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      Expression and Distribution of Ectonucleotidases in Mouse Urinary Bladder

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      PLoS ONE
      Public Library of Science

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          Abstract

          Background

          Normal urinary bladder function requires bidirectional molecular communication between urothelium, detrusor smooth muscle and sensory neurons and one of the key mediators involved in this intercellular signaling is ATP. Ectonucleotidases dephosphorylate nucleotides and thus regulate ligand exposure to P2X and P2Y purinergic receptors. Little is known about the role of these enzymes in mammalian bladder despite substantial literature linking bladder diseases to aberrant purinergic signaling. We therefore examined the expression and distribution of ectonucleotidases in the mouse bladder since mice offer the advantage of straightforward genetic modification for future studies.

          Principal Findings

          RT-PCR demonstrated that eight members of the ectonucleoside triphosphate diphosphohydrolase (NTPD) family, as well as 5′-nucleotidase (NT5E) are expressed in mouse bladder. NTPD1, NTPD2, NTPD3, NTPD8 and NT5E all catalyze extracellular nucleotide dephosphorylation and in concert achieve stepwise conversion of extracellular ATP to adenosine. Immunofluorescent localization with confocal microscopy revealed NTPD1 in endothelium of blood vessels in the lamina propria and in detrusor smooth muscle cells, while NTPD2 was expressed in cells localized to a region of the lamina propria adjacent to detrusor and surrounding muscle bundles in the detrusor. NTPD3 was urothelial-specific, occurring on membranes of intermediate and basal epithelial cells but did not appear to be present in umbrella cells. Immunoblotting confirmed NTPD8 protein in bladder and immunofluorescence suggested a primary localization to the urothelium. NT5E was present exclusively in detrusor smooth muscle in a pattern complementary with that of NTPD1 suggesting a mechanism for providing adenosine to P1 receptors on the surface of myocytes.

          Conclusions

          Ectonucleotidases exhibit highly cell-specific expression patterns in bladder and therefore likely act in a coordinated manner to regulate ligand availability to purinergic receptors. This is the first study to determine the expression and location of ectonucleotidases within the mammalian urinary bladder.

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          Most cited references54

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          The E-NTPDase family of ectonucleotidases: Structure function relationships and pathophysiological significance

          Ectonucleotidases are ectoenzymes that hydrolyze extracellular nucleotides to the respective nucleosides. Within the past decade, ectonucleotidases belonging to several enzyme families have been discovered, cloned and characterized. In this article, we specifically address the cell surface-located members of the ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase/CD39) family (NTPDase1,2,3, and 8). The molecular identification of individual NTPDase subtypes, genetic engineering, mutational analyses, and the generation of subtype-specific antibodies have resulted in considerable insights into enzyme structure and function. These advances also allow definition of physiological and patho-physiological implications of NTPDases in a considerable variety of tissues. Biological actions of NTPDases are a consequence (at least in part) of the regulated phosphohydrolytic activity on extracellular nucleotides and consequent effects on P2-receptor signaling. It further appears that the spatial and temporal expression of NTPDases by various cell types within the vasculature, the nervous tissues and other tissues impacts on several patho-physiological processes. Examples include acute effects on cellular metabolism, adhesion, activation and migration with other protracted impacts upon developmental responses, inclusive of cellular proliferation, differentiation and apoptosis, as seen with atherosclerosis, degenerative neurological diseases and immune rejection of transplanted organs and cells. Future clinical applications are expected to involve the development of new therapeutic strategies for transplantation and various inflammatory cardiovascular, gastrointestinal and neurological diseases.
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            NPP-type ectophosphodiesterases: unity in diversity.

            Nucleotide pyrophosphatase/phosphodiesterase (NPP)-type ectophosphodiesterases are found at the cell surface as type-I or type-II transmembrane proteins, but are also found extracellularly as secreted or shedded enzymes. They hydrolyze pyrophosphate or phosphodiester bonds in a variety of extracellular compounds including nucleotides, (lyso)phospholipids and choline phosphate esters. Despite their structurally related catalytic domain, each enzyme has well-defined substrate specificity. Catalysis by NPPs affects processes as diverse as cell proliferation and motility, angiogenesis, bone mineralization and digestion. In addition, there is emerging evidence for non-catalytic functions of NPPs in cell signaling. NPP-type ectophosphodiesterases are also implicated in the pathophysiology of cancer, insulin resistance and calcification diseases, and they hold great promise as easily accessible therapeutic targets.
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              Targeted disruption of cd39/ATP diphosphohydrolase results in disordered hemostasis and thromboregulation.

              CD39, or vascular adenosine triphosphate diphosphohydrolase, has been considered an important inhibitor of platelet activation. Unexpectedly, cd39-deficient mice had prolonged bleeding times with minimally perturbed coagulation parameters. Platelet interactions with injured mesenteric vasculature were considerably reduced in vivo and purified mutant platelets failed to aggregate to standard agonists in vitro. This platelet hypofunction was reversible and associated with purinergic type P2Y1 receptor desensitization. In keeping with deficient vascular protective mechanisms, fibrin deposition was found at multiple organ sites in cd39-deficient mice and in transplanted cardiac grafts. Our data indicate a dual role for adenosine triphosphate diphosphohydrolase in modulating hemostasis and thrombotic reactions.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2011
                14 April 2011
                : 6
                : 4
                : e18704
                Affiliations
                [1 ]Renal Research Division, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts, United States of America
                [2 ]Division of Gastroenterology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts, United States of America
                [3 ]Matrix Biology Division, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts, United States of America
                Florida State University, United States of America
                Author notes

                Conceived and designed the experiments: WY SCR WGH. Performed the experiments: WY. Analyzed the data: WY WGH. Contributed reagents/materials/analysis tools: SCR. Wrote the paper: WY SCR WGH.

                Article
                PONE-D-11-01313
                10.1371/journal.pone.0018704
                3077397
                21533188
                fab1259d-5b9e-4995-9762-fd17d15b9316
                Yu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                History
                : 11 January 2011
                : 8 March 2011
                Page count
                Pages: 12
                Categories
                Research Article
                Biology
                Genetics
                Human Genetics
                Model Organisms
                Animal Models
                Mouse
                Molecular Cell Biology
                Signal Transduction
                Membrane Receptor Signaling
                Nucleotide Receptor Signaling
                Medicine
                Nephrology
                Bladder and Ureteric Disorders
                Urology
                Bladder and Ureteric Disorders

                Uncategorized
                Uncategorized

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