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Mucosal immunization with recombinant homologous or heterologous urease prevents H. Heilmannii infection in mice
April 1998
Esophageal, Gastric, and Duodenal Disorders A103
• G0420 MUCOSAL IMMUNIZATION WITH RECOMBINANT HOMOLOGOUS OR HETEROLOGOUS UREASE PREVENTS H. HEILMANNII INFECTION IN MICE. C, Dieterich, H. Bouzonr~ne*, A, L. Blum, Corth~sv-Theulaz. Division of Gastroenterology, *Pathology Institut, CHUV, Lausanne Switzerland
H. heilmannii (Hh) is a gastric pathogen widely spread in cats, dogs and pigs. It can also infect humans and lead to gastric erosions and cancer, similarly to
H. pylori (Hp). Objectives: To determine whether Hh infection can be prevented by mucosal vaccination and to evaluate the potential benefit to use homologous antigens to immunize against helicobacter spp able to infect humans. Methods: BALB/c mice were nasally immunized 4 times on a weekly schedule with 30lag Hh UreB or 30pg Hp UreAB (kindly provided by OraVax, Ltd, Cambridge, USA), together with 5lag Cholera toxin (CT) or CT only. Twenty days after the last immunization, mice were infected once orally with gastric tissue from Hh infected mice or twice intragastrically with 5x107 n. Jelis (HO. One month later, Helicobacter infection was assessed by Rapid Urease Test (RUT at OD550nrn) and histology. Results: All CT immunized mice were found infected by Hh or H)'~ In contrast, nasal immunization with Hh UreB protected 80% of the mice subsequently exposed to either Hh or Hf. Nasal immunization with tip UreAB elicited protection (90%) a ainst Hh infection. Immunogen [ Hh UreB Hp UreAB i Hh UreB Adjuvant [ CT ~ CT ~ CT i CT . CT ......'in?ection'"'"I...... ]7);............. "fib............. "fiT,............... t/I" .............. i / y ....... Protected/N I 0/9 8/10' 9/10" 0.8 8/10" RUT [ 0.4±0.0 0.0 ± 0.0 0.0 + 0.1 0.4±0.1 0.0 ± 0.1 * p < 0.002 versus CT immunized animals, Fisher's exact test. Conclusions: Recombinant Hh UreB as well as recombinant Hp UreAB protect mice from 1-1. heilmannii and H. Jelis infection. Prevention of H. heilmannii infection can therefore be achieved with the same immunization regimen, i.e. Hp urease, used to prevent 11. pylori and H. Jelis infections. These results might have important implications in veterinarian medicine and lead to a vaccine to prevent H. heilmannii infection in pets. Supported by SNF grants n ° 31-46858.96 and 31-43240.95 G0421 ENDOSCOPY
endoscopic gastritis in detecting the histologic gastritis. As for the chronic erosion with very low PPV for Hp and gastritis, we suggest that their inclusion in the endoscopic classification of gastritis its to be reconsidered. G0422 COMBINATION OF RANITIDINE BISMUTH CITRATE AND CLARITHROMYCIN IN ERADICATING HELICOBACTER PYLORI IN PATIENTS WITH ATROPHIC GASTRITIS. G. Di Felice, M. Schiavo, A. Schicchi, G. Calarco, G. Viola, A. Barbieri, P. Spinelli, National Cancer Institute, Milan (Italy)
It is well established that Helicobacter pylori (Hp) is implicated as a causal agent in active chronic gastritis, and progression of gastritis to chronic atrophic gastritis has been documented by Kujipers (1995). The purpose of this study was to evaluate, in the eradication of Hp in symptomatic patients with atrophic gastritis, the efficacy of a dual therapy (Ranitidine bismuth citrate [RBC] plus clarithromycin), which allows the use of the mncosal protective and antibacterial properties of bismuth-containing compounds. Methods: At pre-study endoscopy, Hp infection was determined by ureasetest and antral - with/without corpus - biopsies, which also allowed histologic assessment. All patients, with histologically documented atrophic gastritis and Hp infection, were included in the study. Hp positive patients received RBC 400 mg bd with clarithromycin 500 mg bd for 14 days followed by 14 days of RBC 400 mg hd. Eight weeks after the end of treatment, a second endoscopy was performed and Hp infection was evaluated with the same modalities. Eradication was assumed if both the tests were negative for Hp. Results: Two hundred and twenty-nine patients with the required characteristics entered the study. Eight weeks from the end of treatment, 187 (81.7%) of them resulted eradicated, 23 (10.0%) had not been eradicated, while 19 (8.3%) were lost to follow-up. If the 19 patients lost to follow-up are assumed "not eradicated" (LF least favourable approach), the percentage of non-eradicated is 18.3% (421229). Considering only the 210 patients analyzable for efficacy (PP approach), the percentage of eradicated patients is 89.1%, while the percentage of non-eradicated ones is 10.9%. No severe adverse events occurred and none of the patients were withdrawn from the study. Mild side-effects were observed in 31 cases (14,7%). Conclusions: The results of our study demonstrate that a dual therapy with RBC and clarithromycin for 2 weeks is effective in the eradication of H. pylori in patients with atrophic gastritis. • G0423
FAILS
IN IDENTIFYING THE PRESENCE OF HELICOBACTER PYLORI (Hp) IN GASTRIC ANTRUM OF ADULTS. G. Di Febo. C. Calabrese, G. Brandi, A.M. Morselli-Labate, A. Areni, M. Gandolfi, G. Biasco, M. Miglioli. Department of Internal Medicine and Gastroenterology, University of Bologna, Italy.
EXTRACELLULAR MATRIX FACTORS LAMININ, FIBRONECTIN AND COLLAGEN IV MODULATE THE ADHESION, RESTITUTION AND PROLIFERATION OF RAT INTESTINAL EPITHELIAL CELLS. A.U. Dignass, A. Becker, H. Goebell. Division of Gastroenterology, Department Of Medicine, University of Essen, Germany
BACKGROUND. Although some studies have suggested that the Hp infection can he determined on the basis of particular endoscopic features only, the relationship between Hp and macroscopic features is still controversial. We carried out a large prospective study to correlate the main endoscopic features of antral mucosa both to the Hp status defined by histology, and to histological aspects. PATIENTS AND METHODS. All patients admitted to our endoscopic service over 3 consecutive months were considered for the study. Patients with previous gastric resection, recent upper GI bleeding, gastric cancer, liver cirrhosis, recent assumption of NSAID or PPI, previous Hp suppressive therapy were excluded. In all patients, the Hp status was unknown. Gastritis was assessed according to Sydney's classification both for endoscopy and for histology. The endoscopic patterns were: normal, erythematous/exudative, chronic erosive, atrophic, nodular. In our work nodularity was considered separately according to its capacity in detecting Hp, as shown in recent study. Four antral biopsies were examined by two pathologists unaware of endoscopic findings. RESULTS. Of the 364 eligible patients (males: 185, females: 179; age: mean 52.4 year, range: 1685), 186 (51.1%) were Hp positive. The prevalence of Hp in five endoscopic patterns was: normal 30.9%, erythematous/exudative 51.9%, chronic erosive 46%, atrophic 46.7%, nodular 69.9%. The distribution of Hp infection was significantly different among the 5 endoscopic patterns (~2=20.1, p < 0.001), being significantly less frequent in the normal (z=2.96, p=0.003) and significantly higher in the nodular (z=3.72, p < 0.001). The presence of endoscopic "gastritis" was significantly higher in Hp positive as compared to Hp negative subjects (90.9 vs. 78.8%; Z2=9.64, p=0.002), with sensitivity, specificity, PPV and NPV of 90.9%, 21.3%, 54.7% and 69.1% respectively. The histological findings were: normal 98 cases (26.9%), chronic non atrophic gastritis 201 (55.2%), atrophic gastritis 65 (17.9%). The capacity of both all gastritis and each abnormal endoscopic pattern to identify an underlying histologic damage was assessed through the Z2 test vs the histologic and macroscopic pattern of normality. A significant correlation between all gastritis vs normality was observed (~2=30.3; p < 0.001), with PPV and NPV of 78.6% (243/309)-and 58.2% (32/55) respectively. In the erythematous-exudative PPV was 70.5%; in chronic erosions PPV=61.7%; in atrophic gastritis PPV=96.7% and in nodularity PPV=91.8% (p<0.001). CONCLUSIONS. Despite the significant relation between some endoscopic patterns and Hp the PPV is low, thus making irrelevant its use as a marker of bacterial presence in clinical practice. We confirm the low specificity of
Backeround: Extracellular matrix factors are assumed to modulate wound healing mechanisms, Rapid regeneration of epithelial wounds after various forms of superficial injury is neccesary to reestablish the integrity of the gastrointestinal mucosa. Three processes are involved in epithelial wound healing. First, the adhesion of epithelial cells, second, the migration of epithelial cells into the wound, a process named restitution and third, the proliferation of epithelial cells. Recently, a TGF[3-dependent upregulation of mRNA coding for extracellular matrix was demonstrated in an vitro model of intestinal injury. Aim: The aim of this study was to investigate how the extracellular matrix factors laminin (Ln), flbronectin (Fn) and collagen IV (Coll IV) modulate the adhesion, migration and proliferation of rat intestinal IEC-6 cells in vitro. In addition, we investigated the influence of extracellular matrix factors on TGFI3-expression. Methods: Adhesion of IEC-6 cells to extracellular matrix factors was determined with a hexosaminidase adhesion assay. The migration of IEC-6 cells on extracellular matrix factors was examined using a stainless steel fence migration model. Effects of Ln, Fn and Coll IV on IEC-6 cell proliferation were assessed using a colorimetric MTr assay. Bioactive and latent TGF[3 was measured with an ELISA method. Resglts: The adhesion of IEC-6 cells to Ln, Fn and Coll IV was significantly enhanced compared to plastic matrix (p <0.001, student's t-test). IEC-6 migraton was generally enhanced by Ln and Fn to 137% and 136% (p < 0.05) compared to control. Migration into zones far from the wound margin was higher for all three matrix factors compared to control. IEC-6 proliferation after 24h was significantly higher for Coll IV (130.5%; p < 0.05), after 48h enhanced for Fn and Coll IV (128.3%, 124.6; p < 0.05) and after 72h reduced for Ln (69.4%; p < 0.01). Binactive and latent TGF[3 peptide levels in supematants of IEC-6 cells after an incubation period of 48h on Ln, Fn and Coll IV were not different from plastic. Conclusions: The extracellular matrix factors Ln, Fn and Coli IV modulate different physiological processes involved in epithelial wound healing. The effects of Ln, Fn and Coll IV are not mediated by changes of TGFI3 peptide expression. This research was funded by the Deutsche Forschungsgemeinschaft (Di 477/3-1).
Esophageal, Gastric, and Duodenal Disorders A103
• G0420 MUCOSAL IMMUNIZATION WITH RECOMBINANT HOMOLOGOUS OR HETEROLOGOUS UREASE PREVENTS H. HEILMANNII INFECTION IN MICE. C, Dieterich, H. Bouzonr~ne*, A, L. Blum, Corth~sv-Theulaz. Division of Gastroenterology, *Pathology Institut, CHUV, Lausanne Switzerland
H. heilmannii (Hh) is a gastric pathogen widely spread in cats, dogs and pigs. It can also infect humans and lead to gastric erosions and cancer, similarly to
H. pylori (Hp). Objectives: To determine whether Hh infection can be prevented by mucosal vaccination and to evaluate the potential benefit to use homologous antigens to immunize against helicobacter spp able to infect humans. Methods: BALB/c mice were nasally immunized 4 times on a weekly schedule with 30lag Hh UreB or 30pg Hp UreAB (kindly provided by OraVax, Ltd, Cambridge, USA), together with 5lag Cholera toxin (CT) or CT only. Twenty days after the last immunization, mice were infected once orally with gastric tissue from Hh infected mice or twice intragastrically with 5x107 n. Jelis (HO. One month later, Helicobacter infection was assessed by Rapid Urease Test (RUT at OD550nrn) and histology. Results: All CT immunized mice were found infected by Hh or H)'~ In contrast, nasal immunization with Hh UreB protected 80% of the mice subsequently exposed to either Hh or Hf. Nasal immunization with tip UreAB elicited protection (90%) a ainst Hh infection. Immunogen [ Hh UreB Hp UreAB i Hh UreB Adjuvant [ CT ~ CT ~ CT i CT . CT ......'in?ection'"'"I...... ]7);............. "fib............. "fiT,............... t/I" .............. i / y ....... Protected/N I 0/9 8/10' 9/10" 0.8 8/10" RUT [ 0.4±0.0 0.0 ± 0.0 0.0 + 0.1 0.4±0.1 0.0 ± 0.1 * p < 0.002 versus CT immunized animals, Fisher's exact test. Conclusions: Recombinant Hh UreB as well as recombinant Hp UreAB protect mice from 1-1. heilmannii and H. Jelis infection. Prevention of H. heilmannii infection can therefore be achieved with the same immunization regimen, i.e. Hp urease, used to prevent 11. pylori and H. Jelis infections. These results might have important implications in veterinarian medicine and lead to a vaccine to prevent H. heilmannii infection in pets. Supported by SNF grants n ° 31-46858.96 and 31-43240.95 G0421 ENDOSCOPY
endoscopic gastritis in detecting the histologic gastritis. As for the chronic erosion with very low PPV for Hp and gastritis, we suggest that their inclusion in the endoscopic classification of gastritis its to be reconsidered. G0422 COMBINATION OF RANITIDINE BISMUTH CITRATE AND CLARITHROMYCIN IN ERADICATING HELICOBACTER PYLORI IN PATIENTS WITH ATROPHIC GASTRITIS. G. Di Felice, M. Schiavo, A. Schicchi, G. Calarco, G. Viola, A. Barbieri, P. Spinelli, National Cancer Institute, Milan (Italy)
It is well established that Helicobacter pylori (Hp) is implicated as a causal agent in active chronic gastritis, and progression of gastritis to chronic atrophic gastritis has been documented by Kujipers (1995). The purpose of this study was to evaluate, in the eradication of Hp in symptomatic patients with atrophic gastritis, the efficacy of a dual therapy (Ranitidine bismuth citrate [RBC] plus clarithromycin), which allows the use of the mncosal protective and antibacterial properties of bismuth-containing compounds. Methods: At pre-study endoscopy, Hp infection was determined by ureasetest and antral - with/without corpus - biopsies, which also allowed histologic assessment. All patients, with histologically documented atrophic gastritis and Hp infection, were included in the study. Hp positive patients received RBC 400 mg bd with clarithromycin 500 mg bd for 14 days followed by 14 days of RBC 400 mg hd. Eight weeks after the end of treatment, a second endoscopy was performed and Hp infection was evaluated with the same modalities. Eradication was assumed if both the tests were negative for Hp. Results: Two hundred and twenty-nine patients with the required characteristics entered the study. Eight weeks from the end of treatment, 187 (81.7%) of them resulted eradicated, 23 (10.0%) had not been eradicated, while 19 (8.3%) were lost to follow-up. If the 19 patients lost to follow-up are assumed "not eradicated" (LF least favourable approach), the percentage of non-eradicated is 18.3% (421229). Considering only the 210 patients analyzable for efficacy (PP approach), the percentage of eradicated patients is 89.1%, while the percentage of non-eradicated ones is 10.9%. No severe adverse events occurred and none of the patients were withdrawn from the study. Mild side-effects were observed in 31 cases (14,7%). Conclusions: The results of our study demonstrate that a dual therapy with RBC and clarithromycin for 2 weeks is effective in the eradication of H. pylori in patients with atrophic gastritis. • G0423
FAILS
IN IDENTIFYING THE PRESENCE OF HELICOBACTER PYLORI (Hp) IN GASTRIC ANTRUM OF ADULTS. G. Di Febo. C. Calabrese, G. Brandi, A.M. Morselli-Labate, A. Areni, M. Gandolfi, G. Biasco, M. Miglioli. Department of Internal Medicine and Gastroenterology, University of Bologna, Italy.
EXTRACELLULAR MATRIX FACTORS LAMININ, FIBRONECTIN AND COLLAGEN IV MODULATE THE ADHESION, RESTITUTION AND PROLIFERATION OF RAT INTESTINAL EPITHELIAL CELLS. A.U. Dignass, A. Becker, H. Goebell. Division of Gastroenterology, Department Of Medicine, University of Essen, Germany
BACKGROUND. Although some studies have suggested that the Hp infection can he determined on the basis of particular endoscopic features only, the relationship between Hp and macroscopic features is still controversial. We carried out a large prospective study to correlate the main endoscopic features of antral mucosa both to the Hp status defined by histology, and to histological aspects. PATIENTS AND METHODS. All patients admitted to our endoscopic service over 3 consecutive months were considered for the study. Patients with previous gastric resection, recent upper GI bleeding, gastric cancer, liver cirrhosis, recent assumption of NSAID or PPI, previous Hp suppressive therapy were excluded. In all patients, the Hp status was unknown. Gastritis was assessed according to Sydney's classification both for endoscopy and for histology. The endoscopic patterns were: normal, erythematous/exudative, chronic erosive, atrophic, nodular. In our work nodularity was considered separately according to its capacity in detecting Hp, as shown in recent study. Four antral biopsies were examined by two pathologists unaware of endoscopic findings. RESULTS. Of the 364 eligible patients (males: 185, females: 179; age: mean 52.4 year, range: 1685), 186 (51.1%) were Hp positive. The prevalence of Hp in five endoscopic patterns was: normal 30.9%, erythematous/exudative 51.9%, chronic erosive 46%, atrophic 46.7%, nodular 69.9%. The distribution of Hp infection was significantly different among the 5 endoscopic patterns (~2=20.1, p < 0.001), being significantly less frequent in the normal (z=2.96, p=0.003) and significantly higher in the nodular (z=3.72, p < 0.001). The presence of endoscopic "gastritis" was significantly higher in Hp positive as compared to Hp negative subjects (90.9 vs. 78.8%; Z2=9.64, p=0.002), with sensitivity, specificity, PPV and NPV of 90.9%, 21.3%, 54.7% and 69.1% respectively. The histological findings were: normal 98 cases (26.9%), chronic non atrophic gastritis 201 (55.2%), atrophic gastritis 65 (17.9%). The capacity of both all gastritis and each abnormal endoscopic pattern to identify an underlying histologic damage was assessed through the Z2 test vs the histologic and macroscopic pattern of normality. A significant correlation between all gastritis vs normality was observed (~2=30.3; p < 0.001), with PPV and NPV of 78.6% (243/309)-and 58.2% (32/55) respectively. In the erythematous-exudative PPV was 70.5%; in chronic erosions PPV=61.7%; in atrophic gastritis PPV=96.7% and in nodularity PPV=91.8% (p<0.001). CONCLUSIONS. Despite the significant relation between some endoscopic patterns and Hp the PPV is low, thus making irrelevant its use as a marker of bacterial presence in clinical practice. We confirm the low specificity of
Backeround: Extracellular matrix factors are assumed to modulate wound healing mechanisms, Rapid regeneration of epithelial wounds after various forms of superficial injury is neccesary to reestablish the integrity of the gastrointestinal mucosa. Three processes are involved in epithelial wound healing. First, the adhesion of epithelial cells, second, the migration of epithelial cells into the wound, a process named restitution and third, the proliferation of epithelial cells. Recently, a TGF[3-dependent upregulation of mRNA coding for extracellular matrix was demonstrated in an vitro model of intestinal injury. Aim: The aim of this study was to investigate how the extracellular matrix factors laminin (Ln), flbronectin (Fn) and collagen IV (Coll IV) modulate the adhesion, migration and proliferation of rat intestinal IEC-6 cells in vitro. In addition, we investigated the influence of extracellular matrix factors on TGFI3-expression. Methods: Adhesion of IEC-6 cells to extracellular matrix factors was determined with a hexosaminidase adhesion assay. The migration of IEC-6 cells on extracellular matrix factors was examined using a stainless steel fence migration model. Effects of Ln, Fn and Coll IV on IEC-6 cell proliferation were assessed using a colorimetric MTr assay. Bioactive and latent TGF[3 was measured with an ELISA method. Resglts: The adhesion of IEC-6 cells to Ln, Fn and Coll IV was significantly enhanced compared to plastic matrix (p <0.001, student's t-test). IEC-6 migraton was generally enhanced by Ln and Fn to 137% and 136% (p < 0.05) compared to control. Migration into zones far from the wound margin was higher for all three matrix factors compared to control. IEC-6 proliferation after 24h was significantly higher for Coll IV (130.5%; p < 0.05), after 48h enhanced for Fn and Coll IV (128.3%, 124.6; p < 0.05) and after 72h reduced for Ln (69.4%; p < 0.01). Binactive and latent TGF[3 peptide levels in supematants of IEC-6 cells after an incubation period of 48h on Ln, Fn and Coll IV were not different from plastic. Conclusions: The extracellular matrix factors Ln, Fn and Coli IV modulate different physiological processes involved in epithelial wound healing. The effects of Ln, Fn and Coll IV are not mediated by changes of TGFI3 peptide expression. This research was funded by the Deutsche Forschungsgemeinschaft (Di 477/3-1).