-associated 1 family, member 2 expression. Corrigendum in /ijo/46/4/1858

The innate immune system plays an important role as the first line of defense against many types of microbes. Accumulating reports suggest that human β-defensins (hBDs) are expressed by and have certain roles in some cancer cells. In this study, we investigated the roles of hBD-3 in colon cancer cells. The expression of hBD-3 was examined by reverse transcriptase-polymerase chain reaction analysis of colon cancer cell lines and immunohistochemical staining of colon cancer tissues. The effect of hBD-3 on proliferation of colon cancer was assessed using the MTT assay and a real-time cell analyzer, and the effect of hBD-3 on the migration of colon cancer cells was also examined. The results showed that hBD-3 is not expressed in colon cancer cells but is produced by tumor-infiltrating monocytes. Migration of colon cancer cells was significantly inhibited by hBD-3 in a dose-dependent manner, although proliferation of colon cancer cells was not affected by administration of hBD-3. Moreover, reduced expression of metastasis-associated 1 family, member 2 (MTA2) mRNA in colon cancer cells was associated with exposure to hBD-3. In conclusion, progression of colon cancer was inhibited by hBD-3 in a paracrine fashion. Therefore, hBD-3 may be a potent new agent for treating colon cancer.

Resveratrol sensitizes HepG2 cells to TRAIL-induced apoptosis.

Resveratrol is a natural polyphenol found in a wide variety of plants, including grapes, berries, and peanuts. Resveratrol can modulate a wide spectrum of molecular targets, including those involved in cancer signaling pathways. Here, we evaluated the role of resveratrol in tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and examined the molecular mechanisms in the human hepatocellular carcinoma cell line HepG2. We used the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay to assess cell viability, flow cytometry to analyze cell cycle and apoptosis, and immunoblotting to detect protein expression. Resveratrol decreased cell viability at a concentration of 100 &mgr;mol/l or higher. At a concentration of 50 &mgr;mol/l, resveratrol induced S phase arrest of the cell cycle without apoptosis. In addition, phospho-AMPK increased significantly in a dose-dependent manner. Resveratrol was found to synergistically augment TRAIL-induced apoptosis. The rates of early apoptosis were 3.4, 9.6, and 49.6% on treatment with 50 &mgr;mol/l resveratrol, 10 ng/ml TRAIL, and both reagents, respectively. Resveratrol significantly downregulated the expression of survivin in a dose-dependent manner. In conclusion, we found that that resveratrol could augment TRAIL sensitivity by downregulating survivin. These results suggest that combination resveratrol with TRAIL may be an effective new strategy for the treatment of hepatocellular carcinoma.

The long-term effects of splenectomy and subsequent interferon therapy in patients with HCV-related liver cirrhosis

Partial splenic embolization (PSE) or splenectomy is widely performed to increase platelet counts for interferon (IFN) therapy. The aim of the present study was to evaluate the long-term effects of splenectomy and subsequent IFN therapy in patients with hepatitis C virus (HCV)-related liver cirrhosis (LC). The present study included 19 patients with HCV-related LC who underwent splenectomy for thrombo-cytopenia caused by hypersplenism. IFN therapy was performed in all 19 patients. The effects of splenectomy and subsequent IFN therapy on peripheral blood counts, liver function, carcinogenesis and survival rates were evaluated. Splenectomy was safely performed in all patients without major complications with the exception of portal thrombosis, which, however, it did not affect liver function when treated appropriately. Thrombocytopenia improved and IFN therapy could be performed in all the patients. A sustained virological response (SVR) was not observed in patients with genotype 1 although it was observed in 75% of patients with genotype 2. Due to severe side effects, five patients did not undergo scheduled IFN therapy. Over 5 years, the mean platelet number increased from 5.2 x 10(4) to 16.8 x 10(4)/mm3 (P<0.01) and liver function improved following splenectomy (albumin, Alb: 3.5‑3.8 g/dl; total bilirubin, T-Bil: 1.0‑0.7 mg/dl; prothrombin time, PT: 74.1‑97.7%; total cholesterol; T-cho: 140‑168 mg/dl; P<0.05). Hepatocellular carcinoma (HCC) occurred in only one patient during long‑term observation and follow‑up of the patients not presenting with HCC at entry. The results of the present study demonstrate that splenectomy followed by interferon therapy could be beneficial in patients with HCV-related LC.

Epigenetic regulation of proliferation and invasion in hepatocellular carcinoma cells by CBP/p300 histone acetyltransferase activity

Altered epigenetic control of gene expression plays a substantial role in tumor development and progression. Accumulating studies suggest that somatic mutations of CREB binding proteins (CBP)/p300 occur in some cancer cells. CBP/p300 possess histone acetyltransferase (HAT) activity, and are involved in many cellular processes. In this study, we investigated the expression and functional role of CBP/p300 in hepatocellular carcinoma (HCC) using the specific inhibitor C646 of CBP/p300 HAT activity. We examined its effect on several apoptosis-related proteins and invasion-related genes. The results showed that CBP/p300 were highly expressed in HCC tissues and that expression of p300, but not of CBP, was strongly correlated with the malignant character of HCC. C646 inhibited proliferation of HCC cell lines in a dose dependent manner. C646 significantly augmented TRAIL-induced apoptotic sensitivity, which was accompanied by reduced levels of survivin, in HepG2, HLE and SK-HEP1 cells. C646 significantly inhibited invasion of Huh7, HLE and SK-HEP1 cells. The level of matrix metallopeptidase 15 (MMP15) mRNA expression was significantly reduced, whereas the level of laminin alpha 3 (LAMA3) and secreted phosphoprotein 1 (SPP1) mRNA expression was significantly increased in Huh7 cells following exposure to C646. In conclusion, our results suggest that CBP/p300 HAT activity has an important role in malignant transformation, proliferation, apoptotic sensitivity and invasion in HCC. CBP/p300 could be a promising therapeutic target in HCC.

Magnetic resonance laparoscopy: A new non‐invasive technique for the assessment of chronic viral liver disease

Laparoscopy‐guided liver biopsy is the most accurate method for assessing liver fibrosis but have several limitations. We designed a non‐invasive method, called magnetic resonance laparoscopy (MRL), based on gadolinium‐ethoxybenzyl‐diethylenetriamine pentaacetic acid‐enhanced magnetic resonance imaging, to assess liver fibrosis in patients with chronic hepatitis B and C virus.

Changes in liver function and body composition by direct-acting antiviral therapy for hepatitis C virus infection.

Management of low skeletal muscle mass (LSM) is a very important topic as LSM affects patient mortality in liver diseases. Changes in body composition are unexplored in chronic hepatitis C virus (HCV) patients, including those with liver cirrhosis, who receive direct‐acting antiviral (DAA) therapy. Body composition measurements and liver function tests were carried out before and after DAA therapy.

Ratio between estimated glomerular filtration rates of creatinine and cystatin C predicts overall survival in patients with hepatocellular carcinoma: Creatinine/cystatin predicts survival in HCC

Hepatocellular carcinoma (HCC) patients with sarcopenia have a poor survival, but there are no predictive markers for survival relating to muscle mass and liver function. Therefore, we investigated whether the ratio between estimated glomerular filtration rates of serum creatinine (Scre) and serum cystatin C (Scys) (eGFRcre/eGFRcys) can be used as a predictive marker of survival in HCC patients.

Su2049 Radiofrequency Ablation Therapy for the Treatment of Metastatic Lung Tumor From Primary Hepatocellular Carcinoma

Background: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide. The prevalence of HCC varies widely in different regions of the world. The impact of specific causes of chronic liver disease to HCC prevalence is well-established. In contrast, the contribution of the underlying genetic architecture to the risk of HCC remains undefined. Genome wide association studies have identified genetic polymorphisms associated with increased susceptibility to HCC. Our aim was to characterize evolutionary trends in the genetic profile of HCC. We hypothesized that negative selection would reduce susceptibility to genetic polymorphisms that increase risk of HCC. Methods: SNPs identified from genome wide association studies with an increased risk of HCC associated with HBV (rs17401966) or HCV (rs9275572, rs2596542) were analyzed. Genetic risk was determined from the risk odds ratio and the frequency of the risk alleles across 53 indigenous human populations across all continents using the Human Genome Diversity Project and ALFRED databases. The fixation index (Fst ) was used to describe the degree of population differentiation (low 1.5) as a measure of gene flow and similarity across subpopulations. A genetic risk score for HCC was determined based on the combined risk of five SNPs associated with increased risk of HCC [rs1800562, rs2596542, rs2267716, rs9275572 and rs17401966]. The frequency of each risk allele and the genetic risk score were correlated with geographic location and with temporal and spatial patterns of human migration. Results: A moderate increase in differentiation was noted for rs2596542 (Fst = 0.106) and rs17401966 (Fst =0.116). Both of these SNPs show an increase in allelic frequency with the most recent human migrations from East Asia, to Oceania and the Americas. In contrast, rs9275572 lacked differentiation (Fst=0.09) with stable allelic expression across populations. The percentile genetic risk score for HCC was greatest in populations from Africa (e.g. Mbuti pygymies 100, San 97), and decreased with subsequent migration into Europe and Asia through to East Asia (e.g. Lahu 1, Yi 1.5). However, a major increase was noted with most recent migrations into Oceania and the Americas (e.g. Karitiana 93.9). Conclusions: Population based studies of genomic variations provide insight into the role of migration on genetic risk for hepatocarcinogenesis. There are differences in directional differentiation of HCC risk alleles across human populations, indicating that susceptibility to HCC can be modified by factors other than genetic drift. Variations in allelic frequency of HCC associated SNPs can contribute to population-based differences in HCC prevalence, and need to be considered for developing regional strategies for screening and surveillance for HCC.