Yoko Yamagiwa

Characterization of the epithelial cell adhesion molecule (EpCAM)+ cell population in hepatocellular carcinoma cell lines

Accumulating evidence suggests that cancer stem cells (CSC) play an important role in tumorigenicity. Epithelial cell adhesion molecule (EpCAM) is one of the markers that identifies tumor cells with high tumorigenicity. The expression of EpCAM in liver progenitor cells prompted us to investigate whether CSC could be identified in hepatocellular carcinoma (HCC) cell lines. The sorted EpCAM+ subpopulation from HCC cell lines showed a greater colony formation rate than the sorted EpCAM− subpopulation from the same cell lines, although cell proliferation was comparable between the two subpopulations. The in vivo evaluation of tumorigenicity, using supra‐immunodeficient NOD/scid/γcnull (NOG) mice, revealed that a smaller number of EpCAM+ cells (minimum 100) than EpCAM− cells was necessary for tumor formation. The bifurcated differentiation of EpCAM+ cell clones into both EpCAM+ and EpCAM− cells was obvious both in vitro and in vivo, but EpCAM− clones sustained their phenotype. These clonal analyses suggested that EpCAM+ cells may contain a multipotent cell population. Interestingly, the introduction of exogenous EpCAM into EpCAM+ clones, but not into EpCAM− clones, markedly enhanced their tumor‐forming ability, even though both transfectants expressed a similar level of EpCAM. Therefore, the difference in the tumor‐forming ability between EpCAM+ and EpCAM− cells is probably due to the intrinsic biological differences between them. Collectively, our results suggest that the EpCAM+ population is biologically quite different from the EpCAM− population in HCC cell lines, and preferentially contains a highly tumorigenic cell population with the characteristics of CSC. (Cancer Sci 2010)

Translational Regulation of X-Linked Inhibitor of Apoptosis Protein by Interleukin-6 A Novel Mechanism of Tumor Cell Survival

Interleukin-6 (IL-6) is a pleiotropic cytokine with diverse biological effects. IL-6 has been implicated in autocrine signaling pathways promoting tumor progression and chemoresistance in some human tumors. However, the mechanisms by which IL-6 modulates these responses are unknown. Aberrant apoptosis has been implicated as a fundamental mechanism of chemotherapeutic resistance. Thus, we investigated whether IL-6 alters the expression of apoptosis regulatory proteins as a mechanism of drug resistance. We provide evidence that IL-6 rapidly phosphorylates the translation initiation factor eukaryotic initiation factor-4E and triggers antiapoptotic responses in cholangiocarcinoma cells. Reduction of cellular eukaryotic initiation factor-4E by RNA interference decreases IL-6-induced effects on cytotoxic drug-induced caspase activation and apoptosis. Furthermore, IL-6 increases expression of the endogenous X-linked inhibitor of apoptosis protein expression by translation at an internal ribosome entry site. Our findings that IL-6 translationally regulates X-linked inhibitor of apoptosis protein expression reveal a novel mechanism by which IL-6 mediates tumor cell survival that may be targeted therapeutically to decrease tumor progression and chemoresistance.

Expression of galectin-3 involved in prognosis of patients with hepatocellular carcinoma.

Aims:  Galectins are multifunctional lectins binding to the β‐galactoside of glycoproteins that affect diverse physiological and pathophysiological processes such as development, inflammation and tumor growth. In hepatocellular carcinoma (HCC), the over‐expression of galectin‐1, 3, and 4 has been reported, although their function and correlation with tumor progression remain unknown. Thus, we aimed to assess the role of galectin‐3 during HCC progression.

Four‐year study of lamivudine and adefovir combination therapy in lamivudine‐resistant hepatitis B patients: influence of hepatitis B virus genotype and resistance mutation pattern

Summary.  To investigate the efficacy of long‐term lamivudine (3TC) and adefovir dipivoxil (ADV) combination therapy in 3TC‐resistant chronic hepatitis B virus (HBV) infected patients, we analysed 28 3TC‐resistant patients treated with the combination therapy during 47 months (range, 9–75). At 12, 24, 36, and 48 months, the rates of virological response with undetectable HBV DNA (≤2.6 log copies/mL) were 56, 80, 86, and 92%, respectively. Among 17 hepatitis B e antigen (HBeAg)‐positive patients, HBeAg disappeared in 24% at 12 months, 25% at 24 months, 62% at 36 months, and 88% at 48 months. When HBV genotypes were compared, patients with genotype B achieved virological response significantly more rapidly than those with genotype C (P = 0.0496). One patient developed virological breakthrough after 54 months, and sequence analysis of HBV obtained from the patient was performed. An rtA200V mutation was present in the majority of HBV clones, in addition to the 3TC‐resistant mutations of rtL180M+M204V. The rtN236T ADV‐resistant mutation was observed in only 25% clones. In vitro analysis showed that the rtA200V mutation recovered the impaired replication capacity of the clone with the rtL180M+M204V mutations and induced resistance to ADV. Moreover, rtT184S and rtS202C, which are known entecavir‐resistant mutations, emerged in some rtL180M+M204V clones without rtA200V or rtN236T. In conclusion, 3TC+ADV combination therapy was effective for most 3TC‐resistant patients, especially with genotype B HBV, but the risk of emergence of multiple drug‐resistant strains with long‐term therapy should be considered. The mutation rtA200V with rtL180M+M204V may be sufficient for failure of 3TC+ADV therapy.

Lack of evidence that bone marrow cells contribute to cholangiocyte repopulation during experimental cholestatic ductal hyperplasia

Abstract: Background: Ductopenia is observed in end‐stage human cholestatic diseases. The limited capability of cholangiocytes for proliferation is suggested to be the principal reason. Recently, bone marrow cells (BMCs) have been reported to behave as hepatic stem cells; however, their capability to differentiate into cholangiocytes in cholestasis remains unclear.

The model for end-stage liver disease score is useful for predicting economic outcomes in adult cases of living donor liver transplantation

BackgroundThe model for end-stage liver disease (MELD) is useful for assessing the recipients of liver transplants, namely, deceased-donor transplantation. The application of MELD for living donor liver transplantation (LDLT) is under investigation. Thus, the aim of this study was to analyze the impact of the MELD score in LDLT in Japan.MethodsSeventeen adult cases of LDLT during 2001 to 2005 were enrolled. Indications for LDLT were primary biliary cirrhosis, seven; liver cirrhosis, two; hepatocellular carcinoma (HCC), three; metabolic liver disease, one; primary sclerosing cholangitis, two; Carolis disease, one; and biliary atresia, one. Total medical charges during the operative periods were retrospectively evaluated. The united network of organ sharing (UNOS) modified was obtained using preoperative clinical data.ResultsThe average medical expense of the 17 cases was approximately

Amniotic epithelial cell-derived cholangiocytes in experimental cholestatic ductal hyperplasia

97 901. The UNOS-modified MELD score was 22.1. A statistically significant positive correlation was found between the MELD score and medical expense (P = 0.0086, ρ = 0.657), and between the MELD score and the length of stay in the intensive care unit (ICU) (P = 0.0396, ρ = 0.515). The cause of the liver disease leading to transplantation was not related to MELD score, medical expense, or length of ICU stay.ConclusionsAlthough not originally designed for the application to LDLT, the MELD score is useful for predicting medical expenses in LDLT. Similar to those of deceased-donor liver transplantation, the disadvantage of high medical expenses associated with a high MELD score allow consideration of an earlier elective operation in suitable cases.

A case of severe recurrent hepatitis with common variable immunodeficiency

Aim:  Bile duct paucity, ductopenia, is a feature of end‐stage chronic cholangiopathies such as primary biliary cirrhosis. The limited proliferative ability of cholangiocytes after specific injury is thought to be the principal cause of ductopenia, although the detailed mechanisms involved are unclear. It has been reported that human amniotic epithelial cells (AEC) express differentiation markers of hepatic parenchymal cells, suggesting a resemblance of AEC to hepatic progenitor cells. The aim of the present study was to develop a mouse model of experimental cholestasis to assess the capability of mouse AEC to trans‐differentiate into cholangiocytes.

Correlation between p21waf1 and p16INK4a expression in hepatocellular carcinoma

Severe hepatitis with an indistinct etiology manifested in a 16‐year‐old boy who had no particular history. The histological features of the liver and clinical course of the patient were similar to those of patients with autoimmune hepatitis characterized by interface hepatitis and severe lobular inflammation of the liver and recurrent exacerbations of hepatitis. We administered intravenous glycyrrhizin preparation daily or three times a week combined with the oral administration of ursodeoxycholic acid daily throughout the term after the initial onset of disease for the control of disease activity. The normalization of the concentration of alanine aminotransferase in serum was achieved in response to the therapy during the course. The serum concentration of immunoglobulins of the patient gradually decreased from the onset of the disease to an unacceptable level without globulin preparation during the following period of 17 months. Immunological tests revealed impairment of immunoglobulin production bythe B cell population of the patient, which led to the diagnosis of the patient as common variable immunodeficiency (CVID). The patient, with improved liver histology after 27 months from the onset of disease, benefited from the current combination therapy without severe infection through the avoidance of overimmunosuppression. CVID is defined as a heterogeneous syndrome characterized by various degrees of hypogammaglobulinemia without any specific predisposing causes, frequently associated with autoimmunity. Diagnostic criteria and therapeutic options of persistent hepatitis with CVID are to be established, as discussed in the current report.

A case of HIV co-infected with hepatitis B virus precore/core deletion mutant treated by entecavir.

Abstract Background: The cyclin dependent kinase p21waf1 plays a crucial role in the regulation of cell cycle. The family of p53 proteins has the ability to induce p21waf1, whereas p16INK4a modulates post-transcriptionally the expression of p21waf1. Methods: Total 36 hepatocellular carcinomas (HCCs) and 24 paired adjacent liver tissues were evaluated for the following: (1) expression of p21waf1 and p16INK4a; (2) that of p21waf1, p73 and p63 mRNAs; (3) genomic mutations and the loss of heterozygosity of p73 and p53; and (4) frequency of methylation in the 5′CpG promoter region of p16INK4a. Results: In HCCs compared with the adjacent non-cancerous liver tissues, the expression of p21waf1 and p16INK4a was reduced. Indeed, p21waf1 was not detected in 36% (8/22) of HCCs in spite of the presence of p21waf1 mRNA: among them, mutations of p53 gene were found in 50%, whereas a lack of p16INK4a expression in all of them. p21waf1 and p16INK4a were reduced in proportion to the degree of methylation in p16INK4a gene. p73 did not mutated, and p63 did not expressed in HCCs. Conclusion: Methylation status of p16INK4a gene will play a part for reducing constitutive expression of p16INK4a and of p21waf1 coordinately in HCCs.