Table Of Content

M E T H O D S I N M O L E C U L A R M E D I C I N ETM IInntteerrlleeuukkiinn PPrroottooccoollss EEddiitteedd bbyy LLuukkee AA.. JJ.. OO’’NNeeiillll AAnnddrreeww BBoowwiiee HHuummaannaa PPrreessss Interleukin Protocols M E T H O D S I N M O L E C U L A R M E D I C I N ETM John M. Walker, SERIES EDITOR 69. Gene Therapy Protocols, 2nd ed., edited 51. Angiotensin Protocols, edited by Donna byJeffrey R. Morgan, 2002 H. Wang, 2001 68. Molecular Analysis of Cancer, edited by 50. Colorectal Cancer: Methods and Jacqueline Boultwood and Carrie Fidler, 2002 Protocols, edited by Steven M. Powell, 2001 67. Meningococcal Disease: Methods and Protocols,edited by Andrew J. Pollard 49. Molecular Pathology Protocols, edited by and Martin C. J. Maiden, 2001 Anthony A. Killeen, 2001 66. Meningococcal Vaccines: Methods and 48. Antibiotic Resistance Methods and Protocols,edited by Andrew J. Pollard and Protocols,edited by Stephen H. Gillespie, Martin C. J. Maiden, 2001 2001 65. Nonviral Vectors for Gene Therapy: 47. Vision Research Protocols,edited by P. Methods and Protocols, edited by Mark A, Elizabeth Rakoczy, 2001 Findeis,2001 46. AngiogenesisProtocols,edited byJ. 64. Dendritic Cell Protocols, edited by Stephen Clifford Murray, 2001 P. Robinson and Andrew J. Stagg, 2001 45. Hepatocellular Carcinoma: Methods and 63. Hematopoietic Stem Cell Protocols, Protocols, edited by Nagy A. Habib, 2000 edited by Christopher A. Klug and Craig T. 44. Asthma: Mechanisms and Protocols, edited Jordan,2001 byK. Fan Chung and Ian Adcock, 2001 62. Parkinson’s Disease: Methods and Protocols, 43. Muscular Dystrophy: Methods and edited by M. Maral Mouradian, 2001 Protocols,edited by Katherine B. Bushby 61. Melanoma Techniques and Protocols: and Louise Anderson, 2001 Molecular Diagnosis, Treatment, and 42. Vaccine Adjuvants: Preparation Methods Monitoring,edited by Brian J. Nickoloff, 2001 and Research Protocols, edited by Derek T. O’Hagan, 2000 60. Interleukin Protocols, edited by Luke A. J. O’Neill and Andrew Bowie, 2001 41. Celiac Disease: Methods and Protocols, edited by Michael N. Marsh, 2000 59. Molecular Pathology of the Prions, edited by Harry F. Baker, 2001 40. Diagnostic and Therapeutic Antibodies, edited by Andrew J. T. George and 58. Metastasis Research Protocols: Volume 2, Cell Behavior In Vitro and In Vivo, Catherine E. Urch, 2000 edited by Susan A. Brooks and Udo 39. Ovarian Cancer: Methods and Protocols, Schumacher,2001 edited by John M. S. Bartlett, 2000 57. Metastasis Research Protocols: Volume 1, 38. Aging Methods and Protocols, edited by Analysis of Cells and Tissues,edited by Yvonne A. Barnett and Christopher R. Susan A. Brooks and Udo Schumacher, 2001 Barnett, 2000 56. Human Airway Inflammation: Sampling 37. Electrochemotherapy, Electrogenetherapy, Techniques and Analytical Protocols, edited and Transdermal Drug Delivery: byDuncan F. Rogers and Louise E. Donnelly, Electrically Mediated Delivery of Molecules to 2001 Cells, edited by Mark J. Jaroszeski, Richard 55. Hematologic Malignancies: Methods and Heller, and Richard Gilbert, 2000 Protocols,edited by Guy B. Faguet, 2001 36. Septic Shock Methods and Protocols, 54. Mycobacterium tuberculosis Protocols, edited edited by Thomas J. Evans, 2000 byTanya Parish and Neil G. Stoker, 2001 35. Gene Therapy of Cancer: Methods and 53. Renal Cancer: Methods and Protocols, Protocols, edited by Wolfgang Walther and edited by Jack H. Mydlo, 2001 Ulrike Stein, 2000 52. Atherosclerosis: Experimental Methods and 34. Rotaviruses:Methods and Protocols, edited Protocols,edited by Angela F. Drew, 2001 byJames Gray and Ulrich Desselberger M E T H O D S I N M O L E C U L A R M E D I C I N ETM Interleukin Protocols Edited by Luke A. J. O'Neill Department of Biochemistry and Biotechnology Institute, Trinity College, Dublin, Ireland and Andrew Bowie Department of Pharmacology, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Dublin, Ireland Humana Press Totowa, New Jersey © 2001 Humana Press Inc. 999 Riverview Drive, Suite 208 Totowa, New Jersey 07512 All rights reserved. No part of this book may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, microfilming, recording, or otherwise without written permission from the Publisher. Methods in Molecular Medicine™is a trademark of The Humana Press Inc. The content and opinions expressed in this book are the sole work of the authors and editors, who have warranted due diligence in the creation and issuance of their work. 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Printed in the United States of America. 10 9 8 7 6 5 4 3 2 1 Library of Congress Cataloging in Publication Data Interleukin protocols / edited by Luke A. J. O'Neill and Andrew Bowie. p. cm. -- (Methods in molecular medicine ; 60) Includes bibliographical references and index. ISBN 0-89603-738-X (alk. paper) 1. Interleukins--Laboratory manuals. I. O'Neill, Luke A. J. II. Bowie, Andrew. III. Series. QR185.8.I56 I587 2001 616.07'9--dc21 2001016960 Preface Interleukins are a family of proteins that regulate the maturation, differ- entiation, or activation of cells involved in immunity and inflammation, and belong to a broader family termed cytokines. Collectively these proteins are the key orchestrators of host defense and the response to tissue injury. There are currently 23 different interleukins (numbered from IL-1 to IL-23), although the full extent of the interleukin family will only become clear upon analysis of the human genome sequence. Most important, interleukins are central to the pathogenesis of a wide range of diseases that involve an immune compo- nent, including such conditions as rheumatoid arthritis, multiple sclerosis, ulcerative colitis, psoriasis, and asthma. Interleukins have also been impli- cated in other conditions, including cancer, migraine, myocardial infarction, and depression. In essence, when cells are activated by interleukins, a program of gene expression is initiated in the target cell that alters the cell’s phenotype, leading to enhanced immune reactivity, inflammation, and/or proliferation. Interleukins are therefore at the core of the cellular basis for many diseases. They are the subject of intense investigation by biomedical researchers and the targeting or use of interleukins in the clinic is proceeding apace. Approaches such as targeting IL-4 in asthma or IL-1 in joint disease are being pursued, and it is likely that in the next 5–10 years a number of new therapies based on either inhibit- ing or administering interleukins will be available. In addition, the assaying of interleukins has a role in the diagnosis and prognosis of disease, and polymorphisms in interleukin genes may well be found to predispose individuals to disease. The basis for these many advances in interleukin research lies in the use of a range of methodologies for their study. In Interleukin Protocolswe have brought together a critical mass of chapters covering the major techniques currently available to researchers in this area. The book is divided into five sections. Parts I and II concern a range of methods for assaying interleukin protein and mRNA. The ELISA is the mainstay of assaying interleukin protein production and the chapters here cover the basic methodologies, where to purchase reagents and also recent developments in the use of ELISA. The use v vi Preface of FACS as a method of assaying interleukins intracellularly has been an important advance and is also covered. The ability to measure interleukin mRNA is another important technique in interleukin research and several chapters describe quantitative methods that mainly rely on RT-PCR and RNAse protection. Part II gives examples of how to measure specific interleukins in order to illustrate the approaches that can be used for investigators interested in a particular interleukin. Part III covers the assays of interleukins in specific pathologies, including breast cancer, depression, psoriasis, Grave's disease, migraine, and myocardial infarction. Part IV is related to Part III in that it also concerns pathologies, but has as its focus the assaying of interleukins in different biological fluids rel- evant to disease. These include peritoneal fluids, sputum from asthma patients, synovial fluid from arthritic joints, and cerebrospinal fluid from patients with meningitis. More important, this section covers the difficulties associated with the measuring of interleukins in such fluids. Finally, Part V concerns newer methods in the study of interleukin signal transduction, analysis of polymorphisms in interleukin genes, and the use of cDNA arrays, areas that will surely expand greatly in the next years as the feasibility of assaying the consequences of interleukin action in disease becomes more apparent. Interleukin Protocols will therefore be of interest to a wide range of investigators, from molecular and cell biologists to immunologists to clinical investigators. The discovery of interleukins and the analysis of their role in disease represent major advances in molecular medicine. The methods described will help researchers continue to advance, ultimately leading to bet- ter diagnosis, prognosis, and treatment of many diseases where there remains an unmet medical need. Luke A. J. O'Neill Andrew Bowie Contents Preface .............................................................................................................v Contributors.....................................................................................................xi PART I: GENERAL METHODS FOR ASSAYING INTERLEUKIN PROTEIN AND MRNA................................................................................................. 1 1 ELISAs and Interleukin Research Catherine Greene................................................................................... 3 2 ELISPOT Technique for Assaying Interleukins Almuth Christine Hauer and Mona Bajaj-Elliott...............................17 3 Measurement of Interleukins by ELISPOT Assay with Particular Application to Dual-Color Analysis (Stardust Assay) Yoshihiro Okamoto and Mikio Nishida.............................................29 4 Assaying Interleukins Intracellularly by FACS Liam O'Mahony and Maurice O’Donoghue.......................................41 5 Basic RT-PCR for Measurement of Cytokine Expression Mona Bajaj-Elliott and Almuth Christine Hauer...............................49 6 Quantitative PCR for Measurement of Cytokine Expression Ellen M. Nilsen, Finn-Eirik Johansen, and Per Brandtzaeg...........59 7 Single Cell RT-PCR for the Detection of Cytokine mRNA Moshe Ligumsky, Hovav Nechushtan, and Ehud Razin.................77 8 Quantitation of Cytokine mRNA by Flash-Type Bioluminescence Jeffrey K. Actor....................................................................................83 9 Measurement of Interleukin mRNA by Northern Blotting Using a Nonradioactive Detection Method Ginette Tardif, John A. Di Battista, Jean-Pierre Pelletier, and Johanne Martel-Pelletier.........................................................99 10 Quantification of Mouse IL-6 and TNF-(cid:95) mRNA Using Xplore Assays Scott W. Van Arsdell, Kevin P. Murphy, Csaba Pazmany, Diane Erickson, and Mark D. Moody...........................................111 vii viii Contents 11 Measurement of Cytokine and Chemokine mRNA Using Nonisotopic Multiprobe RNase Protection Assay Zheng-Ming Wang and Roman Dziarski.........................................125 12 Isolation of Total and Bioactive Interleukins by Immunoaffinity-ReceptorAffinity Chromatography Terry M. Phillips and Benjamin F. Dickens....................................145 13 Detection of Cytokines by Immunohistochemistry Mahmoud Huleihel.............................................................................157 14 Whole Blood Assays and the Influence of Circadian Rhythmicity on Human Cytokine Measurement Nikolai Petrovsky...............................................................................163 PART II: ILLUSTRATION OF METHODOLOGIES USING INDIVIDUAL INTERLEUKINS..........175 15 Assaying Interleukin-8 Paul N. Moynagh................................................................................177 16 Measurement of Interleukin-2 by Bioassay Lyanne Weston..................................................................................185 17 Assaying Interleukin-15 Amos Douvdevani, Michal Weiler, and Eli Lewis..........................193 PART III: ASSAYING INTERLEUKINS IN PARTICULAR PATHOLOGIES...........................205 18 Assaying Interleukin-6 in Breast Cancer Judith Harmey, Amanda Haverty, Deirdre Foley, and David Bouchier-Hayes...........................................................207 19 Cytokines, Stress, and Depressive Illness Hymie Anisman, Shawn Hayley, Arun V. Ravindran, Zul Merali, and Jenna Griffiths........................................................................217 20 Measurement of Interleukins in Cutaneous Disorders: Competitive RT-PCR for Determination of IL-10 mRNA Expression in Psoriasis Khusru Asadullah, Antje Haeussler-Quade, Wolf Dietrich Doecke, Wolfram Sterry, and Hans-Dieter Volk.........................................239 21 Interleukins in Graves’ Disease Jan Komorowski................................................................................251 22 Evaluation of Cytokines in Migraine Patients by ELISA Irene Munno, Mariarosaria Marinaro, Giuseppe Lacedra, Antonia Bassi, and Vincenzo Centonze.....................................263 23 Assaying Interleukins in Plasma in the Course of Acute Myocardial Infarction Radek Pudil........................................................................................273 Contents ix PART IV: ASSAYING INTERLEUKINS IN PARTICULAR BIOLOGICAL FLUIDS.................283 24 Measurement of Cytokines in Peritoneal Fluids Reijo Punnonen and Juha Punnonen.............................................285 25 Measurement of Cytokines in Seminal Plasma Marek Glezerman, Eitan Lunenfeld and Mahmoud Huleihel........293 26 Measurement of Cytokines in Induced Sputum: Application to the Investigation of Asthma and COPD Peter G. Gibson and Jodie L. Simpson..........................................305 27 Cytokines in Blister Fluids Franco Ameglio and Luciano D’Auria.............................................313 28 Measurement of Cytokines in Synovial Fluid Fabrizio De Benedetti........................................................................327 29 Measurement of Cytokines in Cerebrospinal Fluid Toshiyuki Yokoyama.........................................................................337 30 Detection of Cytokines in Tears Archana Thakur and Mark D. P. Willcox.........................................345 PART V: NEWER TECHNIQUES:INTERLEUKIN SIGNAL TRANSDUCTION, RECOMBINANT INTERLEUKIN PRODUCTION, INTERLEUKIN GENE POLYMORPHISMS, ANDTHE USEOFCDNA ARRAYS...............................................................355 31 Measurement of Phosphoinositide 3-Kinase Activation by Interleukins Stephen G. Ward................................................................................357 32 Analysis of Interleukin-2 Signaling Using Affinity Precipitations and Polyacrylamide Gel Electrophoresis Paul Brennan and Verónica Athié-Morales....................................369 33 Applications for Green Fluorescent Protein in Cell Signaling Paul Brennan and Emmanuelle Astoul...........................................385 34 Use of Pichia pastoris for Production of Recombinant Cytokines Kevin P. Murphy, Csaba Pazmany, and Mark D. Moody...............393 35 Analysis of Single Nucleotide Polymorphisms in the Interleukin-1B Gene Using 5' Nuclease Assays Emad M. El-Omar...............................................................................409 36 Measurement of Cytokine and Chemokine Gene Expression Patterns Using cDNA Array Zheng-Ming Wang and Roman Dziarski.........................................419 Index............................................................................................................439