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Su1969 Interleukin-33 and Its Receptor, ST2, Are Dysregulated in Gastritis and Gastric Cancer and May Influence Gastric Epithelial Proliferation
AGA Abstracts
p=0.678; Day 21: 89.81 ± 30.00 vs. 85.58 ± 17.27, p=0.744). RT-PCR detection of cytokine transcripts demonstrated that IL-10 expression prominently decreased at the time of SPEM development, and normalization of IL-10 expression was accompanied by reversible histologic change. Immunofluorescence staining for IL-10 showed cytoplasmic localization IL10. There was no significant change of (INF-γ, IL-12p70, IL-4, IL-5, IL-6, TNF-α, IL17A, and IL-1β level according to development of SPEM. Conclusion: IL-10 may play a pivotal role in tamoxifen-induced acute development of gastric SPEM.
total of 581 patients (323 males and 258 females) revisited for follow-up endoscopy. The mean duration of follow up was 6.0±4.6 years. Out of 581 patients, new gastric cancer developed in 21 cases (20 intestinal-type and 1 diffuse-type). Fifteen were from male, and six were from female. Cumulative incidence of gastric cancer at 5 year was 2.9% in total, 1.5%, 5.2%, and 10% in IM groups A, B, and C, and 0.7%, 1.9% and 11% in mild, moderate, and severe endoscopic atrophy groups, respectively. IM group B, IM group C, and severe endoscopic atrophy were identified as independent predictive factors by multivariate analysis. Compared with IM group A, the Hazard Ratio for IM group B was 4.2 (95% confidence interval: 1.5-13), and that for IM group C was3.5 (1.03-12). Compared with none or mild endoscopic atrophy group, the Hazard Ratio for severe atrophy was 4.5 (1.1-31). Conclusion; Patients with histological intestinal metaplasia or severe endoscopic atrophy were at increased risk for gastric cancer development after H. pylori eradication. These findings may be useful to clarify the high risk patients after H. pylori eradication.
Su1969 Interleukin-33 and Its Receptor, ST2, Are Dysregulated in Gastritis and Gastric Cancer and May Influence Gastric Epithelial Proliferation Laura F. Pisani, Gian Eugenio Tontini, Davide Bona, Luigi Bonavina, Maurizio Vecchi, Luca Pastorelli
Dual immunoflourescent staining co-localized IL-10 expression with parietal cell marker (VEGF-β) and mucus neck cell marker (GSII)
Background and aims: Gastric cancer is still a relevant cause of death. Interleukin (IL)-1 family cytokines play key roles in promoting gastric cancer growth. IL-33, a novel member of the IL-1 family, binds to the receptor ST2 and induces prominent epithelial hyperplasia in the gut. Aim of this study is to characterize IL-33 and ST2 expression modifications throughout the progression from healthy gastric mucosa to cancer and obtain insights on the potential involvement of the IL-33/ST2 axis in regulating gastric epithelial proliferation. Methods: Biopsies from normal gastric mucosa (CONT) (N=7), Helicobacter pylori (Hp)(N=8) and non Hp-related gastritis (N=8), gastric intestinal metaplasia (IM) (N=4), gastric cancer (GC) (N=11) and normal tissue close to cancer lesions (N=11) were collected from patients undergoing upper gastrointestinal endoscopies. IL-33 and ST2 expression was evaluated by WB and qRT-PCR. AGS gastric epithelial cell were cultured with growing concentration of rIL-33 (0 ng/ml-10 ng/ml). Cell proliferation was assessed by means of XTT and CFSE assay using 0, 6 and 24h, or 4, 6 and 8 days' timepoints, respectively. Wound-healing scratch test was performed on AGS monolayers. Caspase-Glo 3/7 luminescent assay evaluated AGS apoptosis. Results: WB for IL-33 showed both full-length (30 kD) and cleaved (22 kD) forms in all the experimental groups; IL-33 mRNA transcripts were increased in non Hp-related gastritis (9.1±3.2 fold increase; p=0.04), compared to CONT (set as 1), not significantly changed in gastric cancer, whereas a trend towards reduction was detected in Hp-related gastritis and IM. WB for ST2 demonstrated both the soluble sST2 and the transmembrane ST2L variants in all the experimental groups; qRT-PCR for sST2 showed no significant changes in the different experimental groups; ST2L mRNA was more abundant in the normal tissue close to cancer lesions (34.2±4.3 fold increase, p=0.05) and in gastric cancer (9.7±4.6 fold increase, p=0.14), compared to CONT (set as 1). The XTT proliferation assay, wound healing experiments and CFSE assay showed an inhibitory effect of IL-33 on AGS in a dose-dependent way. In addition, apoptosis increased consensually to IL-33 concentrations. Conclusions: These data suggest that IL-33/ST2 axis is dysregulated during the progression from gastritis to gastric cancer and may inhibit pathologic gastric epithelial growth.
Su1971 Experimental Validation of a Novel Mutation Gene by Whole Exome Sequence Indicates Poor Survival Outcome of Gastric Cancer Patients Tian-Tian Sun, Haoyan Chen, Jie Hong, Jing-Yuan Fang INTRODUCTION: Genetic modifications play important roles in gastric carcinogenesis. We analyzed the whole exome sequence from 24 gastric cancer tumors and identified a highly mutated gene, temporarily termed as GASX. The role of GASX in gastric cancer is still unclear. AIMS&METHODS: Gene expression profile analyses were performed and Gene Set Enrichment Analysis (GSEA) was used to explore its gene signatures. The expression of GASX in human gastric tissues were measured by Western Blot and immunohistochemical staining. Gastric cancer cells BGC823 and MKN45 cells, which display a lower expression of GASX, were transfected with GASX wild type (WT), GASX (V123L), GASX(R345W), GASX(V346D) or control plasmids and analyzed for the functional roles of GASX, including proliferation and metastasis. The roles of GASX were also clarified by establishing the gastric cancer xenograft model and metastasis models in vivo. RESULTS: Integrated analysis revealed that GASX expression was gradually decreased from normal gastric tissue through to precancerous lesions and then to GC and its expression was negatively correlated with the poor pathologic stage and poor prognosis. Gene Set Enrichment Analysis (GSEA) revealed that cell proliferation and metastasis-related genes were enriched in GASX-low expression patients. Gain of GASX WT function decreased cell proliferation and reduced the cell invasion in BGC823 and MKN45 cells. Overexpression of GASX mutant (V123L, R345W or V346D) resulted in a significant increase in proliferation in BGC823 and MKN45 cells compared with control or GASX WT plasmids transfection. The experiments in vivo showed the same results with the experiments in vitro. The tumours injected with GASX mutant (V123L, R345W or V346D) adenoviruses were bigger than those in control or WT group. More lung metastasis and shorter overall survival were detected after overexpression of GASX mutant (V123L, R345W or V346D) compared with control or WT in vivo. CONCLUSION: We firstly revealed the role of GASX in the progression of gastric cancer and demonstrated for the first time that GASX may be a potential biomarker to predict gastric carcinogenesis. Mutations of GASX gene were associated with gastric carcinogenesis. GASX acts as a tumor suppressor and inhibited cell proliferation and metastasis.
Su1970 Cytokine Profile in Tamoxifen-Induced Murine Model for Spasmolytic Polypeptide-Expressing Metaplasia Hyuk Lee, Sung Noh Hong, Chang Mo Moon, Seoyun Hwang, Young-Ho Kim, Jae J Kim
Su1972
Background and aims: Gastric cancer evolves in the setting of a pathologic mucosal milieu characterized by both loss of acid-secreting parietal cells and mucous cell metaplasia, called as spasmolytic polypeptide-expressing metaplasia (SPEM). Cytokines, such as IL-10, IL-1β, and IL-6 play a key role in the gastric carcinogenesis and their roles have been confirmed in murine models. However, the change of cytokine profile in SPEM, early reversible stage of gastric carcinogenesis, has not been evaluated. Recently it was reported that induction of acute parietal cell loss with tamoxifen leads to the rapid, reversible SPEM in mouse stomach. Because tamoxifen-induced murine model showed SPEM changes occurs within 3 days and gastric histology returns to nearly normal by 3 weeks, it is thought to be suitable to evaluation of changes with cytokine profiling in SPEM and reversible stage. Methods: Intra-peritoneal administration of tamoxifen was performed in eighteen C57BL/6 mice at 8 weeks of age, and 18 control mice served as vehicle only. The mice were sacrificed at 3, 10, and 21 days after treatment. Emerging SPEM lineages in mice treated with tamoxifen were examined for immunolocalization of GS-II and VEGF-β. Multiplex bead array assay for detection of 9 cytokines (INF-γ, IL-12p70, IL-4, IL-5, IL-6, TNF-α, IL-10, IL17A, and IL-1β) performed in the stomach of tamoxifen-treated/control mice. Cytokine expression was confirmed using RT-PCR and immunofluorescence Results: Administration of tamoxifen led to rapid development of SPEM at 3 days, and this murine model showed reversible change followed by normalization of phenotype at 10 days after administration. Multiplex assay revealed that significant decline in level of IL-10 at 3 days of tamoxifen administration (58.38 ± 34.44 pg/ml vs. 94.09 ± 4.98 pg/ml, p=0.031) and normalized at 10 days and 21 days of tamoxifen administration (Day 10: 85.10 ± 31.08 pg/ml vs. 78.99 ± 16.09 pg/ml,
AGA Abstracts
Urine Metabolomics As a Novel Screening Tool for Early Gastric Cancer Ji Won Park, Yun Gyoung Park, Hyuk Lee, Byung-Hoon Min, Jun Haeng Lee, Young-Ho Kim, Sunghyouk Park, Jae J Kim Background/Aims Current the best screening tool of gastric cancer is endoscopy. However it needs adequate facilities, expensive equipment and skilled staff and it is time-consuming and unpleasant to patients. Serum tumor markers such as CEA, CA 19-9, CA 72-4 can be easily used but they are not satisfactory in making a diagnosis and lack clinical efficacy. The purpose of this study was to investigate the differences of urine metabolic profiles between gastric cancer patients and healthy volunteers using a NMR-based metabolomic approach and to provide an effective screening tool for gastric cancer. Methods Urine samples were collected from gastric cancer patients (n=139) and healthy volunteers (n= 156). Nuclear magnetic resonance (NMR) spectra of these urine samples were analyzed using orthogonal partial least square discriminant analysis (OPLS-DA). Using training set composed of gastric cancer patients (n=93) and healthy volunteers (n=105), prediction model was established. Then, the validity of prediction model was assessed using an external validation set consisting of gastric cancer patients (n=46) and healthy volunteers (n=51). In addition, analysis with early stage of gastric cancer (stage Ia, n=79 and stage I, n=97) in the same way were performed. Results The metabolomic 2-D score plot and OPLS-DA multivariate analysis showed good separation between gastric cancer and normal group (Q2 of 72.4%). The prediction model using validation set exhibited 95.7% sensitivity and 94.1% specificity
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p=0.678; Day 21: 89.81 ± 30.00 vs. 85.58 ± 17.27, p=0.744). RT-PCR detection of cytokine transcripts demonstrated that IL-10 expression prominently decreased at the time of SPEM development, and normalization of IL-10 expression was accompanied by reversible histologic change. Immunofluorescence staining for IL-10 showed cytoplasmic localization IL10. There was no significant change of (INF-γ, IL-12p70, IL-4, IL-5, IL-6, TNF-α, IL17A, and IL-1β level according to development of SPEM. Conclusion: IL-10 may play a pivotal role in tamoxifen-induced acute development of gastric SPEM.
total of 581 patients (323 males and 258 females) revisited for follow-up endoscopy. The mean duration of follow up was 6.0±4.6 years. Out of 581 patients, new gastric cancer developed in 21 cases (20 intestinal-type and 1 diffuse-type). Fifteen were from male, and six were from female. Cumulative incidence of gastric cancer at 5 year was 2.9% in total, 1.5%, 5.2%, and 10% in IM groups A, B, and C, and 0.7%, 1.9% and 11% in mild, moderate, and severe endoscopic atrophy groups, respectively. IM group B, IM group C, and severe endoscopic atrophy were identified as independent predictive factors by multivariate analysis. Compared with IM group A, the Hazard Ratio for IM group B was 4.2 (95% confidence interval: 1.5-13), and that for IM group C was3.5 (1.03-12). Compared with none or mild endoscopic atrophy group, the Hazard Ratio for severe atrophy was 4.5 (1.1-31). Conclusion; Patients with histological intestinal metaplasia or severe endoscopic atrophy were at increased risk for gastric cancer development after H. pylori eradication. These findings may be useful to clarify the high risk patients after H. pylori eradication.
Su1969 Interleukin-33 and Its Receptor, ST2, Are Dysregulated in Gastritis and Gastric Cancer and May Influence Gastric Epithelial Proliferation Laura F. Pisani, Gian Eugenio Tontini, Davide Bona, Luigi Bonavina, Maurizio Vecchi, Luca Pastorelli
Dual immunoflourescent staining co-localized IL-10 expression with parietal cell marker (VEGF-β) and mucus neck cell marker (GSII)
Background and aims: Gastric cancer is still a relevant cause of death. Interleukin (IL)-1 family cytokines play key roles in promoting gastric cancer growth. IL-33, a novel member of the IL-1 family, binds to the receptor ST2 and induces prominent epithelial hyperplasia in the gut. Aim of this study is to characterize IL-33 and ST2 expression modifications throughout the progression from healthy gastric mucosa to cancer and obtain insights on the potential involvement of the IL-33/ST2 axis in regulating gastric epithelial proliferation. Methods: Biopsies from normal gastric mucosa (CONT) (N=7), Helicobacter pylori (Hp)(N=8) and non Hp-related gastritis (N=8), gastric intestinal metaplasia (IM) (N=4), gastric cancer (GC) (N=11) and normal tissue close to cancer lesions (N=11) were collected from patients undergoing upper gastrointestinal endoscopies. IL-33 and ST2 expression was evaluated by WB and qRT-PCR. AGS gastric epithelial cell were cultured with growing concentration of rIL-33 (0 ng/ml-10 ng/ml). Cell proliferation was assessed by means of XTT and CFSE assay using 0, 6 and 24h, or 4, 6 and 8 days' timepoints, respectively. Wound-healing scratch test was performed on AGS monolayers. Caspase-Glo 3/7 luminescent assay evaluated AGS apoptosis. Results: WB for IL-33 showed both full-length (30 kD) and cleaved (22 kD) forms in all the experimental groups; IL-33 mRNA transcripts were increased in non Hp-related gastritis (9.1±3.2 fold increase; p=0.04), compared to CONT (set as 1), not significantly changed in gastric cancer, whereas a trend towards reduction was detected in Hp-related gastritis and IM. WB for ST2 demonstrated both the soluble sST2 and the transmembrane ST2L variants in all the experimental groups; qRT-PCR for sST2 showed no significant changes in the different experimental groups; ST2L mRNA was more abundant in the normal tissue close to cancer lesions (34.2±4.3 fold increase, p=0.05) and in gastric cancer (9.7±4.6 fold increase, p=0.14), compared to CONT (set as 1). The XTT proliferation assay, wound healing experiments and CFSE assay showed an inhibitory effect of IL-33 on AGS in a dose-dependent way. In addition, apoptosis increased consensually to IL-33 concentrations. Conclusions: These data suggest that IL-33/ST2 axis is dysregulated during the progression from gastritis to gastric cancer and may inhibit pathologic gastric epithelial growth.
Su1971 Experimental Validation of a Novel Mutation Gene by Whole Exome Sequence Indicates Poor Survival Outcome of Gastric Cancer Patients Tian-Tian Sun, Haoyan Chen, Jie Hong, Jing-Yuan Fang INTRODUCTION: Genetic modifications play important roles in gastric carcinogenesis. We analyzed the whole exome sequence from 24 gastric cancer tumors and identified a highly mutated gene, temporarily termed as GASX. The role of GASX in gastric cancer is still unclear. AIMS&METHODS: Gene expression profile analyses were performed and Gene Set Enrichment Analysis (GSEA) was used to explore its gene signatures. The expression of GASX in human gastric tissues were measured by Western Blot and immunohistochemical staining. Gastric cancer cells BGC823 and MKN45 cells, which display a lower expression of GASX, were transfected with GASX wild type (WT), GASX (V123L), GASX(R345W), GASX(V346D) or control plasmids and analyzed for the functional roles of GASX, including proliferation and metastasis. The roles of GASX were also clarified by establishing the gastric cancer xenograft model and metastasis models in vivo. RESULTS: Integrated analysis revealed that GASX expression was gradually decreased from normal gastric tissue through to precancerous lesions and then to GC and its expression was negatively correlated with the poor pathologic stage and poor prognosis. Gene Set Enrichment Analysis (GSEA) revealed that cell proliferation and metastasis-related genes were enriched in GASX-low expression patients. Gain of GASX WT function decreased cell proliferation and reduced the cell invasion in BGC823 and MKN45 cells. Overexpression of GASX mutant (V123L, R345W or V346D) resulted in a significant increase in proliferation in BGC823 and MKN45 cells compared with control or GASX WT plasmids transfection. The experiments in vivo showed the same results with the experiments in vitro. The tumours injected with GASX mutant (V123L, R345W or V346D) adenoviruses were bigger than those in control or WT group. More lung metastasis and shorter overall survival were detected after overexpression of GASX mutant (V123L, R345W or V346D) compared with control or WT in vivo. CONCLUSION: We firstly revealed the role of GASX in the progression of gastric cancer and demonstrated for the first time that GASX may be a potential biomarker to predict gastric carcinogenesis. Mutations of GASX gene were associated with gastric carcinogenesis. GASX acts as a tumor suppressor and inhibited cell proliferation and metastasis.
Su1970 Cytokine Profile in Tamoxifen-Induced Murine Model for Spasmolytic Polypeptide-Expressing Metaplasia Hyuk Lee, Sung Noh Hong, Chang Mo Moon, Seoyun Hwang, Young-Ho Kim, Jae J Kim
Su1972
Background and aims: Gastric cancer evolves in the setting of a pathologic mucosal milieu characterized by both loss of acid-secreting parietal cells and mucous cell metaplasia, called as spasmolytic polypeptide-expressing metaplasia (SPEM). Cytokines, such as IL-10, IL-1β, and IL-6 play a key role in the gastric carcinogenesis and their roles have been confirmed in murine models. However, the change of cytokine profile in SPEM, early reversible stage of gastric carcinogenesis, has not been evaluated. Recently it was reported that induction of acute parietal cell loss with tamoxifen leads to the rapid, reversible SPEM in mouse stomach. Because tamoxifen-induced murine model showed SPEM changes occurs within 3 days and gastric histology returns to nearly normal by 3 weeks, it is thought to be suitable to evaluation of changes with cytokine profiling in SPEM and reversible stage. Methods: Intra-peritoneal administration of tamoxifen was performed in eighteen C57BL/6 mice at 8 weeks of age, and 18 control mice served as vehicle only. The mice were sacrificed at 3, 10, and 21 days after treatment. Emerging SPEM lineages in mice treated with tamoxifen were examined for immunolocalization of GS-II and VEGF-β. Multiplex bead array assay for detection of 9 cytokines (INF-γ, IL-12p70, IL-4, IL-5, IL-6, TNF-α, IL-10, IL17A, and IL-1β) performed in the stomach of tamoxifen-treated/control mice. Cytokine expression was confirmed using RT-PCR and immunofluorescence Results: Administration of tamoxifen led to rapid development of SPEM at 3 days, and this murine model showed reversible change followed by normalization of phenotype at 10 days after administration. Multiplex assay revealed that significant decline in level of IL-10 at 3 days of tamoxifen administration (58.38 ± 34.44 pg/ml vs. 94.09 ± 4.98 pg/ml, p=0.031) and normalized at 10 days and 21 days of tamoxifen administration (Day 10: 85.10 ± 31.08 pg/ml vs. 78.99 ± 16.09 pg/ml,
AGA Abstracts
Urine Metabolomics As a Novel Screening Tool for Early Gastric Cancer Ji Won Park, Yun Gyoung Park, Hyuk Lee, Byung-Hoon Min, Jun Haeng Lee, Young-Ho Kim, Sunghyouk Park, Jae J Kim Background/Aims Current the best screening tool of gastric cancer is endoscopy. However it needs adequate facilities, expensive equipment and skilled staff and it is time-consuming and unpleasant to patients. Serum tumor markers such as CEA, CA 19-9, CA 72-4 can be easily used but they are not satisfactory in making a diagnosis and lack clinical efficacy. The purpose of this study was to investigate the differences of urine metabolic profiles between gastric cancer patients and healthy volunteers using a NMR-based metabolomic approach and to provide an effective screening tool for gastric cancer. Methods Urine samples were collected from gastric cancer patients (n=139) and healthy volunteers (n= 156). Nuclear magnetic resonance (NMR) spectra of these urine samples were analyzed using orthogonal partial least square discriminant analysis (OPLS-DA). Using training set composed of gastric cancer patients (n=93) and healthy volunteers (n=105), prediction model was established. Then, the validity of prediction model was assessed using an external validation set consisting of gastric cancer patients (n=46) and healthy volunteers (n=51). In addition, analysis with early stage of gastric cancer (stage Ia, n=79 and stage I, n=97) in the same way were performed. Results The metabolomic 2-D score plot and OPLS-DA multivariate analysis showed good separation between gastric cancer and normal group (Q2 of 72.4%). The prediction model using validation set exhibited 95.7% sensitivity and 94.1% specificity
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