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M1639 Reciprocal Regulation of E-Cadherin and SRC Homology Phosphotyrosyl Phosphatase 2 (SHP2) By Colonic Extracellular Calcium Sensing Receptor (CaSR)
M1639
AGA Abstracts
Reciprocal Regulation of E-Cadherin and SRC Homology Phosphotyrosyl Phosphatase 2 (SHP2) By Colonic Extracellular Calcium Sensing Receptor (CaSR) Amanda E. Vanderlee, R. John MacLeod BACKGROUND: Colon cancer is characterized by the progressive loss of E-cadherin, a calcium dependent tight-junction component and epithelial marker. Furthermore, inhibition of the phosphatase SHP2, essential in the cell survival signalling via the epidermal growth factor receptor pathway, has been shown to upregulate E-cadherin in breast cancer cell lines. This suggests that SHP2 may promote a cancer cell phenotype. AIM: Define molecular mechanisms by which dietary calcium is chemoprotective against colon cancer by determining the CaSR-mediated determinants responsible for changes in E-cadherin and SHP-2 protein expression. Because CaSR activation inhibits defective Wnt signalling in colon cancer cells, we speculated that there was a relationship between CaSR activation and E-cadherin and SHP2 expression levels. METHODS: A colon cancer cell line which lacks endogenous Ecadherin expression, SW480, was used as a model cell. CaSR levels were manipulated by transient transfection and E-cadherin and SHP2 expression levels were determined by Western blotting. E-cadherin expression was also assessed with RT-PCR and immunohistochemistry. RESULTS: We found that after CaSR activation via calcium [EC50 ~2.6 mM} or other known CaSR agonists (neomycin sulphate, spermine), E-cadherin expression was stimulated and increased and SHP2 expression was substantially decreased. Furthermore, pharmacological inhibition of JNK or ERK had no effect on CaSR-mediated increases of E-cadherin protein in SW480 cells, while inhibition of p38 MAP kinase inhibited Ca2+-induced increases. In contrast, SHP2 decreases observed after Ca2+-treatment were obliterated after pharmalogical inhibition of JNK in the presence of high Ca2+. However, this prevention of SHP2 decrease was not seen after inhibition of ERK or PKC in the presence of Ca2+. CONCLUSION: We conclude that CaSR activation in colonic epithelial cancer cells stimulated increases in Ecadherin transcript and protein, through a mechanism involving p38 MAPK, and reciprocally inhibited SHP2 expression, through a JNK-mediated mechanism. Given the findings of this study, we propose that the chemoprotective nature of dietary calcium supplements against colon cancer may involve reciprocal regulation of E-cadherin and SHP2 via the CaSR. M1640 Helicobacter pylori Infection and Plasma Ghrelin and Gastrin Levels in Gastric Cancer Patients Before and After Gastrectomy Anna Zub, Kazimierz Rembiasz, Stanislaw Konturek, Bartosz Jenner, Peter C. Konturek
The percent change in POD1 adiponectin levels was significantly more for those who developed postoperative infections compared to patients without infections (expressed as the mean ±SEM) (-28.5 ± 2.7 v. -15.8 ± 1.6, p<0.0001, t-test)
Ghrelin and gastrin are produced by the endocrine Gr and G cells of the oxyntic and antral mucosa, respectively, and their plasma levels were reported to be altered by the gastric Helicobacter pylori (Hp) infection but the results were found to be inconclusive. The aim of this study was to determine the alterations in plasma ghrelin and gastrin in gastric cancer (GC) patients before and after gastrectomy and in duodenal ulcer (DU) patients with Hp infection. Methods: The following groups were included; 1. GC patients with Hp infection (n=23), 2. patients with total gastrectomy performed about 5 years ago (n=10), 3. patients with active DU and Hp infection. (n=18), 4. patients with atrophic gastritis without Hp infection (n=10) and 5.control group consisting of 25 healthy volunteers without Hp matched with BMI and age with studied groups. The ghrelin and gastrin concentrations were measured by RIA before and 60 min after the meal. In GC group the hormone measurements were made before and after gastrectomy. Results: Our study shows that presence of Hp significantly influences the ghrelin and gastrin release both in GC and DU groups. Adjusted for age, BMI, sex and the time of measurements (meals) back transformed means of ghrelin and gastrin concentration are: patients with DU have highest level of ghrelin (average 502.6 pg/ ml 95% CI: 285.1 - 886.2), while those with atrophic gastritis and gastrectomy performed in the past had the lowest and similar levels of the hormone (atrophic gastritis 143.7 pg/ ml CI: 93.0 - 222.0; gastrectomy; 150.5 pg/ml CI: 105.3 - 215.2). In GC patients ghrelin concentration was 203.5 pg/ml CI: 160.8 - 257.4, and in control group the value was 255.4 pg/ml CI: 160.5 - 406.3. In cancer group after total gastrectomy ghrelin level was 135 pg/ ml 95% CI: 83-220.2 and after peripheral gastrectomy 220.3 pg/ml 95% CI: 103-273.2. For gastrin, the highest level was detected in group of patients with duodenal ulcer (average 56.3 pM 95% CI: 25-127.3) and in atrophic gastritis group (average 56 pM 95% CI: 30105). In cancer group before operation gastrin value was 29.3 pM 95% CI: 21- 41.3, after total gastrectomy gastrin median level fell to 15.6 pM 95% CI: 11.5-21.3; and after peripheral gastrectomy to17.3 pM 95% CI: 10.7-28.1. In patients gastrectomized in the past, the level of gastrin was 25.4 pM 95% CI: 15.2-42.4. The lowest gastrin level was in control group, average 20.4 pM 95% CI: 14.2-29.2. Conclusions: This study shows that the presence of Hp in the stomach with peptic ulcer increases plasma ghrelin and gastrin levels, whereas in gastric cancer and atrophic gastritis it causes a marked decrease in plasma levels of ghrelin.
M1638 Serotonin Is Not Abnormally Expressed in Uncomplicated Diverticulosis and Has No Association to Bowel Symptoms Santhini Jeyarajah, Nuzhat Akbar, Jane Moorhead, Amyn Haji, Savvas Papagrigoriadis INTRODUCTION: Bowel mucosa contains 95% body serotonin(5-HT). Its action on enteric physiology is unclear.Altered colonic motility due to neurotransmitter imbalance is hypothesised as a pathogenetic mechanism in Diverticular Disease(DD).We previously reported increased 5-HT in sigmoid mucosa of colon resected for complicated DD.We aimed to identify if abnormal 5-HT expression is associated with uncomplicated DD and bowel symptoms. METHODS: This was a prospective comparative study and follow-up survey of symptoms.We examined the differences in 5-HT in normal vs DD patients and bowel symptoms at endoscopy and from 2 years post.Sigmoid biopsies were collected at colonoscopy. Immunocytochemical staining for 5-HT cells was performed on 4μm tissue sections with polyclonal antibody(NCL-SEROTp).Mean number of 5-HT-positive cells per microscopic field was obtained.Staining distribution was defined as low(0-33%),moderate(3366%) and high(>66%) by percentage contrast material/cell.ANOVA and the Kruskall-Wallis tests were used for analysis of parametric and non-parametric data. RESULTS:37(42.5%) DD and 50(57.5%) controls were recruited.No patients underwent surgery.Median age was 55.6(Range 29.3-87.5).Endoscopy was undertaken for rectal bleeding in 39(44.8%),diarrhoea in 23(26.4%),constipation in 14(16.1%),abdominal pain in 26(29.9%).There was no significant difference in mean number of 5-HT cells in DD compared to controls nor between patients and controls with symptoms.Mean 5-HT counts(low & total),confidence intervals(CI) and p values are tabulated.41 patients(47.1%) responded to questionnaires at median 57.8 months from biopsy(Range 20.3-67.3).18(43.9%) were DD and 23(56.1%) controls.10(24.3%) had PR bleeding,6(14.6%) diarrhoea,14(34.2%) constipation,12(29.2%) pain.5-HT counts showed no significant association to persistence of these symptoms. CONCLUSION: Although 5-HT expression in sigmoid mucosa has been found to be increased in complicated DD this does not occur in uncomplicated disease nor is it associated with bowel symptoms. This raises the suggestion that 5-HT may be involved in the development of complications rather than creation of diverticulae. More research may define a role for 5-HT antagonists in preventing DD complications. Table 1: Mean 5-HT counts
M1641 Trefoil Factors: Useful Biomarkers to Differentiate Intestinal and Diffuse Types of Gastric Carcinoma Kristina A. Matkowskyj, Sambasiva Rao, Guang Yu Yang Background: Trefoil peptide/factor family (TFF) is comprised of three peptides which contain 3 intrachain disulfide bonds, forming the trefoil motif. These peptides are secreted by mucusproducing cells of the gastrointestinal tract to protect from injury/promote repair. TFF1 is known to be expressed in the mucous cells of the gastric fundus and basal cells of the antrum/pylorus, while TFF3 is identified within the goblet cells of the small/large bowel. It may play a role in carcinogenesis via involvement in cell scattering, apoptosis and angiogenesis. The alterations of TFF expression in human gastric cancer remain unclear, with conflicting data as one transitions from a benign to malignant phenotype. We set out to analyze the expression pattern and potential diagnostic role of TFF1 and TFF3 in human gastric carcinomas. Design: One hundred six cases including 29 early and 77 advanced stage gastric carcinomas were randomly selected from our Tumor Bank. All specimens were immunohistochemically probed using antibodies recognizing TFF1 and TFF3 and chromogen
AGA Abstracts
A-400
AGA Abstracts
Reciprocal Regulation of E-Cadherin and SRC Homology Phosphotyrosyl Phosphatase 2 (SHP2) By Colonic Extracellular Calcium Sensing Receptor (CaSR) Amanda E. Vanderlee, R. John MacLeod BACKGROUND: Colon cancer is characterized by the progressive loss of E-cadherin, a calcium dependent tight-junction component and epithelial marker. Furthermore, inhibition of the phosphatase SHP2, essential in the cell survival signalling via the epidermal growth factor receptor pathway, has been shown to upregulate E-cadherin in breast cancer cell lines. This suggests that SHP2 may promote a cancer cell phenotype. AIM: Define molecular mechanisms by which dietary calcium is chemoprotective against colon cancer by determining the CaSR-mediated determinants responsible for changes in E-cadherin and SHP-2 protein expression. Because CaSR activation inhibits defective Wnt signalling in colon cancer cells, we speculated that there was a relationship between CaSR activation and E-cadherin and SHP2 expression levels. METHODS: A colon cancer cell line which lacks endogenous Ecadherin expression, SW480, was used as a model cell. CaSR levels were manipulated by transient transfection and E-cadherin and SHP2 expression levels were determined by Western blotting. E-cadherin expression was also assessed with RT-PCR and immunohistochemistry. RESULTS: We found that after CaSR activation via calcium [EC50 ~2.6 mM} or other known CaSR agonists (neomycin sulphate, spermine), E-cadherin expression was stimulated and increased and SHP2 expression was substantially decreased. Furthermore, pharmacological inhibition of JNK or ERK had no effect on CaSR-mediated increases of E-cadherin protein in SW480 cells, while inhibition of p38 MAP kinase inhibited Ca2+-induced increases. In contrast, SHP2 decreases observed after Ca2+-treatment were obliterated after pharmalogical inhibition of JNK in the presence of high Ca2+. However, this prevention of SHP2 decrease was not seen after inhibition of ERK or PKC in the presence of Ca2+. CONCLUSION: We conclude that CaSR activation in colonic epithelial cancer cells stimulated increases in Ecadherin transcript and protein, through a mechanism involving p38 MAPK, and reciprocally inhibited SHP2 expression, through a JNK-mediated mechanism. Given the findings of this study, we propose that the chemoprotective nature of dietary calcium supplements against colon cancer may involve reciprocal regulation of E-cadherin and SHP2 via the CaSR. M1640 Helicobacter pylori Infection and Plasma Ghrelin and Gastrin Levels in Gastric Cancer Patients Before and After Gastrectomy Anna Zub, Kazimierz Rembiasz, Stanislaw Konturek, Bartosz Jenner, Peter C. Konturek
The percent change in POD1 adiponectin levels was significantly more for those who developed postoperative infections compared to patients without infections (expressed as the mean ±SEM) (-28.5 ± 2.7 v. -15.8 ± 1.6, p<0.0001, t-test)
Ghrelin and gastrin are produced by the endocrine Gr and G cells of the oxyntic and antral mucosa, respectively, and their plasma levels were reported to be altered by the gastric Helicobacter pylori (Hp) infection but the results were found to be inconclusive. The aim of this study was to determine the alterations in plasma ghrelin and gastrin in gastric cancer (GC) patients before and after gastrectomy and in duodenal ulcer (DU) patients with Hp infection. Methods: The following groups were included; 1. GC patients with Hp infection (n=23), 2. patients with total gastrectomy performed about 5 years ago (n=10), 3. patients with active DU and Hp infection. (n=18), 4. patients with atrophic gastritis without Hp infection (n=10) and 5.control group consisting of 25 healthy volunteers without Hp matched with BMI and age with studied groups. The ghrelin and gastrin concentrations were measured by RIA before and 60 min after the meal. In GC group the hormone measurements were made before and after gastrectomy. Results: Our study shows that presence of Hp significantly influences the ghrelin and gastrin release both in GC and DU groups. Adjusted for age, BMI, sex and the time of measurements (meals) back transformed means of ghrelin and gastrin concentration are: patients with DU have highest level of ghrelin (average 502.6 pg/ ml 95% CI: 285.1 - 886.2), while those with atrophic gastritis and gastrectomy performed in the past had the lowest and similar levels of the hormone (atrophic gastritis 143.7 pg/ ml CI: 93.0 - 222.0; gastrectomy; 150.5 pg/ml CI: 105.3 - 215.2). In GC patients ghrelin concentration was 203.5 pg/ml CI: 160.8 - 257.4, and in control group the value was 255.4 pg/ml CI: 160.5 - 406.3. In cancer group after total gastrectomy ghrelin level was 135 pg/ ml 95% CI: 83-220.2 and after peripheral gastrectomy 220.3 pg/ml 95% CI: 103-273.2. For gastrin, the highest level was detected in group of patients with duodenal ulcer (average 56.3 pM 95% CI: 25-127.3) and in atrophic gastritis group (average 56 pM 95% CI: 30105). In cancer group before operation gastrin value was 29.3 pM 95% CI: 21- 41.3, after total gastrectomy gastrin median level fell to 15.6 pM 95% CI: 11.5-21.3; and after peripheral gastrectomy to17.3 pM 95% CI: 10.7-28.1. In patients gastrectomized in the past, the level of gastrin was 25.4 pM 95% CI: 15.2-42.4. The lowest gastrin level was in control group, average 20.4 pM 95% CI: 14.2-29.2. Conclusions: This study shows that the presence of Hp in the stomach with peptic ulcer increases plasma ghrelin and gastrin levels, whereas in gastric cancer and atrophic gastritis it causes a marked decrease in plasma levels of ghrelin.
M1638 Serotonin Is Not Abnormally Expressed in Uncomplicated Diverticulosis and Has No Association to Bowel Symptoms Santhini Jeyarajah, Nuzhat Akbar, Jane Moorhead, Amyn Haji, Savvas Papagrigoriadis INTRODUCTION: Bowel mucosa contains 95% body serotonin(5-HT). Its action on enteric physiology is unclear.Altered colonic motility due to neurotransmitter imbalance is hypothesised as a pathogenetic mechanism in Diverticular Disease(DD).We previously reported increased 5-HT in sigmoid mucosa of colon resected for complicated DD.We aimed to identify if abnormal 5-HT expression is associated with uncomplicated DD and bowel symptoms. METHODS: This was a prospective comparative study and follow-up survey of symptoms.We examined the differences in 5-HT in normal vs DD patients and bowel symptoms at endoscopy and from 2 years post.Sigmoid biopsies were collected at colonoscopy. Immunocytochemical staining for 5-HT cells was performed on 4μm tissue sections with polyclonal antibody(NCL-SEROTp).Mean number of 5-HT-positive cells per microscopic field was obtained.Staining distribution was defined as low(0-33%),moderate(3366%) and high(>66%) by percentage contrast material/cell.ANOVA and the Kruskall-Wallis tests were used for analysis of parametric and non-parametric data. RESULTS:37(42.5%) DD and 50(57.5%) controls were recruited.No patients underwent surgery.Median age was 55.6(Range 29.3-87.5).Endoscopy was undertaken for rectal bleeding in 39(44.8%),diarrhoea in 23(26.4%),constipation in 14(16.1%),abdominal pain in 26(29.9%).There was no significant difference in mean number of 5-HT cells in DD compared to controls nor between patients and controls with symptoms.Mean 5-HT counts(low & total),confidence intervals(CI) and p values are tabulated.41 patients(47.1%) responded to questionnaires at median 57.8 months from biopsy(Range 20.3-67.3).18(43.9%) were DD and 23(56.1%) controls.10(24.3%) had PR bleeding,6(14.6%) diarrhoea,14(34.2%) constipation,12(29.2%) pain.5-HT counts showed no significant association to persistence of these symptoms. CONCLUSION: Although 5-HT expression in sigmoid mucosa has been found to be increased in complicated DD this does not occur in uncomplicated disease nor is it associated with bowel symptoms. This raises the suggestion that 5-HT may be involved in the development of complications rather than creation of diverticulae. More research may define a role for 5-HT antagonists in preventing DD complications. Table 1: Mean 5-HT counts
M1641 Trefoil Factors: Useful Biomarkers to Differentiate Intestinal and Diffuse Types of Gastric Carcinoma Kristina A. Matkowskyj, Sambasiva Rao, Guang Yu Yang Background: Trefoil peptide/factor family (TFF) is comprised of three peptides which contain 3 intrachain disulfide bonds, forming the trefoil motif. These peptides are secreted by mucusproducing cells of the gastrointestinal tract to protect from injury/promote repair. TFF1 is known to be expressed in the mucous cells of the gastric fundus and basal cells of the antrum/pylorus, while TFF3 is identified within the goblet cells of the small/large bowel. It may play a role in carcinogenesis via involvement in cell scattering, apoptosis and angiogenesis. The alterations of TFF expression in human gastric cancer remain unclear, with conflicting data as one transitions from a benign to malignant phenotype. We set out to analyze the expression pattern and potential diagnostic role of TFF1 and TFF3 in human gastric carcinomas. Design: One hundred six cases including 29 early and 77 advanced stage gastric carcinomas were randomly selected from our Tumor Bank. All specimens were immunohistochemically probed using antibodies recognizing TFF1 and TFF3 and chromogen
AGA Abstracts
A-400