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Su1904 Effect of REG Protein on Mucosal Permeability and Adhesion Molecule Expression in the Small Intestine
cells reduced TEER by 34.3%, and increased dextran permeability (p<0.05), while lentiviral AFTPH overexpression reduced permeability (p<0.05). Conclusions: Our results indicate that gene silencing of AFTPH in human colonocytes increased epithelial permeability, possibly by regulating expression and localization of genes related to epithelial cell adhesion signaling. These are the first results suggesting that AFTPH may be a new gene regulating intestinal epithelial permeability. Acknowledgement: UCLA Vector Core and UCLA Clinical Microarray Core. Supported by NIH grant DK60729 (CP), P50 DK64539 (Project 2, CP), and the Blinder Research Foundation for Crohn's Disease (IKML).
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Background and aims: Non-invasive methods to screen for Barrett's esophagus (BE) and esophageal adenocarcinoma (EAC) are desirable. Metabolomics involves the systematically study of small molecule metabolites that form unique chemical fingerprints associated with specific cellular or biological processes. We aimed to explore the potential role of metabolomic profiling in differentiating BE and EAC from population controls with and without reflux symptoms. Methods: Fasting plasma samples from 40 patients: 10 each with EAC, BE, Gastroesophageal reflux disease (GERD) and Non-GERD controls were obtained for comprehensive metabolomic profiling. All subjects were residents of Olmsted County, MN. Subjects with EAC and BE had both endoscopic and histologic confirmation of diagnosis. EAC subjects had early (6, 60 %) and locally advanced (4, 40 %) disease. All controls had endoscopy which was negative for BE and EAC. All subjects provided information on reflux symptoms by filling out validated reflux symptoms questionnaires. Metabolomic profiles were generated using liquid chromatography- mass spectrometry (LC-MS) analysis with standard methods by investigators who were blinded to the classification of subjects. Data from the plasma samples corresponding to each of the patient groups were analyzed using an untargeted metabolic profiling approach. Data processing and statistical analysis were performed using Masshunter DA reprocessor and Mass Profiler Professional software (Agilent Technologies Inc.) respectively. Results: Demographic details of subjects in all four groups are shown in table 1. Preliminary analyses showed a separation of the metabolomic profile of subjects with EAC when compared to BE or control subjects. Similar separation was also seen between the metabolomic profile of BE and controls. Figure 1 shows Principal component Analysis (PCA) plots of all the four patient groups and their expressed metabolites. Some of the differentially expressed metabolites were identified using accurate mass obtained by Time of flight- Mass spectrometry analysis (TOF-MS) and by comparing them against the METLIN (Metabolite and Tandem Mass spectrometry) database. These include 2-Tetrahydrothiopheneacetic acid and N-stearoyl tyrosine in patients with EAC which were upregulated when compared with BE as well as non GERD controls. Also, Dihydroshikonofuran, 2-(6'Methylthio)hexylmalic acid, Gibberellin A15 and N,N,N′,N′-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) were upregulated in the BE patient group when compared with non GERD controls. Conclusion: Metabolic profiling of plasma from BE,EAC and control subjects is able to identify distinct metabolites associated with each of the disease states.This may potentially reflect differences in regulatory pathways associated with these conditions and could be utilized in early detection strategies for BE and EAC. Table 1: Baseline characteristics of subjects in the BE, EAC, GERD and Non GERD study groups
Su1904 Effect of REG Protein on Mucosal Permeability and Adhesion Molecule Expression in the Small Intestine Yoshitaka Kitayama, Hirokazu Fukui, Takahisa Yamasaki, Takashi Kondo, Tomoaki Kono, Fumihiko Toyoshima, Hisatomo Ikehara, Yoshio Ohda, Toshihiko Tomita, Tadayuki Oshima, Jiro Watari, Hiroto Miwa Backgrounds: The disturbance of mucosal barrier is a key mechanism to explain how NSAID induces the mucosal injury in the gastrointestinal tract. We have previously reported that REG protein, a tissue regeneration-associated molecule, plays a cytoprotective role in NSAIDinduced mucosal injury in the small intestine. However, it still remains unclear how REG protein confers the resistance to NSAID-induced mucosal injury. Therefore, focusing on the mucosal barrier function, we examined the effect of REG protein on mucosal permeability and adhesion molecule expression in the small intestine. Methods: The expression of adhesion molecules in the small intestine were examined by real-time RT-PCR and immunohistochemistry in Reg I-knockout (KO) and wild-type littermates. Reg I-KO and WT mice received subcutaneous indomethacin (10 mg/kg). Mice were orally administrated with FITCdextran 24 hours after the treatment of indomethacin, and blood samples were collected from the treated mice. The effect of REG protein on epithelial (Caco2 cell) permeability was examined by trans-epithelial electrical resistance (TEER) method. Moreover, Caco2 cells were treated with REG protein and examined in adhesion molecules expression by western blot. Results: The expression of E-cadherin was decreased in Reg I-KO mice compared with WT mice. Serum FITC level was significantly elevated in Reg I-KO mice than in WT one when mice were treated with indomethacin. The level of TEER was significantly increased in REG-treated group than in control. REG protein treatment (10 nM) promoted the phosphorylation of Akt in Caco2 cells and furthermore enhanced the expression of claudin 15, E-cadherin and desmoglein 2 in those cells. Conclusions: REG protein is involved in adhesion molecules expression and maintenance of mucosal permeability in the small intestine. Su1905 Intestinal Adiponectin Receptor 1 (AdipoR1) Modulates Inflammation During Colitis: a Potential Link in Adipose Tissue-Intestinal Crosstalk During Inflammatory Bowel Disease Aristea Sideri, Hon Wai Koon, David Q. Shih, Charalabos Pothoulakis, Iordanis Karagiannidis Background and aims: We presented evidence suggesting that the adiponectin-AdipoR1 axis may play a role in the communication between mesenteric preadipocytes and the intestine during IBD (Gastroenterology 2014;146:S-823). Here, we expanded these studies and examined whether mesenteric whole fat induces inflammatory responses in human colonic epithelial NCM460 cells. The potential intracellular signaling pathways regulating AdipoR1 expression and the role of AdipoR1 in mice with colitis were also investigated. Methods: Conditioned medium from mesenteric fat from Ulcerative colitis (UC), Crohn's disease (CD) and control subjects (n=8-11 individual samples per group) were added to human colonic epithelial NCM460 cell monolayers and cytokine and PPARγ mRNA levels were measured using FlexScript LDA and RT-PCR, respectively. Colonic biopsies of control, UC and CD patients (n=4) were stained for AdipoR1. AdipoR1 mRNA levels in colon tissues from TNBS and vehicle-exposed mice (n=4) were evaluated with RT-PCR. The effect of intracolonic silencing of AdipoR1 (by si- AdipoR1) in the severity (weight, colon length, histologic colitis score) of TNBS-induced colitis (48 hrs) in mice (n=4-6 per group) was also evaluated. Results: VEGFA (p=0.0698), TGFB2 (p<0.05), G-CSF (p=0.0538), and IL8 (p<0.05) were increased while CCL4 (p=0.0541), IFNγ (p=0.0981), and IL-5 (p=0.0814) mRNAs were decreased in colonocytes exposed to UC vs control fat media. IL-8 (p<0.05) and IL-12A (p<0.05) were increased, while IL-1β (p=0.0764), IL-5 (p=0.0732), IL-15 (p= 0.0785), CCL3 (p<0.0897), and PDGFA (p<0.01) mRNAs were decreased in NCM460 cells exposed to CD vs control fat media. Interestingly, VEGFA (p<0.05), TGFB2 (p<0.05), CCL4 (p<0.05) and IL-12A (p=0.0721) levels were different in colonocytes exposed to UC vs CD media. Human colonic biopsies of UC and CD patients and mice with TNBS colitis show higher AdipoR1 levels (p<0.05). PPARγ mRNA levels were decreased in colonocytes exposed to media from UC (p<0.01) and CD (p=0.0832) whole fat media, mirroring the regulation of AdipoR1. Mice with intracolonic AdipoR1 silencing lost more weight and had worse colitis scores (p<0.05). Conclusion: Media from human mesenteric fat of UC and CD patients induce disease-dependent inflammation-related responses in colonocytes, including changes in AdipoR1 levels that may be related to down regulation of PPARγ, a known regulator of AdipoR1. The exacerbation of colitis in mice after intra-colonic silencing of AdipoR1suggests a novel protective role for this receptor in the development of colitis and IBD. Supported by DK047343 (CP), P50 DK64539 (Project 2, CP), The Broad Foundation (BMRP) (IK), and the Blinder Research Foundation for Crohn's Disease (AS).
Figure 1: 3D Score plot of Principal Component Analysis (PCA) of the study groups. [ Key: Gp1 (in red) - Non-GERD controls, Gp2 (in blue) - GERD controls, Gp3 (in brown) - BE and Gp4 (in grey) - EAC group ] Su1913 Activin a Stifles Esophageal Squamous Cell Invasion in a Premalignant but Not Malignant Microenvironment Holli A. Loomans, Chase Taylor, Claudia D. Andl Background: The interactions of cancer cells with their microenvironment can promote invasion and metastasis. Within the tumor microenvironment, fibroblasts facilitate extracellular matrix remodeling, which is a critical component of not only cancer cell invasion. Activin A (Act A) signaling is intertwined with TGFβ signaling and has been implicated as a novel
S-549
AGA Abstracts
AGA Abstracts
Utility of Plasma Metabolomic Profiles in Differentiating Barrett's Esophagus and Esophageal Adenocarcinoma From Control Subjects: A Pilot Study Swarup Kumar, Dhananjay Sakrikar, Michele L. Johnson, Charles Ford, Prasad G. Iyer
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Background and aims: Non-invasive methods to screen for Barrett's esophagus (BE) and esophageal adenocarcinoma (EAC) are desirable. Metabolomics involves the systematically study of small molecule metabolites that form unique chemical fingerprints associated with specific cellular or biological processes. We aimed to explore the potential role of metabolomic profiling in differentiating BE and EAC from population controls with and without reflux symptoms. Methods: Fasting plasma samples from 40 patients: 10 each with EAC, BE, Gastroesophageal reflux disease (GERD) and Non-GERD controls were obtained for comprehensive metabolomic profiling. All subjects were residents of Olmsted County, MN. Subjects with EAC and BE had both endoscopic and histologic confirmation of diagnosis. EAC subjects had early (6, 60 %) and locally advanced (4, 40 %) disease. All controls had endoscopy which was negative for BE and EAC. All subjects provided information on reflux symptoms by filling out validated reflux symptoms questionnaires. Metabolomic profiles were generated using liquid chromatography- mass spectrometry (LC-MS) analysis with standard methods by investigators who were blinded to the classification of subjects. Data from the plasma samples corresponding to each of the patient groups were analyzed using an untargeted metabolic profiling approach. Data processing and statistical analysis were performed using Masshunter DA reprocessor and Mass Profiler Professional software (Agilent Technologies Inc.) respectively. Results: Demographic details of subjects in all four groups are shown in table 1. Preliminary analyses showed a separation of the metabolomic profile of subjects with EAC when compared to BE or control subjects. Similar separation was also seen between the metabolomic profile of BE and controls. Figure 1 shows Principal component Analysis (PCA) plots of all the four patient groups and their expressed metabolites. Some of the differentially expressed metabolites were identified using accurate mass obtained by Time of flight- Mass spectrometry analysis (TOF-MS) and by comparing them against the METLIN (Metabolite and Tandem Mass spectrometry) database. These include 2-Tetrahydrothiopheneacetic acid and N-stearoyl tyrosine in patients with EAC which were upregulated when compared with BE as well as non GERD controls. Also, Dihydroshikonofuran, 2-(6'Methylthio)hexylmalic acid, Gibberellin A15 and N,N,N′,N′-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) were upregulated in the BE patient group when compared with non GERD controls. Conclusion: Metabolic profiling of plasma from BE,EAC and control subjects is able to identify distinct metabolites associated with each of the disease states.This may potentially reflect differences in regulatory pathways associated with these conditions and could be utilized in early detection strategies for BE and EAC. Table 1: Baseline characteristics of subjects in the BE, EAC, GERD and Non GERD study groups
Su1904 Effect of REG Protein on Mucosal Permeability and Adhesion Molecule Expression in the Small Intestine Yoshitaka Kitayama, Hirokazu Fukui, Takahisa Yamasaki, Takashi Kondo, Tomoaki Kono, Fumihiko Toyoshima, Hisatomo Ikehara, Yoshio Ohda, Toshihiko Tomita, Tadayuki Oshima, Jiro Watari, Hiroto Miwa Backgrounds: The disturbance of mucosal barrier is a key mechanism to explain how NSAID induces the mucosal injury in the gastrointestinal tract. We have previously reported that REG protein, a tissue regeneration-associated molecule, plays a cytoprotective role in NSAIDinduced mucosal injury in the small intestine. However, it still remains unclear how REG protein confers the resistance to NSAID-induced mucosal injury. Therefore, focusing on the mucosal barrier function, we examined the effect of REG protein on mucosal permeability and adhesion molecule expression in the small intestine. Methods: The expression of adhesion molecules in the small intestine were examined by real-time RT-PCR and immunohistochemistry in Reg I-knockout (KO) and wild-type littermates. Reg I-KO and WT mice received subcutaneous indomethacin (10 mg/kg). Mice were orally administrated with FITCdextran 24 hours after the treatment of indomethacin, and blood samples were collected from the treated mice. The effect of REG protein on epithelial (Caco2 cell) permeability was examined by trans-epithelial electrical resistance (TEER) method. Moreover, Caco2 cells were treated with REG protein and examined in adhesion molecules expression by western blot. Results: The expression of E-cadherin was decreased in Reg I-KO mice compared with WT mice. Serum FITC level was significantly elevated in Reg I-KO mice than in WT one when mice were treated with indomethacin. The level of TEER was significantly increased in REG-treated group than in control. REG protein treatment (10 nM) promoted the phosphorylation of Akt in Caco2 cells and furthermore enhanced the expression of claudin 15, E-cadherin and desmoglein 2 in those cells. Conclusions: REG protein is involved in adhesion molecules expression and maintenance of mucosal permeability in the small intestine. Su1905 Intestinal Adiponectin Receptor 1 (AdipoR1) Modulates Inflammation During Colitis: a Potential Link in Adipose Tissue-Intestinal Crosstalk During Inflammatory Bowel Disease Aristea Sideri, Hon Wai Koon, David Q. Shih, Charalabos Pothoulakis, Iordanis Karagiannidis Background and aims: We presented evidence suggesting that the adiponectin-AdipoR1 axis may play a role in the communication between mesenteric preadipocytes and the intestine during IBD (Gastroenterology 2014;146:S-823). Here, we expanded these studies and examined whether mesenteric whole fat induces inflammatory responses in human colonic epithelial NCM460 cells. The potential intracellular signaling pathways regulating AdipoR1 expression and the role of AdipoR1 in mice with colitis were also investigated. Methods: Conditioned medium from mesenteric fat from Ulcerative colitis (UC), Crohn's disease (CD) and control subjects (n=8-11 individual samples per group) were added to human colonic epithelial NCM460 cell monolayers and cytokine and PPARγ mRNA levels were measured using FlexScript LDA and RT-PCR, respectively. Colonic biopsies of control, UC and CD patients (n=4) were stained for AdipoR1. AdipoR1 mRNA levels in colon tissues from TNBS and vehicle-exposed mice (n=4) were evaluated with RT-PCR. The effect of intracolonic silencing of AdipoR1 (by si- AdipoR1) in the severity (weight, colon length, histologic colitis score) of TNBS-induced colitis (48 hrs) in mice (n=4-6 per group) was also evaluated. Results: VEGFA (p=0.0698), TGFB2 (p<0.05), G-CSF (p=0.0538), and IL8 (p<0.05) were increased while CCL4 (p=0.0541), IFNγ (p=0.0981), and IL-5 (p=0.0814) mRNAs were decreased in colonocytes exposed to UC vs control fat media. IL-8 (p<0.05) and IL-12A (p<0.05) were increased, while IL-1β (p=0.0764), IL-5 (p=0.0732), IL-15 (p= 0.0785), CCL3 (p<0.0897), and PDGFA (p<0.01) mRNAs were decreased in NCM460 cells exposed to CD vs control fat media. Interestingly, VEGFA (p<0.05), TGFB2 (p<0.05), CCL4 (p<0.05) and IL-12A (p=0.0721) levels were different in colonocytes exposed to UC vs CD media. Human colonic biopsies of UC and CD patients and mice with TNBS colitis show higher AdipoR1 levels (p<0.05). PPARγ mRNA levels were decreased in colonocytes exposed to media from UC (p<0.01) and CD (p=0.0832) whole fat media, mirroring the regulation of AdipoR1. Mice with intracolonic AdipoR1 silencing lost more weight and had worse colitis scores (p<0.05). Conclusion: Media from human mesenteric fat of UC and CD patients induce disease-dependent inflammation-related responses in colonocytes, including changes in AdipoR1 levels that may be related to down regulation of PPARγ, a known regulator of AdipoR1. The exacerbation of colitis in mice after intra-colonic silencing of AdipoR1suggests a novel protective role for this receptor in the development of colitis and IBD. Supported by DK047343 (CP), P50 DK64539 (Project 2, CP), The Broad Foundation (BMRP) (IK), and the Blinder Research Foundation for Crohn's Disease (AS).
Figure 1: 3D Score plot of Principal Component Analysis (PCA) of the study groups. [ Key: Gp1 (in red) - Non-GERD controls, Gp2 (in blue) - GERD controls, Gp3 (in brown) - BE and Gp4 (in grey) - EAC group ] Su1913 Activin a Stifles Esophageal Squamous Cell Invasion in a Premalignant but Not Malignant Microenvironment Holli A. Loomans, Chase Taylor, Claudia D. Andl Background: The interactions of cancer cells with their microenvironment can promote invasion and metastasis. Within the tumor microenvironment, fibroblasts facilitate extracellular matrix remodeling, which is a critical component of not only cancer cell invasion. Activin A (Act A) signaling is intertwined with TGFβ signaling and has been implicated as a novel
S-549
AGA Abstracts
AGA Abstracts
Utility of Plasma Metabolomic Profiles in Differentiating Barrett's Esophagus and Esophageal Adenocarcinoma From Control Subjects: A Pilot Study Swarup Kumar, Dhananjay Sakrikar, Michele L. Johnson, Charles Ford, Prasad G. Iyer