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Pancreatic ductal fluid- and HCO3-secretion is reduced in aquaporin 1 and 4 knock-out mice
S22
Abstracts / Pancreatology 16 (2016) S1eS130
seems advisable and future studies should identify risk factors and early treatment strategies for NOPD.
Abstract ID: 1547. Genome-wide RNAi screening identified metastasis suppressor genes in an orthotopic pancreatic cancer mouse model Yangchao Chen The Chinese University of Hong Kong, Hong Kong S.A.R., China Introduction: Pancreatic cancer is an aggressive malignancy with extremely poor prognosis. It is usuallly diagnosed when metastases are already present. Aims: To identify genes that play critical roles in the process of pancreatic cancer metastasis, a whole genome RNAi screening was performed. Materials & methods: An shRNA library targeting all human genes was introduced into a human pancreatic cancer cell line Capan-2. The infected cells were then transplanted into the pancreas of nude mice. Because Capan-2 is of low metastatic potential, we hypothesized that knocking down of metastasis suppressor genes would facilitate Capan-2 cells to spread to the liver. By retrieving shRNA templates from the liver metastatic nodules, several candidate genes were found. One fo them Sox9 has been validated as a metastasis suppressor gene in vivo, implying that loss of expression of Sox9 promoted pancreatic cnacer metastasis. Results: In the validation experiments, 10 and 11 nude mice were used for control shCTRL and shSox9 group respectively. All of them had primary tumors. 7 out of 11 mice in shSox9 group developed liver metastasis, whereas only 2 out of 10 metastasized in the control group. Knocking down Sox9 promoted the orthotopic transplanted Capan-2 cells to metastasize to the liver. Conclusion: Our genome wide RNAi screening identified Sox9 as a metastasis suppressor gene in pancreatic cancer.
Abstract ID: 1549. The utility of minichromosome maintenance proteins as novel diagnostic markers in pancreaticobiliary malignancy Margaret G. Keane 1, Matthew T. Huggett 1, Tu Vinh Luong 2, Douglas Thorburn 1, Gavin J. Johnson 3, Michael H. Chapman 3, George J. Webster 3, James Mackay 4, Gareth Williams 5, Stephen P. Pereira 1 1 University College London, Institute for Liver and Digestive Health, United Kingdom 2 Department of Cellular Pathology, Royal Free London NHS Foundation Trust, United Kingdom 3 University College London, NHS Foundation Trust, United Kingdom 4 University College London, Department of Genetics Evolution and Environment, United Kingdom 5 University College London, Wolfson Institute for Biomedical Research, United Kingdom
Introduction: Pancreatic cancer is the fourth commonest cause of cancer death, with a 5-year survival of <4%. Earlier diagnosis and detection of precursor lesions with significant malignant potential would enable timely surgical resection. Minichromosome maintenance proteins (Mcm 2e7) are expressed by actively cycling malignant tumours. Aims: Evaluate if Mcm protein expression can differentiate premalignant and malignant pancreaticobiliary disease. Materials & methods: 1.) Mcm2 protein expression was assessed in formalin-fixed paraffin embedded (FFPE) tissue samples. Sections were immunostained and protein expression analysis performed by determining the labelling index (positive cells / total number of cells). 2.) An automated immunocolormetric assay was used to measure levels of Mcm5 in: (i) 97 biliary brush (BB) samples obtained at endoscopic retrograde cholangiopancreatography and (ii) 28 pancreatic cyst fluid (PCF) samples obtained during endoscopic ultrasound.
Results: FFPE tissue samples from 73 patients with pancreatic cancer, 44 patients with a range of cystic tumours and 9 patients with benign pancreatic disease (normal pancreas or chronic pancreatitis) were stained for Mcm2. Mcm2 protein expression in pancreatic cancer, mucinous cysts, serous cystadenomas and benign disease was 62.2%, 35.5%, 9.3% and 0.4% respectively (p<0.05). Mcm5 levels in BB samples were more sensitive than brush cytology for the detection of malignancy (62% vs. 47%; p<0.05) in patients with an indeterminate biliary stricture. In pancreatic cysts Mcm5 detected malignancy with a sensitivity of 50% (specificity 73%) and measurement was feasible in small and acellular samples. Conclusion: Mcm protein expression can differentiate malignant, premalignant and benign pancreaticobiliary diseases. For BB samples, the Mcm5 assay was superior to cytology.
Abstract ID: 1551, Oral-1. Pancreatic ductal fluid- and HCO3-secretion is reduced in aquaporin 1 and 4 knock-out mice zi 1, Petra Pallagi 2, Anita Bala zs 2, Matthias Sendler 3, Jens Eszter Becskeha n Rakonczay, Jr. 2, P ria Venglovecz 1 eter Hegyi 4, Vikto Kühn 3, Zolta 1 Department of Pharmacology and Pharmacotherapy, University of Szeged, Hungary 2 First Department of Medicine, University of Szeged, Hungary 3 Ernst-Moritz-Arndt-University, Greifswald, Germany 4 cs, Institute for Translational Medicine & Department University of Pe of Translational Medicine/1st Department of Medicine, MTA-SZTE Lendület Translational Gastroenterology Research Group, University of Szeged, Hungary
Introduction: Acute pancreatitis (AP) is a multicellular disease which usually associated with impaired fluid secretion by the ductal cells. Contribution of aquaporins (AQPs) to this fluid secretion is less characterized. Aims: our aim in this study was to determine the role of AQPs in ductal fluid and HCO-3 secretion using AQP knock out (KO) mice. Materials & methods: Intra/interlobular pancreatic ducts were isolated from mice by collagenase digestion. Pancreatic HCO-3 secretion was measured using the Cl- withdrawal technique, whereas the rate of ductal fluid secretion was determined by video microscopy. Pancreatic juice secretion was examined in vivo in anesthetized mice using magnetic resonance imaging cholangiopancreatography. Results: Intracellular pH alkalinisation induced by removal of luminal Cl- was significantly reduced in AQP-1 (42±3,2%) and AQP-4 (52±4,5%) KO vs. wild type (WT) mice. There were no significant changes in the basal fluid secretion between WT and KO ducts. In contrast, stimulation of the ducts with 5 mM forskolin caused swelling of the WT ducts, whereas ducts from AQP-1 and AQP-4 KO mice showed no or only a slight response to forskolin. On retro-orbital injection of 10 U/kg body weight secretin, the increase in total excreted volume (TEV) in WT animals (0.023 TEV/cm3) was significantly higher than in AQP-1 (0.0041 TEV/ cm3) and AQP-4 KO animals (0.0068 TEV/ cm3). Conclusion: Pancreatic fluid and HCO-3 secretion secretion significantly reduced in AQP-1 and AQP-4 KO mice, indicating that AQPs play an essential role in pancreatic fluid secretion. Supported by OTKA (NF105758, NF100677, K109756) and MTA-SZTE Momentum Grant (LP2014-10/2014)
Abstract ID: 1553. Cigarette smoke extract inhibits fluid and HCO3 activity in guinea pig pancreatic ductal cells
secretion and CFTR
ria Venglovecz 2, Krisztina To th 1, Andrea Schnúr 1, Petra Pallagi 1, Vikto n Rakonczay, Jr. 4, Kata zsef Mal } Csupor 3, Zolta th 1, Jo eth 1, Dezso Emese To 5 5 6 } Cseko , Zsuzsanna Helyes , Peter Hegyi
Abstracts / Pancreatology 16 (2016) S1eS130
seems advisable and future studies should identify risk factors and early treatment strategies for NOPD.
Abstract ID: 1547. Genome-wide RNAi screening identified metastasis suppressor genes in an orthotopic pancreatic cancer mouse model Yangchao Chen The Chinese University of Hong Kong, Hong Kong S.A.R., China Introduction: Pancreatic cancer is an aggressive malignancy with extremely poor prognosis. It is usuallly diagnosed when metastases are already present. Aims: To identify genes that play critical roles in the process of pancreatic cancer metastasis, a whole genome RNAi screening was performed. Materials & methods: An shRNA library targeting all human genes was introduced into a human pancreatic cancer cell line Capan-2. The infected cells were then transplanted into the pancreas of nude mice. Because Capan-2 is of low metastatic potential, we hypothesized that knocking down of metastasis suppressor genes would facilitate Capan-2 cells to spread to the liver. By retrieving shRNA templates from the liver metastatic nodules, several candidate genes were found. One fo them Sox9 has been validated as a metastasis suppressor gene in vivo, implying that loss of expression of Sox9 promoted pancreatic cnacer metastasis. Results: In the validation experiments, 10 and 11 nude mice were used for control shCTRL and shSox9 group respectively. All of them had primary tumors. 7 out of 11 mice in shSox9 group developed liver metastasis, whereas only 2 out of 10 metastasized in the control group. Knocking down Sox9 promoted the orthotopic transplanted Capan-2 cells to metastasize to the liver. Conclusion: Our genome wide RNAi screening identified Sox9 as a metastasis suppressor gene in pancreatic cancer.
Abstract ID: 1549. The utility of minichromosome maintenance proteins as novel diagnostic markers in pancreaticobiliary malignancy Margaret G. Keane 1, Matthew T. Huggett 1, Tu Vinh Luong 2, Douglas Thorburn 1, Gavin J. Johnson 3, Michael H. Chapman 3, George J. Webster 3, James Mackay 4, Gareth Williams 5, Stephen P. Pereira 1 1 University College London, Institute for Liver and Digestive Health, United Kingdom 2 Department of Cellular Pathology, Royal Free London NHS Foundation Trust, United Kingdom 3 University College London, NHS Foundation Trust, United Kingdom 4 University College London, Department of Genetics Evolution and Environment, United Kingdom 5 University College London, Wolfson Institute for Biomedical Research, United Kingdom
Introduction: Pancreatic cancer is the fourth commonest cause of cancer death, with a 5-year survival of <4%. Earlier diagnosis and detection of precursor lesions with significant malignant potential would enable timely surgical resection. Minichromosome maintenance proteins (Mcm 2e7) are expressed by actively cycling malignant tumours. Aims: Evaluate if Mcm protein expression can differentiate premalignant and malignant pancreaticobiliary disease. Materials & methods: 1.) Mcm2 protein expression was assessed in formalin-fixed paraffin embedded (FFPE) tissue samples. Sections were immunostained and protein expression analysis performed by determining the labelling index (positive cells / total number of cells). 2.) An automated immunocolormetric assay was used to measure levels of Mcm5 in: (i) 97 biliary brush (BB) samples obtained at endoscopic retrograde cholangiopancreatography and (ii) 28 pancreatic cyst fluid (PCF) samples obtained during endoscopic ultrasound.
Results: FFPE tissue samples from 73 patients with pancreatic cancer, 44 patients with a range of cystic tumours and 9 patients with benign pancreatic disease (normal pancreas or chronic pancreatitis) were stained for Mcm2. Mcm2 protein expression in pancreatic cancer, mucinous cysts, serous cystadenomas and benign disease was 62.2%, 35.5%, 9.3% and 0.4% respectively (p<0.05). Mcm5 levels in BB samples were more sensitive than brush cytology for the detection of malignancy (62% vs. 47%; p<0.05) in patients with an indeterminate biliary stricture. In pancreatic cysts Mcm5 detected malignancy with a sensitivity of 50% (specificity 73%) and measurement was feasible in small and acellular samples. Conclusion: Mcm protein expression can differentiate malignant, premalignant and benign pancreaticobiliary diseases. For BB samples, the Mcm5 assay was superior to cytology.
Abstract ID: 1551, Oral-1. Pancreatic ductal fluid- and HCO3-secretion is reduced in aquaporin 1 and 4 knock-out mice zi 1, Petra Pallagi 2, Anita Bala zs 2, Matthias Sendler 3, Jens Eszter Becskeha n Rakonczay, Jr. 2, P ria Venglovecz 1 eter Hegyi 4, Vikto Kühn 3, Zolta 1 Department of Pharmacology and Pharmacotherapy, University of Szeged, Hungary 2 First Department of Medicine, University of Szeged, Hungary 3 Ernst-Moritz-Arndt-University, Greifswald, Germany 4 cs, Institute for Translational Medicine & Department University of Pe of Translational Medicine/1st Department of Medicine, MTA-SZTE Lendület Translational Gastroenterology Research Group, University of Szeged, Hungary
Introduction: Acute pancreatitis (AP) is a multicellular disease which usually associated with impaired fluid secretion by the ductal cells. Contribution of aquaporins (AQPs) to this fluid secretion is less characterized. Aims: our aim in this study was to determine the role of AQPs in ductal fluid and HCO-3 secretion using AQP knock out (KO) mice. Materials & methods: Intra/interlobular pancreatic ducts were isolated from mice by collagenase digestion. Pancreatic HCO-3 secretion was measured using the Cl- withdrawal technique, whereas the rate of ductal fluid secretion was determined by video microscopy. Pancreatic juice secretion was examined in vivo in anesthetized mice using magnetic resonance imaging cholangiopancreatography. Results: Intracellular pH alkalinisation induced by removal of luminal Cl- was significantly reduced in AQP-1 (42±3,2%) and AQP-4 (52±4,5%) KO vs. wild type (WT) mice. There were no significant changes in the basal fluid secretion between WT and KO ducts. In contrast, stimulation of the ducts with 5 mM forskolin caused swelling of the WT ducts, whereas ducts from AQP-1 and AQP-4 KO mice showed no or only a slight response to forskolin. On retro-orbital injection of 10 U/kg body weight secretin, the increase in total excreted volume (TEV) in WT animals (0.023 TEV/cm3) was significantly higher than in AQP-1 (0.0041 TEV/ cm3) and AQP-4 KO animals (0.0068 TEV/ cm3). Conclusion: Pancreatic fluid and HCO-3 secretion secretion significantly reduced in AQP-1 and AQP-4 KO mice, indicating that AQPs play an essential role in pancreatic fluid secretion. Supported by OTKA (NF105758, NF100677, K109756) and MTA-SZTE Momentum Grant (LP2014-10/2014)
Abstract ID: 1553. Cigarette smoke extract inhibits fluid and HCO3 activity in guinea pig pancreatic ductal cells
secretion and CFTR
ria Venglovecz 2, Krisztina To th 1, Andrea Schnúr 1, Petra Pallagi 1, Vikto n Rakonczay, Jr. 4, Kata zsef Mal } Csupor 3, Zolta th 1, Jo eth 1, Dezso Emese To 5 5 6 } Cseko , Zsuzsanna Helyes , Peter Hegyi