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531 Laparoscopic Approach to Median Arcuate Ligament Syndrome
531
623
Laparoscopic Approach to Median Arcuate Ligament Syndrome Yves Borbely, Branislav Risti, Othmar Schoeb
Induced Expression of Interaction Molecule 1 (STIM1) Sensitizes Intestinal Epithelial Cells to Apoptosis By Modulating Store-Operated CA2+ Influx Jennifer A. Timmons, Jaladanki N. Rao, Tongtong Zou, Lan Liu, Lan Xiao, Pengyuan Wang, Douglas J. Turner, Jian-Ying Wang
The median arcuate ligament syndrome is a rare entity, characterized by weight loss, chronic abdominal pain and epigastric bruit. It is caused by extrinsic compression on the celiac tripod either by an unusually low inserting fibrous band uniting the cura on either side of the aortic hiatus - the median arcuate ligament- or by a hypertrophic celiac plexus. Illustrated by the case of a 28-year-old woman, we demonstrate the management via laparoscopic approach. After diagnostic laparoscopy, preparation of the celiac tripod with division of the median arcuate ligament and resection of the neural fibers compressing the artery was performed via five trocar sites. Operative time was 110 min.
Apoptosis plays a critical role in the maintenance of gut mucosal epithelial homeostasis and is tightly regulated by numerous factors including intracellular Ca2+. Transient receptor potential canonical-1 (TRPC1) is expressed in intestinal epithelial cells (IECs) and functions as a store-operated Ca2+ channel. We have recently demonstrated that increased TRPC1 activity sensitizes IECs to apoptosis, but the upstream signaling initiating TRPC1 activation remains elusive. The novel protein, STIM1, is shown to act as a store Ca2+ sensor and it can rapidly translocate to the plasma membrane (PM) where it directly interacts with TRPC1. The current study determined whether STIM1 plays an important role in the regulation of IEC apoptosis by activating TRPC1 channel activity. Methods: Studies were conducted in IEC-6 cells (derived from rat intestinal crypts) and stable TRPC1-transfected IECs (IECTRPC1). Apoptosis was induced by tumor necrosis factor-α (TNFα)/cycloheximide (CHX), and intracellular free Ca2+ concentration ([Ca2+]cyt) was measured by fluorescence digital imaging analysis. Functions of STIM1 were investigated by specific siRNA (siSTIM1) and ectopic overexpression of the constitutively active STIM1 EF-hand mutants. Results: Stable STIM1-transfected IEC-6 cells (IEC-STIM1) highly expressed STIM1 protein (~5-folds) and displayed a sustained increase in Ca2+ influx after Ca2+ store depletion (~2-folds). Susceptibility of IEC-STIM1 cells to TNFα/CHX-induced apoptosis increased significantly as measured by changes in morphological features, DNA fragmentation, and caspase-3 activity. Apoptotic cells were increased from ~30% in parental IEC-6 cells to ~70% in stable IEC-STIM1 cells 4 h after exposure to TNFα/CHX. In addition, stable IEC-TRPC1 cells also exhibited an increased sensitivity to TNFα/CHX-induced apoptosis, which was prevented by STIM1 silencing through siSTIM1 transfection. STIM1 silencing by siSTIM1 also decreased Ca2+ influx after store depletion in cells overexpressing TRPC1. Levels of Ca2+ influx due to store depletion were decreased by ~70% in STIM1-silenced populations. Similarly, exposure of IEC-STIM1 cells to the Ca2+ free medium also blocked increased sensitivity to apoptosis. Conclusions: These results indicate that 1) STIM1 plays an important role in the regulation of IEC apoptosis by altering TRPC1 activity and 2) ectopic STIM1 expression sensitizes IECs to apoptosis through induction in TRPC1-mediated Ca2+ influx.
532 Laparoscopic Sigmoid Resection and Rectopexy for Rectal Prolapse: A 3 Step Procedure Philippe Bouchard, Jacques Heppell, Jonathan E. Efron, Tonia Young-Fadok This video illustrates the 3 key steps for a laparoscopic approach for rectal prolapse: complete rectal mobilization, sigmoid resection and rectopexy. The video demonstrates rectal mobilization to the level of the pelvic floor, with nerve preservation. Technical aspects such as vaginal retraction and control of middle hemorrhoidal artery bleeding are presented. Thereafter, the video illustrates sigmoid resection and handsewn anastomosis performed via a small periumbilical incision. Before the rectopexy is performed, important structures and surgical landmarks are shown. The rectopexy technique with the use of a surgical tacker is reviewed. 622
SSAT Abstracts
Triptolide and Anti-Death Receptor 5: An Effective Combination That Activates Both the Intrinsic and Extrinsic Apoptotic Pathways in Pancreatic Cancer Cells Daniel Borja-Cacho, Yumi Yokoyama, Rohit Chugh, Vikas Dudeja, Ashok K. Saluja, Selwyn M. Vickers BACKGROUND Death receptor therapy induces apoptosis in different tumors but pancreatic cancer is resistant to this therapy. We have shown how triptolide decreases apoptosis resistance in pancreatic cancer. We hypothesized that triptolide increases the effectiveness of death receptor therapy in pancreatic cancer. Our aims were to evaluate the effects of combined therapy with triptolide and anti-Death Receptor 5 Antibody (anti-DR5Ab) on a) cell viability and apoptosis and b) anti-apoptotic protein expression in pancreatic cancer cells. METHODS 5 pancreatic cancer cell lines were exposed to triptolide (0-400 nM), antiDR5Ab (0-100 ng/ml), or both (triptolide 100 nM + anti-DR5Ab 12.5 ng/ml). We assessed the effects of both drugs on cell viability, apoptosis, caspase-3, -8 and -9 activity, PARP and BID cleavage. XIAP protein levels were measured after 24 hours of treatment. RESULTS Pancreatic cancer cells were resistant to anti-DR5Ab. Combined therapy decreased cell viability, triggered apoptosis and caspase -3, -8 and -9 activity in all cell lines(Table) PARP and BID cleavage were only seen in combined therapy. Triptolide decreased XIAP levels, an inhibitor of caspase -3 and -9.(Figure) CONCLUSIONS Triptolide decreased XIAP protein levels. Pancreatic cancer cells are resistant to anti-DR5Ab but combined therapy with triptolide and anti-DR5Ab triggers apoptosis because it induces both apoptotic pathways. This effect is seen in anti-DR5 resistant cell lines.
624 Influence of Colonic Bacteria On the Expression of Host Intestinal Tract Genes Which Regulate Water and Electrolyte Absorption Gail Cresci, Muthusamy Thangaraju, Kebin Liu, John D. Mellinger, Vadivel Ganapathy Introduction: We identified a Na+-coupled transporter (SLC5A8) and a G-protein-coupled receptor (GPR109A) for butyrate, a short-chain fatty acid generated in the colon by bacterial fermentation of dietary fiber. GPR109A is coupled to Gi and decreases cellular levels of cAMP. cAMP is diarrheagenic as it inhibits the Na+/H+ and Cl-/HCO3-exchanger activities and stimulates the Cl--channel activity. Butyrate may function as an antidiarrheal agent in the colon through its role as a substrate for SLC5A8 to promote Na+ absorption and as a ligand for GPR109A to reduce cellular levels of cAMP. Since butyrate is produced in the colonic lumen by bacterial fermentation, the presence or absence of bacteria in the colon might influence the expression of SLC5A8 and GPR109A. Hypothesis: SLC5A8 and GPR109A expression in the intestine/colon varies between conventional (CONV) and germ-free (GF) mice. Methods: Age-matched CONV and GF mice were obtained from Taconic and used for experiments upon arrival at our institution. GF mice were also kept in conventional conditions for 0.5-4 weeks for studies involving bacterial re-colonization. RNA and tissue sections were prepared from the intestinal tract and the expression of SLC5A8 and GPR109A was assessed by RT-PCR and immunohistochemistry. Differences in global gene expression among the CONV, GF, and re-colonized mice were explored by microarray analysis. Results: SLC5A8 and GPR109A mRNA and protein levels in colon and ileum were markedly reduced in GF compared to CONV mice, but the expression levels were completely reversed in the re-colonized GF mice. As a positive control, the expressions of SGLT1 and FIAF were simultaneously monitored. It's known that SGLT1 mRNA levels are reduced while FIAF mRNA levels are increased in the ileum in GF compared to CONV mice. Our studies confirmed these earlier findings in the ileum and also showed a similar trend in the colon. DNA microarray analysis indicated altered expression of ~700 genes by more than 2fold in GF compared to CONV mouse colon. These changes were reversed following recolonization. Regarding electrolyte and water absorption, we found down-regulation of DRA, which codes for a Cl-/HCO3- exchanger, and aquaporin 4, which is the primary colonic water channel. Conclusions: These studies show that commensal bacteria promote the expression of several genes whose protein products are responsible for water and electrolyte absorption in the gut (SLC5A8, GPR109A, SGLT1, DRA, AQP4). Thus colonic bacteria are critical for regulating gut water and electrolyte absorption which could have a clinical impact relating to gut inflammation, infectious diarrhea and antibiotic therapy. 625 Expression of Chemokine Receptor CCR9 in Pancreatic Intraepithelial Neoplasia and Its Progression to Invasive Cancer and Liver Metastasis Brian A. Mailey, Xiaoming Shen, Joshua Khalili, Nicelio Sanchez-luege, Andrew M. Lowy, Joseph Kim Introduction: Chemokine receptors may have a critical role in the metastasis and progression of invasive malignancies. There is little data, however, regarding chemokine receptors and premalignant lesions. Our objective was to determine the potential role of chemokine receptors in pancreatic intraepithelial neoplasia (PanIN), the precursor lesion to invasive pancreatic duct cancer. Methods: The PanIN cell line, established from the PDX-1-Cre;LSLK-RASG12D mouse, was assessed by cDNA microarray analysis (Affymetrix Mouse Genome 430 2.0 array) for chemokine receptor expression. The invasive pancreatic duct cancer (5143PDA) and liver metastasis (5143LM) cell lines from the PDX-1-Cre;LSL-K-RASG12D/
SSAT Abstracts
A-874
623
Laparoscopic Approach to Median Arcuate Ligament Syndrome Yves Borbely, Branislav Risti, Othmar Schoeb
Induced Expression of Interaction Molecule 1 (STIM1) Sensitizes Intestinal Epithelial Cells to Apoptosis By Modulating Store-Operated CA2+ Influx Jennifer A. Timmons, Jaladanki N. Rao, Tongtong Zou, Lan Liu, Lan Xiao, Pengyuan Wang, Douglas J. Turner, Jian-Ying Wang
The median arcuate ligament syndrome is a rare entity, characterized by weight loss, chronic abdominal pain and epigastric bruit. It is caused by extrinsic compression on the celiac tripod either by an unusually low inserting fibrous band uniting the cura on either side of the aortic hiatus - the median arcuate ligament- or by a hypertrophic celiac plexus. Illustrated by the case of a 28-year-old woman, we demonstrate the management via laparoscopic approach. After diagnostic laparoscopy, preparation of the celiac tripod with division of the median arcuate ligament and resection of the neural fibers compressing the artery was performed via five trocar sites. Operative time was 110 min.
Apoptosis plays a critical role in the maintenance of gut mucosal epithelial homeostasis and is tightly regulated by numerous factors including intracellular Ca2+. Transient receptor potential canonical-1 (TRPC1) is expressed in intestinal epithelial cells (IECs) and functions as a store-operated Ca2+ channel. We have recently demonstrated that increased TRPC1 activity sensitizes IECs to apoptosis, but the upstream signaling initiating TRPC1 activation remains elusive. The novel protein, STIM1, is shown to act as a store Ca2+ sensor and it can rapidly translocate to the plasma membrane (PM) where it directly interacts with TRPC1. The current study determined whether STIM1 plays an important role in the regulation of IEC apoptosis by activating TRPC1 channel activity. Methods: Studies were conducted in IEC-6 cells (derived from rat intestinal crypts) and stable TRPC1-transfected IECs (IECTRPC1). Apoptosis was induced by tumor necrosis factor-α (TNFα)/cycloheximide (CHX), and intracellular free Ca2+ concentration ([Ca2+]cyt) was measured by fluorescence digital imaging analysis. Functions of STIM1 were investigated by specific siRNA (siSTIM1) and ectopic overexpression of the constitutively active STIM1 EF-hand mutants. Results: Stable STIM1-transfected IEC-6 cells (IEC-STIM1) highly expressed STIM1 protein (~5-folds) and displayed a sustained increase in Ca2+ influx after Ca2+ store depletion (~2-folds). Susceptibility of IEC-STIM1 cells to TNFα/CHX-induced apoptosis increased significantly as measured by changes in morphological features, DNA fragmentation, and caspase-3 activity. Apoptotic cells were increased from ~30% in parental IEC-6 cells to ~70% in stable IEC-STIM1 cells 4 h after exposure to TNFα/CHX. In addition, stable IEC-TRPC1 cells also exhibited an increased sensitivity to TNFα/CHX-induced apoptosis, which was prevented by STIM1 silencing through siSTIM1 transfection. STIM1 silencing by siSTIM1 also decreased Ca2+ influx after store depletion in cells overexpressing TRPC1. Levels of Ca2+ influx due to store depletion were decreased by ~70% in STIM1-silenced populations. Similarly, exposure of IEC-STIM1 cells to the Ca2+ free medium also blocked increased sensitivity to apoptosis. Conclusions: These results indicate that 1) STIM1 plays an important role in the regulation of IEC apoptosis by altering TRPC1 activity and 2) ectopic STIM1 expression sensitizes IECs to apoptosis through induction in TRPC1-mediated Ca2+ influx.
532 Laparoscopic Sigmoid Resection and Rectopexy for Rectal Prolapse: A 3 Step Procedure Philippe Bouchard, Jacques Heppell, Jonathan E. Efron, Tonia Young-Fadok This video illustrates the 3 key steps for a laparoscopic approach for rectal prolapse: complete rectal mobilization, sigmoid resection and rectopexy. The video demonstrates rectal mobilization to the level of the pelvic floor, with nerve preservation. Technical aspects such as vaginal retraction and control of middle hemorrhoidal artery bleeding are presented. Thereafter, the video illustrates sigmoid resection and handsewn anastomosis performed via a small periumbilical incision. Before the rectopexy is performed, important structures and surgical landmarks are shown. The rectopexy technique with the use of a surgical tacker is reviewed. 622
SSAT Abstracts
Triptolide and Anti-Death Receptor 5: An Effective Combination That Activates Both the Intrinsic and Extrinsic Apoptotic Pathways in Pancreatic Cancer Cells Daniel Borja-Cacho, Yumi Yokoyama, Rohit Chugh, Vikas Dudeja, Ashok K. Saluja, Selwyn M. Vickers BACKGROUND Death receptor therapy induces apoptosis in different tumors but pancreatic cancer is resistant to this therapy. We have shown how triptolide decreases apoptosis resistance in pancreatic cancer. We hypothesized that triptolide increases the effectiveness of death receptor therapy in pancreatic cancer. Our aims were to evaluate the effects of combined therapy with triptolide and anti-Death Receptor 5 Antibody (anti-DR5Ab) on a) cell viability and apoptosis and b) anti-apoptotic protein expression in pancreatic cancer cells. METHODS 5 pancreatic cancer cell lines were exposed to triptolide (0-400 nM), antiDR5Ab (0-100 ng/ml), or both (triptolide 100 nM + anti-DR5Ab 12.5 ng/ml). We assessed the effects of both drugs on cell viability, apoptosis, caspase-3, -8 and -9 activity, PARP and BID cleavage. XIAP protein levels were measured after 24 hours of treatment. RESULTS Pancreatic cancer cells were resistant to anti-DR5Ab. Combined therapy decreased cell viability, triggered apoptosis and caspase -3, -8 and -9 activity in all cell lines(Table) PARP and BID cleavage were only seen in combined therapy. Triptolide decreased XIAP levels, an inhibitor of caspase -3 and -9.(Figure) CONCLUSIONS Triptolide decreased XIAP protein levels. Pancreatic cancer cells are resistant to anti-DR5Ab but combined therapy with triptolide and anti-DR5Ab triggers apoptosis because it induces both apoptotic pathways. This effect is seen in anti-DR5 resistant cell lines.
624 Influence of Colonic Bacteria On the Expression of Host Intestinal Tract Genes Which Regulate Water and Electrolyte Absorption Gail Cresci, Muthusamy Thangaraju, Kebin Liu, John D. Mellinger, Vadivel Ganapathy Introduction: We identified a Na+-coupled transporter (SLC5A8) and a G-protein-coupled receptor (GPR109A) for butyrate, a short-chain fatty acid generated in the colon by bacterial fermentation of dietary fiber. GPR109A is coupled to Gi and decreases cellular levels of cAMP. cAMP is diarrheagenic as it inhibits the Na+/H+ and Cl-/HCO3-exchanger activities and stimulates the Cl--channel activity. Butyrate may function as an antidiarrheal agent in the colon through its role as a substrate for SLC5A8 to promote Na+ absorption and as a ligand for GPR109A to reduce cellular levels of cAMP. Since butyrate is produced in the colonic lumen by bacterial fermentation, the presence or absence of bacteria in the colon might influence the expression of SLC5A8 and GPR109A. Hypothesis: SLC5A8 and GPR109A expression in the intestine/colon varies between conventional (CONV) and germ-free (GF) mice. Methods: Age-matched CONV and GF mice were obtained from Taconic and used for experiments upon arrival at our institution. GF mice were also kept in conventional conditions for 0.5-4 weeks for studies involving bacterial re-colonization. RNA and tissue sections were prepared from the intestinal tract and the expression of SLC5A8 and GPR109A was assessed by RT-PCR and immunohistochemistry. Differences in global gene expression among the CONV, GF, and re-colonized mice were explored by microarray analysis. Results: SLC5A8 and GPR109A mRNA and protein levels in colon and ileum were markedly reduced in GF compared to CONV mice, but the expression levels were completely reversed in the re-colonized GF mice. As a positive control, the expressions of SGLT1 and FIAF were simultaneously monitored. It's known that SGLT1 mRNA levels are reduced while FIAF mRNA levels are increased in the ileum in GF compared to CONV mice. Our studies confirmed these earlier findings in the ileum and also showed a similar trend in the colon. DNA microarray analysis indicated altered expression of ~700 genes by more than 2fold in GF compared to CONV mouse colon. These changes were reversed following recolonization. Regarding electrolyte and water absorption, we found down-regulation of DRA, which codes for a Cl-/HCO3- exchanger, and aquaporin 4, which is the primary colonic water channel. Conclusions: These studies show that commensal bacteria promote the expression of several genes whose protein products are responsible for water and electrolyte absorption in the gut (SLC5A8, GPR109A, SGLT1, DRA, AQP4). Thus colonic bacteria are critical for regulating gut water and electrolyte absorption which could have a clinical impact relating to gut inflammation, infectious diarrhea and antibiotic therapy. 625 Expression of Chemokine Receptor CCR9 in Pancreatic Intraepithelial Neoplasia and Its Progression to Invasive Cancer and Liver Metastasis Brian A. Mailey, Xiaoming Shen, Joshua Khalili, Nicelio Sanchez-luege, Andrew M. Lowy, Joseph Kim Introduction: Chemokine receptors may have a critical role in the metastasis and progression of invasive malignancies. There is little data, however, regarding chemokine receptors and premalignant lesions. Our objective was to determine the potential role of chemokine receptors in pancreatic intraepithelial neoplasia (PanIN), the precursor lesion to invasive pancreatic duct cancer. Methods: The PanIN cell line, established from the PDX-1-Cre;LSLK-RASG12D mouse, was assessed by cDNA microarray analysis (Affymetrix Mouse Genome 430 2.0 array) for chemokine receptor expression. The invasive pancreatic duct cancer (5143PDA) and liver metastasis (5143LM) cell lines from the PDX-1-Cre;LSL-K-RASG12D/
SSAT Abstracts
A-874