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Sastre J, Pérez S, Sabater L, Rius-Pérez S. Redox signaling in the pancreas in health and disease. Physiol Rev 2025; 105:593-650. [PMID: 39324871 DOI: 10.1152/physrev.00044.2023] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2023] [Revised: 09/11/2024] [Accepted: 09/17/2024] [Indexed: 09/27/2024] Open
Abstract
This review addresses oxidative stress and redox signaling in the pancreas under healthy physiological conditions as well as in acute pancreatitis, chronic pancreatitis, pancreatic cancer, and diabetes. Physiological redox homeodynamics is maintained mainly by NRF2/KEAP1, NF-κB, protein tyrosine phosphatases, peroxisome proliferator-activated receptor-γ coactivator 1α (PGC1α), and normal autophagy. Depletion of reduced glutathione (GSH) in the pancreas is a hallmark of acute pancreatitis and is initially accompanied by disulfide stress, which is characterized by protein cysteinylation without increased glutathione oxidation. A cross talk between oxidative stress, MAPKs, and NF-κB amplifies the inflammatory cascade, with PP2A and PGC1α as key redox regulatory nodes. In acute pancreatitis, nitration of cystathionine-β synthase causes blockade of the transsulfuration pathway leading to increased homocysteine levels, whereas p53 triggers necroptosis in the pancreas through downregulation of sulfiredoxin, PGC1α, and peroxiredoxin 3. Chronic pancreatitis exhibits oxidative distress mediated by NADPH oxidase 1 and/or CYP2E1, which promotes cell death, fibrosis, and inflammation. Oxidative stress cooperates with mutant KRAS to initiate and promote pancreatic adenocarcinoma. Mutant KRAS increases mitochondrial reactive oxygen species (ROS), which trigger acinar-to-ductal metaplasia and progression to pancreatic intraepithelial neoplasia (PanIN). ROS are maintained at a sufficient level to promote cell proliferation, while avoiding cell death or senescence through formation of NADPH and GSH and activation of NRF2, HIF-1/2α, and CREB. Redox signaling also plays a fundamental role in differentiation, proliferation, and insulin secretion of β-cells. However, ROS overproduction promotes β-cell dysfunction and apoptosis in type 1 and type 2 diabetes.
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Affiliation(s)
- Juan Sastre
- Department of Physiology, Faculty of Pharmacy, University of Valencia, Valencia, Spain
| | - Salvador Pérez
- Department of Physiology, Faculty of Pharmacy, University of Valencia, Valencia, Spain
| | - Luis Sabater
- Liver, Biliary and Pancreatic Unit, Hospital Clínico, Department of Surgery, Faculty of Medicine, University of Valencia, Valencia, Spain
| | - Sergio Rius-Pérez
- Department of Physiology, Faculty of Pharmacy, University of Valencia, Valencia, Spain
- Department of Cell Biology, Functional Biology and Physical Anthropology, Faculty of Biology, University of Valencia, Valencia, Spain
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Swetha K, Indumathi MC, Kishan R, Siddappa S, Chen CH, Marathe GK. Selenium Mitigates Caerulein and LPS-induced Severe Acute Pancreatitis by Inhibiting MAPK, NF-κB, and STAT3 Signaling via the Nrf2/HO-1 Pathway. Biol Trace Elem Res 2025:10.1007/s12011-025-04531-2. [PMID: 39907886 DOI: 10.1007/s12011-025-04531-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/29/2024] [Accepted: 01/20/2025] [Indexed: 02/06/2025]
Abstract
Severe acute pancreatitis (SAP) leads to systemic inflammation, resulting in multiorgan damage. Acute lung injury and acute respiratory distress syndrome develop in one-third of SAP patients, with a high mortality rate of 60% due to secondary complications. Patients with pancreatitis often have selenium deficiency, and selenium supplements may provide beneficial effects. This study examined the protective role of selenium in a model of SAP induced by caerulein + lipopolysaccharide (cae + LPS). Mice were administered selenium (1 mg/kg) before being challenged with caerulein (6 injections of 50 μg/kg) and LPS (10 mg/kg). At 3 h after the last caerulein injection, blood was collected for estimating pancreatic enzymes and cytokine levels, and the mice were euthanized. We performed morphological and histological studies, measured levels of protease and oxidative stress markers and conducted western blot, ELISA, and RT-qPCR analyses. We examined lung tissue histologically and estimated myeloperoxidase levels. Selenium pretreatment significantly reduced pancreatic enzyme levels such as amylase, lipase, and proteases (specifically MMPs) and reversed tissue injury in the pancreas and lungs caused by cae + LPS. In addition, selenium-treated mice showed decreased levels of inflammatory markers and chemokines. Examination of the downstream inflammatory pathways confirmed the protective effect of selenium, which mediates its anti-inflammatory and antioxidant action by inhibiting the major inflammatory signaling pathways (MAPKs, NF-κB, and STAT3) and activating the phosphorylation of Nrf2 via Nrf2/HO-1 pathways. These findings suggest that selenium may be a potential therapeutic option for treating SAP-associated secondary complications.
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Affiliation(s)
- Kamatam Swetha
- Department of Studies in Biochemistry, University of Mysore, Manasagangotri Mysore, 570006, India
| | | | - Raju Kishan
- Department of Studies in Molecular Biology, University of Mysore, Manasagangotri Mysore, 570006, India
| | - Shiva Siddappa
- Division of Biochemistry, School of Life Sciences, JSS Academy of Higher Education and Research, Mysore, 570015, India
| | - Chu-Huang Chen
- Vascular and Medicinal Research, The Texas Heart Institute, Houston, TX, 77030, USA
| | - Gopal K Marathe
- Department of Studies in Biochemistry, University of Mysore, Manasagangotri Mysore, 570006, India.
- Department of Studies in Molecular Biology, University of Mysore, Manasagangotri Mysore, 570006, India.
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Williams JA. Cholecystokinin (CCK) Regulation of Pancreatic Acinar Cells: Physiological Actions and Signal Transduction Mechanisms. Compr Physiol 2019; 9:535-564. [PMID: 30873601 DOI: 10.1002/cphy.c180014] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Pancreatic acinar cells synthesize and secrete about 20 digestive enzymes and ancillary proteins with the processes that match the supply of these enzymes to their need in digestion being regulated by a number of hormones (CCK, secretin and insulin), neurotransmitters (acetylcholine and VIP) and growth factors (EGF and IGF). Of these regulators, one of the most important and best studied is the gastrointestinal hormone, cholecystokinin (CCK). Furthermore, the acinar cell has become a model for seven transmembrane, heterotrimeric G protein coupled receptors to regulate multiple processes by distinct signal transduction cascades. In this review, we briefly describe the chemistry and physiology of CCK and then consider the major physiological effects of CCK on pancreatic acinar cells. The majority of the review is devoted to the physiologic signaling pathways activated by CCK receptors and heterotrimeric G proteins and the functions they affect. The pathways covered include the traditional second messenger pathways PLC-IP3-Ca2+ , DAG-PKC, and AC-cAMP-PKA/EPAC that primarily relate to secretion. Then there are the protein-protein interaction pathways Akt-mTOR-S6K, the three major MAPK pathways (ERK, JNK, and p38 MAPK), and Ca2+ -calcineurin-NFAT pathways that primarily regulate non-secretory processes including biosynthesis and growth, and several miscellaneous pathways that include the Rho family small G proteins, PKD, FAK, and Src that may regulate both secretory and nonsecretory processes but are not as well understood. © 2019 American Physiological Society. Compr Physiol 9:535-564, 2019.
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Affiliation(s)
- John A Williams
- University of Michigan, Departments of Molecular & Integrative Physiology and Internal Medicine (Gastroenterology), Ann Arbor, Michigan, USA
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Yang Z, Yang W, Lu M, Li Z, Qiao X, Sun B, Zhang W, Xue D. Role of the c-Jun N-terminal kinase signaling pathway in the activation of trypsinogen in rat pancreatic acinar cells. Int J Mol Med 2017; 41:1119-1126. [PMID: 29207022 DOI: 10.3892/ijmm.2017.3266] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2016] [Accepted: 11/08/2017] [Indexed: 11/06/2022] Open
Abstract
Bile acid causes trypsinogen activation in pancreatic acinar cells through a complex process. Additional research is required to further elucidate which signaling pathways affect trypsinogen activation when activated. the changes in the whole‑genome expression profile of AR42J cells under the effect of taurolithocholic acid 3‑sulfate (TLC‑S) were investigated. Furthermore, gene groups that may play a regulatory role were analyzed using the modular approach of biological networks. The aim of the present study was to improve our understanding of the changes in TLC‑S‑stimulated AR42J cells through a genetic functional modular analysis. whole‑genome expression profile chip arrays were applied to detect genes that were differentially expressed in pancreatic acinar AR42J cells treated with TLC‑S for 20 min. Based on the human protein reference database, a protein‑protein interaction network was obtained, which was then processed by CFinder software to derive 14 modules. Among these 14 modules, the gene ontology biological processes enrichment analysis identified two as modules of interest. Kyoto encyclopedia of genes and genomes map analysis revealed that MAP2K4, MAPK8 and FLNA are part of the c-Jun N-terminal kinase (JNK) pathway. The JNK signaling pathway is involved in regulating trypsinogen activation in rat pancreatic AR42J cells. Next, a regulatory network of seven kinase inhibitors was constructed. SP600125 is an ATP‑competitive, efficient, selective and reversible inhibitor of JNK. the results were verified by four sets of experiments and demonstrated that trypsinogen activation is mediated by the JNK signaling pathway in the pathogenesis of acute pancreatitis (AP). The present study provided a useful reference for better understanding the pathogenesis of AP and identifying new targets to regulate trypsinogen activation, in addition to providing valuable information for the treatment of AP.
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Affiliation(s)
- Zhengpeng Yang
- Department of General Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China
| | - Weiguang Yang
- Department of General Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China
| | - Ming Lu
- Department of Surgery, David Geffen School of Medicine, University of Califonia at Los Angeles, Los Angeles, CA 90095, USA
| | - Zhituo Li
- Department of General Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China
| | - Xin Qiao
- Department of Surgery, David Geffen School of Medicine, University of Califonia at Los Angeles, Los Angeles, CA 90095, USA
| | - Bei Sun
- Department of General Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China
| | - Weihui Zhang
- Department of General Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China
| | - Dongbo Xue
- Department of General Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China
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Steury MD, McCabe LR, Parameswaran N. G Protein-Coupled Receptor Kinases in the Inflammatory Response and Signaling. Adv Immunol 2017; 136:227-277. [PMID: 28950947 DOI: 10.1016/bs.ai.2017.05.003] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
G protein-coupled receptor kinases (GRKs) are serine/threonine kinases that regulate a large and diverse class of G protein-coupled receptors (GPCRs). Through GRK phosphorylation and β-arrestin recruitment, GPCRs are desensitized and their signal terminated. Recent work on these kinases has expanded their role from canonical GPCR regulation to include noncanonical regulation of non-GPCR and nonreceptor substrates through phosphorylation as well as via scaffolding functions. Owing to these and other regulatory roles, GRKs have been shown to play a critical role in the outcome of a variety of physiological and pathophysiological processes including chemotaxis, signaling, migration, inflammatory gene expression, etc. This diverse set of functions for these proteins makes them popular targets for therapeutics. Role for these kinases in inflammation and inflammatory disease is an evolving area of research currently pursued in many laboratories. In this review, we describe the current state of knowledge on various GRKs pertaining to their role in inflammation and inflammatory diseases.
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Affiliation(s)
| | - Laura R McCabe
- Michigan State University, East Lansing, MI, United States
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Gingras D, Bendayan M. Evaluation of Pancreatic Amylase mRNA upon Cholinergic Stimulation of Secretion. J Histochem Cytochem 2016; 53:93-103. [PMID: 15637342 DOI: 10.1177/002215540505300111] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
The primary function of the exocrine pancreas consists of the synthesis and secretion of several digestive enzymes. It is well established that amylase secretion by rat pancreatic tissue or by isolated acinar cells in culture can be stimulated by the cholinergic agonist carbachol. However, the effect of this secretagogue on enzyme synthesis remains unclear. Some studies demonstrated increases in rates of synthesis, whereas others reported increases in secretion with or without decreases in synthesis. We have evaluated changes in pancreatic amylase mRNA and total RNA after a single injection of carbachol and under fasting conditions. Two approaches in molecular morphology were applied on rat pancreatic tissue: in situ hybridization and RNase A-gold. Both revealed decreases in RNA labeling at the level of the rough endoplasmic reticulum (RER) 5 min after stimulation of secretion and after fasting. Gradual recovery was registered 15 and 30 min after stimulation of secretion. Northern blotting confirmed drastic decreases in amylase mRNA 5 min after stimulation and after fasting. The combination of such different approaches has demonstrated drastic decreases in RNA at the RER level, reflecting declines in rates of synthesis at the translational level under all conditions tested.
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Affiliation(s)
- Diane Gingras
- Department of Pathology and Cell Biology, Université de Montréal, CP 6128 Succursale Centre-ville, Montréal, Québec, Canada H3T 1J4
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Nuche-Berenguer B, Ramos-Álvarez I, Jensen RT. Src kinases play a novel dual role in acute pancreatitis affecting severity but no role in stimulated enzyme secretion. Am J Physiol Gastrointest Liver Physiol 2016; 310:G1015-G1027. [PMID: 27033118 PMCID: PMC4935475 DOI: 10.1152/ajpgi.00349.2015] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/07/2015] [Accepted: 03/28/2016] [Indexed: 01/31/2023]
Abstract
In pancreatic acinar cells, the Src family of kinases (SFK) is involved in the activation of several signaling cascades that are implicated in mediating cellular processes (growth, cytoskeletal changes, apoptosis). However, the role of SFKs in various physiological responses such as enzyme secretion or in pathophysiological processes such as acute pancreatitis is either controversial, unknown, or incompletely understood. To address this, in this study, we investigated the role/mechanisms of SFKs in acute pancreatitis and enzyme release. Enzyme secretion was studied in rat dispersed pancreatic acini, in vitro acute-pancreatitis-like changes induced by supramaximal COOH-terminal octapeptide of cholecystokinin (CCK). SFK involvement assessed using the chemical SFK inhibitor (PP2) with its inactive control, 4-amino-7-phenylpyrazol[3,4-d]pyrimidine (PP3), under experimental conditions, markedly inhibiting SFK activation. In CCK-stimulated pancreatic acinar cells, activation occurred of trypsinogen, various MAP kinases (p42/44, JNK), transcription factors (signal transducer and activator of transcription-3, nuclear factor-κB, activator protein-1), caspases (3, 8, and 9) inducing apoptosis, LDH release reflective of necrosis, and various chemokines secreted (monocyte chemotactic protein-1, macrophage inflammatory protein-1α, regulated on activation, normal T cell expressed and secreted). All were inhibited by PP2, not by PP3, except caspase activation leading to apoptosis, which was increased, and trypsin activation, which was unaffected, as was CCK-induced amylase release. These results demonstrate SFK activation is playing a dual role in acute pancreatitis, inhibiting apoptosis and promoting necrosis as well as chemokine/cytokine release inducing inflammation, leading to more severe disease, as well as not affecting secretion. Thus, our studies indicate that SFK is a key mediator of inflammation and pancreatic acinar cell death in acute pancreatitis, suggesting it could be a potential therapeutic target in acute pancreatitis.
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Affiliation(s)
- Bernardo Nuche-Berenguer
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland
| | - Irene Ramos-Álvarez
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland
| | - R T Jensen
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland
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He JP, Tang GD, Qin MB, Shi HR, Fu HZ, Lin SD, Luo Q. Role of myosin light chain kinase in hypertriglyceridemia-associated acute pancreatitis in rats. Shijie Huaren Xiaohua Zazhi 2016; 24:19-27. [DOI: 10.11569/wcjd.v24.i1.19] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the role of myosin light chain kinase (MLCK) in hypertriglyceridemia-associated acute pancreatitis (HTGP).
METHODS: Forty-eight male SD rats were randomly divided into two groups to be fed either a high-fat diet (group A) or a normal diet (group B). After being raised for 4 weeks, blood was harvested from the retroorbital venous plexus to measure serum triglyceride (TG) levels. After that, group A was randomly divided into three subgroups: HTG, HTG + AP, and HTG + AP + ML-7; group B were also randomly divided into three subgroups: C, AP, and AP + ML-7. All rats were sacrificed 24 h after the last injection of cerulein. The pancreases were carefully removed for HE staining and transmission electron microscopy to observe the morphological changes, ultrastructure and tight junction (TJ). Blood was obtained from the heart to measure serum amylase levels. The expression and localization of MLCK and p-JNK in the pancreas were assayed by immunohistochemistry.
RESULTS: Compared with group B, serum TG level was significantly increased in group A after being fed a high fat diet (P < 0.01). The pathologic score of the pancreas and serum amylase (AMY) activity were significantly elevated in the HTG + AP group, compared with the AP group (P < 0.05). The ultrastructure of the pancreas in the HTG + AP and AP groups was damaged and the TJ was broadened (most significant in the HTG + AP group). Besides, MLCK and p-JNK were significantly up-regulated in the HTG + AP group compared with those in the AP group (P < 0.05), and there was a positive correlation between the expression of MLCK and p-JNK in the pancreas and the pathologic score of the pancreas (r1 = 0.795, r2 = 0.789, P < 0.01). ML-7, an inhibitor of MLCK, significantly ameliorated the pathologic signs of the pancreas, down-regulated AMY level (P < 0.01), improved the TJ and decreased the expression of p-JNK (P < 0.05).
CONCLUSION: MLCK may be associated with the severity of HTGP, and involved in the formation of HTGP by broadening cell-cell TJ and activation of the JNK pathway.
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Abstract
G-protein-coupled receptor kinases (GRKs) are serine/threonine protein kinases originally discovered for their role in G-protein-coupled receptor (GPCR) phosphorylation. Recent studies have demonstrated a much broader function for this kinase family including phosphorylation of cytosolic substrates involved in cell signaling pathways stimulated by GPCRs, as well as by non-GPCRs. In addition, GRKs modulate signaling via phosphorylation-independent functions. Because of these various biochemical functions, GRKs have been shown to affect critical physiological and pathophysiological processes, and thus are considered as drug targets in diseases such as heart failure. Role of GRKs in inflammation and inflammatory diseases is an evolving area of research and several studies including work from our lab in the recent years have demonstrated critical role of GRKs in the immune system. In this review, we discuss the classical and the newly emerging functions of GRKs in the immune system and their role in inflammation and disease processes.
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Tripathi S, Flobak Å, Chawla K, Baudot A, Bruland T, Thommesen L, Kuiper M, Lægreid A. The gastrin and cholecystokinin receptors mediated signaling network: a scaffold for data analysis and new hypotheses on regulatory mechanisms. BMC SYSTEMS BIOLOGY 2015. [PMID: 26205660 PMCID: PMC4513977 DOI: 10.1186/s12918-015-0181-z] [Citation(s) in RCA: 41] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
Background The gastrointestinal peptide hormones cholecystokinin and gastrin exert their biological functions via cholecystokinin receptors CCK1R and CCK2R respectively. Gastrin, a central regulator of gastric acid secretion, is involved in growth and differentiation of gastric and colonic mucosa, and there is evidence that it is pro-carcinogenic. Cholecystokinin is implicated in digestion, appetite control and body weight regulation, and may play a role in several digestive disorders. Results We performed a detailed analysis of the literature reporting experimental evidence on signaling pathways triggered by CCK1R and CCK2R, in order to create a comprehensive map of gastrin and cholecystokinin-mediated intracellular signaling cascades. The resulting signaling map captures 413 reactions involving 530 molecular species, and incorporates the currently available knowledge into one integrated signaling network. The decomposition of the signaling map into sub-networks revealed 18 modules that represent higher-level structures of the signaling map. These modules allow a more compact mapping of intracellular signaling reactions to known cell behavioral outcomes such as proliferation, migration and apoptosis. The integration of large-scale protein-protein interaction data to this literature-based signaling map in combination with topological analyses allowed us to identify 70 proteins able to increase the compactness of the map. These proteins represent experimentally testable hypotheses for gaining new knowledge on gastrin- and cholecystokinin receptor signaling. The CCKR map is freely available both in a downloadable, machine-readable SBML-compatible format and as a web resource through PAYAO (http://sblab.celldesigner.org:18080/Payao11/bin/). Conclusion We have demonstrated how a literature-based CCKR signaling map together with its protein interaction extensions can be analyzed to generate new hypotheses on molecular mechanisms involved in gastrin- and cholecystokinin-mediated regulation of cellular processes. Electronic supplementary material The online version of this article (doi:10.1186/s12918-015-0181-z) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Sushil Tripathi
- Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), N-7489, Trondheim, Norway.
| | - Åsmund Flobak
- Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), N-7489, Trondheim, Norway.
| | - Konika Chawla
- Department of Biology, Norwegian University of Science and Technology (NTNU), N-7491, Trondheim, Norway.
| | - Anaïs Baudot
- I2M, Marseilles Institute of Mathematics CNRS - AMU, Case 907, 13288, Marseille, Cedex 9, France.
| | - Torunn Bruland
- Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), N-7489, Trondheim, Norway.
| | - Liv Thommesen
- Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), N-7489, Trondheim, Norway. .,Department of Technology, Sør-Trøndelag University College, N-7004, Trondheim, Norway.
| | - Martin Kuiper
- Department of Biology, Norwegian University of Science and Technology (NTNU), N-7491, Trondheim, Norway.
| | - Astrid Lægreid
- Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), N-7489, Trondheim, Norway. .,Institute of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), N-7489, Trondheim, Norway.
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Abstract
OBJECTIVES In this study, we identified the protein kinases that play the most distinct roles in the occurrence of acute pancreatitis (AP). METHODS Gene expression profile data were downloaded from Gene Expression Omnibus database (GSE3644). The sample was from caerulein-induced AP mice. The intersection of the differentially expressed genes in AP mice taken from a protein kinase database was obtained for screening of the protein kinase encoded genes that were differentially expressed. Database for annotation, visualization, and integrated discovery was used for the functional enrichment analysis. Kinase inhibitors that regulated these kinases were retrieved from PubMed through text mining. RESULTS Twenty-nine differentially expressed kinase encoded genes were identified through screening. The functional enrichment analysis demonstrated that the functions of these genes were primarily enriched in "mitogen-activated protein kinase signaling pathway," followed by "extracellular regulated protein kinases pathway," "neurotrophin signaling pathway," "adherens junction," and "gap junction." SRC and epidermal growth factor receptor (EGFR) were related to extracellular regulated protein kinases pathway and also related to adherens junction as well as gap junction. On the basis of the regulated kinases, the kinase inhibitors reported in the literature were classified into multiple groups. CONCLUSIONS EGFR and SRC may be coexpressed in AP. The kinase inhibitors working together in SRC and EGFR may play better efficacy in the treatment of AP.
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Nuche-Berenguer B, Moreno P, Jensen RT. Elucidation of the roles of the Src kinases in pancreatic acinar cell signaling. J Cell Biochem 2015; 116:22-36. [PMID: 25079913 PMCID: PMC4229413 DOI: 10.1002/jcb.24895] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2014] [Accepted: 07/25/2014] [Indexed: 12/13/2022]
Abstract
Recent studies report the Src-family kinases (SFK's) are important in a number of physiological and pathophysiological responses of pancreatic acinar cells (pancreatitis, growth, apoptosis); however, the role of SFKs in various signaling cascades important in mediating these cell functions is either not investigated or unclear. To address this we investigated the action of SFKs in these signaling cascades in rat pancreatic acini by modulating SFK activity using three methods: adenovirus-induced expression of an inactive dominant-negative CSK (Dn-CSK-Advirus) or wild-type CSK (Wt-CSK-Advirus), which activate or inhibit SFK, respectively, or using the chemical inhibitor, PP2, with its inactive control, PP3. CCK (0.3, 100 nM) and TPA (1 μM) activated SFK and altered the activation of FAK proteins (PYK2, p125(FAK)), adaptor proteins (p130(CAS), paxillin), MAPK (p42/44, JNK, p38), Shc, PKC (PKD, MARCKS), Akt but not GSK3-β. Changes in SFK activity by using the three methods of altering SFK activity affected CCK/TPAs activation of SFK, PYK2, p125(FAK), p130(CAS), Shc, paxillin, Akt but not p42/44, JNK, p38, PKC (PKD, MARCKS) or GSK3-β. With chemical inhibition the active SFK inhibitor, PP2, but not the inactive control analogue, PP3, showed these effects. For all stimulated changes pre-incubation with both adenoviruses showed similar effects to chemical inhibition of SFK activity. In conclusion, using three different approaches to altering Src activity allowed us to define fully for the first time the roles of SFKs in acinar cell signaling. Our results show that in pancreatic acinar cells, SFKs play a much wider role than previously reported in activating a number of important cellular signaling cascades shown to be important in mediating both acinar cell physiological and pathophysiological responses.
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Affiliation(s)
- Bernardo Nuche-Berenguer
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-1804, USA
| | - Paola Moreno
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-1804, USA
| | - R. T. Jensen
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-1804, USA
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Irrera N, Bitto A, Interdonato M, Squadrito F, Altavilla D. Evidence for a role of mitogen-activated protein kinases in the treatment of experimental acute pancreatitis. World J Gastroenterol 2014; 20:16535-16543. [PMID: 25469021 PMCID: PMC4248196 DOI: 10.3748/wjg.v20.i44.16535] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/28/2014] [Revised: 05/23/2014] [Accepted: 07/22/2014] [Indexed: 02/06/2023] Open
Abstract
Acute pancreatitis (AP) is an inflammatory disease characterized by acute inflammation and necrosis of the pancreatic parenchyma. AP is often associated with organ failure, sepsis, and high mortality. The pathogenesis of AP is still not well understood. In recent years several papers have highlighted the cellular and molecular events of acute pancreatitis. Pancreatitis is initiated by activation of digestive enzymes within the acinar cells that are involved in autodigestion of the gland, followed by a massive infiltration of neutrophils and macrophages and release of inflammatory mediators, responsible for the local and systemic inflammatory response. The hallmark of AP is parenchymal cell necrosis that represents the cause of the high morbidity and mortality, so that new potential therapeutic approaches are indispensable for the treatment of patients at high risk of complications. However, not all factors that determine the onset and course of the disease have been explained. Aim of this article is to review the role of mitogen-activated protein kinases in pathogenesis of acute pancreatitis.
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Holtz BJ, Lodewyk KB, Sebolt-Leopold JS, Ernst SA, Williams JA. ERK activation is required for CCK-mediated pancreatic adaptive growth in mice. Am J Physiol Gastrointest Liver Physiol 2014; 307:G700-10. [PMID: 25104499 PMCID: PMC4187068 DOI: 10.1152/ajpgi.00163.2014] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
High levels of cholecystokinin (CCK) can stimulate pancreatic adaptive growth in which mature acinar cells divide, leading to enhanced pancreatic mass with parallel increases in protein, DNA, RNA, and digestive enzyme content. Prolonged release of CCK can be induced by feeding trypsin inhibitor (TI) to disrupt normal feedback control. This leads to exocrine growth in a CCK-dependent manner. The extracellular signal-related kinase (ERK) pathway regulates many proliferative processes in various tissues and disease models. The aim of this study was to evaluate the role of ERK signaling in pancreatic adaptive growth using the MEK inhibitors PD-0325901 and trametinib (GSK-1120212). It was determined that PD-0325901 given two times daily by gavage or mixed into powdered chow was an effective and specific inhibitor of ERK signaling in vivo. TI-containing chow led to a robust increase in pancreatic mass, protein, DNA, and RNA content. This pancreatic adaptive growth was blocked in mice fed chow containing the MEK inhibitors. PD-0325901 blocked TI-induced ERK-regulated early response genes, cell-cycle proteins, and mitogenesis by acinar cells. It was determined that ERK signaling is necessary for the initiation of pancreatic adaptive growth but not necessary to maintain it. PD-0325901 blocked adaptive growth when given before cell-cycle initiation but not after mitogenesis had been established. Furthermore, GSK-1120212, a chemically distinct inhibitor of the ERK pathway that is now approved for clinical use, inhibited growth similar to PD-0325901. These data demonstrate that the ERK pathway is required for CCK-stimulated pancreatic adaptive growth.
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Affiliation(s)
- Bryan J. Holtz
- 1Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, Michigan;
| | - Kevin B. Lodewyk
- 1Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, Michigan;
| | | | - Stephen A. Ernst
- 3Department of Cellular and Developmental Biology, University of Michigan Medical School, Ann Arbor, Michigan; and
| | - John A. Williams
- 1Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, Michigan; ,4Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, Michigan
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15
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Abstract
OBJECTIVE The aim of this study was to evaluate the effects of Opuntia humifusa (OH) on cerulein-induced acute pancreatitis (AP). METHODS Acute pancreatitis was induced via intraperitoneal injection of cholecystokinin analog cerulein (50 μg/kg). In the OH pretreatment group, OH was administered intraperitoneally (100, 250, or 500 mg/kg) 1 hour before first cerulein injection. In the posttreatment group, OH was administered intraperitoneally (500 mg/kg) 1 hour after the first cerulein injection. Furthermore, we isolated the pancreatic acinar cells using collagenase method, then investigated the acinar cell viability, cytokine productions, and the regulating mechanisms. RESULTS The both pretreatment and posttreatment of OH treatment attenuated the severity of AP, as shown by the histology of the pancreas and lung, and inhibited neutrophil infiltration; serum amylase and lipase activities; proinflammatory cytokine expression such as interleukin 1, interleukin 6, and tumor necrosis factor α; and cell death including apoptosis and necrosis. Furthermore, OH inhibited the activation of c-Jun N-terminal kinases. CONCLUSIONS These results suggest that OH reduces the severity of AP by inhibiting acinar cell death through c-Jun N-terminal kinases.
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Guo L, Sans MD, Hou Y, Ernst SA, Williams JA. c-Jun/AP-1 is required for CCK-induced pancreatic acinar cell dedifferentiation and DNA synthesis in vitro. Am J Physiol Gastrointest Liver Physiol 2012; 302:G1381-96. [PMID: 22461029 PMCID: PMC3378092 DOI: 10.1152/ajpgi.00129.2010] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Endogenous CCK plays an important role in pancreatic regeneration after pancreatitis. We used primary culture of mouse pancreatic acinar cells to evaluate the effect of CCK on acinar cell morphology and gene expression and to determine signaling pathways required for proliferation of acinar cells in vitro. Over 4 days in culture, cells grew out from acini and formed patches of monolayer, which displayed a reduced expression of acinar cell markers including digestive enzymes and Mist1 and an increased expression of ductal and embryonic markers, including cytokeratin 7, β-catenin, E-cadherin, pdx-1, and nestin. There was no appearance of stellate cell markers. CCK enhanced cellular spreading, DNA synthesis, and cyclin D1 expression. When signaling pathways were evaluated, CCK stimulation increased c-Jun expression, JNK and ERK activity, and AP-1 activation. Chemical inhibitors of JNK and ERK pathways, dominant-negative JNK and c-Jun, and c-Jun shRNA significantly inhibited CCK-induced DNA synthesis, CCK-induced AP-1 activation, and cyclin D1 expression. Furthermore, dominant-negative c-Jun reduced the increased expression of β-catenin and the decreased expression of amylase during culture. These results show that MAPK/c-Jun/AP-1 pathway plays an important role in pancreatic acinar cell dedifferentiation and proliferation in culture. Monolayer culture can serve as a model to study acinar cell proliferation similar to regeneration after pancreatitis in vivo.
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Affiliation(s)
- Lili Guo
- 1Department of Molecular and Integrative Physiology, The University of Michigan Medical School, Ann Arbor, Michigan,
| | - Maria Dolors Sans
- 1Department of Molecular and Integrative Physiology, The University of Michigan Medical School, Ann Arbor, Michigan,
| | - Yanan Hou
- 1Department of Molecular and Integrative Physiology, The University of Michigan Medical School, Ann Arbor, Michigan,
| | - Stephen A. Ernst
- 2Department of Cellular and Developmental Biology, The University of Michigan Medical School, Ann Arbor, Michigan, and
| | - John A. Williams
- 1Department of Molecular and Integrative Physiology, The University of Michigan Medical School, Ann Arbor, Michigan, ,3Department of Internal Medicine, The University of Michigan Medical School, Ann Arbor, Michigan
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17
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Jensen K, Afroze S, Munshi MK, Guerrier M, Glaser SS. Mechanisms for nicotine in the development and progression of gastrointestinal cancers. TRANSLATIONAL GASTROINTESTINAL CANCER 2012; 1:81-87. [PMID: 22701817 PMCID: PMC3371638 DOI: 10.3978/j.issn.2224-4778.2011.12.01] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Long-term smoking is major risk factor for a variety of cancers, including those of the gastrointestinal (GI) tract. Historically, nicotine and its derivatives are well known for their role in addiction, and have more recently been documented for their carcinogenic role in a number of human cancers. The cellular and molecular pathways activated by nicotine mimic physiological and environmental carcinogenesis in cancers throughout the GI tract potentiating cancer growth and/or inducing the formation of cancer on their own. Thus, it is important to unlock the carcinogenic mechanisms induced by nicotine in these systems, and underscore nicotine's potential as an environmental hazard. This review outlines the specific pathways demonstrated to mediate nicotine's carcinogenic mechanism in the GI tract. The abundance of cell and animal evidence calls for increased epidemiologic and case-control evaluation of nicotine's role in cancer.
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18
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Escobar J, Pereda J, López-Rodas G, Sastre J. Redox signaling and histone acetylation in acute pancreatitis. Free Radic Biol Med 2012; 52:819-37. [PMID: 22178977 DOI: 10.1016/j.freeradbiomed.2011.11.009] [Citation(s) in RCA: 61] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/25/2011] [Revised: 11/08/2011] [Accepted: 11/09/2011] [Indexed: 12/22/2022]
Abstract
Histone acetylation via CBP/p300 coordinates the expression of proinflammatory cytokines in the activation phase of inflammation, particularly through mitogen-activated protein kinases (MAPKs), nuclear factor-κB (NF-κB), and signal transducers and activators of transcription (STAT) pathways. In contrast, histone deacetylases (HDACs) and protein phosphatases are mainly involved in the attenuation phase of inflammation. The role of reactive oxygen species (ROS) in the inflammatory cascade is much more important than expected. Mitochondrial ROS act as signal-transducing molecules that trigger proinflammatory cytokine production via inflammasome-independent and inflammasome-dependent pathways. The major source of ROS in acute inflammation seems to be NADPH oxidases, whereas NF-κB, protein phosphatases, and HDACs are the major targets of ROS and redox signaling in this process. There is a cross-talk between oxidative stress and proinflammatory cytokines through serine/threonine protein phosphatases, tyrosine protein phosphatases, and MAPKs that greatly contributes to amplification of the uncontrolled inflammatory cascade and tissue injury in acute pancreatitis. Chromatin remodeling during induction of proinflammatory genes would depend primarily on phosphorylation of transcription factors and their binding to gene promoters together with recruitment of histone acetyltransferases. PP2A should be considered a key modulator of the inflammatory cascade in acute pancreatitis through the ERK/NF-κB pathway and histone acetylation.
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Affiliation(s)
- Javier Escobar
- Department of Physiology, School of Pharmacy, University of Valencia, Burjasot, Valencia, Spain
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19
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Yun SW, Bae GS, Kim MS, Park KC, Koo BS, Kim BJ, Kim TH, Seo SW, Shin YK, Lee SH, Song HJ, Park SJ. Melittin inhibits cerulein-induced acute pancreatitis via inhibition of the JNK pathway. Int Immunopharmacol 2011; 11:2062-72. [PMID: 21939783 DOI: 10.1016/j.intimp.2011.08.020] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2011] [Revised: 07/21/2011] [Accepted: 08/28/2011] [Indexed: 12/29/2022]
Abstract
The major compound of bee venom, melittin, has been used as an anti-inflammatory reagent for decades. However, the potential of melittin to ameliorate acute pancreatitis (AP) is unknown. Our aim was to investigate the effect of melittin on cerulein-induced AP. Pre- and post-treatment with melittin inhibited histological changes in the pancreas and lungs during cerulein-induced AP. Pancreatic weight/body weight ratios; digestive enzymes, including amylase and lipase; serum and pancreatic cytokine expression; and myeloperoxidase activity were decreased. In addition, treatment with melittin inhibited the activation of c-Jun NH(2)-terminal protein kinase (JNK) in the pancreas during cerulein-induced pancreatitis. In accordance with the results of in vivo experiments, melittin reduced cerulein-induced cell death, and production of inflammatory cytokines. In conclusion, our results suggest that melittin attenuated AP and AP-associated lung injury through the inhibition of JNK activation.
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Affiliation(s)
- Seung-Won Yun
- ChungBuk Oriental Medicine Center, Jecheon, 390-250, ChungBuk, South Korea
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20
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Bae GS, Kim MS, Jeong J, Lee HY, Park KC, Koo BS, Kim BJ, Kim TH, Lee SH, Hwang SY, Shin YK, Song HJ, Park SJ. Piperine ameliorates the severity of cerulein-induced acute pancreatitis by inhibiting the activation of mitogen activated protein kinases. Biochem Biophys Res Commun 2011; 410:382-8. [PMID: 21663734 DOI: 10.1016/j.bbrc.2011.05.136] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2011] [Accepted: 05/25/2011] [Indexed: 12/29/2022]
Abstract
Piperine is a phenolic component of black pepper (Piper nigrum) and long pepper (Piper longum), fruits used in traditional Asian medicine. Our previous study showed that piperine inhibits lipopolysaccharide-induced inflammatory responses. In this study, we investigated whether piperine reduces the severity of cerulein-induced acute pancreatitis (AP). Administration of piperine reduced histologic damage and myeloperoxidase (MPO) activity in the pancreas and ameliorated many of the examined laboratory parameters, including the pancreatic weight (PW) to body weight (BW) ratio, as well as serum levels of amylase and lipase and trypsin activity. Furthermore, piperine pretreatment reduced the production of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 during cerulein-induced AP. In accordance with in vivo results, piperine reduced cell death, amylase and lipase activity, and cytokine production in isolated cerulein-treated pancreatic acinar cells. In addition, piperine inhibited the activation of mitogen-activated protein kinases (MAPKs). These findings suggest that the anti-inflammatory effect of piperine in cerulein-induced AP is mediated by inhibiting the activation of MAPKs. Thus, piperine may have a protective effect against AP.
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Affiliation(s)
- Gi-Sang Bae
- Department of Herbology, School of Oriental Medicine, Wonkwang University, Iksan, 540-749 Jeonbuk, South Korea
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21
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Abstract
OBJECTIVES Nardostachys jatamansi belonging to the family Valerianaceae has been used as a remedy for stomach and skin ailments in Korea. The effect of N. jatamansi on acute pancreatitis (AP) has not been defined. Therefore, we investigated the effect of N. jatamansi on cerulein-induced AP. METHODS In the pretreatment group, N. jatamansi was administered orally to mice at 10 and 20 mg/kg for 5 days, and the mice were intraperitoneally injected with the stable cholecystokinin analogue cerulein hourly for 6 hours. In the posttreatment group, cerulein was injected hourly for 6 hours, and N. jatamansi was administered at the indicated time (1, 3, and 5 hours after the first cerulein injection) and dose (10 and 20 mg/kg) during the cerulein injection. Blood samples were taken 6 hours later to determine the serum amylase, the lipase, and the cytokine levels. The pancreas and the lung were rapidly removed for morphologic examination, myeloperoxidase assay, and real-time reverse transcription polymerase chain reaction. RESULTS Nardostachys jatamansi treatment attenuated the AP, as shown by the histological examination results of the pancreas and the lung, reductions in pancreatic edema, neutrophil infiltration, serum amylase and lipase levels, serum cytokine levels, and messenger RNA expressions of inflammatory mediators. CONCLUSIONS These results suggest that N. jatamansi attenuates the severity of AP and pancreatitis-associated lung injury.
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22
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Cawston EE, Miller LJ. Therapeutic potential for novel drugs targeting the type 1 cholecystokinin receptor. Br J Pharmacol 2009; 159:1009-21. [PMID: 19922535 DOI: 10.1111/j.1476-5381.2009.00489.x] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Cholecystokinin (CCK) is a physiologically important gastrointestinal and neuronal peptide hormone, with roles in stimulating gallbladder contraction, pancreatic secretion, gastrointestinal motility and satiety. CCK exerts its effects via interactions with two structurally related class I guanine nucleotide-binding protein (G protein)-coupled receptors (GPCRs), the CCK(1) receptor and the CCK(2) receptor. Here, we focus on the CCK(1) receptor, with particular relevance to the broad spectrum of signalling initiated by activation with the natural full agonist peptide ligand, CCK. Distinct ligand-binding pockets have been defined for the natural peptide ligand and for some non-peptidyl small molecule ligands. While many CCK(1) receptor ligands have been developed and have had their pharmacology well described, their clinical potential has not yet been fully explored. The case is built for the potential importance of developing more selective partial agonists and allosteric modulators of this receptor that could have important roles in the treatment of common clinical syndromes.
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Affiliation(s)
- Erin E Cawston
- Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic, Scottsdale, AZ, USA
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23
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Ferrés-Masó M, Sacilotto N, López-Rodas G, Dagorn JC, Iovanna JL, Closa D, Folch-Puy E. PAP1 signaling involves MAPK signal transduction. Cell Mol Life Sci 2009; 66:2195-2204. [PMID: 19434369 PMCID: PMC11115593 DOI: 10.1007/s00018-009-0040-4] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2009] [Revised: 03/26/2009] [Accepted: 04/21/2009] [Indexed: 12/21/2022]
Abstract
Pancreatitis-associated protein 1 (PAP1) belongs to the Reg family of secretory proteins. Several important biological roles have been attributed to PAP1 but the signaling pathways activated by this protein remain only partially understood. Here, we describe the intracellular pathways triggered by PAP1 in a pancreatic acinar cell line. Taking advantage of the fact that PAP1 induces its own transcription, we performed ChIP assays to analyze the recruitment of transcriptional factors on its promoter. Our results show that PAP1 increased the transactivation activity of pap1 and the binding on its promoter of the nuclear factors C/EBPbeta, P-CREB, P-ELK1, EGR1, STAT3, and ETS2, which are downstream targets of MAPK signaling. p44/42, p38, and JNK MAPKs activity increased after PAP1 treatment. In addition, pharmacological inhibition of these kinases markedly inhibited the induction of pap1 mRNA. Taken together, these results indicated that the mechanism of PAP1 action involves the activation of the MAPK superfamily.
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Affiliation(s)
- M. Ferrés-Masó
- Department of Experimental Pathology, Institut d’Investigacions Biomèdiques de Barcelona, Consejo Superior de Investigaciones Científicas-Institut d’Investigacions Biomèdiques August Pi i Sunyer, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, c/Rosselló 161, 7º, 08036 Barcelona, Spain
| | - N. Sacilotto
- Department of Biochemistry and Molecular Biology, University of Valencia, Valencia, Spain
| | - G. López-Rodas
- Department of Biochemistry and Molecular Biology, University of Valencia, Valencia, Spain
| | - J. C. Dagorn
- Centre de Recherche INSERM U.624, Stress Cellulaire, Marseille, France
| | - J. L. Iovanna
- Centre de Recherche INSERM U.624, Stress Cellulaire, Marseille, France
| | - D. Closa
- Department of Experimental Pathology, Institut d’Investigacions Biomèdiques de Barcelona, Consejo Superior de Investigaciones Científicas-Institut d’Investigacions Biomèdiques August Pi i Sunyer, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, c/Rosselló 161, 7º, 08036 Barcelona, Spain
| | - E. Folch-Puy
- Department of Experimental Pathology, Institut d’Investigacions Biomèdiques de Barcelona, Consejo Superior de Investigaciones Científicas-Institut d’Investigacions Biomèdiques August Pi i Sunyer, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, c/Rosselló 161, 7º, 08036 Barcelona, Spain
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24
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Abstract
Reactive oxygen and reactive nitrogen species (ROS/RNS) have been implicated in the pathogenesis of acute and chronic pancreatitis. Clinical and basic science studies have indicated that ROS/RNS formation processes are intimately linked to the development of the inflammatory disorders. The detrimental effects of highly reactive ROS/RNS are mediated by their direct actions on biomolecules (lipids, proteins, and nucleic acids) and activation of proinflammatory signal cascades, which subsequently lead to activation of immune responses. The present article summarizes the possible sources of ROS/RNS formation and the detailed signaling cascades implicated in the pathogenesis of pancreatic inflammation, as observed in acute and chronic pancreatitis. A therapeutic ROS/RNS-scavenging strategy has been advocated for decades; however, clinical studies examining such approaches have been inconsistent in their results. Emerging evidence indicates that pancreatitis-inducing ROS/RNS generation may be attenuated by targeting ROS/RNS-generating enzymes and upstream mediators.
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Affiliation(s)
- Po Sing Leung
- Department of Physiology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong, China.
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25
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Namkung W, Yoon JS, Kim KH, Lee MG. PAR2 exerts local protection against acute pancreatitis via modulation of MAP kinase and MAP kinase phosphatase signaling. Am J Physiol Gastrointest Liver Physiol 2008; 295:G886-94. [PMID: 18755806 DOI: 10.1152/ajpgi.00053.2008] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
During acute pancreatitis, protease-activated receptor 2 (PAR2) can be activated by interstitially released trypsin. In the mild form of pancreatitis, PAR2 activation exerts local protection against intrapancreatic damage, whereas, in the severe form of pancreatitis, PAR2 activation mediates some systemic complications. This study aimed to identify the molecular mechanisms of PAR2-mediated protective effects against intrapancreatic damage. A mild form of acute pancreatitis was induced by an intraperitoneal injection of caerulein (40 microg/kg) in rats. Effects of PAR2 activation on intrapancreatic damage and on mitogen-activated protein (MAP) kinase signaling were assessed. Caerulein treatment activated extracellular signal-regulated kinase (ERK) and c-Jun NH(2)-terminal kinase (JNK) within 15 min and maintained phosphorylation of ERK and JNK for 2 h in the rat pancreas. Although PAR2 activation by the pretreatment with PAR2-activating peptide (AP) itself increased ERK phosphorylation in rat pancreas, the same treatment remarkably decreased caerulein-induced activation of ERK and JNK principally by accelerating their dephosphorylation. Inhibition of ERK and JNK phosphorylation by the pretreatment with MAP/ERK kinase (MEK) or JNK inhibitors decreased caerulein-induced pancreatic damage that was similar to the effect induced by PAR2-AP. Notably, in caerulein-treated rats, PAR2-AP cotreatment highly increased the expression of a group of MAP kinase phosphatases (MKPs) that deactivate ERK and JNK. The above results imply that downregulation of MAP kinase signaling by MKP induction is a key mechanism involved in the protective effects of PAR2 activation on caerulein-induced intrapancreatic damage.
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Affiliation(s)
- Wan Namkung
- Dept. of Pharmacology, Yonsei Univ. College of Medicine, Seoul 120-752, Korea
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26
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Abstract
Pathological activation of selective signaling molecules within the pancreatic acinar cell mediates the development of acute pancreatitis. Some of the key early acinar cell events include activation of proteases, inhibition of apical secretion, and elaboration of inflammatory mediators. Previous studies have shown that supraphysiological concentrations of cholecystokinin (CCK) that can cause pancreatitis in vivo, also initiate these pathological responses in dispersed groups of acinar cells (acini). Protein kinase C (PKC) regulates many cellular events and a role for this family of signaling molecules has been described in some of the pathological responses of pancreatitis. Notably, ethanol can activate specific PKC isoforms and sensitize the acinar cells to the pathological effects of CCK. Our preliminary studies in isolated pancreatic acini and a cell-free reconstitution system suggest that PKC can mediate protease activation in the acinar cell. These findings may be relevant to the pathogenesis of pancreatitis from alcohol and other etiologies.
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Affiliation(s)
- Fred Gorelick
- Department of Internal Medicine, Connecticut VA Healthcare, Yale University, West Haven, Connecticut 06516, USA
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27
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Ramnath RD, Sun J, Adhikari S, Bhatia M. Effect of mitogen-activated protein kinases on chemokine synthesis induced by substance P in mouse pancreatic acinar cells. J Cell Mol Med 2007; 11:1326-1341. [PMID: 18205703 PMCID: PMC4401295 DOI: 10.1111/j.1582-4934.2007.00086.x] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2007] [Accepted: 06/21/2007] [Indexed: 11/28/2022] Open
Abstract
Substance P, acting via its neurokinin 1 receptor (NK1 R), plays an important role in mediating a variety of inflammatory processes. Its interaction with chemokines is known to play a crucial role in the pathogenesis of acute pancreatitis. In pancreatic acinar cells, substance P stimulates the release of NFkappaB-driven chemokines. However, the signal transduction pathways by which substance P-NK1 R interaction induces chemokine production are still unclear. To that end, we went on to examine the participation of mitogen-activated protein kinases (MAPKs) in substance P-induced synthesis of pro-inflammatory chemokines, monocyte chemoanractant protein-1 (MCP-I), macrophage inflammatory protein-lalpha (MIP-lalpha) and macrophage inflammatory protein-2 (MIP-2), in pancreatic acini. In this study, we observed a time-dependent activation of ERK1/2, c-Jun N-terminal kinase (JNK), NFkappaB and activator protein-1 (AP-1) when pancreatic acini were stimulated with substance P. Moreover, substance P-induced ERK 1/2, JNK, NFkappaB and AP-1 activation as well as chemokine synthesis were blocked by pre-treatment with either extracellular signal-regulated protein kinase kinase 1 (MEK1) inhibitor or JNK inhibitor. In addition, substance P-induced activation of ERK 112, JNK, NFkappaB and AP-1-driven chemokine production were attenuated by CP96345, a selective NK1 R antagonist, in pancreatic acinar cells. Taken together, these results suggest that substance P-NK1 R induced chemokine production depends on the activation of MAPKs-mediated NFkappaB and AP-1 signalling pathways in mouse pancreatic acini.
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Affiliation(s)
| | - Jia Sun
- Department of Pharmacology, National University of Singapore, Singapore
| | - Sharmila Adhikari
- Department of Pharmacology, National University of Singapore, Singapore
| | - Madhav Bhatia
- Department of Pharmacology, National University of Singapore, Singapore
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28
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Kasugai K, Watson SJ, A Flavell R, J Davis R, Todisco A. Crucial role of c-Jun NH2-terminal kinase 1 (JNK1) in cold-restraint stress-induced gastric lesions in mice. Dig Dis Sci 2007; 52:1698-705. [PMID: 17061159 DOI: 10.1007/s10620-006-9155-8] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/08/2005] [Accepted: 11/14/2005] [Indexed: 12/18/2022]
Abstract
c-Jun NH2-terminal kinase 1 /JNK1, is activated in response to a broad array of cellular stresses. We investigated the role of JNK1 in the pathophysiology of cold-restraint stress-induced gastric lesions in mice. B6/129, wild type (WT) mice, or mutant mice deficient in Jnk1 (Jnk1-/- mice) were exposed to cold-restraint stress for different time periods. Gastric lesions were identified and quantitated by morphometric analysis. JNK1 activity in mucosal homogenates was quantitated by immunoprecipitation and in-vitro kinase assays. JNK1 expression and Akt activation were assessed by Western blots with anti-JNK1 and anti-phospho Akt antibodies, respectively. Gastric mucosal homogenates from Jnk1-/- mice exhibited no significant expression of JNK1 and no detectable level of JNK1 activation. Exposure of WT mice to cold-restraint stress led to the development of significant gastric lesions and to a greater than three-fold induction in JNK1 activity, while no lesions were detected in the gastric mucosa of Jnk1-/- mice. Since cold-restraint stress-induced gastric lesions involve the activation of cholinergic pathways, we tested the effect of atropine on both the development of gastric lesions and JNK1 activation. Pretreatment of WT mice with atropine completely inhibited both cold-restraint stress-induced lesions and JNK1 activation. Cold-restraint stress induced protein kinase B/Akt to a similar level in the gastric mucosa of both WT and Jnk1-/- mice indicating the integrity of other signaling pathways. JNK1 plays a key role in the development of cold-restraint stress-induced gastric lesions in mice through the activation of cholinergic, atropine sensitive pathways.
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Affiliation(s)
- K Kasugai
- Department of Internal Medicine, University of Michigan, Ann Arbor, MI 48109-0682, USA
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29
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Kubisch CH, Logsdon CD. Secretagogues differentially activate endoplasmic reticulum stress responses in pancreatic acinar cells. Am J Physiol Gastrointest Liver Physiol 2007; 292:G1804-12. [PMID: 17431218 DOI: 10.1152/ajpgi.00078.2007] [Citation(s) in RCA: 64] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Endoplasmic reticulum (ER) stress leads to the accumulation of misfolded proteins in the ER lumen and initiates the unfolded protein response (UPR). Components of the UPR are important in pancreatic development, and recent studies have indicated that the UPR is activated in the arginine model of acute pancreatitis. However, the effects of secretagogues on UPR components in the pancreas are unknown. The present study aimed to examine the effects of different types and concentrations of secretagogues on acinar cell function and specific components of the UPR. Rat pancreatic acini were stimulated with the CCK analogs CCK8 (10 pM-10 nM) or JMV-180 (10 nM-10 microM) or with bombesin (1-100 nM). Components of the UPR, including chaperone BiP expression, PKR-like ER kinase (PERK) phosphorylation, X box-binding protein 1 (XBP1) splicing, and CCAAT/enhancer binding protein homologous protein (CHOP) expression, were measured, as were effects on amylase secretion and intracellular trypsin activation. CCK8 generated a biphasic secretion dose-response curve, and high concentrations increased intracellular active trypsin levels. In contrast, JMV-180 and bombesin secretion dose-response curves were monophasic, and high concentrations did not increase intracellular trypsin activity. All three secretagogues increased BiP levels and XBP1 splicing. However, only supraphysiological levels of CCK8 associated with inhibited amylase secretion and trypsin activation stimulated PERK phosphorylation and expression of CHOP. The effects of CCK8 on UPR components were rapid, occurring within 5-20 min. In conclusion, ER stress response mechanisms appear to be involved in both pancreatic physiology and pathophysiology, and future efforts should be directed at understanding the roles of these mechanisms in the pancreas.
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MESH Headings
- Amylases/metabolism
- Animals
- Basic-Leucine Zipper Transcription Factors/genetics
- Basic-Leucine Zipper Transcription Factors/metabolism
- Bombesin/pharmacology
- Cells, Cultured
- DNA-Binding Proteins
- Dose-Response Relationship, Drug
- Endoplasmic Reticulum/drug effects
- Endoplasmic Reticulum/metabolism
- Endoplasmic Reticulum/pathology
- Enzyme Activation
- Heat-Shock Proteins/metabolism
- Male
- Molecular Chaperones/metabolism
- Neoplasm Proteins/genetics
- Neoplasm Proteins/metabolism
- Pancreas, Exocrine/drug effects
- Pancreas, Exocrine/metabolism
- Pancreas, Exocrine/pathology
- Phosphorylation
- Protein Folding
- RNA Splicing
- RNA, Messenger/metabolism
- Rats
- Rats, Sprague-Dawley
- Regulatory Factor X Transcription Factors
- Signal Transduction/drug effects
- Sincalide/analogs & derivatives
- Sincalide/metabolism
- Sincalide/pharmacology
- Stress, Physiological/metabolism
- Stress, Physiological/pathology
- Stress, Physiological/physiopathology
- Transcription Factor CHOP/genetics
- Transcription Factor CHOP/metabolism
- Transcription Factors
- Trypsin/metabolism
- Trypsinogen/metabolism
- X-Box Binding Protein 1
- eIF-2 Kinase/metabolism
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Affiliation(s)
- Constanze H Kubisch
- The Univ. of Texas M. D. Anderson Cancer Center, Dept. of Cancer Biology, Unit 0953, SCRB2.2021, 7435 Fannin St., PO Box 301429, Houston, TX 77230-1429, USA
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30
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Kowalik AS, Johnson CL, Chadi SA, Weston JY, Fazio EN, Pin CL. Mice lacking the transcription factor Mist1 exhibit an altered stress response and increased sensitivity to caerulein-induced pancreatitis. Am J Physiol Gastrointest Liver Physiol 2007; 292:G1123-32. [PMID: 17170023 DOI: 10.1152/ajpgi.00512.2006] [Citation(s) in RCA: 67] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Several animal models have been developed to investigate the pathobiology of pancreatitis, but few studies have examined the effects that altered pancreatic gene expression have in these models. In this study, the sensitivity to secretagogue-induced pancreatitis was examined in a mouse line that has an altered acinar cell environment due to the targeted deletion of Mist1. Mist1 is an exocrine specific transcription factor important for the complete differentiation and function of pancreatic acinar cells. Mice lacking the Mist1 gene [Mist1 knockout (KO) mice] exhibit cellular disorganization and functional defects in the exocrine pancreas but no gross morphological defects. Following the induction of pancreatitis with caerulein, a CCK analog, we observed elevated serum amylase levels, necrosis, and tissue damage in Mist1 KO mice, indicating increased pancreatic damage. There was also a delay in the regeneration of acinar tissue in Mist1 KO animals. Molecular profiling revealed an altered activation of stress response genes in Mist1 KO pancreatic tissue compared with wild-type (WT) tissue following the induction of pancreatitis. In particular, Western blot analysis for activating transcription factor 3 and phosphorylated eukaryotic initiation factor 2alpha (eIF2alpha), mediators of endoplasmic reticulum (ER) stress, indicated limited activation of this pathway in Mist1 KO animals compared with WT controls. Conversely, Mist1 KO pancreatic tissue exhibits increased expression of growth arrest and DNA damage inducible 34 protein, an inhibitor of eIF2alpha phosphorylation, before and after the induction of pancreatitis. These finding suggest that activation of the ER stress pathway is a protective event in the progression of pancreatitis and highlight the Mist1 KO mouse line as an important new model for studying the molecular events that contribute to the sensitivity to pancreatic injury.
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MESH Headings
- Activating Transcription Factor 3/genetics
- Activating Transcription Factor 3/metabolism
- Acute Disease
- Amylases/blood
- Animals
- Antigens, Differentiation/genetics
- Antigens, Differentiation/metabolism
- Apoptosis/drug effects
- Basic Helix-Loop-Helix Transcription Factors/deficiency
- Basic Helix-Loop-Helix Transcription Factors/genetics
- Basic Helix-Loop-Helix Transcription Factors/metabolism
- Cell Cycle Proteins/genetics
- Cell Cycle Proteins/metabolism
- Cells, Cultured
- Ceruletide
- Cholecystokinin/metabolism
- Cholecystokinin/pharmacology
- Disease Models, Animal
- Dose-Response Relationship, Drug
- Endoplasmic Reticulum/metabolism
- Eukaryotic Initiation Factor-2/genetics
- Eukaryotic Initiation Factor-2/metabolism
- Gene Expression
- Immediate-Early Proteins/genetics
- Immediate-Early Proteins/metabolism
- Male
- Mice
- Mice, Inbred C57BL
- Mice, Knockout
- Pancreas, Exocrine/metabolism
- Pancreas, Exocrine/pathology
- Pancreatitis/chemically induced
- Pancreatitis/genetics
- Pancreatitis/metabolism
- Pancreatitis/pathology
- Pancreatitis/physiopathology
- Protein Phosphatase 1
- RNA, Messenger/metabolism
- Regeneration
- Severity of Illness Index
- Stress, Physiological/chemically induced
- Stress, Physiological/genetics
- Stress, Physiological/metabolism
- Stress, Physiological/pathology
- Stress, Physiological/physiopathology
- Time Factors
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Affiliation(s)
- Agnes S Kowalik
- Department of Physiology, The University of Western Ontario, London, Ontario, Canada
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31
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Guo L, Sans MD, Gurda GT, Lee SH, Ernst SA, Williams JA. Induction of early response genes in trypsin inhibitor-induced pancreatic growth. Am J Physiol Gastrointest Liver Physiol 2007; 292:G667-77. [PMID: 17095753 DOI: 10.1152/ajpgi.00433.2006] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Endogenous CCK release induced by a synthetic trypsin inhibitor, camostat, stimulates pancreatic growth; however, the mechanisms mediating this growth are not well established. Early response genes often couple short-term signals with long-term responses. To study their participation in the pancreatic growth response, mice were fasted for 18 h and refed chow containing 0.1% camostat for 1-24 h. Expression of 18 early response genes were evaluated by quantitative PCR; mRNA for 17 of the 18 increased at 1, 2, 4, or 8 h. Protein expression for c-jun, c-fos, ATF-3, Egr-1, and JunB peaked at 2 h. Nuclear localization was confirmed by immunohistochemistry of c-fos, c-jun, and Egr-1. Refeeding regular chow induced only a small increase of c-jun and none in c-fos expression. JNKs and ERKs were activated 1 h after camostat feeding as was the phosphorylation of c-jun and ATF-2. AP-1 DNA binding evaluated by EMSA showed a significant increase 1-2 h after camostat feeding with participation of c-jun, c-fos, ATF-2, ATF-3, and JunB shown by supershift. The CCK antagonist IQM-95,333 blocked camostat feeding-induced c-jun and c-fos expression by 67 and 84%, respectively, and AP-1 DNA binding was also inhibited. In CCK-deficient mice, the maximal response of c-jun induction and AP-1 DNA binding were reduced by 64 and 70%, respectively. These results indicate that camostat feeding induces a spectrum of early response gene expression and AP-1 DNA binding and that these effects are mainly CCK dependent.
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Affiliation(s)
- Lili Guo
- Dept of Molecular and Integrative Physiology, Univ of Michigan Medical School, Ann Arbor, MI 48109-0622, USA
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32
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Chowdhury P, Bose C, Udupa KB. Nicotine-induced proliferation of isolated rat pancreatic acinar cells: effect on cell signalling and function. Cell Prolif 2007; 40:125-41. [PMID: 17227300 PMCID: PMC6496628 DOI: 10.1111/j.1365-2184.2007.00418.x] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2006] [Accepted: 09/13/2006] [Indexed: 01/18/2023] Open
Abstract
OBJECTIVES The aim of the current study was to investigate whether nicotine treatment would induce the proliferation of isolated rat primary pancreatic acinar cells in culture by activating mitogen-activated protein kinase (MAPK) signalling and exocrine secretion. MATERIALS AND METHODS A nicotine dose- and time-response curve was initially developed to determine the optimal dose and time used for all subsequent studies. Proliferation studies were conducted by cell counting and confirmed further by bromodeoxyuridine (BrdU) incorporation and flow cytometry assays. MAPK signalling studies were conducted by Western blot analysis. Localization of ERK1/2 signals, with or without nicotine and the MAPK inhibitor, was visualized by immunofluorescence. RESULTS Nicotine treatment caused dose-dependent activation of extracellular signal-regulated kinases (ERK1/2), the maxima occurring at 100 micro m and at 3 min after treatment; the response was suppressed by the ERK1/2 inhibitor. Maximal nicotine-induced cell proliferation occurred at 24 h, and UO126-treatment significantly reduced this response. Exposure of cells to 100 microm nicotine for 6 min significantly enhanced both baseline and cholecystokinin-stimulated cell function, and these effects were not affected by treatment with the inhibitor of ERK1/2 but were suppressed by mecamylamine, a nicotinic receptor antagonist. CONCLUSIONS Our results suggest that nicotine treatment induced cell proliferation of isolated pancreatic acinar cells and that this is coupled with the activation of MAPK signalling with no effect on its function. Hence, in primary cells, the mechanism of induction and regulation of these two processes, cell proliferation and cell function, by nicotine treatment are independent of each other.
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Affiliation(s)
- P Chowdhury
- Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, College of Medicine, Little Rock, AR 72205, USA.
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33
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Abstract
Cell proliferation is an important process in life for growth of normal and cancer cells. The signal transduction pathways activated during this process are strictly regulated. This editorial focuses on the role of nicotine, a mitogen, in the induction of signaling pathways resulting in proliferation of pancreatic tumor cells and compares these events with those in normal acinar cells isolated from the rat pancreas. The data shows striking similarities between these two cellular systems. In addition, the editorial reviews very recent literature of the contribution of MAPK signaling in cell lines associated with human diseases. A prospective cellular model of nicotine induced activation of MAPK cascade is presented.
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Affiliation(s)
- Parimal Chowdhury
- Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, 4301 W Markham Street, Little Rock, Arkansas 72205, United States.
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34
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Abstract
Cholecystokinin and gastrin receptors (CCK1R and CCK2R) are G protein-coupled receptors that have been the subject of intensive research in the last 10 years with corresponding advances in the understanding of their functioning and physiology. In this review, we first describe general properties of the receptors, such as the different signaling pathways used to exert short- and long-term effects and the structural data that explain their binding properties, activation, and regulation. We then focus on peripheral cholecystokinin receptors by describing their tissue distribution and physiological actions. Finally, pathophysiological peripheral actions of cholecystokinin receptors and their relevance in clinical disorders are reviewed.
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Affiliation(s)
- Marlène Dufresne
- Institut National de la Santé et de la Recherche Médicale U. 531, Institut Louis Bugnard, Centre Hospitalier Universitaire Rangueil, France
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Tashiro M, Dabrowski A, Guo L, Sans MD, Williams JA. Calcineurin-dependent and calcineurin-independent signal transduction pathways activated as part of pancreatic growth. Pancreas 2006; 32:314-20. [PMID: 16628088 DOI: 10.1097/01.mpa.0000218316.12577.c0] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
OBJECTIVE We have recently reported that pancreatic growth driven by cholecystokinin released endogenously by feeding the synthetic trypsin inhibitor camostat requires the Ca-activated phosphatase calcineurin. In the present study, we evaluated a number of signal transduction pathways for their activation as part of the growth response and whether their activation was dependent on calcineurin. METHODS Male ICR mice were fed with either chow or chow plus 1 mg/g of camostat. FK506 was administered at 3 mg/kg. After various times from 12 hours to 10 days, pancreatic samples were prepared and assayed for activity of various signal transduction pathway components. RESULTS Camostat feeding increased the activation of extracellular signal-regulated kinases, c-Jun NH2-terminal kinases, and phosphorylation of the translation factor eukaryotic initiation factor 4E and activated the mammalian target of rapamycin pathway that leads to phosphorylation of the ribosomal protein S6 and of the eukaryotic initiation factor 4E binding protein but with different time courses. Treatment of mice with the calcineurin inhibitor FK506 totally blocked c-Jun NH2-terminal kinase activation, partially blocked the mammalian target of rapamycin pathway, and had no effect on extracellular signal-regulated kinase activation or the phosphorylation of eukaryotic initiation factor 4E. CONCLUSIONS The pancreatic growth response is accompanied by activation of a number of signaling pathways regulating transcription and translation, some of which are dependent on and some independent of calcineurin.
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Affiliation(s)
- Mitsuo Tashiro
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109-0622, USA
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Andreolotti AG, Bragado MJ, Tapia JA, Jensen RT, Garcia-Marin LJ. Adapter protein CRKII signaling is involved in the rat pancreatic acini response to reactive oxygen species. J Cell Biochem 2006; 97:359-367. [PMID: 16187300 DOI: 10.1002/jcb.20624] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Recent studies demonstrate that reactive oxygen species (ROS) are important mediators of acute pancreatitis, whether induced experimentally or in necrotizing pancreatitis in humans; however, the cellular processes involved remain unclear. Adapter protein CrkII, plays a central role for convergence of cellular signals from different stimuli. Cholecystokinin (CCK), which induces pancreatitis, stimulates CrkII tyrosine phosphorylation and CrkII protein complexes, raising the possibility it can be important in the acinar cell responses to ROS. Therefore, our aim was to investigate whether CrkII signaling is involved in the biological response of rat pancreatic acini to H2O2 and the intracellular mediators implicated. Treatment of isolated rat pancreatic acini with H2O2 rapidly stimulates CrkII phosphorylation, measured as electrophoretic mobility shift and by using a phosphospecific antibody (pTyr221). Tyrosine kinase blocker B44 inhibits the higher phosphorylation state, demonstrating that it occurs mainly in tyrosine residues. H2O2-induced CrkII phosphorylation is time- and concentration-dependent, showing maximal effect with 3 mM H2O2 at 5 min. The intracellular pathways induced by H2O2 leading to CrkII tyrosine phosphorylation do not involve PKC, intracellular calcium, PI3-K or the actin cytoskeleton integrity. ROS generation clearly promotes the formation of protein complex CrkII-PYK2. In conclusion, ROS clearly affect the key adapter protein CrkII signaling by two ways: stimulation of CkII phosphorylation and a functional consequence: formation of CrkII-protein complexes. Because of its central role in activating more distal pathways, CrkII might likely play an important role in the ability of ROS to induce pancreatic cellular injury and pancreatitis.
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37
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Bose C, Zhang H, Udupa KB, Chowdhury P. Activation of p-ERK1/2 by nicotine in pancreatic tumor cell line AR42J: effects on proliferation and secretion. Am J Physiol Gastrointest Liver Physiol 2005; 289:G926-34. [PMID: 16051920 DOI: 10.1152/ajpgi.00138.2005] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
The objectives of the present study were to determine the effect of nicotine on MAPK signaling and on the proliferation of AR42J cells as well as to assess the relationship between MAPK activation and exocrine secretion in these cells. AR42J cells were incubated with nicotine and analyzed for the activation of MAPK by Western blot analysis using their respective antibodies and confirmed by immunohistochemistry. The effect of nicotine on cell proliferation was determined by the spectrophotometric method, and cell function was assessed by cholecystokinin (CCK)-stimulated amylase release into the culture medium. Nicotine at a dose of 100 microM induced phospho-ERK1/2 activation maximally in 3 min compared with untreated cells. Furthermore, immunofluorescence study confirmed the nicotine-induced increase in translocation of phospho-ERK1/2 to the nucleus. Activation of phospho-ERK1/2 was inhibited by an ERK1/2 pathway inhibitor but not by a nicotine receptor antagonist. At the same dose, there was significantly enhanced proliferation of AR42J cells until 72 h without toxic effect, as the percentage of lactate dehydrogenase release remained unchanged. Other MAPKs, c-Jun NH2-terminal kinase 1/2 and p38 MAPK, were not affected by nicotine treatment. At a nicotine dose of 100 microM, the CCK-stimulated release of amylase was maximal at 6 min, and, although a nicotinic receptor antagonist inhibited this response, it was not inhibited by the ERK1/2 pathway inhibitor. We conclude that nicotine treatment induced activation of ERK1/2 and increased the proliferation of AR42J cells. The data further indicate that MAPK signaling by nicotine is independent of the secretory response.
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Affiliation(s)
- Chhanda Bose
- Donald W. Reynolds Department of Geriatrics, University of Arkansas for Medical Sciences, College of Medicine, Little Rock, Arkansas 72205, USA
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38
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Perides G, Sharma A, Gopal A, Tao X, Dwyer K, Ligon B, Steer ML. Secretin differentially sensitizes rat pancreatic acini to the effects of supramaximal stimulation with caerulein. Am J Physiol Gastrointest Liver Physiol 2005; 289:G713-21. [PMID: 15920015 DOI: 10.1152/ajpgi.00519.2004] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Supramaximal stimulation of the rat pancreas with CCK, or its analog caerulein, triggers acute pancreatitis and a number of pancreatitis-associated acinar cell changes including intracellular activation of digestive enzyme zymogens and acinar cell injury. It is generally believed that some of these various acinar cell responses to supramaximal secretagogue stimulation are interrelated and interdependent. In a recent report, Lu et al. showed that secretin, by causing generation of cAMP and activation of PKA, sensitizes acinar cells to secretagogue-induced zymogen activation, and, as a result, submaximally stimulating concentrations of caerulein can, in the presence of secretin, trigger intracellular zymogen activation. We found that secretin also sensitizes acinar cells to secretagogue-induced cell injury and to subapical F-actin redistribution but that it did not alter the caerulein concentration dependence of other pancreatitis-associated changes such as the induction of a peak plateau intracellular [Ca(2+)] rise, inhibition of secretion, activation of ERK1/2, and activation of NF-kappaB. The finding that secretin sensitizes acinar cells to both intracellular zymogen activation and cell injury is consistent with the concept that these two early events in pancreatitis are closely interrelated and, possibly, interdependent. On the other hand, the finding that, in the presence of secretin, caerulein can trigger subapical F-actin redistribution without inhibiting secretion challenges the concept that disruption of the subapical F-actin web is causally related to high-dose secretagogue-induced inhibition of secretion in pancreatic acinar cells.
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Affiliation(s)
- G Perides
- Dept. of Surgery, Tufts-New England Medical Center, Boston, MA 02111, USA.
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39
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Gaiser S, Ahler A, Gundling F, Kruse ML, Savkovic V, Selig L, Teich N, Tomasini R, Dagorn JC, Mössner J, Keim V, Bödeker H. Expression of mutated cationic trypsinogen reduces cellular viability in AR4-2J cells. Biochem Biophys Res Commun 2005; 334:721-8. [PMID: 16036133 DOI: 10.1016/j.bbrc.2005.06.148] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2005] [Accepted: 06/26/2005] [Indexed: 01/29/2023]
Abstract
Mutations in the human cationic trypsinogen are associated with hereditary pancreatitis. The cDNA coding for human cationic trypsinogen was subcloned into the expression vector pcDNA3. The mutations R122H, N29I, A16V, D22G, and K23R were introduced by site directed mutagenesis. We constructed an expression vector coding for active trypsin by subcloning the cDNA of trypsin lacking the coding region for the trypsin activating peptide behind an appropriate signal peptide. Expression of protein was verified by Western blot and measurement of enzymatic activity. AR4-2J cells were transiently transfected with the different expression vectors and cell viability and intracellular caspase-3 activity were quantified. In contrast to wild-type trypsinogen, expression of active trypsin and mutated trypsinogens reduced cell viability of AR4-2J cells. Expression of trypsin and R122H trypsinogen induced caspase-3 activity. Acinar cells might react to intracellular trypsin activity by triggering apoptosis.
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Affiliation(s)
- Sebastian Gaiser
- Medizinische Klinik und Poliklinik II, Universitätsklinikum Leipzig AöR, Ph.-Rosenthal-Str. 27, 04103 Leipzig, Germany
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40
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Gallmeier E, Schäfer C, Moubarak P, Tietz A, Plössl I, Huss R, Göke B, Wagner ACC. JAK and STAT proteins are expressed and activated by IFN-gamma in rat pancreatic acinar cells. J Cell Physiol 2005; 203:209-16. [PMID: 15493010 DOI: 10.1002/jcp.20216] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
The development of acute pancreatitis (AP) is triggered by acinar events, but the subsequent extra-acinar events, particularly a distinct immune response, appear to determine its severity. Cytokines modulate this immune response and are derived not only from immunocytes but also from pancreatic acinar cells. We studied whether pancreatic acinar cells were also capable of responding to cytokines. The JAK/STAT-pathway represents the main effector for many cytokines. Therefore, expression and regulation of JAK and STAT proteins were investigated in rat pancreatic acinar cells. Western blotting showed expression of JAK1, JAK2, Tyk2, and STAT1, STAT2, STAT3, STAT5, STAT6. In addition, STAT1 was reversibly tyrosine-phosphorylated upon the procedure of acinar cell isolation. In contrast, STAT3-phosphorylation occurred spontaneously after pancreas removal and was not reversible within 8 h. STAT1 phosphorylation was also observed upon treatment with IFN-gamma but not upon EGF, TNF-alpha or IL-6, and inhibited by the JAK2-inhibitor AG-490. Immunohistochemistry revealed cytoplasmic expression of unphosphorylated STAT1 in untreated acinar cells and nuclear translocation of phosphorylated STAT1 following IFN-gamma-treatment. Interestingly, although CCK leads to the activation of multiple stress pathways in pancreatic acinar cells, we found no influence of CCK on phosphorylation of STAT1, STAT3, or STAT5 in the pancreas. In conclusion, our data provide further evidence that pancreatic acinar cells are able to interact with immune cells. Besides stimulating immune cells via cytokine secretion, acinar cells are in turn capable of responding to IFN-gamma via JAK2 and STAT1 which may have an impact on the development of AP.
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Affiliation(s)
- E Gallmeier
- Department of Internal Medicine II, Klinikum Grosshadern, Ludwig-Maximilians-University, Munich, Germany.
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Morel C, Ibarz G, Oiry C, Carnazzi E, Bergé G, Gagne D, Galleyrand JC, Martinez J. Cross-interactions of two p38 mitogen-activated protein (MAP) kinase inhibitors and two cholecystokinin (CCK) receptor antagonists with the CCK1 receptor and p38 MAP kinase. J Biol Chem 2005; 280:21384-93. [PMID: 15772081 DOI: 10.1074/jbc.m408851200] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Although SB202190 and SB203580 are described as specific p38 MAP kinase inhibitors, several reports have indicated that other enzymes are also sensitive to SB203580. Using a pharmacological approach, we report for the first time that compounds SB202190 and SB203580 were able to directly and selectively interact with a G-protein-coupled receptor, namely the cholecystokinin receptor subtype CCK1, but not with the CCK2 receptor. We demonstrated that these compounds were non-competitive antagonists of the CCK1 receptor at concentrations typically used to inhibit protein kinases. By chimeric construction of the CCK2 receptor, we determined the involvement of two CCK1 receptor intracellular loops in the binding of SB202190 and SB203580. We also showed that two CCK antagonists, L364,718 and L365,260, were able to regulate p38 mitogen-activated protein (MAP) kinase activity. Using a reporter gene strategy and immunoblotting experiments, we demonstrated that both CCK antagonists inhibited selectively the enzymatic activity of p38 MAP kinase. Kinase assays suggested that this inhibition resulted from a direct interaction with both CCK antagonists. Molecular modeling simulations suggested that this interaction occurs in the ATP binding pocket of p38 MAP kinase. These results suggest that SB202190 and SB203580 bind to the CCK1 receptor and, as such, these compounds should be used with caution in models that express this receptor. We also found that L364,718 and L365,260, two CCK receptor antagonists, directly interacted with p38 MAP kinase and inhibited its activity. These findings suggest that the CCK1 receptor shares structural analogies with the p38 MAP kinase ATP binding site. They open the way to potential design of either a new family of MAP kinase inhibitors from CCK1 receptor ligand structures or new CCK1 receptor ligands based on p38 MAP kinase inhibitor structures.
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Affiliation(s)
- Caroline Morel
- Laboratoire des Aminoacides, Peptides, et Protéines, CNRS Unite Mixte de Recherche-5810, UMI et UMII, UFR Pharmacie, 15, Avenue Charles Flahault, 34093 Montpellier Cedex 5, France
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Sharma A, Tao X, Gopal A, Ligon B, Andrade-Gordon P, Steer ML, Perides G. Protection against acute pancreatitis by activation of protease-activated receptor-2. Am J Physiol Gastrointest Liver Physiol 2005; 288:G388-95. [PMID: 15458925 DOI: 10.1152/ajpgi.00341.2004] [Citation(s) in RCA: 62] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Protease-activated receptor-2 (PAR-2) is a widely expressed tethered ligand receptor that can be activated by trypsin and other trypsin-like serine proteases. In the exocrine pancreas, PAR-2 activation modulates acinar cell secretion of digestive enzymes and duct cell ion channel function. During acute pancreatitis, digestive enzyme zymogens, including trypsinogen, are activated within the pancreas. We hypothesized that trypsin, acting via PAR-2, might regulate the severity of that disease, and to test this hypothesis, we examined the effect of either genetically deleting or pharmacologically activating PAR-2 on the severity of secretagogue-induced experimental pancreatitis. We found that experimental acute pancreatitis is more severe in PAR-2(-/-) than in wild-type mice and that in vivo activation of PAR-2, achieved by parenteral administration of the PAR-2-activating peptide SLIGRL-NH2, reduces the severity of pancreatitis. In the pancreas during the early stages of pancreatitis, the MAPK ERK1/2 is activated and translocated to the nucleus, but nuclear translocation is reduced by activation of PAR-2. Our findings indicate that PAR-2 exerts a protective effect on pancreatitis and that activation of PAR-2 ameliorates pancreatitis, possibly by inhibiting ERK1/2 translocation to the nucleus. Our observations suggest that PAR-2 activation may be of therapeutic value in the treatment and/or prevention of severe clinical pancreatitis, and they lead us to speculate that, from a teleological standpoint, PAR-2 may have evolved in the pancreas as a protective mechanism designed to dampen the injurious effects of intrapancreatic trypsinogen activation.
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Affiliation(s)
- Anupriya Sharma
- Department of Surgery, Tufts-New England Medical Center, Tufts University School of Medicine, Boston, Massachusetts, USA
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Pereda J, Sabater L, Cassinello N, Gómez-Cambronero L, Closa D, Folch-Puy E, Aparisi L, Calvete J, Cerdá M, Lledó S, Viña J, Sastre J. Effect of simultaneous inhibition of TNF-alpha production and xanthine oxidase in experimental acute pancreatitis: the role of mitogen activated protein kinases. Ann Surg 2004; 240:108-116. [PMID: 15213626 PMCID: PMC1356382 DOI: 10.1097/01.sla.0000129343.47774.89] [Citation(s) in RCA: 100] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
OBJECTIVE To assess the effects of inhibiting both tumor necrosis factor (TNF)-alpha production and xanthine oxidase activity on the inflammatory response, mitogen-activated protein kinase (MAPK) activation and mortality in necrotizing acute pancreatitis in rats. SUMMARY BACKGROUND DATA Pancreatic injury triggers 2 major pathways involved in the systemic effects of severe acute pancreatitis: pro-inflammatory cytokines and oxidative stress. METHODS Pancreatitis was induced by intraductal infusion of 3.5% sodium taurocholate. We examined whether treatment with oxypurinol, a specific inhibitor of xanthine oxidase, and/or pentoxifylline, an inhibitor of TNF-alpha production, affects pancreatic damage, ascites, lung inflammation, and MAPK phosphorylation. RESULTS Oxypurinol prevented p38 phosphorylation in the pancreas and partially avoided the rise in lung myeloperoxidase activity. Pentoxifylline prevented erk 1/2 and JNK phosphorylation in the pancreas, and it partially reduced ascites and the rise in lung myeloperoxidase activity. Combined treatment with oxypurinol and pentoxifylline almost completely abolished ascites, MAPK phosphorylation in the pancreas, and the increase in lung myeloperoxidase activity. Histology revealed a reduction in pancreatic and lung damage. These changes were associated with a significant improvement of survival. CONCLUSIONS : Simultaneous inhibition of TNF-alpha production and xanthine oxidase activity greatly reduced local and systemic inflammatory response in acute pancreatitis and decreased mortality rate. These effects were associated with blockade of the 3 major MAPKs.
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Affiliation(s)
- Javier Pereda
- Department of Physiology, University of Valencia, Spain
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Pace A, García-Marin LJ, Tapia JA, Bragado MJ, Jensen RT. Phosphospecific site tyrosine phosphorylation of p125FAK and proline-rich kinase 2 is differentially regulated by cholecystokinin receptor type A activation in pancreatic acini. J Biol Chem 2003; 278:19008-19016. [PMID: 12651850 DOI: 10.1074/jbc.m300832200] [Citation(s) in RCA: 20] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
The focal adhesion kinases, p125FAK and proline-rich kinase 2 (PYK2), are involved in numerous processes as adhesion, cytoskeletal changes, and growth. These kinases have 45% homology and share three tyrosine phosphorylation (TyrP) sites. Little information exists on the ability of stimulants to cause TyrP of each kinase site and the cellular mechanism involved. We explored the ability of the neurotransmitter/hormone, CCK, to stimulate TyrP at each site. In rat pancreatic acini, CCK stimulated TyrP at each site in both kinases. TyrP was rapid except for pY397FAK. The magnitude of TyrP differed with the different FAK and PYK2 sites. The CCK dose-response curve for TyrP for sites in each kinase was similar. CCK-JMV, an agonist of the high affinity receptor state and antagonist of the low affinity receptor state, was less efficacious than CCK at each FAK/PYK2 site and inhibited CCK maximal stimulation. Thapsigargin decreased CCK-stimulated TyrP of pY402PYK2 and pY925FAK but not the other sites. GF109203X reduced TyrP of only the PYK2 sites, pY402 and pY580. GF109203X with thapsigargin decreased TyrP of pY402PYK2 and the three FAK sites more than either inhibitor alone. Basal TyrP of pY397FAK was greater than other sites. These results demonstrate that CCK stimulates tyrosine phosphorylation of each of the three homologous phosphorylation sites in FAK and PYK2. However, CCK-stimulated TyrP at these sites differs in kinetics, magnitude, and participation of the high/low affinity receptor states and by protein kinase C and [Ca2+]i. These results show that phosphorylation of these different sites is differentially regulated and involves different intracellular mechanisms in the same cell.
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Affiliation(s)
- Andrea Pace
- Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA
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Abstract
BACKGROUND/AIMS Treatment of isolated pancreatic acini with high concentrations of cholecystokinin (CCK) is known to induce rapid changes in the cellular morphology. The signalling pathways remain to be characterized. METHODS Pancreatic acini were permeabilized by digitonin and incubated with various agents. The acinar morphology was investigated by microscopy. The activation of p125 focal adhesion kinase was determined by Western blot analysis. Amylase was measured photometrically. RESULTS The functionality of the permeabilized acini was tested by measuring stimulated amylase release. 300 microM GTP gamma S was almost as efficient as CCK to stimulate amylase release, while 300 microM GDP beta S inhibited the CCK-stimulated amylase release. Stimulation of permeabilized acini with 0.1 microM CCK induced similar morphological changes as in unpermeabilized acini. Incubation of permeabilized acini with GTP gamma S mimicked the CCK-induced changes, whereas a preincubation with GDP beta S prevented the CCK effects on the acinar morphology. Inhibition of the small G protein rho, which activates p125 focal adhesion kinase, by Clostridium botulinum C3 transferase also prevented the CCK-stimulated morphological changes. Preincubation of intact acini with cell-permeable inhibitors of protein kinase C, MEK or p38MAPK, or with the intracellular calcium chelator BAPTA/AM was without significant effect on the CCK-stimulated changes. CONCLUSION The CCK-induced morphological changes seem to be mediated by G protein signalling via the small G protein rho and the associated activation of p125 focal adhesion kinase.
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Affiliation(s)
- Karlheinz Kiehne
- First Department of Internal Medicine, Christian Albrechts University, Kiel, Germany.
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Tapia JA, Bragado MJ, García-Marín LJ, Jensen RT. Cholecystokinin-stimulated tyrosine phosphorylation of PKC-delta in pancreatic acinar cells is regulated bidirectionally by PKC activation. BIOCHIMICA ET BIOPHYSICA ACTA 2002; 1593:99-113. [PMID: 12431789 DOI: 10.1016/s0167-4889(02)00346-4] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
PKC-delta is important in cell growth, apoptosis, and secretion. Recent studies show its stability is regulated by tyrosine phosphorylation (TYR-P), which can be stimulated by a number of agents. Many of these stimuli also activate phospholipase C (PLC) cascades and little is known about the relationship between these cascades and PKC-delta TYR-P. Cholecystokinin (CCK) stimulates PKCs but it is unknown if it causes PKC-delta TYR-P and if so, the relationship between these cascades is unknown. In rat pancreatic acini, CCK-8 stimulated rapid PKC-delta TYR-P by activation of the low affinity CCK(A) receptor state. TPA had a similar effect. BAPTA did not decrease CCK-stimulated PKC-delta TYR-P but instead, increased it. A23187 did not stimulate PKC-delta TYR-P. Wortmannin and LY 294002 did not alter CCK-stimulated PKC-delta TYR-P. GF 109203X, at low concentrations, increased PKC-delta TYR-P stimulated by CCK or TPA and at higher concentrations, inhibited it. The cPKC inhibitors, Gö 6976 and safingol, caused a similar increase in TPA- and CCK-stimulated PKC-delta TYR-P. These results demonstrate that CCK(A) receptor activation causes PKC-delta TYR-P through activation of only one of its two receptor affinity states. This PKC-delta TYR-P is not directly influenced by changes in [Ca(2+)](i); however, the resultant activation of PKC-alpha has an inhibitory effect. Therefore, CCK activates both stimulatory and inhibitory PKC cascades regulating PKC-delta TYR-P and, hence, likely plays an important role in regulating PKC-delta degradation and cellular abundance.
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Affiliation(s)
- Jose A Tapia
- Departamento de Fisiología, Universidad de Extremadura, Cáceres 10071, Spain
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Williams JA, Sans MD, Tashiro M, Schäfer C, Bragado MJ, Dabrowski A. Cholecystokinin activates a variety of intracellular signal transduction mechanisms in rodent pancreatic acinar cells. PHARMACOLOGY & TOXICOLOGY 2002; 91:297-303. [PMID: 12688372 DOI: 10.1034/j.1600-0773.2002.910606.x] [Citation(s) in RCA: 61] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
Cholecystokinin (CCK) acting through its G protein-coupled receptor is now known to activate a variety of intracellular signaling mechanisms and thereby regulate a complex array of cellular functions in pancreatic acinar cells. The best studied mechanism is the coupling through heterotrimeric G proteins of the Gq family to activate a phospholipase C leading to an increase in inositol trisphosphate and release of intracellular Ca2+. This pathway along with protein kinase C activation in response to the increase in diacylglycerol stimulates the secretion of digestive enzymes by the process of exocytosis. CCK also activates signaling pathways in acini more related to other processes. The three mitogen activated protein kinase cascades leading to ERKs, JNKs and p38 MAPK are all activated by CCK. CCK activates the ERK cascade by PKC activation of Raf which in turn activates MEK and ERKs. JNKs are activated by a distinct mechanism which requires higher concentrations of CCK. Both ERKs and JNKs are presumed to regulate gene expression. CCK activation of p38 MAPK also plays a role in regulating the actin cytoskeleton through phosphorylation of the small heat shock protein HSP27. The PI3K-PKB-mTOR pathway is activated by CCK and plays a major role in regulating protein synthesis at the translational level. This includes both activation of p70 S6K leading to phosphorylation of ribosomal protein S6 and the phosphorylation of the binding protein for initiation factor 4E leading to formation of the mRNA cap binding complex. Other signaling pathways activated by CCK receptors include NF-kappaB and a variety of tyrosine kinases. Further work is needed to understand how CCK receptors activate most of the above pathways and to better understand the biological events regulated by these diverse signaling pathways.
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Affiliation(s)
- John A Williams
- Department of Physiology, University of Michigan, Ann Arbor, MI 48109, USA.
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Clemons AP, Holstein DM, Galli A, Saunders C. Cerulein-induced acute pancreatitis in the rat is significantly ameliorated by treatment with MEK1/2 inhibitors U0126 and PD98059. Pancreas 2002; 25:251-9. [PMID: 12370536 DOI: 10.1097/00006676-200210000-00007] [Citation(s) in RCA: 55] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
INTRODUCTION Both cerulein and cholecystokinin activate mitogen-activated protein (MAP) kinase (ERK1/2) in vivo and in isolated pancreatic acini. AIMS AND METHODOLOGY ERK1/2 in pancreas homogenates was activated in rats rendered pancreatitic by subcutaneous injections of cerulein (5 microg/kg per hour). To determine if blocking ERK1/2 activity might rescue cerulein-induced acute pancreatitis, the "MAP kinase kinase" (also known as MEK1/2) inhibitors PD98059 and U0126 were administered in vivo. RESULTS In rats pretreated with PD98059 (10 mg/kg per i.v. injection) or U0126 (5 mg/kg per i.v. injection) 30 minutes before and then together with hourly cerulein injections for 3 hours, pancreatitis was significantly attenuated on the basis of pancreatic wet weight and histology. Serum amylase concentration was significantly reduced when PD98059 was administered intraperitoneally (10 mg/kg per intraperitoneal injection). PD98059 also ameliorated pancreatitis over a 6-hour cerulein time course. The phosphorylation of pancreatic ERK1/2 was attenuated in PD98059- and U0126-treated animals at both 30 minutes and 3 hours after cerulein injection. Rats rendered neutropenic with vinblastine and pretreated with U0126 still showed attenuated manifestations of cerulein-induced acute pancreatitis, a finding suggesting that pancreatic ERK1/2 is mostly responsible for the effect, rather than infiltrating neutrophils. CONCLUSIONS Inhibition of pancreatic ERK1/2 in vivo affords significant protection against inflammatory sequelae following cerulein-induced acute pancreatitis.
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Affiliation(s)
- Antoinette P Clemons
- Department of Biochemistry, University of Texas Health Science Center at San Antonio, San Antonio, Texas, USA
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Kiehne K, Herzig KH, Fölsch UR. Differential activation of p42ERK2 and p125FAK by cholecystokinin and bombesin in the secretion and proliferation of the pancreatic amphicrine cell line AR42J. Pancreatology 2002; 2:46-53. [PMID: 12120007 DOI: 10.1159/000049448] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
BACKGROUND AR42J rat pancreatic acinar carcinoma cells have retained the potential to secrete digestive enzymes in addition to their ability to proliferate upon stimulation with regulatory peptides. We investigated the involvement of p42ERK2 and p125FAK (extracellular signal-regulated protein kinase and focal adhesion protein kinase, respectively) by cholecystokinin and bombesin stimulation with regard to secretion and mitogenesis. METHODS The p42ERK2 activity was measured by kinase assay and the activation of p125FAK by antiphosphotyrosine Western blot analysis of p125FAK immunoprecipitates. The expression of both kinases was determined by Western blot analysis, the amylase secretion by colorimetry, and the DNA synthesis by [3H]thymidine incorporation. RESULTS p42ERK2 and p125FAK were activated by cholecystokinin and bombesin with maximum stimulation at concentrations above 10 nM. Bombesin was a weaker activator of p42ERK2 and p125FAK, causing only half of the kinase activity induced by stimulation with cholecystokinin. PD98059 was shown to inhibit p42ERK2, while tyrphostin 25 blocked p125FAK tyrosine phosphorylation. Preincubation of AR42J cells with PD98059 or tyrphostin 25 was without influence on cholecystokinin- or bombesin-stimulated secretion in normal or 72-hour dexamethasone-pretreated cells. In contrast, inhibition of both protein kinases leads to reduced cholecystokinin-stimulated [3H]thymidine incorporation rates. CONCLUSIONS Cholecystokinin induced proliferation of AR42J cells by strong activation of p42ERK2 and p125FAK. Bombesin failed to stimulate DNA synthesis, probably due to its reduced potency to stimulate these kinases. Both protein kinases are not implicated in the process of enzyme secretion.
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Affiliation(s)
- Karlheinz Kiehne
- 1st Department of Internal Medicine, Christian Albrechts University, Schittenhelmstrasse 12, D-24105 Kiel, Germany.
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