Mucopolysaccharides - Glycosaminoglycans - of body fluids in health and disease 9783111505688, 9783110084719

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Mucopolysaccharides

Rajendra Varma Ranbir S. Varma

Mucopolysaccharides Glycosaminoglycans of Body Fluids in Health and Disease

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Walter de Gruyter G Berlin · New York 1983 DE

About the Authors Rajendra Varma received undergraduate education at the University of Delhi, India and graduate education at the University of New South Wales, Sydney, Australia with a Ph.D. in 1966. She did postdoctoral work at the Iowa State (1967), Purdue (1968) Universities in U.S.A. and thereafter worked as an assistant professor (Alliance College, Pa.) and as an associate professor (Edinboro State College, Pa.) and from 1971 to the present at Warren State Hospital, Warren, Pennsylvania 16365, U.S.A. She is the director of the biochemistry department at this hospital, a fellow of the American Institute of Chemists, and a member of the New York Academy of Sciences, Sigma Xi, and the American Chemical Society. Ranbir S. Varma earned two graduate degrees at Glancy Medical College, Amritsar, and Panjab University, India, and received postgraduate degrees from the Institute of Basic Medical Sciences, Panjab University, Chandigarh, India, and also the School of Medicine of the State University of New York at Buffalo, N. Y. He worked as an assistant research officer at the All-India Institute of Medical Sciences, New Delhi, India, and as a clinical biochemist/ medical scientist in different hospitals from 1961 to 1970 and at Warren State Hospital, Warren, Pennsylvania 16365, U.S.A. since 1971. He is a bioanalyst certified by the American Board of Bioanalysis, a fellow of the American Institute of Chemists, and a member of the Biochemical Society (U.K.), and Sigma Xi.

CIP-Kurztitelaufnahme der Deutschen Bibliothek Varma, Rajendra: Mucopolysaccharides - Glycosaminoglycans of body fluids in health and disease / Rajendra Varma ; Ranbir S. Varma. - Berlin ; New York . de Gruyter, 1983. ISBN 3-11-008471-6 NE: Varma, Ranbir S.:

Library of Congress Cataloging in Publication Data Varma, R. (Rajendra), 1942 - Mucopolysaccharides (Glycosaminoglycans) of body fluids in health and disease. Bibliography: p. Includes index. 1. Mucopolysaccharides. 2. Glycosaminoglycans. 3. Mucopolysaccharides - Metabolism - Disorders. 4. Body fluids. I. Varma, R. S. (Ranbir S.), 1938 -. II. Title [DNLM: 1. Body fluids Metabolism. 2. Mucopolysaccharides - Metabolism. 3. Glycosaminoglycans - Metabolism. 4. Body fluids -- Metabolism. QU 83 V316m] QP702.M8V37 1983 612'.015782 83-2021 ISBN 3-11008471-6

Copyright © 1983 by Walter de Gruyter & Co., Berlin 30. All rights reserved, including those of translation into foreign languages. No part this book may be reproduced in any form — by photoprint, microfilm or any other means nor transmitted nor translated into a machine language without written permission from the publisher. Printing: Druckerei Gerike GmbH, Berlin. - Binding: Dieter Mikolai, Berlin. - Printed in Germany

Dedicated with love to our sons Rajeev and Sunil

Preface

In recent years there has been a rapid advancement in the knowledge of the structure and function of connective tissues and the importance of the acid mucopolysaccharides (Glycosaminoglycans) in physiological and pathological states. This has been reflected in an increased success in the search for means of amelioration of distressing conditions like coronary arteriosclerosis, rheumatoid arthritis, mucopolysaccharidoses, etc. Also, the measurement of the changes of glycosaminoglycan metabolism in connective tissue reflected in the biological fluids have provided useful parameters for diagnosis and prognosis of the patients. These studies have been approached on a multidisciplinary basis and the contribution to the literature has been enormous and scattered in original articles and reviews in different specialty journals. Since no recent single volume that covers the various aspects of all the biological fluids was available, this monograph was prompted to meet this pressing need. The comprehensive, up-to-date review of the literature presented in this monograph will not only provide a starting point for a new investigator entering the field but will also serve the expert in broadening his/her interest and acquiring a greater depth of knowledge. Because of the broad scope of the field, the subject matter has been organized systematically in a logical sequence of chapters to be of use to the multidisciplinary readers. The first two chapters cover the basic background without involvement of too much of chemistry. The chapter 3 reviews the clinical therapeutic uses and biological roles of glycosaminoglycans. The succeeding chapters relate to the information of glycosaminoglycan metabolism in connective tissues and how these changes in tissue glycosaminoglycans reflected in biological fluids are monitored and interpreted for understanding the mechanisms underlying the physiology and pathology of the tissues and for diagnosis and prognosis of

VIII

diseases. Each biological fluid has been treated separately and virtually all the disease entities reported in literature have been dealt to create a reference work in depth. The authors have tried to be comprehensive and systematic but at some places, where changes in glycosaminoglycans in a disease occur simultaneously in two or more biological fluids, overlap became inevitable and we tried to be repetitious rather than risking omission of a related information. The authors hope that this will give the readers a more complete picture of metabolic changes in glycosaminoglycans occurring in a disease. To ensure an up-to-date bibliography a reference list with title and the first and last pages of the articles has been presented at the end of each chapter. In spite of the tremendous amount of work carried out in the field of connective tissue metabolism, information on glycosaminoglycans in some of the biological fluids is incomplete. Although strictly speaking glycosaminoglycans have not been identified in some minor biological fluids, the authors have digressed to present the information available on other glycoconjugates, like glycoproteins, reported in those biological fluids to serve as a starting point for further exploration. It is important to clarify that the glycosaminoglycans present in the biological fluids are actually the degradation products of the proteoglycans and their aggregates (Chapter 1) in tissues, although in a few instances proteoglycans per se may appear in a biological fluid. Thus a proteoglycan of hyaluronic acid was detected in cyst fluid of cystic mucoid degeneration (Chapter 13) and chondroitin sulfate proteoglycans were found in plasma (Chapter 16) and in synovial fluid from rheumatoid arthritis patients (Chapter 12). In addition to the proteoglycans, aggregates between glycosaminoglycans and glycoproteins were observed in urine (Chapter 17). The connective tissue and glycosaminoglycan metabolism problems are the concerns of people in many scientific specialties. The authors sincerely believe that the integrated, comprehensive survey of the information presented in this monograph will be very useful to diversified groups of individuals, including chemists, biochemists, physiologists, pharmacologists, pathologists, physicians and individuals in the speciality of neurological sciences, orthopedics and laboratory medicine. R. Varma R. S. Varma

Contents

Preface 1.

Chemistry and Metabolism of Glycosaminoglycans and Proteoglycans 1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 1.10 1.11 1.12 1.13 1.14

2.

Nature and classification Modern nomenclature Development of terminology Other related terms in literature Occurrence Chemical structures Organization as proteoglycans and aggregates Role of silicon in molecular organization of glycosaminoglycans Interactions with basic compounds and cations Interaction with other macromolecules Biosynthesis of glycosaminoglycans and proteoglycans Degradation of proteoglycans and glycosaminoglycans Effects of vitamins, hormones and minerals, etc Bibliography

3 3 4 5 10 12 14 23

. .

25 28 29 32 35 39 41

General Methods for Isolation, Fractionation and Analysis . . .

51

2.1 2.2 2.3 2.4 2.5 2.6

51 52 54 59 60

2.7 2.8

Introduction Isolation procedures Fractionation of GAG mixtures Molecular weight determination Fractionation by preparative electrophoresis Characterization by microscopic, optical and spectrometric methods Analysis without hydrolysis Analysis of components in hydrolysates

61 63 64

2.9 2.10 2.11 2.12 2.13 3.

Biological Functions and Therapeutic Uses of Glycosaminoglycans 3.1 3.2 3.3 3.4 3.5 3.6 3.7 3.8 3.9 3.10 3.11 3.12 3.13 3.14 3.15 3.16 3.17

4.

Microfractionation and microanalysis of glycosaminoglycans in mixtures 70 Keratan sulfate determination 70 Hyaluronic acid determination 71 Determination of dermatan and chondroitin sulfates . . . . 71 Bibliography 73

Introduction Control of collagen formation and organization Control of water in extracellular tissues Role in urine concentrating process of kidney Role in immunity, infection and inflammation Role in wound healing Role in eye Role in central nervous system Role in biological lubrication and biomechanics Role in stone formation (lithiasis) Role in hair growth Role as ion-exchangers Role in morphogenesis Role in cell proliferation Role in calcification Therapeutic uses of glycosaminoglycans Bibliography

91 91 92 93 95 96 97 99 99 100 101 103 104 106 106 107 107 110

Gastrointestinal Tract Fluids

123

4.1 4.2 4.3 4.4 4.5 4.6 4.7

123 124 125 134 137 137 139

Introduction Methods of analysis and fractionation Gastric juice Intestinal fluids Aging-related changes in gastrointestinal fluids Gastrointestinal changes in cystic fibrosis Bibliography

XI

5.

Glycosaminoglycans in Fluids and Fluid-Associated Tissues of the Oral Cavity

153

5.1 5.2

Introduction Salivary apparatus and salivary secretion

153 153

5.3

Mucosubstances in saliva and salivary glands

156

5.4 5.5 5.6

Glycosaminoglycans of gingival fluid and gingiva Glycosaminoglycans in oral cyst fluid Relationship between dental plaque, periodontal disease and salivary flow Age-related changes in polysaccharides of salivary glands and oral mucosa Bibliography

163 169

5.7 5.8 6.

Mucopolysaccharides in Fluids and Fluid-Associated Membranes of Respiratory Tract 6.1 6.2 6.3 6.4 6.5 6.6 6.7 6.8 6.9 6.10 6.11 6.12

7.

171 174 175

187

Introduction 187 Mucus production and mucociliary function 187 Mucopolysaccharides in nasal secretion 189 Factors affecting nature and quantity of nasal secretion . 190 Nasal secretion in cystic fibrosis 191 Glycosaminoglycans of nasal and paranasal mucous membranes 192 Acidic glycosaminoglycans in paranasal sinus mucosa . . . 193 Mucopolysaccharides in nasal and paranasal carcinoma . . 194 Mucopolysaccharides of sputum 195 Mucopolysaccharides in bronchial secretion 203 Factors affecting bronchial secretion 205 Bibliography 206

Ocular Fluids

217

7.1 7.2 7.3 7.4 7.5 7.6

217 217 221 226 231 234

Introduction Lacrimal secretion or tears Aqueous humor Vitreous humor Subretinal fluid Bibliography

XII

8.

Bile 8.1 8.2 8.3 8.4 8.5 8.6

9.

10.

Physiological aspects Source and functions of bile mucins Mucosubstances in bile Hexuronic acid conjugates in bile Changes in physiological and pathological states Bibliography

243 243 244 247 250 255

Fluids Associated with Embryonic Membranes

261

9.1 9.2 9.3 9.4 9.5 9.6

261 261 270 271 273 274

Introduction Wharton's jelly of umbilical cord Glycosaminoglycans in Allantoic fluid Glycosaminoglycans in eggs Glycosaminoglycans of cardiac jelly Bibliography

Glycosaminoglycans of Fluids and Fluid-Associated Tissues of Ear 281 10.1 10.2 10.3 10.4 10.5 10.6 10.7 10.8 10.9 10.10 10.11 10.12

11.

243

General consideration Secretion in external ear: Cerumen Pathological middle ear fluid Inner ear fluids: Perilymph and endolymph Source of inner ear fluids Glycosaminoglycans in delicate membranes of inner ear Effects of hormones and vitamins in inner ear Experimental studies in myxedema (hypothyroidism) . . Pathological glycosaminoglycan changes in the inner ear Role of glycosaminoglycans in inner ear function . . . Addendum Bibliography

. . . .

281 282 283 285 290 291 295 296 298 301 303 305

Amniotic Fluid (Liquor amnii)

315

11.1 11.2 11.3

315 315 318

Introduction Origin, volume and fate of amniotic fluid Relationship of amniotic fluid to urine

XIH

11.4 11.5 11.6 11.7 11.8 11.9 11.10 12.

Synovial Fluid 12.1 12.2 12.3 12.4 12.5 12.6 12.7 12.8 12.9

13.

318 325 326 327 330 332 336 345

Introduction 345 Volume and general composition . . 347 Glycosaminoglycan composition 348 Source of Glycosaminoglycans 351 Analytical and fractionation methods 352 Alterations in physiological and pathological states . . . 354 Alterations in Viscosity 361 Synovial membrane in rheumatoid arthritis 362 Bibliography 363

Mucopolysaccharides of Physiological and Pathological Fluid Accumulations (Effusions) 13.1 13.2 13.3 13.4 13.5 13.6 13.7 13.8 13.9

14.

Methods of analysis and fractionation Normal glycosaminoglycan composition Source of glycosaminoglycans in amniotic fluid Glycosaminoglycans variation with gestational age . . . Pathological changes in amniotic fluid glycosaminoglycans Amniocentesis and intrauterine diagnosis of mucopolysaccharidoses: Problems and prospects . . . . Bibliography

Introduction Glycosaminoglycans in extracellular fluids of tumors Blister fluids Cyst fluids Glycosaminoglycans of edema fluids Edema fluid from sexual skin Leprous infiltrates Mucopolysaccharides of serous effusions Bibliography

373 373 . . 373 376 376 378 379 380 381 392

Fluids of the Reproductive System

403

14.1 14.2

403 405

Introduction Semen or seminal fluid

XIV

14.3 14.4 15.

16.

409 421

Cerebrospinal Fluid

433

15.1 15.2 15.3 15.4 15.5 15.7 15.8 15.9

433 434 435 436 437 441 442 443

Introduction Location, origin and fate of CSF Glycosaminoglycans in normal CSF Source of CSF glycosaminoglycans Relationship of CSF glycosaminoglycans to age Lysosomal GAG hydrolyzing enzymes in CSF Effects of hyaluronidase injection into CSF Bibliography

Glycosaminoglycans of Blood

451

16.1 16.2 16.3

451 451

16.4 16.5 16.6 16.7 16.8 17.

Female reproductive system Bibliography

Introduction Source of Plasma Glycosaminoglycans Relationship between glycosaminoglycans in blood and urine Methods of analysis and fractionation Glycosaminoglycans of plasma and serum Glycosaminoglycans of formed elements Physiological and pathological variations in blood glycosaminoglycans Bibliography

453 455 459 463 468 489

Glycosaminoglycans of Urine

511

17.1 17.2 17.3 17.4 .7,5

Introduction Source of urinary glycosaminoglycans Analysis and fractionation procedures Glycosaminoglycans of normal urine Nondialy zable, polymeric and dialyzable, low molecular weight fractions Relationship between plasma, urine and amniotic fluid levels Relationship between glycosaminoglycan and

511 513 514 517

17-hydroxycorticosteroid excretion

521

17.6 17.7

518 520

XV

17.8 17.9 17.10 17.11

Urinary glycosaminoglycans and renal lithiasis Effects of physiological factors Alterations in pathological states Bibliography

18. Secretions of the Skin 18.1 18.2 18.3 18.4 18.5 18.6 18.7 19.

General Views Glycosaminoglycans in normal sweat Glycosaminoglycans of sweat glands in pathological states Variations with age and sex Glycosaminoglycans in sebaceous secretions Hormonal control of sebaceous secretion Bibliography

521 522 534 566 597 597 598 602 604 605 606 607

Other Miscellaneous Body Fluids

613

19.1

Milk

615

19.2

Interstitial fluid

616

19.3

Lymph

618

19.4 19.5

Cartilage Fluid Bibliography

619 621

Subject Index

623

Chapter 1

Chemistry and Metabolism of Glycosaminoglycans and Proteoglycans

1 . 1 Nature and classification The acid mucopolysaccharides (or glycosaminoglycans) are high molecular weight linear heteropolysaccharides formed by polymerization of repeating disaccharide units made up of a hexosamine (glucosamine or galactosamine) and a hexuronic acid (D-glucuronic or/and L-iduronic acid) linked by glycosidic bonds. Acetyl and sulfate substituent groups may be present on either or both residues of the polymer. The glycosaminoglycans (GAG) are present as a component of the extracellular ground substance of the connective tissue. These are synthesized by the connective tissue cells. Besides the cells and the ground substance, the connective tissue also contains collagen and elastin fibers. The connective tissue originates from the differentiation of the embryonic mesenchymal cells. The relative proportions of the cells, fibers and the extracellular ground substance vary in different connective tissues. These variations produce differences in the texture of the connective tissues, as an adaptation to the function performed by different connective tissues. Various GAG show heterogeneity with respect to replacement of a glycosyl residue by another, differences in linkage regions and presence of hybrid polymers. Also, these may show variability in the number and sequence of groups (like sulfate and acetyl) present on the outer parts of the polymer chain. The glycosaminoglycans ( G A G ) may be classified into groups in different ways: On the basis of the presence or absence of sulfate gorups, these can be divided into: a) Non-sulfated GAG include hyaluronic acid which contains equimolar amounts of N-acetylglucosamine and D-glucuronic acid and chondroitin, which is the galactosamine-containing counterpart of hyaluronic acid.

b) Sulfated

GAG contain sulfate groups. Examples are the chondroitin sul-

fates, heparin, heparan sulfate, dermatan sulfate and keratan sulfate. Another classification based on the nature of the hexosamine divides GAG into: a) Glucosaminoglycans are those containing glucosamine e. g., heparin, heparan sulfate and hyaluronic acid and keratan sulfate. b) Galactosaminoglycans are those containing galactosamine. Examples are chondroitin 4- and 6-sulfates and dermatan sulfate. On the basis of the polyanionic character and charge densities, the GAG are classified into: a) Polycarboxylates containing uronic acid but no sulfate groups, e. g., hyaluronic acid and chondroitin b) Polysulfates

contain sulfate groups but no hexuronic acid, e. g. , kera-

tan sulfate in which D-galactose replaces hexuronic acid. c) Polycarboxylsulfates have both carboxyl groups of hexuronic acid and sulfate groups. The chondroitin sulfates, heparan sulfate, heparin and dermatan sulfate belong to this group.

1.2 Modern nomenclature Throughout the different sections of this monograph certain old and new terms related to mucopolysaccharides (glycosaminoglycans) used by different investigators will be mentioned. A survey of both the modern, systematic and old, trivial terminology will be presented first. Meyer (1938) coined the word mucopolysaccharides to describe hexosaminecontaining polysaccharides of animal origin present in free state or as protein salts and which gave viscous (= muco) solutions and like the epithelial secretions had the ability to form "mucin clots" with proteins at a low pH. Since then several investigators have used the prefix "muco" for several types of carbohydrate-containing macromolecules in several different ways as: mucoproteins, mucolipids, mucoids, mucoitin, etc., which created a

great confusion. To ease up the confusing state of nomenclature, Jeanloz (1960) proposed the name glycosaminoglycuronans for mucopolysaccharides containing a hexosamine and hexuronic acid as repeating disaccharide units and other systematic names for members of the glycosaminoglycan family. The old names and systematic names along with their abbreviations are given in Table I. A more detailed systematic nomenclature can be found elsewhere (Jeanloz, 1960; Balazs and Jeanloz, 1965; Balazs, 1970). Although the proposed systematic names are being widely used, the term glycosaminoglycuronans is cumbersome and excludes keratan sulfate which lacks uronic acid. For this reason the term has been replaced by "glycosaminoglycans". In the medical literature the term "mucopolysaccharides1' is still used extensively and the genetic defects of degradation of mucopolysaccharides are called "mucopolysaccharidoses" and not "glycosaminoglycanoses" or "glycosaminoglycuronoses". For this reason we have included the term mucopolysaccharides in the title of this monograph along with the term glycosaminoglycans. The former term is related to the physical properties of the solutions and the latter to the chemical structure and composition of these macromolecules. Although the terms chondroitin sulfates A and C have been replaced by chondroitin 4- and 6-sulfates, the bacterial enzymes for their degradation are still called chondroitinases AC and ABC. Also, there is no systematic name for the oversulfated chondroitin sulfate types D and E. The trend is changing; it appears that the terms like "hypermucopolysacchariduria" will eventually be replaced by "hyperglycosaminoglycanuria".

1.3 Development of terminology

Before the systematic modern nomenclature was suggested (Jeanloz, 1960), various other terms were used for the acidic mucopolysaceharides (acidic glycosaminoglycans). Many of these terms have become obsolete and were actually misleading. The nomenclature went through several changes. A survey of these is presented below not with a view to confuse the reader but to help in interpretation of the older literature.

The amino sugar-containing heteropolysaccharides have been known for a long time. Levene (1925) reviewed the earlier work in a monograph. He considered mucoproteins as conjugated proteins with chondroitin sulfate or mucoitin sulfate as prosthetic groups. Karl Meyer of the Columbia University coined the name "mucopolysaccharides" in 1920. These complexes were later classified and reviewed (Meyer, 1938; Stacey, 1946). Originally the term mucopolysaccharide was chosen to indicate the presence of these macromolecules in viscous solutions. Later it was realized that these macromolecules were not only present in biological fluids or excretions but were also widely distributed in mammalian tissues. Meyer (1938) then defined mucopolysaccharides as "hexosamine-sugar-containing polysaccharides of animal origin occurring in a pure state or as their salts with proteins". Blix (1940) used the term "acidoglucoproteins" for the heteropolysaccharides which contained acid carbohydrate radicals: uronic acid, sialic acid and ester sulfate, in comparison to "neutroglucoproteins" which lacked acidic groups and contained hexosamines and neutral sugars. Meyer (1953) in discussing the amino sugar-containing compounds defined mucopolysaccharides as those containing hexosamine. The mucopolysaccharides were present in mucoproteins and glycoproteins in firm chemical union with protein and contained over 4 % hexosamine in mucoproteins and less than 4 % hexosamine in glycoproteins. Stacey (1946) divided the protein-carbohydrate compounds into mucopolysaccharides and mucoproteins. The term mucopolysaccharide was used for heteropolysaccharide containing a low protein content and with or without hexosamine which gave viscous solutions. The name "mucoproteins" was then used for the carbohydrate-protein compounds in which the protein content predominated. Meyer (1953) gave the following classification on the basis of relative percentage composition of carbohydrates and proteins: Glycoproteins trace-4 % carbohydrate Mucoids 4-15 % carbohydrate Mucoproteins 10-85 % carbohydrate Mucopolysaccharides 100 % carbohydrate. Meyer (1953) considered mucoproteins as ionic or easily split complexes between acid aminopolysaccharides and protein. The impact of Meyer's definition of mucopolysaccharides as 100 % carbohydrates was so strong that the earlier workers used and devised drastic procedures for isolation of

"pure" mucopolysaccharides from tissues and that any presence of protein was considered an impurity. Kent and Whitehouse (1955) used "aminopolysaccharide" for nitrogenous polysaccharides which contained no protein and in which aminosugars (hexosamines) accounted for all the nitrogen. The mucopolysaccharides, though solely polysaccharides in character, conjugate with proteins and lipids to form complexes which may be called mucoproteins and mucolipids, respectively. They called mucoproteins and mucopolysaccharides collectively as mucocomplexes or mucosubstances and these were complexes of protein with carbohydrates containing hexosamine but no glucuronic acid or sulfate. They distinguished mucopolysaccharides from mucoproteins in having aminosugar as the major component in the former and with protein as the major component in mucoproteins. Masamune (1956) used the name glucidamins for proteins having 10-50 % neutral sugars and hexosamines and glycoproteins for the complexes of proteins with acidic polysaccharides. Bettelheim-Jevons (1958) described the complexes of acidic aminopolysaccharides with protein which he believed were through ionic linkages and could be easily split at high pH or ionic strength. From the aforementioned survey of nomenclature of protein-carbohydrate complexes, it is clear that several systems have been proposed and different names have been used for the same compounds. This added a great deal of confusion. To overcome this problem Jeanloz (1960) suggested semisystematic names for the family of mucopolysaccharides as "glycosaminoglycuronans" and the individual names for other members of the family except hyaluronic acid, chondroitin and heparin. This systematized nomenclature has been further elaborated (Balazs and Jeanloz, 1965; Balazs, 1970). In general this nomenclature (Table I) has been widely accepted. However, for the sake of simplicity and to include keratan sulfate, which contains no glucuronic acid "glycosaminoglycans" is the preferred term instead of "glycosaminoglycuronans ".

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