Blood Cell Biochemistry Volume 2 Megakaryocytes, Platelets, Macrophages, and Eosinophils

Blood Cell Biochemistry Series Editor J. R. Harris, North East Thames Regional Transfusion Centre, Brentwood, Essex, England

Volume I Erythroid Cells Edited by J. R. Harris

Volume 2 Megakaryocytes, Platelets, Macrophages, and Eosinophils Edited by J. R. Harris

Volume 3 Lymphocytes and Granulocytes Edited by J. R. Harris

Volume 4 Basophil and Mast Cell Degranulation and Recovery Ann M. Dvorak

A Continuation Order Plan is available for this series. A continuation order will bring delivery of each new volume immediately upon publication. Volumes are billed only upon actual shipment. For further informa­tion please contact the publisher.

Blood Cell Biochemistry Volume 2 Megakaryocytes, Platelets, Macrophages, and Eosinophils

Edited by J. R. Harris North East Thames Regional Transfusion Centre Brentwood, Essex, England

Springer Science+Business Media, LLC

Library of Congress Cataloging-in-Publication Data

Megakaryocytes, platelets, macrophages, and eosinophils I edited by J.R. Harris.

p. em.-- <Blood cell biochemistry: v. 21 Includes bibliographical references and index. ISBN 978-1-4757-9533-2 ISBN 978-1-4757-9531-8 (eBook) DOI 10.1007/978-1-4757-9531-8

1. Blood platelets. 2. Megakaryocytes. 3. Macrophages. 4. Eosinophi Is. I. Harris, James R. II. Ser1es. QP97.M4 1990 612.1'1--dc20

© 1991 Springer Science+Business Media New York Originally published by Plenum Press, New York in 1991 Softcover reprint of the hardcover edition 1991

All rights reserved

90-41741 CIP

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

1st

Contributors

Yoshimitsu Abiko Department of Biochemistry, Nihon University School of Dentis-try at Matsudo, 2-870-1, Sakaecho-Nishi, Matsudo, Chiba 271, Japan

S. J. Ackerman Departments of Pathology and Medicine, Beth Israel Hospital and Harvard Medical School, and Charles A. Dana Research Institute, Beth Israel Hospi­tal, Boston, Massachusetts 02215, USA

Michael Aviram Lipid Research Laboratory, Rambam Medical Center, and Rap-paport Institute for Research in the Medical Sciences, Technion Faculty of Medicine, Haifa, Israel

Rolande Berthier DRF/Laboratory of Hematology, Unite 217 INSERM, Center for Nuclear Studies, F38041 Grenoble Cedex, France

Janine Breton-Gorius France

Unite 91 INSERM, Hospital Henri Mondor, 94010 Creteil,

Alain Duperray DRF/Laboratory of Hematology, Unite 217 INSERM, Center for Nuclear Studies, F38041 Grenoble Cedex, France

Evelyne Dupuy Unite 150 INSERM, Hospital Lariboisiere, 75010 Paris, France

Ann M. Dvorak Departments of Pathology and Medicine, Beth Israel Hospital and Harvard Medical School, and Charles A. Dana Research Institute, Beth Israel Hospi­tal, Boston, Massachusetts 02215, USA

Simon Karpatkin 10016, USA

New York University Medical School, New York, New York

Gerard Marguerie DRF /Laboratory of Hematology, Unite 217 INSERM, Center for Nuclear Studies, F38041 Grenoble Cedex, France

Francine Rendu Unite 150 INSERM, Hospital Lariboisiere, 75010 Paris, France

Yasuko Shibata Department of Biochemistry, Nihon University School of Dentistry at Matsudo, 2-870-1, Sakaecho-Nishi, Matsudo, Chiba 271, Japan

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vi Contributors

Hisashi Takiguchi Department of Biochemistry, Nihon University School of Dentis-try at Matsudo, 2-870-1, Sakaecho-Nishi, Matsudo, Chiba 271, Japan

P. F. Weller Departments of Pathology and Medicine, Beth Israel Hospital and Harvard Medical School, and Charles A. Dana Research Institute, Beth Israel Hospi­tal, Boston, Massachusetts 02215, USA

James G. White Departments of Laboratory Medicine and Pathology, and Pediatrics, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA

J, Michael Wilkinson Department of Biochemistry and Cell Biology, Hunterian Institute, Royal College of Surgeons, Lincoln's Inn Fields, London WC2A 3PN, England

Preface

Blood Cell Biochemistry was initially conceived as part of the Plenum series Subcellular Biochemistry, from which it has developed into a separate series. The present volume is devoted primarily to contributions on megakaryocytes and platelets and, to a lesser extent, to macrophages and eosinophils. The book does not attempt a rigorous or total coverage of the particular topics; it represents the areas of current scientific activity and interest that were selected by the editor at the commencement of this project. In general, the approach has been similar to that adopted for Volume 1 of the series (Erythroid Cells); the same approach will be followed subsequently in Volume 3 (Lymphocytes and Granulocytes).

This book opens with a developmentally oriented chapter by Janine Breton-Gorius on megakaryocyte maturation and platelet release in normal conditions, which serves to set the scene ultrastructurally for much of the data that follow. The biosynthesis and process­ing of platelet glycoproteins in megakaryocytes is dealt with by Alain Duperray and his colleagues, and thereby provides an in-depth biochemical survey of the megakaryocyte. The applications and strengths of crossed immunoelectrophoresis for the study of platelet membrane proteins is then covered by Simon Karpatkin, and a detailed account of the heredity disorders of platelet function is provided by Francine Rendu and Evelyne Dupuy. An impressive ultrastructural presentation, by James G. White, on the cytoskeleton of human blood platelets is appropriately complemented by the chapter by J. Michael Wilkinson, which describes immunological studies on the platelet cytoskeleton. The two following chapters, on macrophages, deal with the effect of lipoproteins and platelets on macrophage cholesterol metabolism (by Michael Aviram) and on macrophage membrane and macrophage structure and function (by Yasuko Shibata and his colleagues). The final chapter (by Ann M. Dvorak and her colleagues) is a detailed ultrastructural presentation on the morphology and function of human eosinophils.

The combination of biochemical and ultrastructural data included within this book, as in the first volume of the series, indicates the strength of this methodological combina­tion as a means for providing an overall view to assist our understanding of blood cell structure and function. Although it must be acknowledged that some relevant topics are not included (due either to their absence in the initial list or to authors backing out during production), it is expected that future volumes of the series will rectify such omissions. It

vii

viii Preface

is hoped that this volume will be of use to hematologists, biochemists, and cell biologists whose areas of research fall within the sphere of the topics covered and, in a broader context, to all those interested in blood cells.

J. R. Harris Brentwood, Essex, England

Contents

Chapter 1 Megakaryocyte Maturation and Platelet Release in Normal and Pathologic Conditions

Janine Breton-Gorius

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I 2. Normal Differentiation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

2.1. Recognition of Megakaryocyte Progenitors and Precursors . . . . . . . . . 2 2.2. Maturational Stages and Ploidy Distribution . . . . . . . . . . . . . . . . . . . . 3 2.3. In Vitro Megakaryocyte Development . . . . . . . . . . . . . . . . . . . . . . . . . 6 2.4. Ultrastructural Aspect of Bone Marrow

Megakaryocyte Maturation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I 0 2. 5. Platelet Release . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17

3. Pathological Megakaryocytopoiesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18 3.1. Hereditary Syndromes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18 3.2. Acquired Malignant Syndromes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20 3.3. Megakaryoblastic Cell Lines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

4. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Chapter 2 Biosynthesis and Processing of Platelet Glycoproteins in Megakaryocytes

Alain Duperray, Rolande Berthier, and Gerard Marguerie

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37 2. Cellular Systems Used in the Study of Megakaryocyte Biochemistry . . . . . . 38

2.1. Purification of Megakaryocytes from Human and Murine Bone Marrow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 38

2.2. Purification of Megakaryocytes Obtained in Culture . . . . . . . . . . . . . . 39 2.3. Murine and Human Megakaryocytic Cell Lines . . . . . . . . . . . . . . . . . . 40

3. Glycoproteins of the Platelet Membrane . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

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3.1. GPib-GPIX............................................... 43 3.2. GPIIb-IIIa . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44

4. Glycoproteins of a-Granules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46 4.1. Fibrinogen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46 4.2. Von Willebrand Factor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48 4.3. Factor V . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49 4.4. GMP-140 (PADGEM Protein) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50 4.5. ProteinS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50

5. Concluding Remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51 6. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51

Chapter 3 Crossed Immunoelectrophoresis for the Study of Platelet Membrane Proteins

Simon Karpatkin

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59 2. Detection oflntrinsic Biologic Activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63 3. Sensitivity and Quantitation of Procedure . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66

3.1. Sensitivity of Antigen-Antibody Immunoprecipitate Arcs . . . . . . . . . 66 3.2. Quantitation of lmmunoprecipitate Arcs . . . . . . . . . . . . . . . . . . . . . . . . 67

4. lmmunoaffinoelectrophoresis with Antigen Ligands . . . . . . . . . . . . . . . . . . . . 68 5. Relative Surface Location of Various Membrane Antigens . . . . . . . . . . . . . . . 69 6. Recognition of Amphiphilic Proteins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71 7. Identification of Charged Antigenic Components

after Enzyme Degradation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72 8. Methodology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72

8.1. Preparation of Platelet Membranes . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72 8.2. Preparation of Anti-Platelet Membrane Antibody . . . . . . . . . . . . . . . . 73 8.3. Crossed Immunoelectrophoresis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73 8.4. Crossed Immunoaffinoelectrophoresis . . . . . . . . . . . . . . . . . . . . . . . . . . 73

9. References..................................................... 74

Chapter 4 Hereditary Disorders of Platelet Function

Francine Rendu and Evelyne Dupuy

1. Classification of Platelet Disorders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77 1.1. Clinical Features of Platelet Disorders . . . . . . . . . . . . . . . . . . . . . . . . . 77 1.2. Normal Platelet Physiology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78 1. 3. Laboratory Screening of Platelet Defects . . . . . . . . . . . . . . . . . . . . . . . 81

2. Thrombocytopathia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83 2.1. Defects in Platelet Adhesion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83 2.2. Defects in Platelet Activation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86 2.3. Defects in Platelet Release Reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . 88

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2.4. Defects of Platelet Aggregation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 96 2.5. Abnormal Platelet Procoagulant Activities . . . . . . . . . . . . . . . . . . . . . . 97

3. Thrombocytopenia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98 3 .1. Megakaryocytopenia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98 3.2. Abnormal Megakaryocyte Fragmentation . . . . . . . . . . . . . . . . . . . . . . . 99 3.3. Wiskott-Aldrich Syndrome . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100

4. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101 5. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 1

Chapter 5 The Cytoskeleton of Human Blood Platelets

James G. White

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113 2. General Aspects of Platelet Structure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 114 3. The Term "Cytoskeleton" . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 114 4. Procedures for Cytoskeletal Preservation . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121 5. The Discoid Platelet . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126 6. Platelet Shape Change . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 7. Dendritic Forms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 8. Spread Forms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130 9. Surface Membrane Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139

10. Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144 11 . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 146

Chapter 6 Immunological Studies of the Platelet Cytoskeleton

J. Michael Wilkinson

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149 2. Morphological Studies of the Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . 150 3. Biochemical Studies of the Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151 4. The Microtubular Ring . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 152 5. Microfilament-Associated Proteins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 154

5.1. Filamin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 154 5.2. Talin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155 5.3. Myosin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156 5 .4. Vinculin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156 5.5. u-Actinin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157 5.6. Gelsolin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 158 5.7. Caldesmon . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 158 5.8. Tropomyosin. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 159 5.9. Profilin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160

6. Cytoskeleton-Associated Surface Receptors . . . . . . . . . . . . . . . . . . . . . . . . . . 161 6.1. GPIIb-llla . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161

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6.2. Other Integrins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161 6.3. GPib . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 162

7. The Membrane Skeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 162 8. Assembly of the Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 164 9. Cytoskeletal Organization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 166

9.1. Membrane Linkages with the Cytoskeleton . . . . . . . . . . . . . . . . . . . . . 166 9.2. The Pseudopodal Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167 9.3. The Contractile Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168

10. Future Directions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169 11. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169

Chapter 7 Effect of Lipoproteins and Platelets on Macrophage Cholesterol Metabolism

Michael Aviram

1. Foam Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 179 2. Plasma Lipoproteins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 181 3. Platelets . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 185 4. Uptake of Cholesterol by Macrophages . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 186

4.1. Non-Receptor-Mediated LDL Uptake . . . . . . . . . . . . . . . . . . . . . . . . . . 186 4.2. LDL (ApoB,E) Receptor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 187 4.3. Acetyl-LDL or Scavenger Receptor . . . . . . . . . . . . . . . . . . . . . . . . . . . 188 4.4. Lipase-Modified LDL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192 4.5. Proteoglycan-Modified LDL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193 4.6. Inflammatory Fluid-Modified LDL . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194 4. 7. Cholesteryl Ester-Protein Complexes

from Atherosclerotic Lesions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194 4.8. "13-VLDL Receptor" . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194 4.9. Uptake of Triglyceride-Rich Lipoprotein . . . . . . . . . . . . . . . . . . . . . . . 195 4.10. Uptake of Platelet-Derived Cholesterol . . . . . . . . . . . . . . . . . . . . . . . . 197

5. Intracellular Metabolism of Cholesterol by Macrophages . . . . . . . . . . . . . . . 198 6. Removal of Cholesterol from Macrophages . . . . . . . . . . . . . . . . . . . . . . . . . . 200 7. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201 8. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202

Chapter 8 Macrophage Membrane: Structure and Function

Yasuko Shibata, Yoshimitsu Abiko, and Hisashi Takiguchi

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 209 2. Macrophage Isolation and Preparation of Its Plasma Membrane . . . . . . . . . . 209 3. Receptors on Plasma Membranes of Macrophages . . . . . . . . . . . . . . . . . . . . . 210

3 .I. Receptors Involved in Phagocytosis . . . . . . . . . . . . . . . . . . . . . . . . . . . 210 3.2. Surface Receptors Involved in Regulation . . . . . . . . . . . . . . . . . . . . . . 213

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3. 3. Receptors for Cytokines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215 3 .4. Other Receptors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218

4. Membrane Lipid-Associated Macrophage Functions . . . . . . . . . . . . . . . . . . . 219 4.1. Arachidonic Acid Metabolites and Phospholipases . . . . . . . . . . . . . . . 219 4.2. Platelet-Activating Factor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 220 4.3. H20 2 Production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 220

5. Cell Surface Antigens and Monoclonal Antibodies for Macrophages . . . . . . 220 5 .1. Cell Surface Antigens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 220 5.2. Monoclonal Antibodies to Macrophages . . . . . . . . . . . . . . . . . . . . . . . 222

6. Membrane-Associated Signal Transmission . . . . . . . . . . . . . . . . . . . . . . . . . . 222 6.1. Ca2 +.................................................... 223 6.2. Protein Kinase C . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 223 6.3. G protein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224

7. Concluding Comments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 224 8. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225

Chapter 9 Subcellular Morphology and Biochemistry of Eosinophils

A.M. Dvorak, S. J. Ackerman, and P. F. Weller

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237 2. Morphology of Eosinophils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 238

2.1. Mature Human Eosinophils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 238 2.2. Mature Eosinophils of Other Species . . . . . . . . . . . . . . . . . . . . . . . . . . 245 2.3. Morphology of Eosinophilic Myelocytes . . . . . . . . . . . . . . . . . . . . . . . 245

3. Ultrastructural Analysis of Eosinophils Using Specialized Ultrastructural Techniques . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 252 3.1. Ultrastructural Autoradiography of [3H]-Arachidonic

Acid Incorporation and Subcellular Localization in Mature Human Eosinophils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 252

3.2. Ultrastructural, Cytochemical, and Immunocytochemical Demonstration of Subcellular Contents in Mature Human Eosinophils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 256

4. Functional Morphology of Human Eosinophils in Vivo . . . . . . . . . . . . . . . . . 267 4.1. Eosinophil Activation Morphology . . . . . . . . . . . . . . . . . . . . . . . . . . . . 267 4.2. Morphologic Release Reactions Observed in Human Eosinophils in

Vivo..................................................... 274 4.3. Expression of Increased Secretory Synthetic Structures and/or

Morphologic Evidence of Cellular Immaturity in Human Tissue Eosinophils in Vivo . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 288

4.4. Necrosis of Tissue Eosinophils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 295 4.5. Mechanisms of Resolution from the Microenvironment of Released

Eosinophil Products . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 298 5. Morphology of Growth Factor-Supplemented Cultures of Human

Eosinophils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 310

xiv Contents

5.1. Basophil and/or Mast Cell (IL-3) Growth Factor-Containing, Human or Murine Cell-Conditioned Supernatant-Supplemented Cultures of Human Cord Blood Mononuclear Cells: Evidence for Different Effects on Human Eosinophil and Basophil Maturation . . . . . . . . . . . 311

5.2. Recombinant Human Interleukin-Supplemented Human Cord Blood Mononuclear Cell or Bone Marrow Cell Cultures: Effects on Eosinophil and Basophil Maturation and Activation Morphologies . . 314

6. Concluding Remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 335 7. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 337

Index............................................................. 345