- Email: [email protected]
The positive rates of H. Pylori declined markedly during the first two years of hemodialysis
GASTROENTEROLOGYVoL 114, No. 4
A338 AGA ABSTRACTS
• G1378 FREQUENT ALLELIC IMBALANCE AT CHROMOSOME 13q IN GASTRIC CANCER IN KOREA. Y. Yamaoka, D. Y. Graham, A. R. Sepulveda VA Medical Center and Baylor College of Medicine, Houston, TX; J. G. Kim. Guru Hospital, Korea University College of Medicine, Seoul, Korea. Background: Gastric cancer is one of the most frequent cancers in the world. The genetic mechanisms that result in the development of gastric neoplasia are poorly understood. Several studies have suggested that intestinal type and diffuse carcinomas may have different genetic lesions. H. pylori has been implicated as a co-factor in gastric carcinoma, but only a small number of individuals infected by H. pylori develops this cancer. Specific types of host genetic makeup may in part explain this outcome. The availability of highly polymorphic microsatellite markers throughout the genome can accelerate the search for new genes involved in cancer development. A high percentage of gastric cancers display allelic imbalance (AI) at various loci, characterized both by replicator errors (RER) and loss of heterozignsity (LOH). Aims: To identify genetic loci frequently altered in gastric carcinoma, by using allelic imbalance analysis in intestinal and diffuse gastric carcinomas. Methods: Biopsies from 15 gastric carcinomas from Korean patients were stained with the Genta stain, evaluated and graded for the presence of H. pylori and carcinoma. Genomic DNA was extracted from paraffin embedded tissues. End-labeled microsatellite markers D1S191, D2S123, D13S170 and TP53 were used to amplify tumor and non-neoplastic DNA. PeR products were separated in 6% denaturing polyacrylamide gels. Microsatellite instability (MI) was defined as a band shift in either of the two alleles in two or more markers. LOH was determined by a loss or decrease in intensity of a band present in the tumor, relative to the same band in the normal allele. Results: Eleven tumors were of intestinal type and four were diffuse carcinomas. Microsatellite instability was found less frequently in non-signet ring cell diffuse carcinomas. Six out of fifteen tumors (40%) displayed MI at 2 or more loci. D13S170 was the marker with more frequent MI 9/15 tumors (60%). In addition, there was frequent loss of heterozigosity (LOH) at the D 13S 170 locus in carcinoma tissues. Conclusions: The microsatellite marker D13S170 is very sensitive to detect allelic instability in gastric carcinomas in a population in Korea. This marker revealed a high level of LOH in this region of the genome (13q22-q31) suggesting that a tumor suppressor gene located in this region may be involved in gastric carcinogenesis. • G1379 HOW GOOD ARE THE SEROLOGICAL TESTS THAT HAVE BEEN USED TO MAKE H. PYLORI-DISEASE ASSOCIATIONS WITH CagA OR VacA? Y. Yamaoka, T. Kodama, K. Kashima, Third Department of Internal Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan; D.Y. Graham, Veterans Affairs Medical Center and Baylor College of Medicine, Houston, TX. Background: There has been considerable interest in the putative virulence factors VacA and CagA. One approach to investigate whether they have any association with a particular H. pylori-related disease is to do a seroepidemiologic study for the presence of anti-CagA or anti-VacA antibodies. Interpretation of the results is critically dependent on the accuracy of the tests used. A number of immunoblotting tests have been introduced and have been used for making associations. Aim: To compare 4 such tests for antibodies to CagA or VacA. Methods: Sera from patients with H. pylori infection with H. pylori isolates known to be CagA-positive (by PeR and immunoblotting for 1-1.pylori) or VacA-positive (by vacuolating cytotoxin activities using Vero cells) were tested by immunoblotting with the Helicoblot 2.0 (Genelabs Diagnostics, Singapore), RIDA-Blot Helieobacter (R-Biopharm GmbH, Germany) and RIBA 11. pylori SIA (Chiron), and by ELISA with recombinant CagA antigen (OraVax). Results: False negative tests for anti-CagA were found (78 positive isolates) with the OraVax test (13%), the RIBA test (15%) and the HB 2.0 tests (10%). The RIDA-BH had no false negative tests. Only 2 cases were CagA negative and although no false positive tests were found, the sample is too small to exclude a high percentage of false positive tests (95% CI for 0/2 = 0-84%). Immunoblotting for VacA showed both false negatives and false positives (13 neg. and 67 pos. cases). The HB 2.0 had 49% false negatives and no false positives, RIDA-BH had no false negatives but 92% false positives, RIBA had 12% false negatives and 15% false positives. Conclusion: None of the available serological (immunoblots) tests for antibody to the putative virulence factors CagA and VacA can be recommended. Associations made using these tests should be viewed with caution. • G1380 TEST OF
THE
HELICOBACTER
HYPOTHESIS THAT vacA GENOTYPES OF PYLORI CORRELATE WITH CagA STATUS,
CYTOTOXIN PRODUCTION, OR CLINICAL OUTCOME. Y. Yamaoka. T. Kodama, M. Kita, J. Imanishi, K. Kashima, Third Department of Internal Medicine and Department of Microbiology, Kyoto Prefectural University of Medicine, Kyoto, Japan; D.Y. Graham, Veterans Affairs Medical Center and Baylor College of Medicine, Houston, TX. Background: Recently, mosaicism in vacA alleles with three distinct families of vacA signal sequences (s I a, s lb and s2) and two distinct families of middle region alleles (ml and m2) were reported and, based on a small number of isolates, proposed to be associated with cytotoxin activity, histological, and clinical findings. Aim: To test whether this hypothesis could be confirmed. In addition, we hypothesized that if the vacA or cagA genotypes were to be associated with a
specific H. pylori-related disease, the proportion of the population with the genotype would fall in parallel with the decline in incidence of peptic ulcer and gastric cancer in this population. Methods: Three hundred and forty-three Japanese isolates of [t. pylori from individuals with gastritis alone, duodenal ulcer, gastric ulcer, or gastric cancer were characterized by vacA typing by polymerase chain reaction (PER) and DNA sequencing. Vacuolating cytotoxin activity was measured by in vitro assay using the Vero cell. Results: Three hundred and eighteen strains (92.7%) isolated were classified as sla/ml, 2% were sla/m2, 0.9% were slb/ml, and 0.3% was slb/m2; no type s2 strains were found. Cytotoxin activity was found with both ml and m2 vacA genotypes. There was no association between the vacA genotype or clinical outcome (e.g., gastritis, ulcer, or cancer). In the gastritis only individuals there was no fall in predominant vacA or cagA genotype in younger age individuals compared to the elderly populations. Conclusion: cagA gene-positive, sl/ml strains are the dominant type of H. pylori in Japan. The hypotheses that ml and not m2 vacA genotype was associated with cytotoxin activity, that cagA positivity was associated with genotype sla, or that cagA or vacA genotypes could be used to identify H. pylori of increased virulence related to clinical outcome were not supported. G1381 THE POSITIVE RATES OF 1t. PYLORI DECLINED MARKEDLY DURING TIlE FIRST TWO YEARS OF HEMODIALYSIS. Y. Yamaoka, T. Kodama, K. Kashima, Third Departments of Internal Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan. Background: Since patients with end-stage renal failure have an increased gastric urea concentration they may be more susceptible to colonization with H. pylori, however, the data to support that hypothesis is controversial. Aim: To identify the roles of/-/, pylori in hemodialysis patients. Methods: Four hundred and seventy-five hemodialysis patients were enrolled in this study. H. pylori status was determined by serological tests. Furthermore, 168 hemodialysis patients had undergone an endoscopy. As a control, 486 patients without chronic renal failure who had undergone serological test and endoscopy were also enrolled in this study. Results: Only 48% of hemodialysis patients were positive for H. pylori, significantly lower rates compared with control (71%). There were no relationships between the infection rates and clinical outcome (e.g., chronic gastritis: 46%, duodenal ulcer 50%). Diabetic derived patients were infected with H. pylori significantly higher than glomerulonephritis derived patients (n=89, 59.6% vs. n=339, 44.5% p < 0.01). In both hemodiaiysis patients and control, the positive rates of H. pylori were increased with age. Positive rates of H. pylori in patients having undergone hemodialysis within 2 years were significantly higher than those more than 3 years [duration < 1 years; 74.1%(n=27), 2 years; 75.0% (n=32), 3 years; 51.2%(n=43), > 3 years; 43.4% (n=373)]. Conclusion: Hemodialysis patients have similar positive rates of H. pylori infection to control in the first 1 to 2 years of hemodialysis. Certain factors prevent the infection of 11. pylori during the first 2 years of hemodialysis. • G1382
THE DEVELOPMENT OF "VIRTUAL ENDOSCOPY" BY TRANSABDOMINAL ULTRASONOGRAPHY. N. Yamashita. J. Hata, K. Haruma, S. Aoki, S. Yoshida, H. Kusunoki, M. Hananoki, E. Okamoto, S. Tanaka, M. Yoshihara, K. Sumii, G. Kajiyama. First Dept. of Internal Medicine, Hiroshima University School of Medicine, Hiroshima, Japan. Three-dimensional ultrasonography (3D-US) has been developed recently. However, 3D-US by transabdominal ultrasonography (US) for the evaluation of gastrointestinal diseases has not yet been reported. The AIM of this study was to evaluate the usefulness of this method. We advocate the term "virtual endoscopy" for the demonstration of virtual endoscopic figures by 3D-US. METHODS: Sixteen patients (9 men, 7 women, mean age 52.0 years) with 2 early and 4 advanced gastric cancer, 1 advanced colon cancer, 4 gastric submucosal tumors, and 5 Crohn's disease (2 colonic type, 3 small intestinal type) entered the study. The gastric disease patients were scanned immediately after the ingestion of distilled water (300 mL), and the small intestinal / colonic disease patients were scanned an hour after the ingestion of Golytely solution (1~2 L). The US equipment employed was SSA-380A (Toshiba, Japan) with the 3.0~5.0 MHz transducer, and 3D-US equipment was COMPACT 3D (Tomtec, Germany) with freehand acquisition device using an electromagnetic sensor system. Virtual endoscopic images were obtained by the surface mode, which is the accumulation of the echogenic lines in each sonographic cross-sections with the site recognition. When virtual endoscopic figures (e.g. size, shape, location) were similar to those of the real endoscopy, we classified these figures as being well, otherwise virtual endoscopic figures were classified as being poor. RESULTS: Results are shown in Table. V'rtu Disease
early
colon Ca
~mhn's tltseas¢ Small intestinal donic type I type
GC
adv~d GC
gastric SM'T
WELL
1
4
3
1
2
2
POOR
1
0
1
0
0
1
Good images were obtained in 13 of 16 patients (81.2%). Poor images of 2 cases with gastric disease were due to floating mucous in the stomach, and poor image of one case with small intestinal disease was due to bowel residua and highly bended intestine. CONCLUSION: Virtual endoscopy can be an alternative modality to endoscopy to obtain the non-invasive images of gastrointestinal diseases. Also, this method is useful in the evaluation of
A338 AGA ABSTRACTS
• G1378 FREQUENT ALLELIC IMBALANCE AT CHROMOSOME 13q IN GASTRIC CANCER IN KOREA. Y. Yamaoka, D. Y. Graham, A. R. Sepulveda VA Medical Center and Baylor College of Medicine, Houston, TX; J. G. Kim. Guru Hospital, Korea University College of Medicine, Seoul, Korea. Background: Gastric cancer is one of the most frequent cancers in the world. The genetic mechanisms that result in the development of gastric neoplasia are poorly understood. Several studies have suggested that intestinal type and diffuse carcinomas may have different genetic lesions. H. pylori has been implicated as a co-factor in gastric carcinoma, but only a small number of individuals infected by H. pylori develops this cancer. Specific types of host genetic makeup may in part explain this outcome. The availability of highly polymorphic microsatellite markers throughout the genome can accelerate the search for new genes involved in cancer development. A high percentage of gastric cancers display allelic imbalance (AI) at various loci, characterized both by replicator errors (RER) and loss of heterozignsity (LOH). Aims: To identify genetic loci frequently altered in gastric carcinoma, by using allelic imbalance analysis in intestinal and diffuse gastric carcinomas. Methods: Biopsies from 15 gastric carcinomas from Korean patients were stained with the Genta stain, evaluated and graded for the presence of H. pylori and carcinoma. Genomic DNA was extracted from paraffin embedded tissues. End-labeled microsatellite markers D1S191, D2S123, D13S170 and TP53 were used to amplify tumor and non-neoplastic DNA. PeR products were separated in 6% denaturing polyacrylamide gels. Microsatellite instability (MI) was defined as a band shift in either of the two alleles in two or more markers. LOH was determined by a loss or decrease in intensity of a band present in the tumor, relative to the same band in the normal allele. Results: Eleven tumors were of intestinal type and four were diffuse carcinomas. Microsatellite instability was found less frequently in non-signet ring cell diffuse carcinomas. Six out of fifteen tumors (40%) displayed MI at 2 or more loci. D13S170 was the marker with more frequent MI 9/15 tumors (60%). In addition, there was frequent loss of heterozigosity (LOH) at the D 13S 170 locus in carcinoma tissues. Conclusions: The microsatellite marker D13S170 is very sensitive to detect allelic instability in gastric carcinomas in a population in Korea. This marker revealed a high level of LOH in this region of the genome (13q22-q31) suggesting that a tumor suppressor gene located in this region may be involved in gastric carcinogenesis. • G1379 HOW GOOD ARE THE SEROLOGICAL TESTS THAT HAVE BEEN USED TO MAKE H. PYLORI-DISEASE ASSOCIATIONS WITH CagA OR VacA? Y. Yamaoka, T. Kodama, K. Kashima, Third Department of Internal Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan; D.Y. Graham, Veterans Affairs Medical Center and Baylor College of Medicine, Houston, TX. Background: There has been considerable interest in the putative virulence factors VacA and CagA. One approach to investigate whether they have any association with a particular H. pylori-related disease is to do a seroepidemiologic study for the presence of anti-CagA or anti-VacA antibodies. Interpretation of the results is critically dependent on the accuracy of the tests used. A number of immunoblotting tests have been introduced and have been used for making associations. Aim: To compare 4 such tests for antibodies to CagA or VacA. Methods: Sera from patients with H. pylori infection with H. pylori isolates known to be CagA-positive (by PeR and immunoblotting for 1-1.pylori) or VacA-positive (by vacuolating cytotoxin activities using Vero cells) were tested by immunoblotting with the Helicoblot 2.0 (Genelabs Diagnostics, Singapore), RIDA-Blot Helieobacter (R-Biopharm GmbH, Germany) and RIBA 11. pylori SIA (Chiron), and by ELISA with recombinant CagA antigen (OraVax). Results: False negative tests for anti-CagA were found (78 positive isolates) with the OraVax test (13%), the RIBA test (15%) and the HB 2.0 tests (10%). The RIDA-BH had no false negative tests. Only 2 cases were CagA negative and although no false positive tests were found, the sample is too small to exclude a high percentage of false positive tests (95% CI for 0/2 = 0-84%). Immunoblotting for VacA showed both false negatives and false positives (13 neg. and 67 pos. cases). The HB 2.0 had 49% false negatives and no false positives, RIDA-BH had no false negatives but 92% false positives, RIBA had 12% false negatives and 15% false positives. Conclusion: None of the available serological (immunoblots) tests for antibody to the putative virulence factors CagA and VacA can be recommended. Associations made using these tests should be viewed with caution. • G1380 TEST OF
THE
HELICOBACTER
HYPOTHESIS THAT vacA GENOTYPES OF PYLORI CORRELATE WITH CagA STATUS,
CYTOTOXIN PRODUCTION, OR CLINICAL OUTCOME. Y. Yamaoka. T. Kodama, M. Kita, J. Imanishi, K. Kashima, Third Department of Internal Medicine and Department of Microbiology, Kyoto Prefectural University of Medicine, Kyoto, Japan; D.Y. Graham, Veterans Affairs Medical Center and Baylor College of Medicine, Houston, TX. Background: Recently, mosaicism in vacA alleles with three distinct families of vacA signal sequences (s I a, s lb and s2) and two distinct families of middle region alleles (ml and m2) were reported and, based on a small number of isolates, proposed to be associated with cytotoxin activity, histological, and clinical findings. Aim: To test whether this hypothesis could be confirmed. In addition, we hypothesized that if the vacA or cagA genotypes were to be associated with a
specific H. pylori-related disease, the proportion of the population with the genotype would fall in parallel with the decline in incidence of peptic ulcer and gastric cancer in this population. Methods: Three hundred and forty-three Japanese isolates of [t. pylori from individuals with gastritis alone, duodenal ulcer, gastric ulcer, or gastric cancer were characterized by vacA typing by polymerase chain reaction (PER) and DNA sequencing. Vacuolating cytotoxin activity was measured by in vitro assay using the Vero cell. Results: Three hundred and eighteen strains (92.7%) isolated were classified as sla/ml, 2% were sla/m2, 0.9% were slb/ml, and 0.3% was slb/m2; no type s2 strains were found. Cytotoxin activity was found with both ml and m2 vacA genotypes. There was no association between the vacA genotype or clinical outcome (e.g., gastritis, ulcer, or cancer). In the gastritis only individuals there was no fall in predominant vacA or cagA genotype in younger age individuals compared to the elderly populations. Conclusion: cagA gene-positive, sl/ml strains are the dominant type of H. pylori in Japan. The hypotheses that ml and not m2 vacA genotype was associated with cytotoxin activity, that cagA positivity was associated with genotype sla, or that cagA or vacA genotypes could be used to identify H. pylori of increased virulence related to clinical outcome were not supported. G1381 THE POSITIVE RATES OF 1t. PYLORI DECLINED MARKEDLY DURING TIlE FIRST TWO YEARS OF HEMODIALYSIS. Y. Yamaoka, T. Kodama, K. Kashima, Third Departments of Internal Medicine, Kyoto Prefectural University of Medicine, Kyoto, Japan. Background: Since patients with end-stage renal failure have an increased gastric urea concentration they may be more susceptible to colonization with H. pylori, however, the data to support that hypothesis is controversial. Aim: To identify the roles of/-/, pylori in hemodialysis patients. Methods: Four hundred and seventy-five hemodialysis patients were enrolled in this study. H. pylori status was determined by serological tests. Furthermore, 168 hemodialysis patients had undergone an endoscopy. As a control, 486 patients without chronic renal failure who had undergone serological test and endoscopy were also enrolled in this study. Results: Only 48% of hemodialysis patients were positive for H. pylori, significantly lower rates compared with control (71%). There were no relationships between the infection rates and clinical outcome (e.g., chronic gastritis: 46%, duodenal ulcer 50%). Diabetic derived patients were infected with H. pylori significantly higher than glomerulonephritis derived patients (n=89, 59.6% vs. n=339, 44.5% p < 0.01). In both hemodiaiysis patients and control, the positive rates of H. pylori were increased with age. Positive rates of H. pylori in patients having undergone hemodialysis within 2 years were significantly higher than those more than 3 years [duration < 1 years; 74.1%(n=27), 2 years; 75.0% (n=32), 3 years; 51.2%(n=43), > 3 years; 43.4% (n=373)]. Conclusion: Hemodialysis patients have similar positive rates of H. pylori infection to control in the first 1 to 2 years of hemodialysis. Certain factors prevent the infection of 11. pylori during the first 2 years of hemodialysis. • G1382
THE DEVELOPMENT OF "VIRTUAL ENDOSCOPY" BY TRANSABDOMINAL ULTRASONOGRAPHY. N. Yamashita. J. Hata, K. Haruma, S. Aoki, S. Yoshida, H. Kusunoki, M. Hananoki, E. Okamoto, S. Tanaka, M. Yoshihara, K. Sumii, G. Kajiyama. First Dept. of Internal Medicine, Hiroshima University School of Medicine, Hiroshima, Japan. Three-dimensional ultrasonography (3D-US) has been developed recently. However, 3D-US by transabdominal ultrasonography (US) for the evaluation of gastrointestinal diseases has not yet been reported. The AIM of this study was to evaluate the usefulness of this method. We advocate the term "virtual endoscopy" for the demonstration of virtual endoscopic figures by 3D-US. METHODS: Sixteen patients (9 men, 7 women, mean age 52.0 years) with 2 early and 4 advanced gastric cancer, 1 advanced colon cancer, 4 gastric submucosal tumors, and 5 Crohn's disease (2 colonic type, 3 small intestinal type) entered the study. The gastric disease patients were scanned immediately after the ingestion of distilled water (300 mL), and the small intestinal / colonic disease patients were scanned an hour after the ingestion of Golytely solution (1~2 L). The US equipment employed was SSA-380A (Toshiba, Japan) with the 3.0~5.0 MHz transducer, and 3D-US equipment was COMPACT 3D (Tomtec, Germany) with freehand acquisition device using an electromagnetic sensor system. Virtual endoscopic images were obtained by the surface mode, which is the accumulation of the echogenic lines in each sonographic cross-sections with the site recognition. When virtual endoscopic figures (e.g. size, shape, location) were similar to those of the real endoscopy, we classified these figures as being well, otherwise virtual endoscopic figures were classified as being poor. RESULTS: Results are shown in Table. V'rtu Disease
early
colon Ca
~mhn's tltseas¢ Small intestinal donic type I type
GC
adv~d GC
gastric SM'T
WELL
1
4
3
1
2
2
POOR
1
0
1
0
0
1
Good images were obtained in 13 of 16 patients (81.2%). Poor images of 2 cases with gastric disease were due to floating mucous in the stomach, and poor image of one case with small intestinal disease was due to bowel residua and highly bended intestine. CONCLUSION: Virtual endoscopy can be an alternative modality to endoscopy to obtain the non-invasive images of gastrointestinal diseases. Also, this method is useful in the evaluation of