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Prevention of the development of autoimmune gastritis by Helicobacter pylori infection in neonatally thymectomized mice
A744 AGA ABSTRACTS
4022 AMMONIA ENHANCES EMOTIONAL STRESS·INDUCED MOUSE GASTRIC MUCOSAL DAMAGE IN COMMUNICATION BOX PARADIGM. Shoichi Nagahashi, Hidekazu Suzuki, Chikako Watanabe, Masaharu Miyazawa, Chizuko Kawaguchi, Masayuki Suzuki, Soichiro Miura, Hiromasa Ishii, Keio Univ Sch of Medicine, Tokyo, Japan; National Tokyo Med Ctr, Tokyo, Japan; National Defense Med Coll, Tokorozawa, Japan. Background. The relationship between Hipylori (Hpj-colonization and the formation of stress-induced gastric mucosal injury has remained unknown. In the Hp-infected gastric mucosa, urease-dependent ammonia (NH3 ) production is reportedly important to induce gastric lesions. The present study was designed to investigate whether NHrpretreatment enhances the formation of gastric lesion in response to emotional stress. Method. Twelve CS7BL/6 mice were treated with 0.01%-NH3 through the gastric tube once a day for a week before applying emotional stress, and other II mice were treated with distilled water. To provide emotional stress, the communication box paradigm, in which each mouse (responder) was placed in transparent plastic compartment adjacent to mice receiving electrical stimulation (3 hours (19:30-22:30) per day for 3 days; sender), was used. After the overnight fasting, the stomach of each animal was examined. Macroscopic and microscopic evaluation were performed in each stomach. Mucosal activity of myeloperoxidase (MPO), an index of neutrophil accumulation, as well as the contents of thiobarbitulate reactive substances (TBARS), an index of mucosal lipid peroxidation were measured (Free Radie. Bioi. Med. 26:679, 1999). Results. The responder mice pretreated with 0.01%-NH3 (NH 3-responder) developed more severe gastric lesions than those pretreated with distilled water (control-responder). MPO activity was enhanced in NH3-responder (0.72±0.3S mU/mg protein) more than that in the control-responder (0.3S±0.09 mU/mg protein) (p
GASTROENTEROLOGY Vol. 118, No.4
ever, this phenomenon was seen only in Mongolian gerbils. Furthermore, only a very small proportion of H.pylori infected population will develop gastric cancer. These suggest the involvement of host factor other than Hipylori itself in the development of gastric cancer. To prove whether host factor is important or not, Hipylori colonization and inflammation induced in the stomach are examined in several animal models. Methods: Five strains of animals, which are 6-week-old male (I) MGSfSea, (2) CS7/BL6, (3) BALB/c, (4) Wistar, (5) SD, were inoculated with Hipylori (TN2GF4) orally and scarified at 4 and 6 weeks after infection to assess colonization of Hipylori and macroscopic and histological changes of the stomach. Hipylori strain, TN2GF4, is one of the strains that induced adenocarcinoma in Mongolian gerbils. Results: From 4 until 6 weeks, colonization of Htpylori was maintained in the stomach of almost animals. Macroscopically, the antral mucosa showed edema and hemorrhagic spots in only Mongolian gerbils. Histologically, chronic gastritis was also observed in only Mongolian gerbils. Namely, there was infiltration of lymphocytes, macrophages and neutrophils into the lamina propria and submucosan and lymphoid follicles had formed in only Mongolian gerbils. Conclusion: These results suggest that host factor causes the difference in the pathological changes induced with Hpylori infection. No. ofanimals colonized
(1) (4weeks) (6weeks)
515 515
No. ofanimals with chronic gastritis (1) 4/5 (4weeks) (6weeks) 5/5
(2) 515 515
(3) 414 213
(4) 215 515
(5) 415 5/5
(2) 0/5 0/5
(3) 014 013
(4) 015 0/5
(5) 0/5 0/5
4025
rr-
GASTRIC FOLLICULAR FORMATION BY HELICOBACTER LORI INFECTION DEPENDS ON TH2 TYPE IMMUNE RE· SPONSES AND STRAIN OF THE HOST. Masaya Ohana, Kazuichi Okazaki, Chikashi Oshima, Toshiki Nishi, Andra's Debreceni, Kazushige Uchida, Suguru Uose, Hiroshi Nakase, Yumi Matsushima, Tsutomu Chiba, Kyoto Univ, Kyoto, Japan. Background; The normal gastric mucosa has no lymphoid organs. The formation of gastric lymphoid follicle is often observed in the H pyloripositive gastritis, which is important in the development of gastric MALT lymphoma. However, it is still unclear how lymphoid follicles arise in the gastric mucosa after H. pylori infection. We analyzed the host factors by using several strains of thymectomized mice infected with H pylori. Materials and Methods; BALB/c, CS7BL/6, C3H1He,DBA/2 mice, which underwent thymectomy on day 3 after birth (d3-Tx) at 6 week old, were orally infected with H pylori. The changes of the gastric mucosa and systemic immune responses were evaluated histologically and immunologically. Results; Seven of 12 (58.3%) infected d3-Tx BALB/c mice developed gastric lymphoid follicle 4 months after infection: no follicular formation was observed in other strains of d3-Tx mice including DBA/2 in which MHC class II is the same as BALB/c. No follicular formation was observed in each strain of normal (no d3-Tx) mice including BALB/c, even 6 months after H. pylori infection. Serum levels of anti-parietal cell antibody were elevated in the d3-Tx BALB/c mice, but not in other strains of d3-Tx mice. Thymectomy induced significant higher levels of anti-H. pylori antibody in BALB/c, but not in other strains. In the infected d3-Tx BALB/c mice with follicular gastritis, the expression of IL-4 in the gastric mucosa was upregulated by RT-PCR as well as IFN--y. Conclusions; These findings suggested genetic factors independent of MHC, and hyper-immune state by d3-Tx may be involved in the gastric follicular formation in H pylori infected mice. 4026 PREVENTION OF THE DEVELOPMENT OF AUTOIMMUNE GASTRITIS BY HELICOBACTER PYLORI INFECTION IN NEONATALLY THYMECTOMIZED MICE. Masaya Ohana, Kazuichi Okazaki, Chikashi Oshima, Toshiki Nishi, Andra's Debreceni, Kazushige Uchida, Suguru Uose, Hiroshi Nakase, Yumi Matsushima, Chiba Tsutomu, Kyoto Univ, Kyoto, Japan. Background and Aim; Patients with autoimmune gastritis (AIG) are often complicated with pernicious anemia, gastric cancer, and gastric carcinoid tumor. There is little agreement as to the relationship between H pylori infection and AIG. We studied whether H. pylori infection affects the pathophysiology of AIG or not. Materials and Methods; BALB/c mice which underwent thymectomy on day 3 after birth (d3-Tx) and developed autoimmune gastritis at the age of 6 weeks, were orally infected with H. pylori. BALB/c nu/nu mice, in which AIG was induced by transfer with splenocytes from the d3-Tx mice, were infected with H pylori. The histological findings of the gastric mucosa, and serum levels of anti-parietal cell and H pylori antibodies were evaluated. Results; Seventy percent of infected d3-Tx mice developed follicular gastritis in addition to autoimmune gastritis. Atrophic scores, reflecting the degree of AIG, showed that parietal cells were significantly preserved in the infected d3-Tx mice (atrophic score; 0.40±0.25) compared with infected normal mice (2.30±0.30). Serum levels of anti-parietal cell antibody were serially decreased in the infected d3-Tx mice compared with the non-infected
April 2000
d3-Tx mice. Serum levels of anti H. pylori antibody were higher in the infected d3-Tx mice than in the infected normal mice. The expression of lPN-gamma was upregulated in the gastric mucosa in the control d3-Tx mice. In the infected d3-Tx mice, the expression of IL-4 was upregulated in addition to IFN-gamma. Conclusions;H. pylori infection may inhibit the destruction of parietal cells in mice with autoimmune gastritis. Th2-type immune responses in the gastric microenvironment may be involved in the inhibitory effects of H. pylori infection on the development of AIG, in which Thl-type responses play an important role.
4027 SERUM PEPSINOGENS IN RELATION TO H. PYLORI STATUS AND GENOTYPE AND GASTRIC INFLAMMATORY CELL INFILTRATION. Guillermo I. Perez-Perez, Andrea Legath, Richard M. Peek, K. Tham, Martin 1. Blaser, Vanderbilt Univ, Nashville, TN; Vanderbilt, Nashville, TN; Dept of Veterans Affairs, Nashville, TN. Background: Chronic atrophic gastritis increases risk for the development of distal gastric cancer. H. pylori colonization increases risk for both of these conditions and appears to decrease risk for GERD and its sequelae. Strains of the cagA+ vacA ml genotype in particular appear to be most associated with these risks. Analysis of serum pepsinogens is a noninvasive means of assessing gastric atrophy and for the trends favoring its development. We sought to examine in a US population the relationship between carriage of H. pylori strains of particular genotypes and infiltration of the gastric mucosa with inflammatory cells, with levels of serum pepsinogen I (PGI) and II (PG II), as well as their ratio, which are markers for the development of gastric atrophy. Methods: 142 dyspeptic Nashville VA patients (98.6% male) undergoing upper endoscopy were studied. H. pylori status was determined by culture, histology, or CLO test. The cagA, vacA, and iceA genotypes of the strains were determined by PCR. PGI and PGII levels were determined using the PEPSINOGEN RIABEAD kits (Dainabot, Ltd; Tokyo). Results: As expected, the PGIIPGII ratio (2.1 ±2.7) for H. pylori" patients was significantly (p=0.036) lower than for the H. pylori" (3.1 ±4.3) patients. Also as expected, among H. pylori" patients, those with higher MNC infiltration scores in the gastric corpus had significantly higher PGI (p=0.035), and PGII (p=O.OO4) scores than did those with little or no infiltration. Amon~ patients with little or no PMN infiltration in the gastric corpus, H. pylori patients had significantly lower PGIIII ratios (1.8± 2.1) than H. pylori" (3.3± 4.4; p=0.02) patients. Parallel differences (p=0.035) were seen in those with little or no MNC infiltration in the gastric corpus. Among the H. pylori" patients, those with vacA ml strains had higher mean PGI (24.2) and PG II (11.5) levels than did patients with vacA m2 strains (12.0 and 5.3; p=0.045, and O.Q2, respectively). No significant differences were observed with respect to cagA, vacA s type, and iceA type. Conclusions: In this US population, markers of the development of atrophy are significantly more common in H. pylori" persons, and are associated with more substantial inflammatory cell infiltrates in the gastric corpus. This may help explain why the falling prevalence of H. pylori is temporally associated with increases in GERD. Persons with vacA ml strains, which are associated with increased risk of gastric cancer, also have PG profiles consistent with increased atrophy. 4028 INCREASED EXPRESSION AND PRODUCTION OF IL-18 IN
HEUCOBACTER PYLORI INFECTED GASTRIC MUCOSA. Kyoko Sakai, Yoshio Yamaoka, Tomoyuki Ohno, Naoki Sawai, Syoji MItsufuji, Tadashi Kodama, Kei Kshima, Masakzu Kita, Jiro Imanishi, Kyoto prefectural Univ of medicine, Kyoto, Japan. Background and Aims: Interleukin- I 8 (lL-I8) is a recently described proinfla mmatory cytokine that enhances IFN-g production and is required for a Thl type cellular response. We previously reported that in animal models, IFN1' plays an important role in H. pylori infection not only for defense against infection, but also for induction of inflammation. However, there were few reports about the induction of IL-I8 from the gastric mucosa. The aim of this study was to examine the role of IL-18 in the stomach and it's relationship to IFN"expression. Methods: mRNA levels of IFN-g and IL-I8 in antral biopsy specimens were meas ured in 81 patients (33: chronic gastritis, 21: gastric ulcer, 26: duodenal ulcer and 1: gastric cancer) by RT-PCR. Protein levels of IL-I8 were measur ed by ELISA. In 23 patients, eradication therapy was performed and biopsy specimens were taken before and after the therapy for measurement of cytokine levels. Results: There was no relationship between IFN"mRNA levels and H. pylori status. Mucosal expression of IL-I8 mRNA were significantly higher in H. pylori positive (66%) than in H. pylori -negative (20%) patients (p < 0.01). Mucosal levels of IL-I8 protein were also significantly higher in H. pylori positive (mean + SD = 34 I + 248 pg/mL) than in H. pylori -negative (30 + 21 pg/ml) patients (p < 0.01). No correlation was observed between IL-18 or IFN"expression and histological findings and endoscopic diagnosis. There was no association between IFN"and IL-I8 mRNA expression. In patients cured of H. pylori infection, IL-18 production decreased significantly after thera py (before: 230 + 131, after 84 + 66 pg/ml, P < 0.01). Conclusions: These findings demonstrate that infection with H. pylori induces IL- I8 expression and production from the gastric mucosa. Increased IL- I8 levels in the gastric mucosa may play an important role in the gastric inflammatory response that is independent of IFN"expression.
AGAA745
4029 EXPERIMENTAL MODEL OF INFECTION OF HEUCOBACTER PYLORI IN THE MOUSE. A STUDY OF GASTRIC DAMAGE CAUSED BY INDOMETHACIN. Mauricio Schraier, Laura Petrelli, Mercedes Bonfanti, Andrea Di Bella, Pablo Pest, Fernando Man, Jose Adami, Hosp Posadas, Buenos Aires, Argentina. Introduction: The action of non-steroidal anti-inflammatory drugs (NSAIDs) on the Helicobacter pylori (Hp) infected mucosa is a matter of debate. Some authors consider them to cause additive iatrogeny whilst others attribute a purportedly protective action to them. The development of an experimental animal model could help clarify this phenomenon. Objectives: I.To develop an animal model of Hp gastric infection. 2.To evaluate the aggressiveness of NSAIDs in this mode!. Materials and Methods Male 6-month old BALB/C mice weighing 38 g were studied. Prior Hp infection was ruled out. On three occasions, in the same week, 18 mice were inoculated intra-gastrically with 0.6 ml of Hp culture broth (brain-heart infusion) containing Ix108-IxI09 CFU/m!. Another group of mice were inoculated with sterile saline. After two months the mice were killed and their stomachs studied. They were divided into groups: a)6 Hp negative control mice b)8 Hp negative mice with prior intra-peritoneal injection of 25 mg/Kg indomethacin (24hr). c)8 mice inoculated with Hp, with indomethacin. d)8 mice inoculated with Hp, without indomethacin The stomachs were opened along the greater curvature and photographed macroscopically in order to map the necrotic area. The antrums were biopsied to test for urease and separate antrum and body specimens were sent for staining with Warthin-Slarry H&E and histopathology. Results All the mice inoculated with Hp acquired the infection. The necrotic area was larger in Group B: 55.5 ± 7.87 mm than in Group C: 15 ± 1.82 mm P
4030 HEUCOBACTER PYLORI INFECTION UPREGULATES IQGAP-l EXPRESSION IN PRIMARY CULTURES OF HUMAN ANTRAL EPITHELIAL CELLS. Valerie C. Smith, Mark Ring, Susan B. Curtis, David Scriven, Edwin Dw Moore, Carrie Rosenberger, Brett B. Finlay, Mark R. Meloche, Alison Mj Buchan, Univ of British Columbia, Vancouver, BC, Canada. In normal epithelial cells J3-cateninis located at cell-cell adherens junctions with E-cadherin and o-carenin. In a number of gastric cancers J3-catenin is concentrated in the nucleus and activates the tcf/lef transcription factor stimulating cell proliferation and the production of the matrix rnetaloproteinase family of enzymes. In cell lines the newly identified protein IQGAP-I forms a complex with e-carenm resulting in dissociation of E-cadherin and loss of cell-cell contacts resulting in increased cytosolic J3-catenin.We are currently investigating how H. pylori infection alters the function of antral epithelial cells to cause either ulceration or gastric cancer. For the present experiments primary cultures of human antral epithelial cells (N = 5, 3 male and 2 female, II - 48 yr) were infected with the CCUG 17874 strain of H. pylori for 48h prior to collection of mRNA for gene array experiments, protein for Western blot analysis or fixation of the cells for immunocytochemical studies using deconvolution microscopy. We used the Neuroscience array (Clontech) to monitor changes in expression of genes encoding cell adherence and signal transduction proteins. Antibodies to E-cadherin (Santa Cruz), IQGAP-l (Transduction), J3-and a-catenin (Transduction) were used for the Westerns and immunocytochemistry. H. pylori infection resulted in a 12 fold increase in IQGAP-I and a 3 fold increase in matrix metalloproteinase 16 mRNA levels with no change in E-cadherin. The Western blot analysis showed a 5 fold increase in IQGAP- I protein levels with no change in E-cadherin, 13- or o-catenin. In control cells immunoreactivity for E-cadherin, IQGAP-I, 13and a-catenin was concentrated to the plasma membrane. In infected cells J3-catenin and IQGAP-l were re-distributed to intracellular tubulovesicles with lower levels of immunoreactivity at the plasma membrane. In addition low levels of J3-catenin were observed within the nucleus. Nuclear boundaries was defined using DAPI labeling. E-cadherin staining remained at the plasma membrane, although at a reduced leveL These data indicated that the increase in IQGAP-I levels in cells infected with H. pylori resulted in a re-distribution of J3-catenin within the cells. The presence of J3-cateninin the nucleus would activate the lef/tcf transcription pathway in turn increasing the expression of matrix metalloproteinase 16. The ability of H. pylori to cause nuclear translocation of J3-catenin may be important in the development of gastric cancer.
4022 AMMONIA ENHANCES EMOTIONAL STRESS·INDUCED MOUSE GASTRIC MUCOSAL DAMAGE IN COMMUNICATION BOX PARADIGM. Shoichi Nagahashi, Hidekazu Suzuki, Chikako Watanabe, Masaharu Miyazawa, Chizuko Kawaguchi, Masayuki Suzuki, Soichiro Miura, Hiromasa Ishii, Keio Univ Sch of Medicine, Tokyo, Japan; National Tokyo Med Ctr, Tokyo, Japan; National Defense Med Coll, Tokorozawa, Japan. Background. The relationship between Hipylori (Hpj-colonization and the formation of stress-induced gastric mucosal injury has remained unknown. In the Hp-infected gastric mucosa, urease-dependent ammonia (NH3 ) production is reportedly important to induce gastric lesions. The present study was designed to investigate whether NHrpretreatment enhances the formation of gastric lesion in response to emotional stress. Method. Twelve CS7BL/6 mice were treated with 0.01%-NH3 through the gastric tube once a day for a week before applying emotional stress, and other II mice were treated with distilled water. To provide emotional stress, the communication box paradigm, in which each mouse (responder) was placed in transparent plastic compartment adjacent to mice receiving electrical stimulation (3 hours (19:30-22:30) per day for 3 days; sender), was used. After the overnight fasting, the stomach of each animal was examined. Macroscopic and microscopic evaluation were performed in each stomach. Mucosal activity of myeloperoxidase (MPO), an index of neutrophil accumulation, as well as the contents of thiobarbitulate reactive substances (TBARS), an index of mucosal lipid peroxidation were measured (Free Radie. Bioi. Med. 26:679, 1999). Results. The responder mice pretreated with 0.01%-NH3 (NH 3-responder) developed more severe gastric lesions than those pretreated with distilled water (control-responder). MPO activity was enhanced in NH3-responder (0.72±0.3S mU/mg protein) more than that in the control-responder (0.3S±0.09 mU/mg protein) (p
GASTROENTEROLOGY Vol. 118, No.4
ever, this phenomenon was seen only in Mongolian gerbils. Furthermore, only a very small proportion of H.pylori infected population will develop gastric cancer. These suggest the involvement of host factor other than Hipylori itself in the development of gastric cancer. To prove whether host factor is important or not, Hipylori colonization and inflammation induced in the stomach are examined in several animal models. Methods: Five strains of animals, which are 6-week-old male (I) MGSfSea, (2) CS7/BL6, (3) BALB/c, (4) Wistar, (5) SD, were inoculated with Hipylori (TN2GF4) orally and scarified at 4 and 6 weeks after infection to assess colonization of Hipylori and macroscopic and histological changes of the stomach. Hipylori strain, TN2GF4, is one of the strains that induced adenocarcinoma in Mongolian gerbils. Results: From 4 until 6 weeks, colonization of Htpylori was maintained in the stomach of almost animals. Macroscopically, the antral mucosa showed edema and hemorrhagic spots in only Mongolian gerbils. Histologically, chronic gastritis was also observed in only Mongolian gerbils. Namely, there was infiltration of lymphocytes, macrophages and neutrophils into the lamina propria and submucosan and lymphoid follicles had formed in only Mongolian gerbils. Conclusion: These results suggest that host factor causes the difference in the pathological changes induced with Hpylori infection. No. ofanimals colonized
(1) (4weeks) (6weeks)
515 515
No. ofanimals with chronic gastritis (1) 4/5 (4weeks) (6weeks) 5/5
(2) 515 515
(3) 414 213
(4) 215 515
(5) 415 5/5
(2) 0/5 0/5
(3) 014 013
(4) 015 0/5
(5) 0/5 0/5
4025
rr-
GASTRIC FOLLICULAR FORMATION BY HELICOBACTER LORI INFECTION DEPENDS ON TH2 TYPE IMMUNE RE· SPONSES AND STRAIN OF THE HOST. Masaya Ohana, Kazuichi Okazaki, Chikashi Oshima, Toshiki Nishi, Andra's Debreceni, Kazushige Uchida, Suguru Uose, Hiroshi Nakase, Yumi Matsushima, Tsutomu Chiba, Kyoto Univ, Kyoto, Japan. Background; The normal gastric mucosa has no lymphoid organs. The formation of gastric lymphoid follicle is often observed in the H pyloripositive gastritis, which is important in the development of gastric MALT lymphoma. However, it is still unclear how lymphoid follicles arise in the gastric mucosa after H. pylori infection. We analyzed the host factors by using several strains of thymectomized mice infected with H pylori. Materials and Methods; BALB/c, CS7BL/6, C3H1He,DBA/2 mice, which underwent thymectomy on day 3 after birth (d3-Tx) at 6 week old, were orally infected with H pylori. The changes of the gastric mucosa and systemic immune responses were evaluated histologically and immunologically. Results; Seven of 12 (58.3%) infected d3-Tx BALB/c mice developed gastric lymphoid follicle 4 months after infection: no follicular formation was observed in other strains of d3-Tx mice including DBA/2 in which MHC class II is the same as BALB/c. No follicular formation was observed in each strain of normal (no d3-Tx) mice including BALB/c, even 6 months after H. pylori infection. Serum levels of anti-parietal cell antibody were elevated in the d3-Tx BALB/c mice, but not in other strains of d3-Tx mice. Thymectomy induced significant higher levels of anti-H. pylori antibody in BALB/c, but not in other strains. In the infected d3-Tx BALB/c mice with follicular gastritis, the expression of IL-4 in the gastric mucosa was upregulated by RT-PCR as well as IFN--y. Conclusions; These findings suggested genetic factors independent of MHC, and hyper-immune state by d3-Tx may be involved in the gastric follicular formation in H pylori infected mice. 4026 PREVENTION OF THE DEVELOPMENT OF AUTOIMMUNE GASTRITIS BY HELICOBACTER PYLORI INFECTION IN NEONATALLY THYMECTOMIZED MICE. Masaya Ohana, Kazuichi Okazaki, Chikashi Oshima, Toshiki Nishi, Andra's Debreceni, Kazushige Uchida, Suguru Uose, Hiroshi Nakase, Yumi Matsushima, Chiba Tsutomu, Kyoto Univ, Kyoto, Japan. Background and Aim; Patients with autoimmune gastritis (AIG) are often complicated with pernicious anemia, gastric cancer, and gastric carcinoid tumor. There is little agreement as to the relationship between H pylori infection and AIG. We studied whether H. pylori infection affects the pathophysiology of AIG or not. Materials and Methods; BALB/c mice which underwent thymectomy on day 3 after birth (d3-Tx) and developed autoimmune gastritis at the age of 6 weeks, were orally infected with H. pylori. BALB/c nu/nu mice, in which AIG was induced by transfer with splenocytes from the d3-Tx mice, were infected with H pylori. The histological findings of the gastric mucosa, and serum levels of anti-parietal cell and H pylori antibodies were evaluated. Results; Seventy percent of infected d3-Tx mice developed follicular gastritis in addition to autoimmune gastritis. Atrophic scores, reflecting the degree of AIG, showed that parietal cells were significantly preserved in the infected d3-Tx mice (atrophic score; 0.40±0.25) compared with infected normal mice (2.30±0.30). Serum levels of anti-parietal cell antibody were serially decreased in the infected d3-Tx mice compared with the non-infected
April 2000
d3-Tx mice. Serum levels of anti H. pylori antibody were higher in the infected d3-Tx mice than in the infected normal mice. The expression of lPN-gamma was upregulated in the gastric mucosa in the control d3-Tx mice. In the infected d3-Tx mice, the expression of IL-4 was upregulated in addition to IFN-gamma. Conclusions;H. pylori infection may inhibit the destruction of parietal cells in mice with autoimmune gastritis. Th2-type immune responses in the gastric microenvironment may be involved in the inhibitory effects of H. pylori infection on the development of AIG, in which Thl-type responses play an important role.
4027 SERUM PEPSINOGENS IN RELATION TO H. PYLORI STATUS AND GENOTYPE AND GASTRIC INFLAMMATORY CELL INFILTRATION. Guillermo I. Perez-Perez, Andrea Legath, Richard M. Peek, K. Tham, Martin 1. Blaser, Vanderbilt Univ, Nashville, TN; Vanderbilt, Nashville, TN; Dept of Veterans Affairs, Nashville, TN. Background: Chronic atrophic gastritis increases risk for the development of distal gastric cancer. H. pylori colonization increases risk for both of these conditions and appears to decrease risk for GERD and its sequelae. Strains of the cagA+ vacA ml genotype in particular appear to be most associated with these risks. Analysis of serum pepsinogens is a noninvasive means of assessing gastric atrophy and for the trends favoring its development. We sought to examine in a US population the relationship between carriage of H. pylori strains of particular genotypes and infiltration of the gastric mucosa with inflammatory cells, with levels of serum pepsinogen I (PGI) and II (PG II), as well as their ratio, which are markers for the development of gastric atrophy. Methods: 142 dyspeptic Nashville VA patients (98.6% male) undergoing upper endoscopy were studied. H. pylori status was determined by culture, histology, or CLO test. The cagA, vacA, and iceA genotypes of the strains were determined by PCR. PGI and PGII levels were determined using the PEPSINOGEN RIABEAD kits (Dainabot, Ltd; Tokyo). Results: As expected, the PGIIPGII ratio (2.1 ±2.7) for H. pylori" patients was significantly (p=0.036) lower than for the H. pylori" (3.1 ±4.3) patients. Also as expected, among H. pylori" patients, those with higher MNC infiltration scores in the gastric corpus had significantly higher PGI (p=0.035), and PGII (p=O.OO4) scores than did those with little or no infiltration. Amon~ patients with little or no PMN infiltration in the gastric corpus, H. pylori patients had significantly lower PGIIII ratios (1.8± 2.1) than H. pylori" (3.3± 4.4; p=0.02) patients. Parallel differences (p=0.035) were seen in those with little or no MNC infiltration in the gastric corpus. Among the H. pylori" patients, those with vacA ml strains had higher mean PGI (24.2) and PG II (11.5) levels than did patients with vacA m2 strains (12.0 and 5.3; p=0.045, and O.Q2, respectively). No significant differences were observed with respect to cagA, vacA s type, and iceA type. Conclusions: In this US population, markers of the development of atrophy are significantly more common in H. pylori" persons, and are associated with more substantial inflammatory cell infiltrates in the gastric corpus. This may help explain why the falling prevalence of H. pylori is temporally associated with increases in GERD. Persons with vacA ml strains, which are associated with increased risk of gastric cancer, also have PG profiles consistent with increased atrophy. 4028 INCREASED EXPRESSION AND PRODUCTION OF IL-18 IN
HEUCOBACTER PYLORI INFECTED GASTRIC MUCOSA. Kyoko Sakai, Yoshio Yamaoka, Tomoyuki Ohno, Naoki Sawai, Syoji MItsufuji, Tadashi Kodama, Kei Kshima, Masakzu Kita, Jiro Imanishi, Kyoto prefectural Univ of medicine, Kyoto, Japan. Background and Aims: Interleukin- I 8 (lL-I8) is a recently described proinfla mmatory cytokine that enhances IFN-g production and is required for a Thl type cellular response. We previously reported that in animal models, IFN1' plays an important role in H. pylori infection not only for defense against infection, but also for induction of inflammation. However, there were few reports about the induction of IL-I8 from the gastric mucosa. The aim of this study was to examine the role of IL-18 in the stomach and it's relationship to IFN"expression. Methods: mRNA levels of IFN-g and IL-I8 in antral biopsy specimens were meas ured in 81 patients (33: chronic gastritis, 21: gastric ulcer, 26: duodenal ulcer and 1: gastric cancer) by RT-PCR. Protein levels of IL-I8 were measur ed by ELISA. In 23 patients, eradication therapy was performed and biopsy specimens were taken before and after the therapy for measurement of cytokine levels. Results: There was no relationship between IFN"mRNA levels and H. pylori status. Mucosal expression of IL-I8 mRNA were significantly higher in H. pylori positive (66%) than in H. pylori -negative (20%) patients (p < 0.01). Mucosal levels of IL-I8 protein were also significantly higher in H. pylori positive (mean + SD = 34 I + 248 pg/mL) than in H. pylori -negative (30 + 21 pg/ml) patients (p < 0.01). No correlation was observed between IL-18 or IFN"expression and histological findings and endoscopic diagnosis. There was no association between IFN"and IL-I8 mRNA expression. In patients cured of H. pylori infection, IL-18 production decreased significantly after thera py (before: 230 + 131, after 84 + 66 pg/ml, P < 0.01). Conclusions: These findings demonstrate that infection with H. pylori induces IL- I8 expression and production from the gastric mucosa. Increased IL- I8 levels in the gastric mucosa may play an important role in the gastric inflammatory response that is independent of IFN"expression.
AGAA745
4029 EXPERIMENTAL MODEL OF INFECTION OF HEUCOBACTER PYLORI IN THE MOUSE. A STUDY OF GASTRIC DAMAGE CAUSED BY INDOMETHACIN. Mauricio Schraier, Laura Petrelli, Mercedes Bonfanti, Andrea Di Bella, Pablo Pest, Fernando Man, Jose Adami, Hosp Posadas, Buenos Aires, Argentina. Introduction: The action of non-steroidal anti-inflammatory drugs (NSAIDs) on the Helicobacter pylori (Hp) infected mucosa is a matter of debate. Some authors consider them to cause additive iatrogeny whilst others attribute a purportedly protective action to them. The development of an experimental animal model could help clarify this phenomenon. Objectives: I.To develop an animal model of Hp gastric infection. 2.To evaluate the aggressiveness of NSAIDs in this mode!. Materials and Methods Male 6-month old BALB/C mice weighing 38 g were studied. Prior Hp infection was ruled out. On three occasions, in the same week, 18 mice were inoculated intra-gastrically with 0.6 ml of Hp culture broth (brain-heart infusion) containing Ix108-IxI09 CFU/m!. Another group of mice were inoculated with sterile saline. After two months the mice were killed and their stomachs studied. They were divided into groups: a)6 Hp negative control mice b)8 Hp negative mice with prior intra-peritoneal injection of 25 mg/Kg indomethacin (24hr). c)8 mice inoculated with Hp, with indomethacin. d)8 mice inoculated with Hp, without indomethacin The stomachs were opened along the greater curvature and photographed macroscopically in order to map the necrotic area. The antrums were biopsied to test for urease and separate antrum and body specimens were sent for staining with Warthin-Slarry H&E and histopathology. Results All the mice inoculated with Hp acquired the infection. The necrotic area was larger in Group B: 55.5 ± 7.87 mm than in Group C: 15 ± 1.82 mm P
4030 HEUCOBACTER PYLORI INFECTION UPREGULATES IQGAP-l EXPRESSION IN PRIMARY CULTURES OF HUMAN ANTRAL EPITHELIAL CELLS. Valerie C. Smith, Mark Ring, Susan B. Curtis, David Scriven, Edwin Dw Moore, Carrie Rosenberger, Brett B. Finlay, Mark R. Meloche, Alison Mj Buchan, Univ of British Columbia, Vancouver, BC, Canada. In normal epithelial cells J3-cateninis located at cell-cell adherens junctions with E-cadherin and o-carenin. In a number of gastric cancers J3-catenin is concentrated in the nucleus and activates the tcf/lef transcription factor stimulating cell proliferation and the production of the matrix rnetaloproteinase family of enzymes. In cell lines the newly identified protein IQGAP-I forms a complex with e-carenm resulting in dissociation of E-cadherin and loss of cell-cell contacts resulting in increased cytosolic J3-catenin.We are currently investigating how H. pylori infection alters the function of antral epithelial cells to cause either ulceration or gastric cancer. For the present experiments primary cultures of human antral epithelial cells (N = 5, 3 male and 2 female, II - 48 yr) were infected with the CCUG 17874 strain of H. pylori for 48h prior to collection of mRNA for gene array experiments, protein for Western blot analysis or fixation of the cells for immunocytochemical studies using deconvolution microscopy. We used the Neuroscience array (Clontech) to monitor changes in expression of genes encoding cell adherence and signal transduction proteins. Antibodies to E-cadherin (Santa Cruz), IQGAP-l (Transduction), J3-and a-catenin (Transduction) were used for the Westerns and immunocytochemistry. H. pylori infection resulted in a 12 fold increase in IQGAP-I and a 3 fold increase in matrix metalloproteinase 16 mRNA levels with no change in E-cadherin. The Western blot analysis showed a 5 fold increase in IQGAP- I protein levels with no change in E-cadherin, 13- or o-catenin. In control cells immunoreactivity for E-cadherin, IQGAP-I, 13and a-catenin was concentrated to the plasma membrane. In infected cells J3-catenin and IQGAP-l were re-distributed to intracellular tubulovesicles with lower levels of immunoreactivity at the plasma membrane. In addition low levels of J3-catenin were observed within the nucleus. Nuclear boundaries was defined using DAPI labeling. E-cadherin staining remained at the plasma membrane, although at a reduced leveL These data indicated that the increase in IQGAP-I levels in cells infected with H. pylori resulted in a re-distribution of J3-catenin within the cells. The presence of J3-cateninin the nucleus would activate the lef/tcf transcription pathway in turn increasing the expression of matrix metalloproteinase 16. The ability of H. pylori to cause nuclear translocation of J3-catenin may be important in the development of gastric cancer.