Table Of ContentVeroffentlichungen aus der
Geomedizinischen Forschungsstelle
(Leiter: Professor Dr. Dr. h. c. multo G. Schettler)
der Heidelberger Akademie der Wissenschaften
Supplement zu den Sitzungsberichten der
Mathematisch -naturwissenschaftlichen Klasse
Ja hrgang 1990
1. Harenberg D. L. Heene
G. Stehle G. Schettler
(Eds.)
New Trends
in Haemostasis
Coagulation Proteins, Endothelium,
and Tissue Factors
With 71 Figures
Springer-Verlag
Berlin Heidelberg New York
London Paris Tokyo
Hong Kong Barcelona
Priv.-Doz. Dr. Job Harenberg
Prof. Dr. Dieter L. Heene
Dr.eJerd Stehle
I. Medizinische Klinik, Fakultat Klinische Medizin Mannheim
Ruprecht-Karls-Universitat Heidelberg
Theodor-Kutzer-Ufer, 6800 Mannheim, FReJ
Prof. Dr. h.c.mult. eJotthard Schettler
Prasident der Heidelberger Akademie der Wissenschaften
(bis 30.91990)
KarlstraBe 4, 6900 Heidelberg, FReJ
ISBN-13 :978-3-540-53275-0 e-ISBN-13 :978-3-642-84318-1
DOl: 10.1007/978-3-642-84318-1
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Table of Contents
Opening Address ....................................................................................................................... VII
Endothelial Functions and Haemostasis ................................................................................... .
Structure, Shape and Function of Glycosaminoglycans
B. Casu ........................................................................................................................................ 2
Structure and Metabolism of Glycosaminoglycans
R.J. Linhardt, D. Loganathan, A. AI-Hakim, SA Ampofo ......................................................... 12
Fibrin Polymerization and Its Role in Regulating Haemostasis
M.W. Mosesson .......................................................................................................................... 27
Function of von Willebrand Factor in Plasma
F.A. Ofosu ................................................................................................................................... 44
Biosynthesis and Assembly of the Factor VIII -von Willebrand Factor Complex
JA van Mourik, A. Leyte, H.B. van Schijndel, J. Voorberg, R.D. Fonteijin,
H. Pannekoek, M.Ph. Verbeet, K. Mertens ................................................................................ 56
Modulation of Endothelial Cell Function by Hypoxia: Perturbation of Barrier
and Anticoagulant Function and Induction of a Novel Procoagulant Activity
S. Ogawa, C. Esposito, M. Matsumoto, J. Brett, D. Stern ........................................................ 66
Tissue Factors and Endothelium ............................................................................................. 79
Platelet Adhesion; A Concerted Action of Hemodynamic and
Biochemical Processes
P.G. de Groot, J.J. Sixma .......................................................................................................... 80
Modulation of Endothelial Functions by Advanced Glycosylation End Products
C. Esposito, S. Ogawa, H. Vlassara, J. Brett, D. Stern ............................................................. 89
Procogulant Activities of Leukocytes
E. Gray, C. Patel, D.E. Marshall, R. Thorpe, T.w. Barrowcliffe ................................................. 97
Cytokine Induced Stimulation of Endothelial Cells in Vitro
P. Nawroth, D. Stern, A. Bierhaus, P. Schumacher, M. Clauss,
H. Ger1ach, S. Stilgenbauer, R. Ziegler .................................................................................... 103
VI
Anti-Platelet Treatment of Metastasis
A. Poggi, L. Beviglia, M.H. Charon, C. Rossi, G. Marguerie, M.B. Donati .............................. 107
The Molecular Anatomy of Vitronectin as Extracellular Regulator Protein:
Multifunctional Properties of the Heparin-Binding Domain
K.T. Preissner ............................................................................................................................ 123
Mode of Action of Annexin V (Vascular Anticoagulant Alpha),
A Protein Synthesized by the Vessel Wall
R. van Gool, HAM. Andree, H.C. Hemker, C.P.M. Reutelingsperger .................................... 136
Coagulation Inhibitors and Haemostasis ................................................................................ 153
Clinical Pharmacologal Profile of Low Molecular Weight Heparins
J. Harenberg ............................................................................................................................. 154
2
Anticoagulant Actions of Sulphated Polysaccharides
T.w. Barrowcliffe, R.E. Merton, J. Watton, E. Gray, D.P. Thomas .......................................... 165
Development of Synthetic Proteinase Inhibitors as Anticoagulant Agents
F. Markwardt ............................................................................................................................. 177
Antithrombin and Its Deficiency States
DA Lane, R. Caso, R.J. Olds, S.L. Thein ................................................................................ 188
"New" Coagulation Inhibitors
G.J. Broze Jr, T.J. Girard, W.F. Novotny, R.P. Smith .............................................................. 202
Biological Function of Vitamin K Antagonists
J.W. Suttie ................................................................................................................................. 208
Plasminogen Activator Inhibitors: Biological Effects
B. Binder, M. Geiger ................................................................................................................. 221
Fibrin-Specific Clot Lysis with Single Chain Urokinase-Type
Plasminogen Activator (scu-PA)
P.J. Declerck, H.R. Lijnen, D. Collen ........................................................................................ 232
Antibody -Mediated Thrombolysis: Promise and Problems
C. Bode, MS. Runge, T. Nordt, T. Eberle. W. KUbler, E. Haber ............................................... 241
Thromboembolism: uetection a,ld ProphylaxiS .................................................................... 249
Causation and Prevention of Coronary Thrombosis: Present Situation
G.V.R. Born ............................................................................................................................... 250
Non-Invasivd Diagnostic Techniques in Deep Vein Thrombosis
D.P.M. Brandjes, H.R. Bueller, J.w. ten Cate, A.WA Lensing,
G.C.F.M. Rutten, H. Heyboer , .................................................................................................. 257
VII
Evaluation of the Bleeding Effects of Antithrombotic Drugs
J.S. G(nsberg ... . .................................................................... . . .......................... 263
List of Participants ................................................................................................................ 2/0
List of Sponsors ........................................................................................................................ 270
Opening Address
by G. Schettler
The Heidelberg Academy for the Humanities and Sciences seems to me an ideal place to hold
international symposia. Important research facilities are located in Heidelberg, which makes it a
very attractive place for meetings like this reflecting the widespread international collaboration of
the scientific community. We are honoured to welcome scientists from all over Europe, such as
Austria, Belgium, France, Great Britain, Italy, and the Netherlands, as well as from further afield
such as Canada and the United States of America. It is a great pleasure and honour for me as
President of the Academy to act as host for these meetings. I hope that the present symposium
on "New Trends in Haemostasis: coagulation proteins, endothelium, and tissue factors" will be
further example of the success of the Heidelberg international workshops, and that you will enjoy
interesting and stimulating scientific presentations and fruitful discussions in our Heidelberg
Academy Centre. We will do our best to give you some ideas of Heidelberg hospitality and
history, especially during the medieval night at the restaurant "GOldenes Schaf." Thank you all for
coming, and special thanks to all who helped make this meeting a good example of international
cooperation. Nothing promotes friendship better than science.
Endothelial Functions and Haemostasis
Structure, Shape and Function of Glycosaminoglycans
B. Casu
Istituto di Chimica e Biochimica "G. Ronzoni", G. Colombo 81
20133 Milano, Italy
INTRODUCTION
Glycosaminoglycans (GAGs) are anionic polysaccharides widely
distributed in animal tissues: hyaluronic acid (HA) is a component
of sinovial fluid and connective tissues, chondroitin sulfates (Ch-4S
and Ch-6S) are abundant in connective tissues, derma tan sulfate (DeS)
in skin and intestinal mucosa, heparan sulfate (HS) is a matrix
constituent of man~ tissues, and seems to be an ubiquitous component
of cell surfaces IFransson, 19851.
In their native state, most GAGs are linked to a polypeptide chain
as proteoglycans, which may be part of larger aggregates. As
schematized in Fig. 1, the linear GAG chains of proteoglycans run
parallel to each other, forming comb-like structures with different
size and shape depending on the type of GAG and on the tissue
Iphelps,19841·
Some of the natural functions of GAGs are exerted only in the
proteoglycan form. Typically, the ChS chains contribute to the
mechanical properties of connective tissues by providing an elastic
network, the elastic properties being associated with the possibility
of entanglement of parallel chains up to the extent their negative
charges repel each other, restoring the original proteoglycan shape
IRees, 19771. On the other hand, heparin (HEP), which is segregated
in mast cell granules, expresses its many biological activities only
when released from these cells, in the form of polysaccharide chains
much shorter than in the original proteoglycan precursor ILindahl,
19891. HEP and the other GAGs used or considered for therapeutic
purposes are obtained by detaching the polysaccharide chains from the
~olypeptide chains by proteolysis or base-catalyzed S-elimination
I Fransson, 19851.
H- -n\ - ~ (d.~al)
DeS
HS De'
(scleral)
macromolecular
cartilage PG
heparin
Ch-4S
Fig. 1. Glycosaminoglycan proteoglycans IPhelps, 19841
3
GLYCOSAMINOGLYCAN STRUCTURES
Regular and irregular regions
The structure of GAGs is usually represented in terms of their preva_
lent repeating disaccharide sequences (Fig. 2). Formally, the most
cornmon GAGs are co-polymers of a hexosamine and an uronic acid, and
should more properly called glycosaminoglycuronans. The hexosamine
is either D-glucosamine (GlcN) or D-galactosamine (GaIN). GAGs can
accordingly be classified as glucosaminoglycans (HS and HEP) , and
galactosaminoglycans (Ch-4S, Ch-6S, and DeS). (The nonsulfated GAG
hyaluronic acid, not mentioned here, is a glucosaminoglycan; also
not discussed is keratan sulfate (KS), a galactosaminoglycan not
containing uronic acid residues.) GAGs can alternatively be
classified on the basis of their major uronic acid. Whereas the
uronic acid of Ch-4S, Ch-6S and DeS is exclusively D-glucuronic acid
(GlcA), that of DeS and HEP is largely L-iduronic acid (IdoA). The
relative content of these two uronic acids in HS varies widely
according to the biological origin of the GAG. However, GlcA is
prevalent in most HS species.
Sequences of Fig. 2 represent the so-called regular regions of GAGs'
Variants of these sequences contribute to the micro-heterogeneity
of these polysaccharides. Apart from the variable content of GlcA
in IdoA-containing GAGs, and of different degrees of N-acetylation
(N-sulfation) in HS and HEP, a major source of heterogeneity of GAGs
is the variability of their sulfation pattern. Some chondroitin
sulfates are hybrid species, consisting of chondroitin chains where
the sulfate group is either at 0-4 or at 0-6. A minor but signifi
cant proportion of chondroitin sulfates and dermatan sulfate is
"oversulfated", with sulfate substituents at both positions 4 and 6
of the same GalNAc residues, or at 0-2 of the uronic acid residue.
GAGs are heterogeneous also in terms of molecular weight, being
constituted of families of polysaccharide chains of different lengths.
The average degree of polymeri za tion of these chains var.ies according
to the type and biological origin of the GAG. (Typical molecular
weight ranges are 20,000-30,000 for galactosaminoglycans, and 10,000-
15,000 for glucosaminoglycans.)
The most heterogeneous GAGs are HS and HEP, the biosynthesis of which
is more complex than that of galactosaminoglycans. As illustrated
in Fig. 3 for HEP, the biosynthesis of these GAGs proceeds through
a series of modifications of the nonsulfated precursor N-acetyl
heparosan (1), which is in sequence N-deacetylated, N-sulfated,
2-0-sulfated, and 6-0-sulfated, to produce sequences 5. Although
these sequences account for a large proportion of HEP (up to 90% o.f
preparations from beef lung, and up to 75% of preparations from pig
or beef mucosa), an incomplete biosynthetic process usually "leaves
behind" some undersulfated, GlcA- and GlcNAc-containing sequences,
which constitute the "irregular regions" of HEP. 1L indahl , 19891.
An important sequence of the irregular regions of HEP is a
pentasaccharide (Fig. 1A), which is the minimal binding site for
antithrombin III (AT). Typical of this pentasaccharide sequence
is an extra sulfate group at position 3,of the central GlcNS03
residue. This group is introduced by a specific sulfotransferase
when the biosynthetic transformation of the following IdoA and GlcNS03
residues is complete, provided the preceeding uronic acid is still
GlcA 1L indahl , 19891.
