Distribution Of Phosphodiesterase Research Articles

222 EXPRESSION AND DISTRIBUTION OF PHOSPHODIESTERASE (PDE) ISOENZYMES IN THE HUMAN URETHRA

You have accessJournal of UrologyBladder and Urethra: Anatomy, Physiology and Pharmacology1 Apr 2011222 EXPRESSION AND DISTRIBUTION OF PHOSPHODIESTERASE (PDE) ISOENZYMES IN THE HUMAN URETHRA George Kedia, Markus Kuczyk, Petter Hedlund, and Stefan Ückert George KediaGeorge Kedia Hannover, Germany More articles by this author , Markus KuczykMarkus Kuczyk Hannover, Germany More articles by this author , Petter HedlundPetter Hedlund Lund, Sweden More articles by this author , and Stefan ÜckertStefan Ückert Hannover, Germany More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.332AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Up untill today, only a very few studies have addressed the physiology/pharmacology of the human urethra (Hassouna M. et al., J. Urol. 129: 1262–1264, 1983; Abdel-Hakim A. et al., J. Urol. 130: 988–991, 1983; Werkström V. et al., BJU Int. 98: 414–423, 2006). An involvement of the nitric oxide (NO)/cyclic GMP pathway in mediating the relaxation of the bladder outlet and urethra during the micturition phase has been supposed. The present study aimed to evaluate in the human male urethra by means of immunohistochemistry the expression and distribution of cyclic AMP and cyclic GMP phosphodiesterase (PDE) isoenzymes in relation to key mediators/proteins of the cyclic GMP and cyclic AMP pathways. METHODS Using conventional immunohistochemistry (double-labelling technique, laser fluorescence microscopy), thin sections (8 μm – 10 μm) of the human male urethra were exposed to anti-PDE1A, -PDE2A, -PDE3A, -PDE4A, -PDE4B, -PDE5A, anti-nNOS (neuronal nitric oxide synthase), -VIP (vasoactive intestinal polypeptide) and -CGRP (calcitonin gene-related peptide) antibodies followed by the application of fluorochrome-labelled secondary antibodies. RESULTS In the smooth muscle (SM) portion of the urethra, immunosignals specific for PDE1A (cyclic AMP/cyclic GMP PDE, calcium/calmodulin-dependent), PDE4A, PDE4B (cyclic AMP PDEs) and PDE5A (cyclic GMP PDE) were registered. SM bundles were seen innervated by varicose nerve fibers characterized by the expression of nNOS. Some of these nerves also presented staining related to VIP or CGRP. No immunoreactivities specific for PDE2A or PDE3A were detected. CONCLUSIONS PDE isoenzymes type 1A, 4A, 4B and 5A are expressed in human urethral smooth muscle. The findings implicate that the cyclic AMP and cyclic GMP pathways work synergistically together in the control of urethral smooth muscle function. This might be of importance with regard to the identification of new pharmacological avenues to facilitate the contraction and relaxation of the human outflow region. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e91 Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information George Kedia Hannover, Germany More articles by this author Markus Kuczyk Hannover, Germany More articles by this author Petter Hedlund Lund, Sweden More articles by this author Stefan Ückert Hannover, Germany More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Phosphodiesterases (PDEs) and PDE inhibitors for treatment of LUTS

Lower urinary tract (LUT) smooth muscle can be relaxed by drugs that increase intracellular concentrations of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). Both of these substances are degraded by phosphodiesterases (PDEs), which play a central role in the regulation of smooth muscle tone. The distribution and functional significance of PDE enzymes vary in different tissues of the LUT. Targeting specific PDE isoenzymes should thus allow organ selectivity. PDE 4 and 5 appear to predominate in the prostate, PDE 1 and 4 are thought to influence detrusor smooth muscle function, and PDE 5 may be functionally important in the urethra and vasculature. Studies on the use of PDE inhibitors to treat various LUT symptoms (LUTS), have yielded favorable results. Thus, positive effects of the PDE 5 inhibitors sildenafil and tadalafil on symptoms and quality of life in men with LUTS, erectile dysfunction, and BPH have also been demonstrated. These effects may be due to effects on cGMP signaling and/or modification of afferent input from bladder, urethral, and prostate tissue. This review gives an update on the distribution of PDEs in structures relevant for LUT function, and discusses how inhibition of these enzymes can contribute to beneficial effects on LUTS. Information for the review was obtained from searches of the PubMed database, and from the authors' files.

Potential role of phosphodiesterases in the development of multiple organ dysfunction

Background. Multiple organ dysfunction syndrome (MODS) is the commonest cause of death in intensive care units. Growing recognition of this disease in the last 30 years may be due to advances in early resuscitation unmasking delayed sequelae in those who would have died previously. MODS occurs after shock of various etiologies, especially sepsis, and may be the result of overactivated systemic inflammation. As yet, therapies directed at preventing or improving MODS have not dramatically altered the outcomes. Phosphodiesterases (PDEs) are enzymes primarily responsible for the breakdown of the intracellular second messengers adenosine 3′,5′-cyclic monophosphate and guanosine 3′,5′-cyclic monophosphate. Although these enzymes were initially described many years ago, new families and subtypes of PDEs continue to be discovered. There is also a wide variation in the distribution of PDEs among tissues or species. This tissue-specific distribution has led to differential pharmacologic actions of PDE inhibitors. E...

Cyclic nucleotide phosphodiesterase (PDE) inhibitors: novel therapeutic agents for progressive renal disease.

Cyclic nucleotides are recognized as critical mediators of many renal functions, including solute transport, regulation of vascular tone, proliferation of parenchymal cells, and inflammation. Although most studies have linked elevated cAMP levels to activation of protein kinase A, cAMP can also directly activate cyclic nucleotide gated ion channels and can signal through activation of GTP exchange factors. Cyclic AMP signaling is highly compartmentalized through plasma membrane localization of adenylyl cyclase and expression of scaffolding proteins that anchor protein kinase A to specific intracellular locations. Cyclic nucleotide levels are largely regulated through catabolic processes directed by phosphodiesterases (PDEs). The PDE superfamily is large and complex, with over 60 distinct isoforms that preferentially hydrolyze cAMP, cGMP, or both. PDEs contribute to compartmentalized cyclic nucleotide signaling. The unique cell- and tissue-specific distribution of PDEs has prompted the development of highly specific PDE inhibitors to treat a variety of inflammatory conditions. In experimental systems, PDE inhibitors have been employed to demonstrate functional compartmentalization of cyclic nucleotide signaling in the kidney. For example, mitogenesis in glomerular mesangial cells and normal tubular epithelial cells is negatively regulated by an intracellular pool of cAMP that is metabolized by PDE3, but not by other PDEs. In Madin-Darby canine kidney cells, an in vitro model of polycystic kidney disease, an intracellular pool of cAMP directed by PDE3 stimulates mitogenesis. In mesangial cells, an intracellular pool of cAMP directed by PDE4 inhibits reactive oxygen species and expression of the potent proin-flammatory cytokine monocyte chemoattractant protein 1. An intracellular pool of cGMP directed by PDE5 regulates solute transport. PDE5 inhibitors ameliorate renal injury in a chronic renal disease model. In this overview, we highlight recent studies to define relationships between PDE expression and renal function and to provide evidence that PDE inhibitors may be effective agents in treating chronic renal disease.

The Effects of Olprinone, a Phosphodiesterase 3 Inhibitor, on Systemic and Cerebral Circulation

Olprinone, a phosphodiesterase (PDE) 3 inhibitor, is used to treat heart failure due to its positive inotropic and vasodilative effects. Selective inhibition of the PDE 3 isozyme increases intracellular adenosine 3;5;-cyclic monophosphate and enhances Ca(2+) influx into cardiac muscle cells. The most significant advantage of PDE 3 inhibitors is their ability not only to enhance myocardial contraction, but to reduce, through vasodilatory action, the stress to which the heart is subjected. In peripheral vessels, the decrease of cytosolic free Ca(2+) induces the vasorelaxation of vascular smooth muscle cells. In this way, olprinone reduces mean aortic and pulmonary artery pressures. Additionally, olprinone exerts differential vasodilatory effects on peripheral vessels in each organ, based on the differences in the distribution of PDE 3 among the organs. With respect to the cerebral circulation, olprinone augments blood flow in the cerebral cortex through direct vasodilatory effects on small cerebral arteries or arterioles. Olprinone increases hepatosplanchnic blood flow and improves oxygen supply. While long-term therapy with PDE 3 inhibitors in patients with chronic heart failure may accelerate the progress of the underlying disease and provoke serious ventricular arrhythmia, olprinone shows good potential for short-term treatment in patients who have experienced severe heart failure or patients who have undergone cardiac surgery.

Cyclic adenosine monophosphate and cyclic guanosine monophosphate-phosphodiesterase isoenzymes in human vagina: Relation to nitric oxide synthase isoforms and vasoactive intestinal polypeptide-containing nerves

Objectives To evaluate the distribution of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) phosphodiesterases (PDEs) in relation to nitric oxide synthase isoforms and vasoactive intestinal polypeptide (VIP) in specimens of the human vagina. Nitric oxide and VIP, mediating biologic signals through cGMP and cAMP, have been assumed to be involved in the control of vaginal smooth muscle. Methods Immunohistochemical techniques were applied to sectioned specimens of the human vaginal wall to evaluate the presence of the PDE isoenzymes 3, 4, 5, and 10 in relation to neuronal nitric oxide synthase (nNOS), endothelial NOS (eNOS), and VIP. Results Immunoreactivity (IR) for cAMP-degrading PDE-4 was observed in the vaginal nonvascular smooth musculature, as well as in the wall of subepithelial arteries. VIP-IR nerves innervated the smooth muscle portion of the vaginal wall and also formed a subepithelial network. Immunoreactivity specific for PDE-5 was also registered in vascular and nonvascular vaginal smooth muscle. Immunosignals for eNOS were detected in the endothelial lining of arteries containing PDE-5-IR smooth muscle cells. These arteries were supplied by nNOS-IR nerve fibers. PDE-10-IR smooth muscle cells were located in muscle bundles of the vaginal wall. Conclusions Our study revealed immunoreactivity specific for PDE-4, PDE-5, and PDE-10 in the vascular and nonvascular smooth muscle of the vagina. Immunosignals for PDE-4 and PDE-5 were also observed in close proximity to nNOS-IR or VIP-IR nerve fibers or to eNOS-IR endothelial cells. The distribution of PDEs may indicate a role of these enzymes in the control of the function of the human vagina.

Distribution of phosphodiesterase I in normal human tissues.

Phosphodiesterase I (PDE I) is an exonuclease capable of hydrolyzing a variety of phosphate ester and pyrophosphate bonds. Cell fractionation and histochemical studies in animal tissues have localized PDE I in the plasma membrane of various epithelia. This suggests a role for the enzyme in active transport. Distribution of PDE I in human tissues has not previously been studied. We have produced a polyclonal antiserum to bovine intestinal PDE I and have demonstrated crossreactivity with the human intestinal enzyme. This polyclonal antiserum was used in PAP immunocytochemistry to localize immunoreactive PDE I in a variety of human tissues. Localization was prominent in the gastrointestinal tract, including the cytoplasm of gastric mucosa parietal cells, cytoplasm of surface epithelium and isolated crypt cells in small intestine, and the colonic epithelial cytoplasm and brush border. Parotid gland acinar cells and scattered ductal cells showed positive cytoplasmic staining. Acinar and scattered pancreatic islet cells contained immunoreactive PDE I, as did Kupffer cells of the liver sinusoids. Immunoreactive PDE I was found in all vascular endothelia. The epithelium of the urinary tract showed extensive immunoreactivity. This included the distal convoluted and collecting tubules of the kidney, and ureteral and bladder urothelium. In previous histochemical studies of animal tissues, no evidence of PDE I activity was noted in male or female reproductive tract. In this study, immunoreactive PDE I was localized to human Sertoli cells and to basal epithelium of the epididymis and prostate acini. Fallopian tube epithelium of female reproductive tract also demonstrated immunoreactive PDI I, as did several cell types in term placenta. Our immunocytochemical results with human tissues differ significantly from previous histochemical studies in animal tissues, principally in the genitourinary system. This may be due in part to the different detection systems employed as well as the higher sensitivity of the immunoperoxidase technique. This underscores the importance of adjunct techniques in tissue surveys. The widespread epithelial distribution of immunoreactive PDE I detected by this polyclonal antibody implies an integral role in cell function, probably in active transport.

Acute in vivo stimulation of low-Km cyclic AMP phosphodiesterase activity by insulin in rat-liver Golgi fractions.

A low-Km phosphodiesterase activity, which is acutely stimulated by insulin in vivo, has been identified in plasma membranes and Golgi fractions prepared from rat liver homogenates in isotonic sucrose. Within seconds after insulin injection (25 micrograms/100 g body weight) cAMP phosphodiesterase activity increases by 30-60% in Golgi fractions and by 25% in plasma membranes; activity in crude particulate and microsomal fractions is unaffected. The increase in activity is short-lived in the light and intermediate Golgi fractions, but persists for at least 10 min in the heavy Golgi fraction. It precedes the translocation of insulin and insulin receptors to these fractions, which is maximal at 5 min. The doses of insulin required for half-maximal and maximal activation are, respectively, 7.5 micrograms/100 g and 25 micrograms/100 g body weight. Golgi-associated cAMP phosphodiesterase activity shows non-linear kinetics; a high-affinity component (Vmax, 13 pmol min-1 mg protein-1; Km, 0.35 microM) is detectable. Insulin treatment increases the Vmax 60-70%, but does not affect the Km. Unlike the low-Km cAMP phosphodiesterase associated with crude particulate fractions, the Golgi-associated activity is not easily extractable by solutions of low or high ionic strength. On analytical sucrose density gradients, low-Km cAMP phosphodiesterase associated with the total particulate fraction equilibrates at lower densities than endoplasmic reticulum and lysosomal markers, but at a higher densities than plasma membrane, Golgi markers and insulin receptors. Insulin treatment increases the specific activity of the enzyme by 20-60% at densities below 1.12 g cm-3, and by 20-40% in the density interval 1.23-1.25 g cm-3. Such treatment also causes a slight, but significant shift in the distribution of phosphodiesterase towards lower densities. It is suggested that Golgi elements or physically similar subcellular structures are a major site of localization of insulin-sensitive cAMP phosphodiesterase in rat liver. However, internalization of the insulin-receptor complex is probably not required for enzyme activation.

Intracellular Localization of Phosphodiesterase Induced by Cyclic Adenosine 3',5'-Monophosphate in Dictyostelium discoideum

The intracellular distribution of phosphodiesterase [EC 3.1.4.17] induced by cyclic adenosine 3',5'-monophosphate (cAMP) in Dictyostelium discoideum was studied. When cAMP-treated cells were homogenized and fractionated according to the method of de Duve et al. ((1955) Biochem, J. 60, 604), the specific activity of phosphodiesterase was highest in the light mitochondrial fraction. Peaks of specific activities of alkaline phosphatase (marker enzyme of membrane) and catalase (marker enzyme of peroxisomes) also appeared in the same fraction as phosphodiesterase. However, after centrifugation of the light mitochondrial fraction in a sucrose density gradient, the activity of phosphodiesterase was clearly separated with that of catalase (density 1.19 g/ml) and showed three peaks at lower density (1.10, 1.13, 1.17 g/ml) with good reproducibility. Some parts (1.13, 1.17 g/ml) of the activity in the gradient overlapped with alkaline phosphatase activity, but in the density fraction of 1.10 g/ml the activity of alkaline phosphatase was hardly detectable. When the light mitochondrial fraction was treated with Emulgen 108, or sonicated, phosphodiesterase was more easily solubilized than alkaline phosphatase and catalase, and was found in supernate after centrifugation at 20,000 X g for 30 min. In order to distinguish the locations of the three enzymes, the supernatant of the light mitochondrial fraction treated with Emulgen 108 was subjected to charge shift electrophoresis. The electrophoretic mobilities of phosphodiesterase and catalase were unaffected by ionic detergent. However, alkaline phosphatase shifted towards the anode in the presence of anionic detergent (sodium deoxycholate), and shifted towards the cathode in cationic detergent (cetyltrimethylammonium bromide), relative to nonionic detergent (Emulgen 108) alone. Thus, some part of the phosphodiesterase induced by cAMP may be associated with the plasma membrane, but the remainder is localized in some kind of intracellular particle of lower density. Moreover, the association with the membrane or particle is more easily dissociated than that of alkaline phosphatase, and the liberated phosphodiesterase is rather hydrophilic.

The Heterogeneous Distribution of 5′-Nucleotidase Among Rabbit Lymphocytes

Abstract Assay of 5′-nucleotidase activity revealed a nonuni-form distribution of enzyme activity among rabbit lymphoid tissues. Although 5′-nucleotidase activity was readily observed in rabbit spleen cells, it was essentially undetectable in cells derived from thymus, blood, bone marrow, and appendix. Popliteal lymph node cells, on the other hand, exhibited approximately 6% of the specific activity observed in spleen. Cytochemical reactions carried out on splenocyte smears showed that only 20% of the spleen cells in the total population possessed demonstrable 5′-nucleotidase activity. The results of cell separation studies indicated that the bulk of the activity was not principally associated with either T cells, B cells, or macrophage cells. However, the association of some enzyme activity with subpopulations of these cell types could not be excluded by this kind of analysis. Autoradiographic studies indicated that 5′-nucleotidase did not afford a selective advantage to those splenocytes possessing the enzyme inasmuch as both enzyme-bearing and enzyme-nonbearing cells, when present in the same population, readily incorporated label derived from extracellular [3H]adenosine-labeled 5′-AMP. Thymus cells, which did not contain a 5′-nucleotidase-bearing population were unable to utilize extracellular 5′-AMP. The distribution of two other plasma membrane-associated enzymes was also explored. Whereas nucleotide pyrophosphatase was more uniformly distributed, the distribution of phosphodiesterase I was quite heterogeneous yet distinguishable from that observed for 5′-nucleotidase.

TRIPHOSPHOINOSITIDE PHOSPHODIESTERASE IN DEVELOPING BRAIN OF THE RAT AND IN SUBCELLULAR FRACTIONS OF BRAIN

Abstract— An assay system for the measurement of triphosphoinositide phosphodiesterase in homogenates of rat brain is described. With triphosphoinositide (TPI) as substrate, and in the presence of 0·1 m‐KCI and saturating amounts of diethyl ether, the activity of phosphodiesterase in myelinated brain was 400–500 μmoles of TPI hydrolysed per g wet wt. per hr. One quarter of the adult level of the enzyme was present in rat brain one day after birth, with the remainder being added prior to and during the early stages of myelination. On subfractionation of brain homogenates, substantial activity of the enzyme was located in the soluble portion and in the paniculate fractions enriched in myelin and synaptosomes. The enzyme associated with the particulate fractions could not be detached from the membranes by any of several methods employed. There was a rough correlation between distribution of phosphodiesterase and that of 5′‐nucleotidase, an enzyme associated with plasma membrane in a number of tissues. Some implications of the results are discussed.