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Cip1 accelerates senescence and acinar-to-ductal metaplasia formation during pancreatitis
S100
Abstracts / Pancreatology 14 (2014) S1eS129
F-044. Effect of lornoxicam therapy on TLR2, TLR4 mRNA expression in patients with SAP systemic complications
phase. Finally, loss of p21 was accompanied by increased DNA damage and development of senescence. Conclusion: Our findings reveal that p21WAF1/Cip1 is a gate-keeper of acinar cell de-differentiation, ADM formation and prevents activation of senescence program during pancreatic regeneration.
Viktor Gorsky, Mikhail Agapov, Marina Khoreva, Igor Leonenko N.I. Pirogov Russian National Research Medical University, Russia Background: Primary pancreatic injury in SAP is associated with SIRS development that may lead to MODS development, patient’s death. TLRs are considered new target for therapeutic interventions in SAP. Aims: To study expression of TLR2, TLR4 mRNA in mononuclear cells of patients with SAP and lornoxicam effects on TLR2, TLR4 mRNA expression, cytokine secretion and treatment results. Materials & methods: 72 SAP patients with systemic complications divided: 41 standard conservative therapy, 31 additionally received lornoxicam as anti-mediator. Lornoxicam was administered IV 5 days. Total mortality, level of TLR2, TLR4 mRNA expression in MNC were used to assess antimediator therapy efficacy. Results: In 1st group 19.1% patients died due to systemic complications, 6,5% in 2 group (p¼0,006). TLR2 mRNA expression in MNCs in both groups on day 1 was statistically significantly higher than in healthy donors (p¼0,00031). In 1st group further increase in TLR2 mRNA expression was observed by admission day 3. TLR2 mRNA expression in this group remained high on day 7 and 12, there was gradual decrease in TLR2 mRNA expression. TLR2 mRNA expression in 2nd group was statistically significantly lower than in 1st group on the same days (p¼0.007 and p¼0.013, respectively). Expression of TLR4 mRNA in MNCs in both groups was statistically significantly higher than in healthy donors. No statistically significant differences in TLR4 mRNA expression between groups. Conclusion: Patients had high level of TLR2, TLR4 mRNA expression in MNCs during 1st week after AP onset . Lornoxicam suppressed TLR2, TLR4 mRNA in MNCs. Addition of lornoxicam to conservative therapy allowed achieve statistically significant patients mortality decrease.
F-045. Loss of p21WAF1/Cip1 accelerates senescence and acinar-to-ductal metaplasia formation during pancreatitis Kamile Grabliauskaite, Sabrina Sonda, Enrica Saponara, Theresia Reding Graf, Rolf Graf
F-046. The assessment of morphological state of pancreas in experimental pancreonecrosis treated with perfluorocarbon emulsions S.O. Abiola a, P.G. Rasputin a, S.D. Andreeva b, O.V. Mashkovstev b a b
General Surgery Department, Kirov State Medical Academy, Russia Pathology Department, Kirov state Agricultural Academy, Russia
Background: Acute pancreatitis is a relatively common abdominal surgical disease with an annual incidence of 10-20 cases per 100,000, population. The majority of cases are alcohol related or due to gallstones, mortality rate is about 10%, but in its most severe form, which is characterized by pancreatic necrosis, 20%-30% of patients die. Aims: The aim of this study is to investigate the effects of Perftoran e the Russian preparation of the group of Perfluorocarbon (PFC) emulsions on the morphological state of pancreas in Pancreonecrosis (PN). Materials & methods: Rats were divided into 4 groups, the 1st group included rats with surgically modeled PN (upper-median laparotomy with a 1.5 minute cold exposure of the splenic segment of the pancreas),the 2ndgroup included rats with perftoran administered intraperitoneally at a dose of 0.5ml per 100g for a period of 5 days after surgically induced PN, the 3rdgroup included rats who were performed just upper-median laparotomy, the 4thgroup included intact rats.Key parameters evaluated: nuclear area, cytoplasmic area and nuclear-cytoplasmic ratio of acinar cells after the administration of Perftoran, duration of the experiment was 14days. Results: The use of PFC Emulsions resulted in reduced necrobiotic and necrotic process in the parenchyma and stroma of pancreas.There was an increase in nuclear area, acinar cell cytoplasm and nuclear cytoplasm ratio which provides the condition for the realization of the reparative regeneration of epithelial and connective tissue structure and restoration of microcirculatory blood vessels and lymphatic flow. Conclusion: The experiment demonstrates that PFC emulsions are suitable for the treatment of Pancreonecrosis.
University Hospital Zurich, Switzerland Background: Transdifferentiation of pancreatic acinar cells into ductallike lesions, a process defined as acinar-to-ductal metaplasia (ADM), is observed during organ regeneration following pancreatitis. ADM is found in association with pre-malignant PanIN lesions and correlates with an increased risk of pancreatic adenocarcinoma (PDAC). Human PDAC samples show down-regulation of p21WAF1/Cip1, a key regulator of cell cycle and cell differentiation. Aims: We investigated whether p21 down-regulation is implicated in controlling the early events of acinar cell trans-differentiation and ADM formation. Materials & methods: Pancreatitis was induced in wild type (WT) and p21WAF1/Cip1 deficient (p21-/-) mice by multiple injections of cerulein. Recovery from pancreatitis was analyzed in mice one week after termination of cerulein treatment. The expression of proliferation markers, cell cycle regulators, and the severity of tissue inflammation and fibrosis were analyzed by immunohistochemistry, western blotting and qRT-PCR. Results: During pancreatitis, we found that p21WAF1/Cip1 was strongly up-regulated in WT acinar cells but absent in cells forming ADM. p21-/mice showed a significant increase in the number and size of ADM. In addition, the loss of p21 did not increase cell replication rates but resulted in a compensatory activation of positive and negative cell cycle regulators. Furthermore, loss of p21 resulted in increased expression and re-localization of b-catenin during both pancreatitis and subsequent recovery
F-047. Orai1 inhibition prevents calcium toxicity and acute pancreatitis Li Wen a, Svetlana Voronina b, David Collier b, M.Ahsan Javed a, Michael Chvanov b, Muhammad Awais a, John Barrett c, Malcom Begg c, David.N. Criddle b, Alexei Tepikin b, Robert Sutton a a NIHR Liverpool Pancreas Biomedical Research Unit, Royal Liverpool University Hospital, University of Liverpool, United Kingdom b Department of Cellular and Molecular Physiology, Institute of Translational Medicine, University of Liverpool, United Kingdom c Respiratory Therapy Area, GlaxoSmithKline, United Kingdom
Background: Prolonged elevations of intracellular calcium trigger pancreatic acinar cell (PAC) injury contributing to experimental acute pancreatitis (EAP). The elevations depend on external Ca2+ influx through store-operated Ca2+ entry channels that may include Orai1. Aims: To determine the effect of Orai1 inhibition with GSK-7975A on PAC injury and of the pro-drug GSK-6288B on hyperstimulation or biliary EAP. Materials & methods: TIRF and confocal microscopy were used to measure cytosolic Ca2+ (Fura-2) and necrotic pathway activation (PI) changes in freshly isolated murine and human PACs following toxin (TLCS, CCK-8, thapsigargin) administration. EAP was induced by cerulein (50 mg/ kg/h IP x 7) or intraductal infusion of TLCS (50 mL 3 mM). GSK-6288B was administered SC by osmotic mini-pump (2001D) and pharmocokinetic (PK)
Abstracts / Pancreatology 14 (2014) S1eS129
F-044. Effect of lornoxicam therapy on TLR2, TLR4 mRNA expression in patients with SAP systemic complications
phase. Finally, loss of p21 was accompanied by increased DNA damage and development of senescence. Conclusion: Our findings reveal that p21WAF1/Cip1 is a gate-keeper of acinar cell de-differentiation, ADM formation and prevents activation of senescence program during pancreatic regeneration.
Viktor Gorsky, Mikhail Agapov, Marina Khoreva, Igor Leonenko N.I. Pirogov Russian National Research Medical University, Russia Background: Primary pancreatic injury in SAP is associated with SIRS development that may lead to MODS development, patient’s death. TLRs are considered new target for therapeutic interventions in SAP. Aims: To study expression of TLR2, TLR4 mRNA in mononuclear cells of patients with SAP and lornoxicam effects on TLR2, TLR4 mRNA expression, cytokine secretion and treatment results. Materials & methods: 72 SAP patients with systemic complications divided: 41 standard conservative therapy, 31 additionally received lornoxicam as anti-mediator. Lornoxicam was administered IV 5 days. Total mortality, level of TLR2, TLR4 mRNA expression in MNC were used to assess antimediator therapy efficacy. Results: In 1st group 19.1% patients died due to systemic complications, 6,5% in 2 group (p¼0,006). TLR2 mRNA expression in MNCs in both groups on day 1 was statistically significantly higher than in healthy donors (p¼0,00031). In 1st group further increase in TLR2 mRNA expression was observed by admission day 3. TLR2 mRNA expression in this group remained high on day 7 and 12, there was gradual decrease in TLR2 mRNA expression. TLR2 mRNA expression in 2nd group was statistically significantly lower than in 1st group on the same days (p¼0.007 and p¼0.013, respectively). Expression of TLR4 mRNA in MNCs in both groups was statistically significantly higher than in healthy donors. No statistically significant differences in TLR4 mRNA expression between groups. Conclusion: Patients had high level of TLR2, TLR4 mRNA expression in MNCs during 1st week after AP onset . Lornoxicam suppressed TLR2, TLR4 mRNA in MNCs. Addition of lornoxicam to conservative therapy allowed achieve statistically significant patients mortality decrease.
F-045. Loss of p21WAF1/Cip1 accelerates senescence and acinar-to-ductal metaplasia formation during pancreatitis Kamile Grabliauskaite, Sabrina Sonda, Enrica Saponara, Theresia Reding Graf, Rolf Graf
F-046. The assessment of morphological state of pancreas in experimental pancreonecrosis treated with perfluorocarbon emulsions S.O. Abiola a, P.G. Rasputin a, S.D. Andreeva b, O.V. Mashkovstev b a b
General Surgery Department, Kirov State Medical Academy, Russia Pathology Department, Kirov state Agricultural Academy, Russia
Background: Acute pancreatitis is a relatively common abdominal surgical disease with an annual incidence of 10-20 cases per 100,000, population. The majority of cases are alcohol related or due to gallstones, mortality rate is about 10%, but in its most severe form, which is characterized by pancreatic necrosis, 20%-30% of patients die. Aims: The aim of this study is to investigate the effects of Perftoran e the Russian preparation of the group of Perfluorocarbon (PFC) emulsions on the morphological state of pancreas in Pancreonecrosis (PN). Materials & methods: Rats were divided into 4 groups, the 1st group included rats with surgically modeled PN (upper-median laparotomy with a 1.5 minute cold exposure of the splenic segment of the pancreas),the 2ndgroup included rats with perftoran administered intraperitoneally at a dose of 0.5ml per 100g for a period of 5 days after surgically induced PN, the 3rdgroup included rats who were performed just upper-median laparotomy, the 4thgroup included intact rats.Key parameters evaluated: nuclear area, cytoplasmic area and nuclear-cytoplasmic ratio of acinar cells after the administration of Perftoran, duration of the experiment was 14days. Results: The use of PFC Emulsions resulted in reduced necrobiotic and necrotic process in the parenchyma and stroma of pancreas.There was an increase in nuclear area, acinar cell cytoplasm and nuclear cytoplasm ratio which provides the condition for the realization of the reparative regeneration of epithelial and connective tissue structure and restoration of microcirculatory blood vessels and lymphatic flow. Conclusion: The experiment demonstrates that PFC emulsions are suitable for the treatment of Pancreonecrosis.
University Hospital Zurich, Switzerland Background: Transdifferentiation of pancreatic acinar cells into ductallike lesions, a process defined as acinar-to-ductal metaplasia (ADM), is observed during organ regeneration following pancreatitis. ADM is found in association with pre-malignant PanIN lesions and correlates with an increased risk of pancreatic adenocarcinoma (PDAC). Human PDAC samples show down-regulation of p21WAF1/Cip1, a key regulator of cell cycle and cell differentiation. Aims: We investigated whether p21 down-regulation is implicated in controlling the early events of acinar cell trans-differentiation and ADM formation. Materials & methods: Pancreatitis was induced in wild type (WT) and p21WAF1/Cip1 deficient (p21-/-) mice by multiple injections of cerulein. Recovery from pancreatitis was analyzed in mice one week after termination of cerulein treatment. The expression of proliferation markers, cell cycle regulators, and the severity of tissue inflammation and fibrosis were analyzed by immunohistochemistry, western blotting and qRT-PCR. Results: During pancreatitis, we found that p21WAF1/Cip1 was strongly up-regulated in WT acinar cells but absent in cells forming ADM. p21-/mice showed a significant increase in the number and size of ADM. In addition, the loss of p21 did not increase cell replication rates but resulted in a compensatory activation of positive and negative cell cycle regulators. Furthermore, loss of p21 resulted in increased expression and re-localization of b-catenin during both pancreatitis and subsequent recovery
F-047. Orai1 inhibition prevents calcium toxicity and acute pancreatitis Li Wen a, Svetlana Voronina b, David Collier b, M.Ahsan Javed a, Michael Chvanov b, Muhammad Awais a, John Barrett c, Malcom Begg c, David.N. Criddle b, Alexei Tepikin b, Robert Sutton a a NIHR Liverpool Pancreas Biomedical Research Unit, Royal Liverpool University Hospital, University of Liverpool, United Kingdom b Department of Cellular and Molecular Physiology, Institute of Translational Medicine, University of Liverpool, United Kingdom c Respiratory Therapy Area, GlaxoSmithKline, United Kingdom
Background: Prolonged elevations of intracellular calcium trigger pancreatic acinar cell (PAC) injury contributing to experimental acute pancreatitis (EAP). The elevations depend on external Ca2+ influx through store-operated Ca2+ entry channels that may include Orai1. Aims: To determine the effect of Orai1 inhibition with GSK-7975A on PAC injury and of the pro-drug GSK-6288B on hyperstimulation or biliary EAP. Materials & methods: TIRF and confocal microscopy were used to measure cytosolic Ca2+ (Fura-2) and necrotic pathway activation (PI) changes in freshly isolated murine and human PACs following toxin (TLCS, CCK-8, thapsigargin) administration. EAP was induced by cerulein (50 mg/ kg/h IP x 7) or intraductal infusion of TLCS (50 mL 3 mM). GSK-6288B was administered SC by osmotic mini-pump (2001D) and pharmocokinetic (PK)