中本 晋吾

We describe a patient infected with hepatitis C virus (HCV) genotype 1 who failed to respond to interferon with or without ribavirin seven times but who then achieved sustained virological response by the combination of interferon-beta plus ribavirin treatment for 48 weeks. He did not respond to stronger treatment such as prolonged peginterferon plus ribavirin treatment for 87 weeks. It might be rare to treat a patient with interferon 8 times considering the cost and time, since patients failing interferon treatment several times are regarded to be refractory to interferon. It is possible that the difference in biological effects between interferon-alfa and interferon-beta might have resulted in a different outcome in our patient. Interferon-beta plus ribavirin therapy might be one of the treatment options for certain patients chronically infected with HCV who do not respond to standard care treatment. © Springer 2013.

Alanine aminotransferase (ALT) elevation was occassionally observed during the treatment with combination peginterferon alpha plus ribavirin. Two forms of peginterferon are currently available as a standard of care with or without direct-acting antivirals against hepatitis C virus (HCV). Until the appearance of interferon-sparing regimen, peginterferon alpha plus ribavirin will play a central role in the eradication of HCV. In the present study, we compared ALT elevations in response to peginterferon alpha-2a plus ribavirin or peginterferon alpha-2b plus ribavirin in HCV genotype-1-infected patients. There were no significant differences in ALT elevations between treatments with the two peginterferons, but in a comparison of the proportions of patients with transient ALT elevation from baseline between the two groups, transient ALT elevation was observed more in sustained virological response (SVR) patients treated with peginterferon alpha-2a than with peginterferon alpha-2b. However, no patients discontinued treatment due to ALT elevation. Patients with transient ALT elevation from baseline during the treatment had less favorable IL28B rs8099917 genotype in the peginterferon alpha-2b group. Patients achieving SVR tended to have lower ALT levels, although some had persistent ALT elevation during treatment. In conclusion, clinicians should pay attention to possible ALT elevation during the treatment of chronic hepatitis C patients.

Viral breakthrough is related to poor adherence to medication in some chronic hepatitis B patients treated with nucleos(t)ide analogues (NAs). Our study aimed to examine how adherence to medication is associated with viral breakthrough in patients treated with NAs. A total of 203 patients (135 ETV and 68 LAM) were analyzed in this retrospective analysis. Physical examination, serum liver enzyme tests, and hepatitis B virus marker tests were performed at least every 3 months. We reviewed medical records and performed medical interviews regarding to patients' adherence to medication. Adherence rates &lt 90% were defined as poor adherence in the present study. Cumulative viral breakthrough rates were lower in the ETV-treated patients than in the LAM-treated patients (P&lt 0.001). Seven ETV-treated (5.1%) and 6 LAM-treated patients (8.8%) revealed poor adherence to medication (P=0.48). Among ETV-treated patients, 4 (3.1%) of 128 patients without poor adherence experienced viral breakthrough and 3 (42.8%) of 7 patients with poor adherence experienced viral breakthrough (P&lt 0.001). Only 3 of 38 (7.8%) LAM-treated patients with viral breakthrough had poor adherence, a lower rate than the ETV-treated patients (P=0.039). Nucleoside analogue resistance mutations were observed in 50.0% of ETV- and 94.1% of LAM-treated patients with viral breakthrough (P=0.047). Viral breakthrough associated with poor adherence could be a more important issue in the treatment with especially stronger NAs, such as ETV. © Ivyspring International Publisher.

Nucleos(t)ide analogues (NAs) lead to viral suppression and undetectable hepatitis B virus (HBV) DNA in some individuals infected with HBV, but the rate of virological rebound has been unknown in such patients. We examined the prevalence of virological rebound of HBV DNA among NA-treated patients with undetectable HBV DNA. We retrospectively analyzed 303 consecutive patients [158 entecavir (ETV)- and 145 lamivudine (LAM)-treated] who achieved HBV DNA negativity, defined as HBV DNA < 3.7 log IU/mL for at least 3 months. They were followed up and their features, including their rates of viral breakthrough, were determined. Viral rebound after HBV DNA negativity was not observed in the ETV-group. Viral rebound after HBV DNA negativity occurred in 38.7% of 62 HBe antigen-positive patients in the LAM-group. On multivariate analysis, age was an independent factor for viral breakthrough among these patients (P = 0.035). Viral rebound after HBV DNA negativity occurred in 29.1% of 79 HBe antigen-negative patients in the LAM-group. Differently from LAM, ETV could inhibit HBV replication once HBV DNA negativity was achieved. In contrast, LAM could not inhibit HBV replication even if HBV negativity was achieved in the early phase. Attention should be paid to these features in clinical practice.

Some patients infected with hepatitis C virus (HCV) genotype 2 could be cured with treatment shorter than 24 weeks using peginterferon plus ribavirin, but there are still treatment- refractory patients. Direct-acting antivirals (DAAs) are not currently available for HCV genotype 2 patients, different from genotype 1 patients, in clinical practice. We investigated 29 HCV genotype 2-infected Japanese patients who had been previously treated and failed to clear HCV. We retreated them with peginterferon alfa-2a plus ribavirin and measured HCV RNA level to assess the efficacy and safety of this treatment in patients who had failed previous therapy. We found that retreatment of HCV genotype 2-infected Japanese patients with peginterferon alfa-2a plus ribavirin for 24-48 weeks led to 60 to 66.6% sustained virological response (SVR) in patients previously treated with (peg-)interferon monotherapy and to 69.9% SVR in relapsers previously treated with peginterferon plus ribavirin. Attention should be paid to certain patients with unique features. Selection of patients according to their previous treatment could lead to optimal therapy in HCV genotype 2 treatment-experienced patients. © Ivyspring International Publisher.

Our aim was to investigate the effects of interferons (IFNs)-lambda (interleukin-29 [IL-29], IL-28A, and IL-28B) on hepatitis C virus (HCV) and hepatitis A virus (HAV) internal ribosomal entry site (IRES)-mediated translation. The effects of these IFNs on HCV/HAV translation from HAV/HCV IRES were investigated using bicistronic reporter constructs. We transfected HCV/HAV IRES constructs into these IFN-expressing cell lines. IL-29 showed stronger inhibition of their IRES-mediated translation. Combining IL-29 with IFN-alpha or amantadine resulted in stronger inhibition of HAV IRES activity. Our findings demonstrated a novel antiviral effect of IFNs-lambda against HAV and HCV through the suppression of IRES-mediated translation.

Background: Receptor-interacting serinel threonine protein kinase-2 (RIPK2) has been reported to be an important regulator of tumor proliferation, differentiation and wound repair. We investigated the effects of RIPK2 knockdown in human hepatoma cells on epithelial-to-mesenchymal transition (EMT)-associated gene expression. Materials and Methods: HepG2 cells stably expressing RIPK2-shRNA (HepG2-shRIPK2) were generated after puromycin selection. Total RNAs from HepG2-shRIPK2 and from HepG2-shcontrol cells were isolated and PER-based arrays were performed to compare the 84 EMT-associated gene expressions. Results: We observed that knockdown of RIPK2 down-regulated mRNA expression of jagged 1 (JAG1); plasminogen activator inhibitor-1 (PAI1); regulator of G-protein signalling 2, 24 kDa (RGS2); E-cadherin (CDH1); fibroblast growth factor binding protein 1 (FGFBP1); snail homolog 2 (SNAI2); protein tyrosine phosphatase type IVA, member 1 (PTP4A1); keratin 19 (KRT19); vimentin (VIM); and survival of motor neuron protein-interacting protein 1 (SIP1). Conclusion: We found that knockdown of RIPK2 down-regulated nuclear factor kappa B (NE-kappa B)-dependent PAH and VIM gene expressions. RIPK2 might play an important role in hepatic cell migration. These findings could shed new light on carcinogenesis and on liver regeneration.

It has been reported that inosine triphosphatase (ITPA) gene variants protect against ribavirin-induced anemia in patients treated for chronic hepatitis C. IL28B variants also influence the treatment response of peginterferon plus ribavirin treatment in these patients. In the present study, we examined how ITPA and IL28B genotypes have clinical impacts on treatment-induced hematotoxicities and treatment response in HCV-infected patients treated with peginterferon plus ribavirin. ITPA genotypes (rs1127354 and rs6051702) and IL28B genotype (rs8099917) were determined by TaqMan SNP assay. We compared clinical background, treatment course and treatment response in terms of these genotypes. Only IL28B rs8099917 major type could predict sustained virological response. ITPA rs1127354 major type leads to significantly greater ribavirin-induced anemia than ITPA rs1127354 minor type between days 0 and 84. We noticed that IL28B rs8099917 minor genotype was associated with higher reduction of neutrophils and platelets. ITPA rs1127354 is useful for the prediction of ribavirin-induced anemia in the early phase after the commencement of peginterferon plus ribavirin treatment and IL28B rs8099917 is useful for the prediction of sustained virological response. Use of the combination of these two genotypes could lead to safe and effective treatment of chronic hepatitis C patients.

We previously reported that hepatitis B virus (HBV) e antigen (HBeAg) inhibits production of interleukin 6 by suppressing NF-kappa B activation. NF-kappa B is known to be activated through receptor-interacting serine/threonine protein kinase 2 (RIPK2), and we examined the mechanisms of interleukin 6 regulation by HBeAg. HBeAg inhibits RIPK2 expression and interacts with RIPK2, which may represent 2 mechanisms through which HBeAg blocks nucleotide-binding oligomerization domain-containing protein 1 ligand-induced NF-kappa B activation in HepG2 cells. Our findings identified novel molecular mechanisms whereby HBeAg modulates intracellular signaling pathways by targeting RIPK2, supporting the concept that HBeAg could impair both innate and adaptive immune responses to promote chronic HBV infection.

Aim: Recently, the number of acute hepatitis A cases has decreased in Japan. However, six patients with acute liver failure caused by hepatitis A virus (HAV) have been admitted to Chiba University Hospital, Japan, in the last 18 months, between 2010 and June 2011. The aim of this study is to characterize the recent HAV genotypes from an urban hospital in Japan and to compare the clinical differences. Methods: Hepatitis A virus RNA was detected by strand-specific reverse transcription. Then, HAV VP1/2A regions were amplified by nested polymerase chain reaction (PCR). Sequences were directly determined and phylogenetic trees were constructed for determining HAV subgenotypes. Results: Analysis of these HAV genomes revealed that 4 and 2 belonged to subgenotypes IA and IIIA, respectively. Conclusions: Fujiwara et al. reported a frequency of HAV subgenotype IIIA of only 2.1% in Japan. We conclude that HAV subgenotype IIIA might be widespread in our country.

Background/Aims: Spontaneous acute exacerbation of chronic hepatitis B virus (HBV) infection occasionally occurs in its natural history, sometimes leading rapidly to fatal hepatic failure. We compared the effects of lamivudine (LAM) with those of entecavir (ETV) treatments in acute exacerbation of chronic hepatitis B with 500 IU/L or higher alanine aminotransferase (ALT) levels. Methods: Thirty-four patients with acute exacerbation were consecutively treated with LAM /ETV. Their clinical improvements were compared. Results: Among LAM-treated and ETV-treated patients, none showed a reduction of <1 log IU/mL in HBV DNA after 1 or 3 months of treatment. Initial virological response, defined as a reduction of 4 log IU/mL in HBV DNA at 6 months, with LAM and ETV, respectively, was 83.3% and 100%. One LAM patient developed hepatic encephalopathy, but all patients in both groups survived. Twelve months after treatment, 41.6% of 24 LAM group patients switched to another drug or added adefovir to their treatment due to the emergence of LAM-resistant mutants. On the other hand, patients receiving ETV did not need to change drugs. Conclusions: ETV appears to be as effective as LAM in the treatment of patients with acute exacerbation of chronic hepatitis B. Clinicians should carefully start to treat these patients as soon as possible.

Background: We investigated associations between signal transducer and activator of transcription (STAT) 1 in pretreated liver tissues, interleukin (IL) 28B polymorphism and treatment response in hepatitis C virus (HCV)-infected patients treated with peginterferon and ribavirin. Methods and Findings: We performed immunostaining analysis of STAT1 in liver tissues and determined IL28B polymorphism at rs8099917. We then compared the results with treatment outcomes in HCV genotype 1 patients with high viral load who were receiving peginterferon plus ribavirin. In univariate analysis, younger age, white blood cell counts, virological responder, early virological responder (EVR), mild activity (A1) of liver inflammation grading, and lower STAT1 nuclear-stain of hepatocytes in zone 1, zone 2 and total zones of liver were associated with sustained virological responder (SVR). Multivariate analysis showed that EVR, age and hepatic STAT1 nuclear-stain in zone 2 of liver were independent predictors of SVR. It was also revealed that IL28B and STAT1-nuclear translocation in hepatocytes are independent predictors of response to treatment with peginterferon and ribavirin in chronic hepatitis C patients. Conclusions: Concomitant assessment of lower STAT1 nuclear-stain of hepatocytes and IL28B polymorphism is useful for prediction of SVR in HCV genotype 1 patients.

Recent studies have implicated nuclear receptors (NRs) in the development of hepatocarcinogenesis. We assumed that hepatitis B virus (HBV) alters the expression of NRs and coregulators, and compared the gene expression profiling for 84 NRs and related genes between HpeG2.2.15, which secretes complete HBV virion, and HepG2 by real-time RT-PCR with SyBr green. Forty (47.6%) genes were upregulated 2-fold or greater, and only 5(5.9%) were down-regulated 2-fold or more, in HepG2.2.15 compared to HepG2. These results suggest that HBV affects NRs and their related signal transduction, and that they play important roles in viral replication and HBV-related hepatocarcinogenesis. (C) 2011 Elsevier Ireland Ltd. All rights reserved.

Background The current standard treatment for patients infected with hepatitis C virus (HCV) of genotype 2 is the combination of peginterferon (PEG-IFN) plus ribavirin (RBV) for 24 weeks.Aims We assessed the sustained virological response (SVR) rates in HCV genotype 2-infected Japanese patients in relation to the duration of treatment.Methods Between 2006 and 2009, among 147 patients with HCV genotype 2-infection in Chiba Prefecture, 138 consecutive patients were finally enrolled. Twenty-one, 97 and 20 patients were treated with PEG-IFN-alfa 2b plus RBV for 16, 24 and 48 weeks, respectively. Epidemiological data and treatment outcomes were retrospectively evaluated. HCV RNA was measured with COBAS AMPLICOR HCV Monitor Test v. 2.0.Results The overall SVR rate was 82.6% (114 of 138): treatment-na < ve patients, 86.4% (89 of 103); patients with history of previous treatment, 71.4% (25 of 35). Patients treated for 16, 24 and 48 weeks obtained SVR rates of 66.6% (14 of 21), 86.5% (84 of 97) and 80.0 (16 of 20), respectively.Conclusions The SVR rates of PEG-IFN-alfa 2b plus RBV in Japanese patients were similar to those in previous studies. Combination treatment for 24 weeks for some patients infected with HCV genotype 2 may be superior to that for 16 weeks. More precise patient selection will be needed to shorten the combination treatment.

Background. Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) has been shown to modulate multiple cellular processes, including apoptosis. The aim of this study was to assess the effects of HCV NS5A on apoptosis induced by Toll- like receptor (TLR) 4 ligand, lipopolysaccharide (LPS). Methods. Apoptotic responses to TLR4 ligands and the expression of molecules involved in TLR signaling pathways in human hepatocytes were examined with or without expression of HCV NS5A. Results. HCV NS5A protected HepG2 hepatocytes against LPS-induced apoptosis, an effect linked to reduced TLR4 expression. A similar downregulation of TLR4 expression was observed in Huh-7-expressing genotype 1b and 2a. In agreement with these findings, NS5A inhibited the expression of numerous genes encoding for molecules involved in TLR4 signaling, such as CD14, MD-2, myeloid differentiation primary response gene 88, interferon regulatory factor 3, and nuclear factor-kappa B2. Consistent with a conferred prosurvival advantage, NS5A diminished the poly(adenosine diphosphate-ribose) polymerase cleavage and the activation of caspases 3, 7, 8, and 9 and increased the expression of anti-apoptotic molecules Bcl-2 and c-FLIP. Conclusions. HCV NS5A downregulates TLR4 signaling and LPS-induced apoptotic pathways in human hepatocytes, suggesting that disruption of TLR4-mediated apoptosis may play a role in the pathogenesis of HCV infection.

Objective. Currently, five nucleos(t)ide analogues (NUCs) are available for the treatment of chronic hepatitis B in the world. We examined the prevalence of hepatitis B virus (HBV) DNA and alanine aminotransferase normalization in patients receiving entecavir (ETV) and the frequency of ETV-resistant mutations during an approximately 27-month use of ETV in chronic hepatitis B patients in an urban hospital in Japan. Materials and methods. A retrospective analysis of 81 NUC-naive chronic hepatitis B patients who received 0.5 mg of ETV daily was performed. HBV DNA was measured and sequence analysis of HBV DNA was performed in virological breakthrough patients. Results. Hepatitis B e antigen (HBeAg)-positive patients with HBV DNA 5.0-7.0 log IU/mL group and all HBeAg-negative patients achieved serum HBV DNA negativity by 12 months. Four patients experienced virological breakthrough during ETV therapy. Two patients had no genotypic mutations, and medical interviews revealed that they had poor adherence to ETV. Conclusions. We found that some of the HBV virological breakthroughs during ETV treatment were related to poor adherence to medication, highlighting that clinicians should pay attention to the emergence of resistant mutants as well as adherence to ETV.

Objective. Several studies recently revealed that single nucleotide polymorphisms (SNPs) in the interleukin28B (IL28B) region are associated with the response to pegylated interferon-alfa (PEG-IFN-alfa) and ribavirin (RBV) treatment among hepatitis C virus (HCV)-infected individuals of European, African and Asian ancestry. The purpose of the study was to establish methods for determining the SNP rs8099917 associated with IL28B, which might be useful for further research of the treatment of HCV. Material and methods. Blood samples obtained from 93 consecutive patients with chronic hepatitis C were examined. On the basis of the sequence data, a new simple genotyping assay based on a PCR-restriction fragment length polymorphism (RFLP) with two enzymes, BsrDI and Tsp45I, was developed. Results. The proportion of null virological responders in the combined TG/GG group was higher than that in the TT group (p = 0.015), suggesting that minor allele is one of the important factors playing crucial roles in IFN-resistance. Genotyping of rs8099917 by our new method showed results identical to PCR and sequence in 98.9% and 98.9% by BsrDI and Tsp45I, respectively. Using two enzymes, BsrDI and Tsp45I, it was possible to distinguish IL28B SNP rs8099917. Conclusion. This simple method using RFLP will provide the framework for further studies of HCV.

Aims. Our aim was to identify the factors that made the specimens inadequate and nondiagnostic in endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) biopsy of suspected submucosal tumors (SMTs). Methods. From August 2001 to October 2009, 47 consecutive patients with subepithelial hypoechoic tumors originating in the fourth sonographic layer of the gastric wall suspected as GIST by standard EUS in Chiba University hospital underwent EUS-FNA for histologic diagnosis. We evaluated patient age, sex, location of lesion, size, pattern of growth in a stomach, and pattern of echography retrospectively. We defined a case of gaining no material or an insufficient material for immunohistological diagnosis as nondiagnostic. Results. The diagnostic yield of EUS-FNA for the diagnosis of gastric SMTs was 74.5%. Multivariate logistic regression analysis identified that age of under 60 years (compared with patients older than 60 years: odds ratio [OR] = 11.91, 95% confidence interval [CI] = 1.761-80.48) and location of SMT at lower third area (compared with upper or middle third area: OR = 10.62, 95% CI = 1.290-87.42) were the predictive factors for inadequate tissue yield in EUS-FNA. Conclusions. The factors associated with inadequate tissue yield in EUS-FNA were younger age and the location of lesion at lower third area in stomach.

Disease activities of hepatitis B are affected by the status of hepatitis B e antigen (HBeAg). The function of the hepatitis B virus (HBV) precore or HBeAg is unknown. We assumed that HBeAg blocks aberrant immune responses, although HBeAg is not required for viral assembly, infection, or replication. We examined the interaction of HBeAg and the immune system, including cytokine production. The inflammatory cytokine TNF, IL-6, IL-8, IL-12A, IFN-alpha 1, and IFN-beta mRNA were downregulated in HBeAg-positive HepG2, which stably expresses HBeAg, compared to HBeAg-negative HepG2 cells. The results of real-time RT-PCR-based cytokine-related gene arrays showed the downregulation of cytokine and IFN production. We also observed inhibition of the activation of NF-kappa B-and IFN-beta-promoter in HBeAg-positive HepG2, as well as inhibition of IFN and IL-6 production in HBeAg-positive HepG2 cell culture fluids. HBeAg might modify disease progression by inhibiting inflammatory cytokine and IFN gene expression, while simultaneously suppressing NF-kappa B-signaling- and IFNb-beta promoter activation.

Aim: Little is known about specific naturally-occurring internal ribosomal entry site (IRES) activities of hepatitis A virus (HAV). We examined these activities using the bicistronic reporter assay and the effects of antiviral amantadine against their activities. Methods: Six HAV IRES clones from three patients with fulminant hepatitis and three with self-limited acute hepatitis were obtained. The activities of their IRES were analyzed using bicistronic reporter assay in hepatocyte- and non-hepatocyte-derived cell lines, and the potential efficaciousness of the amantadine was examined. Results: One clone from fulminant hepatitis had a deletion in domains III-IV of HAV IRES had higher IRES activities than HM175 in HLE and Huh-7 cells. In Huh-7 cells, amantadine is effective for inhibiting HAV IRES activities, and especially fulminant hepatitis-derived ones. Conclusion: HAV IRES derived from clinical isolates have various activities. Bicistronic reporter assay using clinical isolates may be another useful tool for testing antiviral activities like those of amantadine and the new acridines and hydrazones recently reported.

Objective. The effects of hepatitis C virus (HCV) sequence variations on the success of antiviral therapy or the development of hepatocellular carcinoma (HCC) are complex for many reasons. Recently, there have been several reports on the effects of genotype 1b HCV core amino acid substitutions 70 and/or 91 on the outcome of antiviral therapies and the clinical course. The purpose of this study was to establish real-time amplification refractory mutation system (ARMS) reverse transcription (RT)-polymerase chain reaction (PCR) assays for easy detection of these HCV mutations. Material and methods. Plasmids p-core-W, including the wild-type HCV core coding region (70R and 91L), and p-core-M, including the mutant-type HCV core (70Q and 91M), were constructed by cloning and PCR-based mutagenesis for control vector of the wild-type core and that of the mutant core, respectively. Using serially diluted forms of these vectors, SyBr Green-based real-time ARMS RT-PCR detection with each of the specific primer pairs was performed. Results. Each primer could clearly distinguish the difference between p-core-W and p-core-M at the same copy numbers. Concerning substitution 70, the ratios 100:1, 10:1, 1:1, 1:10, and 1:100 of p-core-W versus p-core-M could be distinguished, while for substitution 9 1, the ratios 100:1, 10:1, 1:1, 1:10, 1:100, and 1:1000 could be distinguished, confirming the sensitivity and specificity of the assay. Conclusions. This method could be a useful alternative for the detection of genotype 1b HCV core amino acid substitutions 70 and 91 and be reliably applied for rapid screening.

Background: Conventional endoscopy and chromoendoscopy with indigo carmine dye are usually performed for recognizing adequate tumor-negative lateral margins for succesful endoscopic resection of gastric neoplasia. However, chromoendoscopy with indigo carmine dye added to acetic acid has not been used for this purpose. Objective: Our pirpose was to compare the diagnostic performance of chromoendoscopy with indigo carmine dye added to acetic acid with that of conventional endoscopy and chromoendoscopy with indigo carmine dye or acetic acid alone. Deisgn: Prospective study. Setting: Social Insurance Funabashi Central Hospital. Patients: Forty-seven consecutive patients (53 lesions) with early gastric cancer and gastric adenomas who underwent endoscopic submucosal dissection (ESD) from April 2006 to July 2007 were studied. Interventions: All the lesions were examined by the endoscopic modalities before ESD, and the resected specimens were analyzed histopathologically. Main Outcomes Measurement: Two endoscopists independently evaluated the diagnostic performance of each image in terms of recognition of tumor borders with reference to macroscopic and histopathologic findings of resected specimens. We also conducted a substudy to assess interobserver variability. Results: There was good interobserver agreement between the 2 endoscopists in this study (kappa index = 0.764). The diagnostic performance of chromoendoscopy with indico carmine dye added to acetic acid was significantly greater than that of any of the other modalities (vs each: P &lt; .005). Conclusions: The diagnostic performance of chromoendoscopy with indigo carmine dye added to acetic acid was better compared with conventional endoscopy and chromoendoscopy by using only indigo carmine dye or acetic acid. The applicability of this method for gastric neoplasia merits further investigation.