PROTEASES II Potential Role in Health and Disease

Edited by

Walter H. Hörl University of Freiburg

Freiburg, Federal Republic of Germany

and August Heidland University of Würzburg Würzburg, Federal Republic of Germany

PLENUM PRESS • NEW YORK AND LONDON

/

Library of Congress Cataloging in Publication Data International Symposium on Proteases: Potential Role in Health and Disease (2nd: 1987: Rothenburg ob der Tauber, Germany)

Proteases II: potential role in health and disease. (Advances in experimental medicine and biology; v. 240) "Proceedings of the Second International Symposium on Proteases: Potential Role in

Health and Disease, held May 17-20, 1987 in Rothenburg ob der Tauber, Federal Republic of Germany" —T.p. verso.

Includes bibliographies and index. 1. Proteolytic enzymes —Congresses. 2. Proteolytic enzymes —Pathophysiology —

Congresses. 3. Proteolytic enzyme inhibitors —Congresses. I. Horl, Walter H. II. Heidland, August. III. Title. IV. Series. [DNLM: 1. Peptide Hydrolases —congresses. 2. Protease Inhibitors-congresses. Wl AD559v.240 / QU 136161 1987] QP609.P78I57 1987 612'.01516 88-25549 ISBN 0-306-43018-5

Proceedings of the Second International Symposium on Proteases: Potential Role in Health and Disease, held May 17-20, 1987, in Rothenburg ob der Tauber, Federal Republic of Germany

© 1988 Plenum Press, New York A Division of Plenum Publishing Corporation 233 Spring Street, New York, N.Y. 10013 AH 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

Universität* Bibliothek München

Printed in the United States of America

CONTENTS

I PHYSIOLOGY AND PATHOPHYSIOLOGY OF PROTEASES AND THEIR INHIBITORS

A s p a r t i c P r o t e i n a s e s and I n h i b i t o r s f o r t h e i r C o n t r o l i n H e a l t h and D i s e a s e 1

J. Kay, R. A. Ju p p , C. G. N o r e y , A. D. R i c h a r d s , W. A. R e i d , R. T. T a g g a r t , I . M. S a m l o f f , and B. M . Dunn

Human N e u t r a l E n d o p e p t i d a s e 2 4 . 1 1 (NEP, E n k e p h a l i n a s e ) ; F u n c t i o n , D i s t r i b u t i o n and R e l e a s e 13

E. G. Erdös and R. A. S k i d g e l

N e u t r o p h i l E l a s t a s e and C a t h e p s i n G: S t r u c t u r e , F u n c t i o n and B i o l o g i c a l C o n t r o l 23

W. W a t o r e k , D. F a r l e y , G. S a l v e s e n , and J. T r a v i s

The D e g r a d a t i o n o f C o l l a g e n by a M e t a l l o p r o t e i n a s e f r o m Human L e u c o c y t e s 33

U. K o h n e r t , R. O b e r h o f f , J. F e d r o w i t z , U. Bergmann, J. R a u t e r b e r g , and H. T s c h e s c h e

Plasma Membrane P r o t e a s e s as U s e f u l T o o l i n H i s t o c h e m i c a l T o x i c o l o g y 45

R. G r a f and R. G o s s r a u

A c t i v a t i o n o f L e u k o c y t e s D u r i n g P r o l o n g e d P h y s i c a l E x e r c i s e 57

K. K o k o t , R. M. S c h a e f e r , M. T e s c h n e r , U. G i l g e , R. P l a s s , and A. H e i d l a n d

I n h i b i t i o n o f Human N e u t r o p h i l E l a s t a s e by P o l y g u a n y l i c A c i d and o t h e r S y n t h e t i c P o l y n u c l e o t i d e s 65

S. Simon, M. V e r e d , A. R i n e h a r t , J. C h e r o n i s , and A. J a n o f f

I n h i b i t i o n o f Human N e u t r o p h i l E l a s t a s e by A c i d -S o l u b l e I n t e r - A l p h a - T r y p s i n I n h i b i t o r 75

A. Gast and J. G. B i e t h

D e v e l o p m e n t o f E g l i n c as a D r u g : P h a r m a c o k i n e t i c s 83

H. P. N i c k , A. P r o b s t , and H. P. S c h n e b l i

ix

M o n o c l o n a l A n t i b o d i e s R e c o g n i z i n g I n t e r - A l p h a -T r y p s i n - I n h i b i t o r and i t s R e l a t e d F r a g m e n t s -E v i d e n c e f o r t h e I n v o l v e m e n t o f t h e P r o t e i n a s e I n h i b i t o r i n C u t a n e o u s ( P a t h o - ) P h y s i o l o g y 89

C. J u s t u s , K. H o c h s t r a s s e r , and M. D. Kramer

I n h i b i t i o n o f Human C h y m o t r y p s i n - L i k e P r o t e a s e s by A l p h a - 1 - P r o t e i n a s e I n h i b i t o r and A l p h a - 1 -A n t i c h y m o t r y p s i n 97

A. Hayem, D. Marko, A. L a i n e , and M. D a v r i l

I m m u n o r e a c t i v e P a n c r e a t i c S e c r e t o r y T r y p s i n I n h i b i t o r i n G a s t r o i n t e s t i n a l Mucosa 1 0 1

M. Bohe, C. L i n d s t r ö m , and K. O h l s s o n

I I PROTEASES AND LUNG

S e m i s y n t h e t i c I n h i b i t o r s o f Human L e u k o c y t e E l a s t a s e and t h e i r P r o t e c t i v e E f f e c t on Lung E l a s t i n D e g r a d a t i o n i n v i t r o 107

J. Beckmann, A. M e h l i c h , H. R. W e n z e l , and H. T s c h e s c h e

Human B r o n c h i a l P r o t e i n a s e I n h i b i t o r : R a p i d P u r i f i c a t i o n P r o c e d u r e and I n h i b i t i o n o f L e u c o c y t e E l a s t a s e i n P r e s e n c e and i n Absence o f Human Lung E l a s t i n 115

C. B o u d i e r , D. C a r v a l l o , M. B r u c h , C. R o i t s c h , M. C o u r t n e y , and J. G. B i e t h

F u n c t i o n a l S t u d i e s o f Human S e c r e t o r y L e u k o c y t e P r o t e a s e I n h i b i t o r 123

K. O h l s s o n , M. B e r g e n f e l d t , and P. Björk

The R o l e o f Chymase i n I o n o p h o r e - I n d u c e d H i s t a m i n e R e l e a s e f r o m Human P u l m o n a r y Mast C e l l s 133

T. H u l t s c h , M. E n n i s , and H. H. H e i d t m a n n

P r o t e o l y t i c A c t i v i t i e s i n B r o n c h o a l v e o l a r Lavage F l u i d C o r r e l a t e t o S t a g e and C o u r s e o f I n t e r s t i t i a l Lung D i s e a s e 137

M. S c h m i d t and E. B r u g g e r

B e h a v i o u r o f A n g i o t e n s i n C o n v e r t i n g Enzyme, H y d r o x y p r o l i n e and some P r o t e a s e I n h i b i t o r s i n P u l m o n a r y S a r c o i d o s i s 145

M. M a s i a k , B. P o d w y s o c k i , and A. Gaje w s k a

E x p e r i m e n t a l S t u d i e s on t h e A d u l t R e s p i r a t o r y D i s t r e s s Syndrome: E l a s t a s e I n f u s i o n i n N o r m a l and A g r a n u l o c y t i c M i n i p i g s 149

H. B u r c h a r d i and T. S t o k k e

X

I I I PROTEASES AND LIVER

A r g i n y l a t i o n , S u r f a c e H y d r o p h o b i c i t y and D e g r a d a t i o n o f C y t o s o l P r o t e i n s f r o m Rat H e p a t o c y t e s 159

P. B o h l e y , J. K o p i t z , and G. Adam

P r o t e i n a s e I n h i b i t o r s as A c u t e Phase R e a c t a n t s : R e g u l a t i o n o f S y n t h e s i s and T u r n o v e r 171

A. K o j , D. M a g i e l s k a - Z e r o , A. K u r d o w s k a , and J . B e r e t a

R e g u l a t i o n o f P r o t e i n a s e A c t i v i t y by H i g h M o l e c u l a r W e i g h t I n h i b i t o r s : B i o s y n t h e s i s o f Rat A l p h a -M a c r o g l o b r u l i n s 183

T. G e i g e r , T. Andus, D. Kunz, M. H e i s i g , J. B a u e r , N. N o r t h o f f , F. G a u t h i e r , T.-A. T r a n - T h i , K. D e c k e r , and P. C. H e i n r i c h

I n d u c t i o n o f t h e P r o t e i n a s e I n h i b i t o r A l p h a - 2 -M a c r o g l o b u l i n i n Rat H e p a t o c y t e s by a M o n o c y t e - D e r i v e d F a c t o r 191

T. Andus, H. N o r t h o f f , J. B a u e r , U. G a n t e r , D. M ä n n e l , T.-A. T r a n - T h i , K. D e c k e r , and P. C. H e i n r i c h

A s t r o c y t e s S y n t h e s i z e and S e c r e t e A l p h a - 2 - M a c r o -g l o b u l i n : D i f f e r e n c e s Between t h e R e g u l a t i o n o f A l p h a - 2 - M a c r o j i . . b u l i n S y n t h e s i s i n Rat L i v e r and B r a i n 199

J. B a u e r , P . J . G e o i c k e - H a e r t e r , U. G a n t e r , I . R i c h t e r , and W. Gerok

C h a r a c t e r i z a t i o n o f D i f f e r e n t Forms o f D i p e p t i d y l P e p t i d a s e I V f r o m Rat L i v e r and Hepatoma by M o n o c l o n a l A n t i b o d i e s 207

S. H ä r t e l , C. H a n s k i , R. N e u m e i e r , R. G o s s r a u , and W. R e u t t e r

IV MUSCLE PROTEIN DEGRADATION

N o n - L y s o s o m a l , H i g h - M o l e c u l a r - M a s s C y s t e i n e P r o t e i n a s e s f r o m Rat S k e l e t a l M u s c l e 215

B. Dahlmann, L. Kuehn, F. Kopp, H. R e i n a u e r , and W. T. S t a u b e r

R o l e o f F a c t o r s D e r i v e d f r o m A c t i v a t e d M a c r o p h a g e s i n R e g u l a t i o n o f M u s c l e P r o t e i n T u r n o v e r 225

V . E . B a r a c o s

Responses o f L y s o s o m a l and N o n - L y s o s o m a l P r o t e a s e s t o U n l o a d i n g o f t h e S o l e u s 235

E. J. H e n r i k s e n , S. S a t a r u g , M. E. T i s c h l e r , and P. Fürst

xi

C a t h e p s i n B and D A c t i v i t y i n Human S k e l e t a l M u s c l e i n D i s e a s e S t a t e s 243

G. G u a r n i e r i , G. T o i g o , R. S i t u l i n , M. A. D e l B i a n c o , and L. C r a p e s i

H o r m o n a l R e g u l a t i o n o f M u s c l e P r o t e i n C a t a b o l i s m i n A c u t e l y U r e m i c R a t s : E f f e c t o f A d r e n a l e c t o m y and P a r a t h y r o i d e c t o m y 257

R. M. S c h a e f e r , M. Moser, P. K u l z e r , G. P e t e r , A. H e i d l a n d , W. H. Hö r l , and S. G. Mas s r y

V PROTEASES, KIDNEY AND UREMIA

R e l a t i o n Between U r i n a r y P r o t e i n a s e s and P r o t e i n u r i a i n R a t s w i t h a G l o m e r u l a r D i s e a s e 267

J.-C. D a v i n , M. D a v i e s , J.-M. F o i d a r t , J. B. F o i d a r t , C. A. Dechenne, and P. R. Mahi e u

C h a r a c t e r i z a t i o n and C l i n i c a l R o l e o f G l o m e r u l a r and T u b u l a r P r o t e a s e s f r o m Human K i d n e y 275

J. E. S c h e r b e r i c h , G. W o l f , C. S t u c k h a r d t , P. K u g l e r , and W. Schoeppe

E f f e c t o f G l o m e r u l a r P r o t e i n u r i a on t h e A c t i v i t i e s o f L y s o s o m a l P r o t e a s e s i n I s o l a t e d Segments o f Rat P r o x i m a l T u b u l e 283

C. J. U l b r i c h t

M e p r i n P h e n o t y p e and C y c l o s p o r i n A T o x i c i t y i n Mice 293

J. F. R e c k e l h o f f , S. S. C r a i g , R. J. Beynon, and J. S. Bond

P o t e n t i a l R o l e o f L y s o s o m a l P r o t e a s e s i n G e n t a m i c i n N e p h r o t o x i c i t y 305

C. J. O l b r i c h t , E. G u t j a h r , M. F i n k , and K. M. Koch

U r i n a r y P r o t e i n a s e A c t i v i t y i n P a t i e n t s w i t h A c u t e R e n a l F a i l u r e a f t e r Trauma and K i d n e y T r a n s p l a n t a t i o n 309

C. Wanner, S. G r e i b e r , G. K i r s t e , P. S c h o l l m e y e r , and W. H. Hörl

Mechanisms f o r A c t i v a t i o n o f P r o t e o l y s i s i n Urem i a 315

W. E. M i t c h

E v i d e n c e f o r t h e R o l e o f P r o t e i n a s e s i n U r e m i c C a t a b o l i s m 323

R. M. S c h a e f e r , M. T e s c h n e r , G. P e t e r , J. L e i b o l d , P. K u l z e r , and A. H e i d l a n d

E g l i n C F a i l s t o Reduce C a t a b o l i s m i n A c t u e l y U r e m i c R a t s 3 3 1

M. T e s c h n e r , R. M. S c h a e f e r , C. R u d o l f , P. K u l z e r , G. P e t e r , and A. H e i d l a n d

xii

E v i d e n c e f o r P r o t e i n S p l i t P r o d u c t s i n Plasma o f P a t i e n t s w i t h A c u t e R e n a l F a i l u r e 339

M. Haag, H. E. Meyer, P. S c h o l l m e y e r , and W. H. Hörl

P r o t e a s e s and A n t i p r o t e a s e s a t D i f f e r e n t V a s c u l a r S i t e s i n R e n a l F a i l u r e 345

K. B a u s e w e i n , K. S c h a f f e r h a n s , R. G ö t z , U. G i l g e , E. H e i d b r e d e r , and A. H e i d l a n d

P r o t e a s e H i s t o c h e m i s t r y i n N o r m a l and U r e m i c R a t s 3 5 1

R. G o s s r a u , A. H e i d l a n d , and J. H a u n s c h i l d

T o t a l K i n i n o g e n L e v e l s , Plasma R e n i n A c t i v i t y , D o p a m i n e - B e t a - H y d r o x y l ä s e and Plasma C a t e c h o l a m i n e s i n C h r o n i c and A c u t e R e n a l F a i l u r e 3 6 1

K. M a r c z e w s k i , A. K s i a z e k , J. S o l s k i , and Z. P a c h u c k i

V I PROTEOLYTIC ENZYMES DURING EXTRACORPORAL CIRCULATION

B i o c o m p a t i b i l i t y o f D i a l y s i s Membranes: F a c t o r H B i n d i n g C o r r e l a t e s I n v e r s e l y w i t h Complement A c t i v a t i o n I n d i e - t : :ig a L o c a l I m b a l a n c e o f I n v o l v e d P r o t e a s e s / A n t i - P r o t e a s e s 365

E. W. R a u t e r b e r g and E. R i t z

H e m o d i a l y s i s w i t h C u r o p h a n e Membranes Leads t o A l t e r a t i o n o f G r a n u l o c y t e O x i d a t i v e M e t a b o l i s m and L e u k o c y t e S e q u e s t i o n i n t h e Lung 377

G. K o l b , H. S c h ö n e m a n n , W. F i s c h e r , K. B i t t n e r , H. Lange, H. H ö f f k e n , V. Damann, K. J o s e p h , and K . Havemann

E f f e c t o f I m m u n s u p p r e s s i o n on t h e R e l e a s e o f Main G r a n u l o c y t e Components: I n V i v o and i n V i t r o S t u d i e s 385

C. Wanner, B. Simon, A. G ö s e l e , W. R i e g e l , P. S c h o l l m e y e r , and W. H. Hörl

R e l e a s e o f G r a n u l o c y t e P r o t e i n s D u r i n g C a r d i o p u l m o n a r y B y p a s s : E f f e c t o f D i f f e r e n t P h a r m a c o l o g i c a l I n t e r v e n t i o n s 3 9 1

W. R i e g e l , G. S p i l l n e r , V. S c h l o s s e r , K. Lang, and W. H. Hörl

S i g n i f i c a n t R o l e o f P r o t e a s e I n h i b i t i o n by A p r o t i n i n i n M y o c a r d i a l P r o t e c t i o n f r o m P r o l o n g e d C a r d i o p l e g i a w i t h H y p o t h e r m i a 399

M. S u n a m o r i , R. I n n a m i , H. F u j i w a r a , M. Yokoyama, A. S u z u k i

xiii

F i b r i n o l y s i s Caused by C a r d i o - P u l m o n a r y Bypass and Shed M e d i a s t i n a l B l o o d R e t r a n s f u s i o n -I s i t o f C l i n i c a l R e l e v a n c e ? 405

W. D i e t r i c h , A. B a r a n k a y , P. Wendt, A. S t e m b e r g e r , G. B l ü m e l , M. S p a n n a g l , M. Jochum, and J. A. R i c h t e r

V I I PROTEINASES I N CATABOLIC STATES

N u t r i t i o n and P r o t e a s e A c t i v i t y 4 1 1 J. D. K o p p l e

I n s u l i n D e g r a d a t i o n a f t e r I n j u r y i n Man 4 2 1 S. M. H o a r e , K. N. F r a y n , and R. E. O f f o r d

E n d o t o x i n A b o l i s h e s t h e I n d u c t i o n o f A l p h a - 2 - M a c r o -g l o b u l i n S y n t h e s i s i n C u l t u r e d Human M o n o c y t e s I n d i c a t i n g I n h i b i t i o n o f t h e T e r m i n a l M o n o c y t e M a t u r a t i o n i n t o M a c r o p h a g e s 425

J. B a u e r , U. G a n t e r , and W. Gerok

L o c a l and G e n e r a l D e f e n c e Mechanisms i n B a c t e r i a l and C h e m i c a l P e r i t o n i t i s 433

A. L a s s o n , M. D e l s hammer, and K. O h l s s o n

D e f i c i e n t P h a g o c y t o s i s S e c o n d a r y t o P r o t e o l y t i c B r eakdown o f O p s o n i n s i n P e r i t o n i t i s E x u d a t e 4 4 1

A. B i l l i n g , ü. F r ö h l i c h , M. Jochum, and H. Ko r t m a nn

P r o t e o l y s i s and L i p i d P e r o x i d a t i o n - Two A s p e c t s o f C e l l I n j u r y i n E x p e r i m e n t a l H y p o v o l e m i c -T r a u m a t i c Shock 449

H. R e d l , S. H a l l s t r ö m , C. L i e n e r s , W. F ü r s t , and G. S c h l a g

Plasma L e v e l s o f E l a s t a s e 1 P r o t e a s e I n h i b i t o r Complex i n t h e M o n i t o r i n g o f ARDS and M u l t i -Organ F a i l u r e - A Summary o f T h r e e C l i n i c a l T r i a l s 457

H. R e d l , E. P a u l , R. J. A. G o r i s , R. P a c h e r , W. W o l o s z c z u k , and G. S c h l a g

PMN E l a s t a s e and L e u k o c y t e N e u t r a l P r o t e i n a s e I n h i b i t o r ( L N P I ) f r o m G r a n u l o c y t e s as I n f l a m m a t i o n M a r k e r s i n E x p e r i m e n t a l -S e p t i c e m i a 465

R. G e i g e r , S. S o k a l , G. T r e f z , M. S i e b e c k , and H. H o f f m a n n

Plasma D e r i v a t i v e R e p l a c e m e n t T h e r a p y i n D i s s . I n t r a v a s c . Coag . ( D I C ) I n d u c e d by S e p t i c D i s o r d e r s w i t h h i g h l y E l e v a t e d E l a s t a s e A l p h a - l - A T - C o m p l e x e s 473

R. E g b r i n g , R. S e i t z , M. W o l f , K. Havemann, L. L e r c h , H. B l a n k e , and G. Fuchs

xiv

N e u t r o p h i l E l a s t a s e , T h r o m b i n and P l a s m i n i n S e p t i c Shock 4 8 1

R. S e i t z , M. W o l f , R. E g b r i n g , and K. Havemann

E l a s t a s e - A l p h a - l - P r o t e i n a s e I n h i b i t o r : An E a r l y I n d i c a t o r o f S e p t i c e m i a and B a c t e r i a l M e n i n g i t i s i n C h i l d h o o d 485

C. P. S p e e r , M. R e t h w i l m , and M. Gahr

Serum P a n c r e a t i c S e c r e t o r y T r y p s i n I n h i b i t o r ( P S T I ) i n S e r i o u s l y I n j u r e d and S e p t i c P a t i e n t s 493

H. T a n a k a , M. Ogawa, T. Y o s h i o k a , and T. S u g i m o t o

Changes i n P M N - E l a s t a s e i n B l o o d and i n R e n a l and Plasma K a l l i k r e i n - K i n i n S y s t e m s a f t e r S e v e r e B u r n I n j u r y 499

G. B o n n e r , W. N i e r m a n n , R. F e s t g e , and U. B ü c h s l e r

Serum P a n c r e a t i c S e c r e t o r y T r y p s i n I n h i b i t o r ( P S T I ) i n P a t i e n t s w i t h I n f l a m m a t o r y D i s e a s e s 505

M. Ogawa, T. S h i b a t a , T. N i i n o b u , K. Uda, N. T a k a t a , and T. M o r i

The E f f e c t o f A p r o t i n i n A d m i n i s t r a t i o n on t h e I n t r a o p e r a t i v e H i s t a m i n e R e l e a s e and H a e m o s t a t i c D i s o r d e r s 509

H. H a r k e and S. Rahman

I n c r e a s e d M o r t a l i t y i n S e p t i c R a t s a f t e r L e u p e p t i n A p p l i c a t i o n 515

E. K o v a t s , J. K a r n e r , G. O l l e n s c h l ä g e r , J. K a r n e r , A. S i m m e l , and E. R o t h

L y s o s o m a l Enzymes and G r a n u l o c y t e E l a s t a s e i n S y n o v i a l F l u i d a f t e r M u l t i p l e T r a u m a t i c I n j u r i e s 519

M. Hörl and H. P. B r u c h

A S e r i n e P r o t e i n a s e I n h i b i t o r i n Human A r t i c u l a r C a r t i l a g e - P o s s i b l e R o l e i n t h e P a t h o g e n e s i s o f I n f l a m m a t o r y J o i n t D i s e a s e s 523

H. B u r k h a r d t , M. K a s t e n , and S. R a u l s

D e t e c t i o n o f G r a n u l o c y t e E l a s t a s e S p e c i f i c IgG S p l i t P r o d u c t s i n R h e u m a t o i d S y n o v i a l F l u i d 5 3 1

I . E c k l e , G. K o l b , F. N e u r a t h , and K. Havemann

XV

V I I I PROTEASES AND MALIGNOMA

The T C e l l S p e c i f i c S e r i n e P r o t e i n a s e TSP-1: B i o c h e m i c a l C h a r a c t e r i z a t i o n , G e n e t i c A n a l y s i s , and F u n c t i o n a l R o l e 535

H.-G. Simon, U. F r u t h , R. G e i g e r , M. D. Kramer, and M. M. Simon

P a n c r e a t i c S e c r e t o r y T r y p s i n I n h i b i t o r i n Cancer 547

M. Ogawa, N. T o m i t a , A. H o r i i , N. M a t s u u r a , M. H i g a s h i y a m a , T. Yamamoto, A. M u r a t a , T. M o r i , and K. M a t s u b a r a

P r o t e a s e s and A n t i p r o t e a s e s i n A s c i t e s -D i f f e r e n t i a t i o n o f M a l i g n a n t and Non-m a l i g n a n t A s c i t e s and P r e d i c t i o n o f C o a g u l o p a t h y i n A s c i t e s R e t r a n s f u s i o n 555

J. S c h ö l m e r i c h , E. K ö t t g e n , B. A. V o l k , and W. Gerok

A l p h a - 1 - A n t i t r y p s i n and A l p h a - l - A n t i c h y m o t r y p s i n Serum L e v e l i n R e l a t i o n t o S t a g i n g and P o s t o p e r a t i v e C l i n i c a l C o u r s e o f Human C o l o r e c t a l Cancer 5 6 1

A. K u r y l i s z y n - M o s k a l , K. B e r n a c k a , and S . S i e r a k o w s k i

I n h i b i t i o n o f P r o t e a s e s D u r i n g E x t r a c o r p o r e a l E x t r e m i t y P e r f u s i o n 565

H. W a l t h e r , H. M ü l l e r , and K. R. A i g n e r

INDEX 569

xvi

FIBRINOLYSIS CAUSED BY CARD IO-PULMONARY BYPASS AND SHED

MEDIASTINAL BLOOD RETRANSFUSION - IS IT OF CL IN ICAL RELEVANCE?

W. D i e t r i c h 1 , A. Barankayl, P. Wendt 2, A. stemberger 2, G. Blume! 2, M. Spannagl 3, M. Jochum 3, and J.A. R i c h t e r 1

1 I n s t i t u t e f o r Anesthesiology, German Heart Center, Munich ^Technical U n i v e r s i t y , Munich ^Ludwig Maximilian U n i v e r s i t y , Munich

INTRODUCTION The impo r t a n ce of b lood s a v i n g methods d u r i n g c a r d i a c s u r g e r y

is we l l a c c e p t e d . I t i s commonly known t h a t , i n s p i t e of h e p a r i n t r e a tmen t , s ys t em i c c o a g u l a t i o n a n d f i b r i n o l y s i s w i l l be a c t i v a t e d by c a r d i o - p u I m o n a r y b y pa s s (CPB) . Whether t h i s a c t i v a t i o n is of c l i n i c a l r e l e vance was many t imes a po i n t o f d i s c u s s i o n ( 1 ) . S i m i l a r c hanges were f o u nd i n the shed m e d i a s t i n a l b l ood p o s t o p e r a t i v e l y ( 2 ) , w h i c h was r e t r a n s f u s e d to reduce homologous b l ood r e q u i r emen t ( 3 ) . I t is g e n e r a l l y accep ted now, t h a t t h i s method leads to a r e d u c t i o n of homologous b l ood r e qu i r emen t i n c a r d i a c s u r g e r y ( 4 ) , bu t some r e s e r ­v a t i o n s were made because of the low q u a l i t y of the chest t u be b lood ( 5 ) .

The a im of ou r s t u d y was to i n v e s t i g a t e the deg ree of f i b r i n o ­l y s i s i n the shed b l ood and i n p a t i e n t s ' c i r c u l a t i o n , the i n f l u e n c e of lysosomal enzymes of des i n t e g r a t e d o r a c t i v a t e d g r a n u l o c y t e s upon f i b r i n o l y s e s , a nd to answe r the q u e s t i o n , whe t h e r r e t r a n s f u s i o n of shed med i a s t i n a l b l ood causes c l i n i c a l l y r e l e v a n t changes i n p a ­t i e n t s ' h emos t a s i s .

METHODS In twen t y a d u l t p a t i e n t s u n d e r g o i n g c a r d i a c s u r g e r y p a r ame t e r s

of f i b r i n o l y s i s were a n a l y s e d a t the f o l l o w i n g i n s t a n c e s : 1 . be fo re o p e r a t i o n , 2 . a f t e r CPB ( c ompa r i n g the d a t a o b t a i n e d at these two i n s t ances the e f fec t of CPB on f i b r i n o l y s i s can be e s t i m a t e d ) , 3. in the i n t e n s i v e c a r e u n i t be fo re a n d 4 . 20 m i nu t e s a f t e r r e t r a n s f u s i o n . Samples were t a ken f r om p a t i e n t s ' c i r c u l a t i o n as we l l as f r om the chest tube d r a i n a g e . The a p p l i e d a n a l y t i c a l methods f o r measu remen ts of p l a s m i n o g e n , a n t i p l a s m i n , D - d ime r , e a r l y f i b r i n ( o g e n ) d e g r a d a t i o n p r o du c t s ( N - t e r m i n a l Bß-fragments 1-42 a nd 15-42) a n d e l a s t a s e f r om PMN g r a n u l o c y t e s comp lexed w i t h a ^ - p r o t e i n a s e - i n h i b i t o r ( E - a ^ P I ) a r e shown in Tab l e 1 . Re t r ans f u s i on was o n l y pe r f o rmed when a t leas t 250 cc b lood were co l l e c t ed a nd when i t was i n d i c a t e d by the p a ­t i e n t ' s c i r c u l a t o r y s t a t e .

405

Tab l e 1 . Methods

p l a sm i nogen a n t i p l a s m i n D-d imer N - t e rm i n a l Bß-fragments E-a PI

ch romogen i c s u b s t r a t e ch r omogen i c s u b s t r a t e monoc lona l a n t i b o d i e s monoc lona l a n t i b o d i e s PMNE (EL ISA /Me rck )

S 2251 ( Kab i ) S 2251 ( K ab i ) (EL ISA /MabCo) (EL ISA /N IH )

For s t a t i s t i c a l a n a l y s i s the p a i r e d t - t e s t c ompa r i n g the mea ­su remen t s be f o re a nd a f t e r r e t r a n s f u s i o n was a p p l i e d . A p - v a l u e

^ 0,05 was c on s i d e r e d to be s i g n i f i c a n t . Resu l t s a r e g i v e n as mean + SD.

RESULTS The mean amoun t of the f i r s t ches t b l ood r e t r a n s f us i on was 325

+_ 77 cc ( r a n g i n g f r om 250 - 520 c c ) . The mean hemog l ob i n con ten t of the ches t d r a i n a g e was 69 g / l , t he mean hema t o c r i t 21 % ( r a n g i n g f r om 13 - 29 %) , the p r o t e i n con ten t 40 .1 _+ 5 .4 g / l .

F i g u r e 1 shows the cou rse of p l a sm i nogen a n d a n t i p l a s m i n a c t i ­v i t y , the do t t e d l i n e r e p r e s e n t i n g the v a l u e c o r r e c t e d f o r p r e o p e r a ­t i v e p r o t e i n . When u s i n g t h i s c o r r e c t i o n f o r h emod i l u t i o n no s t a t i s t i c a l l y s i g n i f i c a n t i n f l u e n c e of CPB on these p a r ame t e r s was f o u n d . Howeve r , a n t i p l a s m i n a c t i v i t y i n the d r a i n a g e f l u i d was s i g n i f i c a n t l y r e d u c e d . No d i f f e r e n c e of a c t i v i t y i n the p a t i e n t s ' b l ood c o u l d be demons t r a t e d be fo re a nd a f t e r r e t r a n s f u s i o n . The s u r p r i s i n g f a c t t h a t no r e d u c t i o n of p l a sm i n ogen a c t i v i t y in the ches t d r a i n a g e was measu red m i gh t be e x p l a i n e d by the i n f l u e n c e of f i b r i n a nd f i b r i n ( o g e n ) s p l i t p r o d u c t s on the measurement of p l a sm i nogen ( 6 ) .

% 130

120

110

100

90

80

70

60

40

30

F i g .

plasminogen

antiplasmin

corrected for

preop protein

drainage

preop end of before r e -ECC transfusion

20 ' after r e - * + SD transfusion

1 Course of p l a sm i nogen and a n t i p l a s m i n . The do t t ed l i n e r e p r e sen t s the v a l u e c o r ­r ec t ed f o r p r e o p e r a t i v e p r o t e i n .

406

ng/ml 5000

l a,b

4000

3000

2000

1000 drainage

a=p<0.01 vs preop.

b=p<0.01 vs bef. retr.

preop end of ECC

before r e -transfusion

20' after r e -transfusion

x + SEM

F i g . C r o s s - l i n k e d f i b r i n f r a g m e n t s . R e t r a n s -f u s i o n of of shed b l ood caused a s i g n i f i ­c an t (p ^ 0 ,01) i n c r ea se of the c o n c en ­t r a t i o n in p a t i e n t s ' c i r c u l a t i o n compa red to v a l u e s be fo re r e t r a n s f u s i o n .

The c r o s s - l i n k e d f i b r i n f r a g m e n t s ( F i g u r e 2) showed a w i d e v a ­r i a t i o n a f t e r CPB. The c o n c e n t r a t i o n dec reased a f t e r o p e r a t i o n , bu t de ve l oped a t r emendous i n c r ease up to 5,000 n g / c c i n the d r a i n a g e f l u i d . The r e t r a n s f us i on caused a s i g n i f i c a n t i n c r ease of c r o s s - l i n k e d f i b r i n f r a g m e n t s i n p a t i e n t s ' b l o o d .

pM/ml

droinoge n.s. VS before retrons.

preop end of before r e - 20' after r e - x ± SD

ECC transfusion transfusion

F i g . 3 Bß-related p e p t i d e s . Note the i n t e r r u p t i o n of the Y - a x i s . Desp i t e the h i g h c o n c en ­t r a t i o n in the d r a i n a g e b l ood t he r e was - in c o n t r a s t to t he c r o s s - l i n k e d f i b r i n f r a gmen t s - no s i g n i f i c a n t i n c r ease i n p a t i e n t s ' c i r c u l a t i o n a f t e r r e t r a n s f u s i o n .

407

The Bß-related pep t i d e s ( F i g u r e 3) showed a c ompa r a b l e cou rse d u r i n g CPB w i t h an i n c r ease of t he 1-42 f r a gmen t s w h i c h was not s i g n i f i c a n t . In the ches t t ube d r a i n a g e a more p r onounced i n c r ease of Bß 15-42 p ep t i d e s was f o u n d . Howeve r , t he r e were no d i f f e r e n ce s of Bß-related pep t i d e s i n p a t i e n t s ' b l ood be fo re a n d a f t e r r e t r a n s -f u s i o n .

The PMN E-a^PI comp lex ( F i g u r e 4) showed a t e n f o l d i n c r ease in p a t i e n t s ' b l ood a f t e r CPB, b u t t he r e was an e x t r a o r d i n a r y i n ­c rease in t he d r a i n a g e f l u i d w i t h a mean c o n c e n t r a t i o n of 13,000 n g / c c - more t h a n h u n d r e d f o l d t he p r e o p e r a t i v e v a l u e . Re t r a n s f us ion a l so caused a s i g n i f i c a n t i n c r ease of the E-a^PI comp lex i n p a t i e n t s ' b l o o d . These v a l u e s were s i g n i f i c a n t l y h i g h e r t h a n those o b t a i n e d p r i o r to r e t r a n s f us i on .

No s i g n i f i c a n t c o r r e l a t i o n be tween e i t h e r b l ood loss o r t ime l ag between o p e r a t i o n a nd r e t r a n s f us ion a nd f i b r i n o l y s i s , r e p r e s en t e d by the a n t i p l a s m i n a c t i v i t y , c o u l d be f o u nd i n the shed b l o o d . A f t e r r e -t r a n s f u s i o n b l ood loss d i d not exceed the a v e r a g e of no rma l r a n g e .

before r e -transfusion

20' after r e -transfusion

x + SD

F i g . PMN E-a^PI c omp l e x , a nd d i f f e r e n t s ca l e of

Note the i n t e r r u p t i o n the Y - a x i s . The re

was a t remendous i n c r ease in the shed b l o o d . Re t r ans f u s i on of shed b l ood caused a s i g n i f i c a n t l y i n c r e a sed leve l of the comp lex i n p a t i e n t s ' b l o o d .

DISCUSSION Shed m e d i a s t i n a l b l ood is not c ompa r a b l e to au t o l ogous o r ho ­

mologous (7) b l ood i n r e g a r d to hemog l ob i n c o n t e n t , h ema t o c r i t o r c o a g u l a t i o n p r o p e r t i e s . However , in f o rme r i n v e s t i g a t i o n s ( 8 ) , com­p a r i n g two g r o u p s of p a t i e n t s w i t h a nd w i t h o u t r e t r a n s f u s i on of shed med i a s t i n a l b l o o d , no d i f f e r e n c e i n the p o s t o p e r a t i v e b l ood loss c ou l d be o b s e r v e d .

408

In the p r esen t s t u d y an a c t i v a t i o n of the f i b r i n o l y t i c sys tem i n shed m e d i a s t i n a l b l ood was f o u n d . The m a g n i t u d e of f i b r i n o l y t i c a c t i ­v a t i o n i n shed b l ood was c ompa r a b l e to t h a t measu red d u r i n g CPB. T h i s a c t i v a t i o n of the f i b r i n o l y t i c sys tem was i n d i c a t e d b y c ompa r ­a b l e dec reases of a n t i p l a s m i n a c t i v i t y i n p a t i e n t s ' b l ood a f t e r CPB as we l l as i n the shed b l o o d . Howeve r , r e t r a n s f u s i on of shed b l ood d i d not cause an a d d i t i o n a l dec rease of a n t i p l a s m i n i n the p a t i e n t s ' c i r c u l a t i o n .

Bß-related pep t i d e s a r e a most s e n s i t i v e m a r k e r of t he e ndo ­genous a c t i v a t i o n of the f i b r i n o l y t i c s y s t em , w h i c h can r e v e a l s u b ­c l i n i c a l a c t i v a t i o n of f i b r i n o l y s i s ( 9 ) . A p r onounced i n c r e a se of f i ­b r i n o g e n ) s p l i t p r o d u c t s was f o u n d i n the shed b l o o d . A f t e r r e t r a n s -f u s i o n o n l y the D -d ime rs showed a s i g n i f i c a n t i n c r e a se i n p a t i e n t s ' b l o o d , whe reas the c o n c e n t r a t i o n of Bß-related pep t i d e s showed no i n c r e a s e . T h i s can be e x p l a i n e d by the lower mo l e c u l a r w e i g h t a n d t h e r e f o r e f a s t e r c l e a r i n g r a t e of these p e p t i d e s .

The c o n s t e l l a t i o n of i n c r ea sed f i b r i n ( o g e n ) s p l i t p r o d u c t s a n d s t a b l e a n t i p l a s m i n a c t i v i t y a f t e r r e t r a n s f us i on can be i n t e r p r e t e d as a was t e e f fec t of r e t r a n s f u s i o n . Tha t means , the enhanced f i b r i n o ­l y t i c o r p r o t e o l y t i c a c t i v i t y w i t h i n the shed b l ood is i n h i b i t e d by the p h y s i o l o g i c i n h i b i t o r s . Some of these comp lexes a nd the s p l i t p r o d u c t s o b v i o u s l y r ema i n i n t he c i r c u l a t i o n a f t e r r e t r a n s f us i on w i t h o u t i n d u c i n g f i b r i n o l y t i c a c t i v i t y w i t h i n the p a t i e n t .

In t h i s s t u d y the e l e v a t e d l e ve l s of comp lexed e l a s t a s e a f t e r CPB c o r r e s pond to the r e s u l t s of o t he r a u t h o r s (10) i n d i c a t i n g an a c t i v a t i o n o r des i n t e g r a t i o n of PMN g r a n u l o c y t e s due to CPB. The e x t r a o r d i n a r y i n c r ease of E-a^PI i n the shed b l ood may be e x p l a i n e d by d e s i n t e g r a t i o n of g r a n u l o c y t e s caused by mechan i c a l a l t e r a t i o n due to p e r i c a r d i a l a n d / o r p l e u r a l movement i n the t h o r a x a n d i n the ches t t u b i n g .

Which c on c l u s i o n s can be d r a w n f r om the r e s u l t s of t h i s s t u d y?

The f i b r i n o l y t i c a nd un s p e c i f i c p r o t e o l y t i c a c t i v i t y i n the d r a i n a g e f l u i d does not i n duce f i b r i n o l y t i c a c t i v a t i o n a f t e r r e t r a n s -f u s i o n . The i n c r ea sed leve l of f i b r i n ( o g e n ) s p l i t p r o d u c t s seems not to be c l i n i c a l l y r e l e v a n t , a t l eas t not w i t h the amoun t s h a v i n g been r e t r a n s f u s e d . I t can be con c l u ded t h a t r e t r a n s f u s i on of shed med i a ­s t i n a l b l ood is a save method of au t o l o gous vo lume s u b s t i t u t i o n i n the e a r l y p o s t o p e r a t i v e p e r i o d . However , i t has to be t a k e n i n t o a ccoun t t h a t r e t r a n s f us i on of g r e a t e r v o l umes , of shed b l ood may cause a c o n s i d e r a b l e impa i rmen t of c o a g u l a t i o n a n d / o r the f i b r i n o ­l y t i c s y s t em . T h i s d e t e r i o r a t i o n m i gh t be i n du ced by the i n f u s i o n of f i b r i n ( o g e n ) s p l i t p r o d u c t s a n d / o r t he re l ease of t he con ten t of des i n t e g r a t e d g r a n u l o c y t e s . In t h i s respec t t he pos s i b l e r o l e of p h a r m a c o l o g i c a l i n t e r v e n t i o n by p ro t ease i n h i b i t i o n s h o u l d be ob jec t of f u r t h e r i n v e s t i g a t i o n s .

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