Clinical outcomes of endoscopic resection for nonampullary duodenal high-grade dysplasia and intramucosal carcinoma.

This study retrospectively analyzed the clinical outcomes of endoscopic resection of 26 sporadic (i. e., not associated with polyposis syndrome) nonampullary duodenal lesions representing high-grade dysplasia or intramucosal carcinoma (duodenal HGD/IMC) in 23 patients. No severe complications such as perforation were observed, but three cases of delayed bleeding were seen. The use of endoscopic clips significantly decreased the delayed bleeding rate (0/19, 0%) compared with cases in which clips were not used (3/7, 42.9%; P = 0.013, χ2 test). Eighteen lesions (69.2%) were removed by en bloc resection. The follow-up period after resection was 25.5 ± 23.3 months. Two lesions (7.7%) that recurred locally were detected at the first surveillance endoscopy 3 months after resection. These lesions were 22 and 15 mm in size respectively and were resected piecemeal. Endoscopic resection is an effective and safe procedure for treating duodenal HGD/IMC. En bloc resection and prophylactic clip usage are encouraged.

Longitudinal changes of the laboratory data of chronic hepatitis C patients with sustained virological response on long-term follow-up.

There is no study that follows up longitudinal changes in laboratory data of patients with C-viral chronic liver disease (C-CLD) who achieved sustained virological esponse (SVR) with interferon treatment in a long-term study. We investigated the laboratory data in a long-term retrospective cohort study of 581 patients with C-CLD who underwent liver biopsy between January 1986 and December 2005. 467 were treated with interferon and 207 of these patients achieved SVR with follow-up periods of 8.36 ± 5.13 years. Alanine aminotransferase (ALT) levels, albumin levels, platelet counts, and the aspartate aminotransferase (AST)-to-platelet ratio index (APRI) values were serially examined during the follow-up period. None of the 207 patients with SVR exhibited hepatitis C virus (HCV) RNA positivity more than 6 months after the end of IFN treatment. Platelet counts and albumin levels increased only in those with eradication of HCV. APRI values decreased more in patients with SVR than in those with nonsustained virological responses (non-SVR). Patients who achieved SVR and had fibrosis stage 0-1 and 2-4 at enrolment had platelet counts that longitudinally increased by 2.81 ± 3.95 and 5.49 ± 4.53 × 10(3) /µL during the 10-year follow-up period, respectively. Albumin levels continuously increased during the first 2 years by 0.15 ± 0.31 and 0.33 ± 0.37 in fibrosis stage 0-1 and 2-4, respectively and then plateaued. ALT levels decreased rapidly one year after the start of treatment by 110.3 ± 140.0 and 100.5 ± 123.4 in fibrosis 0-1 and 2-4, respectively. HCV RNA negativity persisted in all patients with SVR, and laboratory data including APRI longitudinally improved during the long-term follow-up period.

Clinical importance of serum hepatitis B surface antigen levels in chronic hepatitis B.

Quantitative serology for hepatitis B surface antigen (HBsAg) is a new candidate marker for prediction of clinical outcome. The aim of this study was to investigate the clinical significance of quantifying HBsAg in patients with hepatitis B virus (HBV) infection. A total of 424 patients who tested positive for HBsAg and were referred to Chiba University Hospital between January 1985 and April 2008 were included in the study, and the following characteristics were analyzed: age, gender, status of hepatitis B e antigen (HBeAg), alanine aminotransferase level (ALT), HBV DNA level, number of platelets and development of hepatocellular carcinoma. Measurement of HBsAg was performed using the chemiluminescent enzyme immunoassay method. The study group consisted of 239 men and 185 women, and their average age was 40.6 ± 14.0 years. HBsAg showed a positive correlation with HBV DNA level (Pearson's product moment correlation, r = 0.586, P < 0.001) and a weak inverse correlation with age (r = 0.3325, P < 0.001). A control study, matched with age and sex, was performed between two groups with and without HBeAg seroconversion during follow-up period. Compared with the age and sex-matched controls, the change in HBsAg levels per year showed a significant decrease 2 years before seroconversion (paired t-test, P < 0.05). The serial measurement of quantitative HBsAg level has the possibility of predicting the occurrence of HBeAg seroconversion.

Hepatitis A virus (HAV) proteinase 3C inhibits HAV IRES-dependent translation and cleaves the polypyrimidine tract-binding protein.

Hepatitis A virus (HAV) infection is still an important issue worldwide. A distinct set of viruses encode proteins that enhance viral cap-independent translation initiation driven by an internal ribosome entry site (IRES) and suppress cap-dependent host translation. Unlike cytolytic picornaviruses, replication of HAV does not cause host cell shut off, and it has been questioned whether HAV proteins interfere with its own and/or host translation. HAV proteins were coexpressed in Huh-7 cells with reporter genes whose translation was initiated by either cap-dependent or cap-independent mechanisms. Among the proteins tested, HAV proteinase 3C suppressed viral IRES-dependent translation. Furthermore, 3C cleaved the polypyrimidine tract-binding protein (PTB) whose interaction with the HAV IRES had been demonstrated previously. The combined results suggest that 3C-mediated cleavage of PTB might be involved in down-regulation of viral translation to give way to subsequent viral genome replication.

Inhibition of subgenomic hepatitis C virus RNA in Huh-7 cells: ribavirin induces mutagenesis in HCV RNA.

Hepatitis C virus (HCV) infection is a major problem throughout the world. Combination therapy of interferon (IFN) and ribavirin is the best treatment for eradication at present, but the mechanism is not completely understood. We used the HCV replicon system to investigate this mechanism. The effects of six drugs (UDCA, glycyrrhizin, TJ-9, bezafibrate, ribavirin, and alpha-IFN 2b) on HCV subgenomic RNA (genotype 1b, NS5B 415Y) were examined by reverse transcription polymerase chain reaction, cloning and sequencing. The HCV replication was inhibited by alpha-IFN 2b (7.39-13.2% at 10 U/mL, 3.29-6.12% at 100 U/mL, 1.3-4.86% at 1000 U/mL) and by ribavirin (4.36-13.9% at 100 microg/mL), but not by the other drugs at 24-72 h after treatment. Furthermore, the combination treatment was superior to IFN monotherapy and to ribavirin monotherapy at 72 h post-treatment. Sequence analyses of the double-stranded RNA-activated protein kinase (PKR)-binding domain and flanking regions within the HCV NS5A region revealed that the total numbers of substitutions caused by ribavirin (n = 36) or combination treatment (n = 57) were more than those of IFN alone (n = 5) and controls (n = 6). The HCV replicon system is the most efficient system for HCV replication and is an excellent choice for testing anti-HCV drugs and disinfectants. Our results further suggested that the combination of alpha-IFN 2b and ribavirin might induce mutations, and inhibit HCV RNA synthesis in hepatocytes to a greater extent than ribavirin monotherapy.

Hepatitis B virus X protein (HBx)-induced apoptosis in HuH-7 cells: influence of HBV genotype and basal core promoter mutations.

BACKGROUND: Hepatitis B virus (HBV) infection is a serious, world-wide problem. HBV genotype and basal core promoter (BCP) mutations affect the clinical course of HBV-infected patients. BCP mutations also lead to mutations at HBV X protein (HBx) codons 130/131. The functional significance of naturally occurring variants of human HBx remains largely unknown. The purpose of the study was to investigate whether HBV genotypes or double mutations affect HBx-induced apoptosis. METHODS: We constructed genotype A, B, C, and D HBx expression vectors and HBx expression vectors with double mutations at HBx codons 130K and 131V or positions 130M and 131I using site-directed mutagenesis. A transient expression system in HuH-7 cells was established and this model was utilized to address the effect of HBx on cell viability. RESULTS: HBx-transfected cells showed a dose-dependent decrease in cell viability by MTS assay. A subset of cells expressing HBx underwent apoptosis according to terminal transferase enzyme-mediated end labeling of DNA and caspase-3 activity. This study demonstrated that HBx can induce cell death by apoptosis in a dose-dependent manner and that HBV genotypes and double mutations did not affect HBx-induced apoptosis. CONCLUSIONS: HBV genotypes and mutation of two amino acids directly adjacent to the conserved Kunitz domain essential for transcription activating activity of HBx did not change the pro-apoptotic activity of HBx. Further study is needed to determine whether HBV genotypes and double mutations have any effect on the function of HBx.

Hepatitis A virus VP3 may activate serum response element associated transcription.

BACKGROUND: Hepatitis A virus (HAV) infection is a major public health problem worldwide. The infection does not induce any visible cytopathic effects or interfere with macromolecular synthesis in host cells. However, the hepatitis B and C viruses have recently been reported to activate intracellular signals. To clarify the effects of HAV infection on intracellular signalling, we examined the influence of 9 FLAG-tagged HAV proteins (VP2, VP3, VP1-2A, 2B, 2C, 3A, 3BC, 3C and 3D) on signal transduction pathways. METHODS: Viral protein expression vectors were co-transfected into HeLa cells with reporter plasmids controlled by a synthetic promoter containing direct repeats of the cyclic AMP response element (CRE), serum response factor (SRF), activator protein 1 (AP-1), nuclear factor kappaB (NF-kappaB) or serum response element (SRE). Cells were harvested 42 h after transfection and luciferase assays were performed. Viral protein activation twice that of the control was defined as significant. RESULTS: VP3 induced an SRE-associated signal 2.2 +/- 0.3 times higher than that of control. VP3 did not activate CRE-, SRF-, AP-1- or NF-kappaB- associated signalling. The other HAV proteins tested also failed to induce these pathways. CONCLUSIONS: HAV interacts with the host signalling mechanism, and HAV VP3, different from HBX and hepatitis C core protein, may activate only SRE-associated intracellular signalling, a pathway associated with cell proliferation and differentiation.

Analysis of full-length hepatitis A virus genome in sera from patients with fulminant and self-limited acute type A hepatitis.

BACKGROUND/AIMS: Type A hepatitis still poses a considerable problem worldwide. Why some patients progress to fulminant type A hepatitis and others do not is still unknown. To examine whether genomic differences of hepatitis A virus (HAV) are responsible for the severity of the disease, we analyzed the whole HAV genomes from patients with fulminant and self-limited acute type A hepatitis. METHODS: Sera from three patients with sporadic type A fulminant hepatitis (FH) and three patients with acute hepatitis (AH) were examined for HAV RNA. Full-length nucleotide sequences were determined using long reverse transcription polymerase chain reaction, 5' and 3' rapid amplification of cDNA ends methods, and direct sequencing. The amino acid sequences were deduced from the nucleotide sequences. RESULTS: HAV RNA was detected in all six patients examined. From the sequence of viral protein 1/2A, all cases were revealed to be genotype IA. By comparing with genotype IA, wild-type HAV strain GBM, the analysis of whole genomes from the six cases showed no specific substitutions between FH and AH. Completely identical nucleotide sequences were observed at 3' non-translated region (NTR) in all six cases. In 5'NTR, less nucleotide substitutions were found in FH than in AH, and in the non-structural protein 2B region, a little more amino acid substitutions seemed to be found in FH than in AH. CONCLUSIONS: This study showed that full-length HAV could be analyzed from serum samples. Although there were no unique nucleotide or amino acid substitutions, possible associations were suggested between the severity of type A hepatitis and the nucleotide substitutions in 5'NTR and the amino acid substitutions in 2B.

State of HBV DNA in HBsAg-negative, anti-HCV-positive hepatocellular carcinoma: existence of HBV DNA possibly as nonintegrated form with analysis by Alu-HBV DNA PCR and conventional HBV PCR.

The role of hepatitis B virus (HBV) in carcinogenesis of hepatitis B surface antigen (HBsAg)-negative, anti-hepatitis C virus (anti-HCV)-positive hepatocellular carcinoma (HCC) remains unknown. To investigate the state of HBV DNA in such HCC, HBV DNA was examined by polymerase chain reaction (PCR) between HBV DNA and human Alu sequence (HBV-Alu PCR), which could detect integrated form of HBV DNA only, and by conventional HBV PCR, which could detect both integrated and episomal forms of HBV DNA. In all the 17 HBsAg-positive HCC, HBV DNA was detected by both HBV-Alu PCR method and conventional HBV PCR method. By contrast, in HBsAg-negative, anti-HCV-positive cases, HBV DNA was detected in 10 of 21 (47.6%) by conventional HBV PCR and in none of 21 (0%) by HBV-Alu PCR method. Thus, integrated form of HBV DNA was not found in most HbsAg-negative, anti-HCV-positive HCC in the current study. The role of episomal form of HBV DNA requires further investigation of its involvement in the process of the development of HBsAg-negative, anti-HCV-positive HCC.

Sequence-motif analysis of 5'-untranslated region of GB virus-C in Japanese patients.

BACKGROUND: GB Virus C (GBV-C) is considered to belong to the Flaviviridae; however, the structures of the N-terminal end of its putative polyprotein are not well known. The internal ribosomal entry site (IRES) at the 5'-untranslated region of GBV-C and an initiating codon at nucleotides (nt) 552-554 have been proposed. We investigated the validity of this proposal. METHODS: The 5'-untranslated region of GBV-C was amplified from serum samples of 17 Japanese patients. Polymerase chain reaction-amplified products were directly sequenced and the obtained sequences were analysed by comparing them with the IRES structure of other viruses. RESULTS: Fifteen of the 17 (88%) GBV-C strains in our patients were classified as being Asian type. The box-A-like sequence (UUUC) and box-B-like sequence (AUCAUGG) observed in the IRES of picornaviruses were highly conserved in all the strains. Based on pair-wise comparisons with the multiple alignment data, overall sequence divergence for the 5'-terminus was 2.9-12%. When compared with the proposed secondary structure of the IRES model, the sequence divergences of the Asian-type GBV-C were higher at the regions of loop structures and lower at the regions of double-stranded RNA. The AUG codons, except for the one located at nt 552-554, produced truncated polyproteins or were not in-frame with the putative protein. CONCLUSIONS: Our examination of the sequence motif of GBV-C supports the proposal that the GBV-C has common structural motifs for IRES at its 5'-untranslated region and the AUG codon at nt 552-554 may be an initiating codon.

Different turnover rate of hepatitis C virus clearance by different treatment regimen using interferon-beta.

BACKGROUND/AIM: Since patients with high viral load and HCV subtype 1b are known to respond poorly to interferon (IFN) therapy, the viral dynamics of HCV RNA after initiation of interferon therapy were examined in the present study with respect to two different administration regimens, once vs. twice a day. METHODS: Twenty-two patients with chronic hepatitis C confirmed by liver biopsy and with >1 Meq/ml of HCV RNA and HCV subtype 1b were randomly assigned to two different IFN administration regimens (6 million units of IFN once a day or 3 million units of IFN twice a day), and the serum HCV RNA level was serially measured. RESULTS: Graphs of HCV RNA levels vs. treatment time showed an initial rapid fall, followed by a slower clearance phase. Fitting the data to a model for HCV decay proposed by Neumann et al. showed that the treatment efficacy was significantly higher with twice daily administration. Negativity for HCV RNA measured by Amplicor assay in the twice-a-day administration group was 18%, 73% and >89% at 1, 2 and 3 weeks, respectively, in contrast to 0%, 0%, and 18%, respectively, with once-a-day administration. However, a significant reduction of platelet count and albumin level, a marked increase in serum aspartate aminotransferase/alanine aminotransferase, and a high incidence of renal toxicity (proteinuria) were found in patients receiving IFN twice a day in comparison with those receiving it once a day. CONCLUSION: The twice-a-day administration of IFN accelerated the clearance of HCV RNA from serum, leading to a more efficient virological response for patients with chronic hepatitis C, but with a high rate of renal toxicity.

Histologic improvement of fibrosis in patients with hepatitis C who have sustained response to interferon therapy.

BACKGROUND: Short-term histologic improvement in hepatitis C-related hepatic fibrosis has been noted in studies with more than 2 years of follow-up, but the long-term effects of interferon therapy on hepatic fibrosis remain unclear. OBJECTIVE: To assess changes in hepatic fibrosis after interferon therapy in patients with chronic hepatitis C. DESIGN: Retrospective cohort study. SETTING: 7 university hospitals and 1 national hospital in Japan. PATIENTS: 593 patients with chronic hepatitis C who underwent a paired liver biopsy from 1987 to 1997. Of these, 487 patients received interferon therapy and 106 patients were untreated. INTERVENTION: Patients in the treatment group received a 2- to 6-month course of interferon within 6 months after the initial biopsy. MEASUREMENTS: Fibrosis and inflammatory activity in paired biopsy samples obtained a median of 3.7 years apart (range, 1 to 10 years) were graded by using the criteria of Desmet and colleagues (F0 to F4) and those of the French METAVIR Cooperative Study Group (A0 to A3), respectively. Changes in fibrosis staging and activity scores and yearly rates of fibrosis progression and regression were calculated. RESULTS: 183 of the 487 interferon-treated patients showed a sustained virologic response. Activity grade was unchanged in most of the untreated patients and improved in 89% (CI, 83% to 93%) of patients with a sustained virologic response. A sustained response to interferon was associated with a mean (+/-SE) reduction in fibrosis score of -0.60+/-0.07 at less than 3 years of follow-up and -0.88+/-0.08 at 3 years or more of follow-up. The rate of fibrosis progression was -0.28+/-0.03 unit/y (regression) in patients with sustained response, 0.02+/-0.02 unit/y in patients with nonsustained response (P< 0.001), and 0.10+/-0.02 unit/y in untreated patients. CONCLUSION: Although the time between biopsies partly affected the patient's clinical course, the differences observed here suggest that in patients with chronic hepatitis C, regression of fibrosis is associated with sustained virologic response to interferon therapy.

Spontaneous negativation of serum hepatitis C virus RNA is a rare event in type C chronic liver diseases: analysis of HCV RNA in 320 patients who were followed for more than 3 years.

BACKGROUND/AIMS: The natural course of hepatitis C virus (HCV) replication in type C liver diseases has not yet been elucidated. The aim of the study was to investigate the spontaneous outcome of the viremia by examining the changes in HCV RNA in patients with chronic type C liver diseases. METHODS: Among patients who visited our liver clinic between June 1981 and December 1993, 320 patients with chronic type C liver diseases were followed for at least 3 years and had no history of interferon treatment. HCV RNA was examined by a highly specific reverse transcription-polymerase chain reaction method in paired serum samples obtained from these patients at the beginning and end of follow-up. RESULTS: Among the 320 cases, HCV RNA was seropositive in 310 (97%) cases at the beginning of follow-up. Of these 310, HCV RNA remained seropositive in 304 (98%) and became seronegative in six (2%) cases by the end of follow-up. All of these six patients had liver cancer. HCV RNA became seronegative after the patients entered the state of liver failure because of the development of tumors or portal thrombosis. The remaining 10 cases who were seronegative for HCV RNA at the beginning were seropositive at the end of follow-up. Among the 320 cases, serum alanine aminotransferase normalized and remained normal for more than 12 months until the end of follow-up in 11 cases (0.6%/year/case), but none became negative for HCV RNA. CONCLUSIONS: Thus, during the natural course of chronic HCV infection, spontaneous negativation of serum HCV RNA seems extremely rare, at least in patients with chronic active hepatitis or cirrhosis of the liver, and may occur primarily at the terminal stage when tumors cause liver failure.

Quantitative evaluation of telomerase activity in small liver tumors: analysis of ultrasonography-guided liver biopsy specimens.

BACKGROUND/AIMS: Telomerase activity which restores the length of telomere repeat arrays is frequently detectable in various malignancies, including hepatocellular carcinoma. The aim of this study was to determine the diagnostic usefulness of the quantitative measurement of telomerase activity in small liver tumors, which has not yet been established. METHODS: Fifty-eight liver specimens from tumorous and non-tumorous portions of 29 small liver tumors equal to or less than 3.0 cm were obtained by ultrasonography-guided liver biopsy, and of these, 25 were diagnosed as hepatocellular carcinoma and four as adenomatous hyperplasia. The telomerase activities in these specimens and control specimens were examined quantitatively by telomeric repeat amplification protocol with standard control. RESULTS: The mean telomerase activity in cirrhosis without liver tumor was 0.4+/-0.6 (+/-S.D.) arbitrary units (AU) and that in 29 non-tumorous parts of the tumors was 4.5+/-7.4 AU. The mean telomerase activity in 13 tumors equal to or less than 2 cm in diameter was 77.1+/-133.7 AU and that in 16 tumors more than 2 cm was 152.7+/-215.2 AU. The mean telomerase activity in the 4 adenomatous hyperplasias was 5.5+/-4.5 AU; those of hepatocellular carcinoma with Edmondson-Steiner classification I, II, III and IV were 49.6+/-47.4 (n = 10), 240.1+/-273.6 (n = 9), 119.2+/-174.6 (n = 4) and 144.6+/-80.2 (n = 2) AU, respectively. CONCLUSIONS: The telomerase activity was significantly higher in hepatocellular carcinoma compared to adenomatous hyperplasia and non-neoplastic tissue, indicating that the quantitation of telomerase activity would be useful for the diagnosis of small hepatocellular carcinoma.

Interferon therapy reduces the risk for hepatocellular carcinoma: national surveillance program of cirrhotic and noncirrhotic patients with chronic hepatitis C in Japan. IHIT Study Group. Inhibition of Hepatocarcinogenesis by Interferon Therapy.

BACKGROUND: Previous studies on the effect of interferon therapy on the incidence of hepatocellular carcinoma have not sufficiently assessed degree of liver fibrosis, a major risk factor for hepatocellular carcinoma. OBJECTIVE: To evaluate the effect of interferon therapy on incidence of hepatocellular carcinoma, adjusting for risk factors, including the degree of liver fibrosis. DESIGN: Retrospective cohort study. SETTING: Seven university hospitals and one regional core hospital in Japan. PATIENTS: 2890 patients with chronic hepatitis C who had undergone liver biopsy since 1986. Of these patients, 2400 received interferon and 490 were untreated. MEASUREMENTS: The degree of liver fibrosis was assessed from stage F0 (no fibrosis) to stage F4 (cirrhosis). Response to interferon was determined virologically and biochemically. Screening for development of hepatocellular carcinoma was performed periodically during an average follow-up of 4.3 years. Effect of interferon therapy on the risk for hepatocellular carcinoma was analyzed by using Cox proportional hazards regression. RESULTS: Hepatocellular carcinoma developed in 89 interferon-treated patients and in 59 untreated patients. Among untreated patients, the annual incidence of hepatocellular carcinoma increased with the degree of liver fibrosis, from 0.5% among patients with stage F0 or F1 fibrosis to 7.9% among patients with stage F4 fibrosis. The cumulative incidence in treated and untreated patients differed significantly for patients with stage F2 fibrosis (P = 0.0128) and for those with stage F3 fibrosis (P = 0.0011). In multivariate analysis, interferon therapy was associated with a reduced risk for hepatocellular carcinoma (adjusted risk ratio, 0.516 [95% CI, 0.358 to 0.742]; P < 0.001), especially among patients with sustained virologic response (risk ratio, 0.197 [CI, 0.099 to 0.392]), among those with persistently normal serum alanine aminotransferase levels (risk ratio, 0.197 [CI, 0.104 to 0.375]), and among those with alanine aminotransferase levels less than two times the upper limit of normal (risk ratio, 0.358 [CI, 0.206 to 0.622]). CONCLUSIONS: Interferon therapy significantly reducesthe risk for hepatocellular carcinoma, especially among virologic or biochemical responders.

Detection of mutations in the enhancer 2/core promoter region of hepatitis B virus in patients with chronic hepatitis B virus infection: comparison with mutations in precore and core regions in relation to clinical status.

To investigate the meaning of the mutations in the enhancer 2/core promoter (Enh2/CP) region of hepatitis B virus (HBV) during the chronic HBV infection, mutations were examined in the Enh2/ CP region (carboxyl half of X region) and their correlation with mutations in the precore and core regions in relation to the presence of chronic liver disease. The entire nucleotide sequences of the Enh2/CP region were determined by direct sequencing of the amplified products derived from 30 cases with chronic HBV infection. The results were compared to the mutations in the precore and core regions. In the Enh2/CP region, 91 generally scattered nucleotide substitutions were detected. There were 11 substitutions in the 10 asymptomatic healthy carriers (mean, 1.1/case) and 80 in the 20 chronic liver disease patients (4.0/case). The most frequent substitutions from A to T at nucleotide 1764 and from G to A at nucleotide 1766 were seen in none of the 10 asymptomatic carriers and in 14 (70%) of the 20 chronic liver disease patients. Comparisons of mutations in the precore and core regions revealed that 14 of 16 patients with mutations in the core region had the mutations in the Enh2/CP region and/or a precore stop codon mutation. These data suggest that mutations in the Enh2/CP and precore regions may affect the expression of the core and HBeAg peptides and might be involved in the pathogenesis of chronic liver disease.

[Molecular mechanism of hepatocarcinogenesis].

Hepatocellular carcinoma (HCC) in Japan is closely associated with the chronic liver diseases of infection with the hepatitis B or C viruses. Analysis of HCC tissues frequently detects loss of heterozygosity at chromosomes 1p, 4, 6q, 8p, 10q, 13q, 16q, 17p, and many genomic and epigenomic abnormalities have been found in p53, beta-catenin, p16CDKI, DNA mismatch repair genes, and others. However, no specific abnormal genetic or epigenetic changes for HCC have been found so far. The development of HCC has been reported in mice transgenic for the hepatitis B virus X gene or the hepatitis C virus core gene, and these viral proteins might play essential roles in hepatocarcinogenesis. Chronic hepatitis and fibrosis due to persistent viral infection might also influence the genomic instability of hepatocytes, leading to accumulation of genomic changes.

Expression of p53 and p21WAF1/CIP1 proteins in gastric and esophageal cancers: comparison with mutations of the p53 gene.

The p53 gene has been shown to be commonly mutated in various human cancers, and mutant p53 can act as a dominant oncogene. The intact p53 protein is also known to induce the cyclin-dependent kinase inhibitor p21WAF1/CIP1 and is implicated in cell cycle arrest. We investigated p53 gene alterations in gastric adenocarcinoma and esophageal squamous cell carcinoma to elucidate the association of the nuclear accumulation of the p53 protein and/or p21WAF1/CIP1 protein. Abnormalities of the tumor suppressor gene p53 protein and the expression of p21WAF1/CIP1 protein were analyzed by immunohistochemical techniques in 32 cases of gastric adenocarcinoma and 15 cases of esophageal squamous cell carcinoma. Twenty cases of gastric cancer and five cases of esophageal cancer were also analyzed for p53 gene mutation by polymerase chain reaction and direct nucleotide sequencing. Overexpression of p53 protein was found in 13/32 (41%) of gastric cancers and 5/15 (33%) of esophageal cancers. We found immunodetectable p53 in 10/14 cases with mutations and in none of 11 cases without mutations in gastric and esophageal cancers. Hence, immunohistochemical and genetic analyses gave concordant results in 84% of 25 cases, revealing a good correlation between immunostaining of p53 and missense mutation of the p53 gene. p53 immunostaining was not observed in cases with frameshift or splicing mutation. The expression of p21WAF1/CIP1 protein was found in 9/32 (29%) of gastric cancers and 4/15 (27%) of esophageal cancers and in 2/14 (14%) cases with alteration of the p53 gene and in 5/11 (45%) without. These results suggest that abnormalities of p53 may be closely associated with the pathogenesis of gastric adenocarcinoma and esophageal squamous cell carcinoma and that the immunoreactivity of p53 protein is a general indicator of the tumors with altered p53 function. The expression of p21WAF1/CIP1 protein was suppressed in the neoplastic tissues with and without p53 gene alteration.

GB virus-C RNA in Japanese patients with hepatocellular carcinoma and cirrhosis.

BACKGROUND/AIMS: The involvement of non-B, non-C virus in the incidence of hepatocellular carcinoma (HCC) is not yet known. We have therefore examined the occurrence of GBV-C RNA in such patients. METHODS: One hundred and eleven patients diagnosed as having HCC and 67 patients with cirrhosis without HCC were examined for the prevalence of GBV-C RNA by nested reverse transcription polymerase chain reaction with primers located at the helicase region. Sera were obtained and kept at -20 degrees C until analysis. RESULTS: GBV-C RNA was positive in 11/111 (9.9%) cases with HCC, in 10/74 (13.5%) anti-HCV positive cases, in 1/25 (4%) HBsAg positive cases, and in 0/8 (0%) anti-HCV and HBsAg negative cases. GBV-C RNA was also positive in 7/67 (10.4%) cases with cirrhosis, in only 1/18 (5.6%) anti-HCV and HBsAg negative cases, in 4/33 (12.1%) anti-HCV positive, and in 2/14 (14.3%) HBsAg positive cases. The clinical background of patients with anti-HCV positive HCC who were also positive for GBV-C RNA did not differ from the background of those negative for GBV-C RNA. CONCLUSIONS: GBV-C is unlikely to be a major etiologic agent of non-B, non-C chronic liver diseases and HCC in Japan.

Telomerase activity and telomere length in hepatocellular carcinoma and chronic liver disease.

BACKGROUND & AIMS: The maintenance of the telomere to a certain length is considered to be vital for cellular immortality. Recently, the presence of telomerase, an enzyme that elongates telomere length, was reported in various malignancies. The aim of this study was to characterize telomerase activity and telomere length in hepatocellular carcinoma and chronic liver diseases to facilitate better understanding and more accurate diagnosis of hepatocellular carcinoma. METHODS: In tumorous and nontumorous tissues from 26 hepatocellular carcinomas and from 20 liver tissues without overt hepatocellular carcinoma, telomerase activity was examined by telomeric repeat amplification protocol and telomere length was detected by Southern blot hybridization method. RESULTS: Telomerase activity was detected in 22 of 26 (85%) hepatocellular carcinoma specimens. In contrast, it was weakly detected in 4 of 46 (9%) nonneoplastic tissues. Telomere length in hepatocellular carcinoma was shorter than that of corresponding nontumorous tissue in 44% and longer in 17%. CONCLUSIONS: The presence of telomerase activity was confirmed in a majority of cases with hepatocellular carcinoma, and alteration of telomere length from nonneoplastic tissue was observed in approximately two thirds of hepatocellular carcinomas. Therefore, these markers might be good indicators for the diagnosis of hepatocellular carcinoma.