Table Of ContentLiver, Nutrition,
and Bi Ie Acids
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o
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Series A: Life SCiences
Liver, Nutrition,
and Bile Acids
Edited by
G. Galli
Institute of Pharmacology and Pharmacognosy
Milan, Italy
and
E. Bosisio
Institute of Pharmacology and Pharmacognosy
Milan, Italy
Plenum Press
New York and London
Published in cooperation with NATO Scientific Affairs Division
Proceedings of a NATO Advanced Study Institute on
Liver, Nutrition, and Bile Acids,
held September 25-0ctober 5, 1983,
in Maratea, Italy
Library of Congress Cataloging in Publication Data
NATO Advanced Study Institute on Liver, Nutrition, and Bile Acids (1983:
Maratea, Italy)
Liver, nutrition, and bile acids.
(NATO ASI series. Series A, Life sciences; v. 90)
"Proceedings of a NATO Advanced Study Institute on Liver, Nutrition, and Bile
Acids, held September 25-0ctober 5, 1983, in Maratea, Italy"-T.p. verso.
"Published in cooperation with NATO Scientific Affairs Division."
Includes bibliographies and index.
1. Liver-Diseases-Congresses. 2. Biliary tract-Diseases-Congresses. 3.
Gallstones-Congresses. 4. Bile acids-Congresses. 5. Cholesterol-Metab
olism-Congresses. 6. Nutrition-Congresses. I. Galli, Giovanni. II. Bosisio, E.
(Enrica) III. North Atlantic Treaty Organization. Scientific Affairs Division. IV. Ti
tle. V. Series. [DNLM: 1. Bile Acids and Salts-physiology-congresses. 2.
Liver-physiology-congresses. 3. Liver-physiopathology-congresses. 4. Nu
trition-congresses. WI 703 N279L 1983·)
RC845.N37 1986 616.3'6 85-9457
ISBN 978-1-4615-9429-1 ISBN 978-1-4615-9427-7 (e8ook)
DOl 10.1007/978-1-4615-9427-7
© 1985 Plenum Press, New York
Softcover reprint of the hardcover 1s t edition 1985
A Division of Plenum Publishing Corporation
233 Spring Street, New York, N.Y. 10013
All rights reserved. No part of this book may be reproduced, stored in a retrieval system,
or transmitted, in any form or by any means, electronic, mechanical, photocopying,
microfilming, recording, or otherwise, without written permission from the Publisher
PREFACE
This volume is a collection of the lectures and commu
nications presented at the Nato Advanced Study Institute on
"Liver, Nutrition and Bile Acids" held in Maratea, Italy,
September 1983.
The research areas covered by the Meeting are very wide,
having to do with the liver and biliary pathophysiology,
nutrition, gallstone disease and atherosclerosis prevention.
Scientists from the major clinical and basic disciplines
provided an update overview of the complex mechanisms regula
ting the biosynthesis and catabolism of cholesterol and the
formation and metabolism of bile acids.
The NATO ASI has given the opportunity for thorough
general discussions on problems of growing interest in
medical sciences in the spirit of the NATO Meetings. This
wealth of information is reflected in the present collection.
We are grateful to all the contributors for the pre
sentation of the results of their research.
A particular thanks is addressed to Mrs. Brita Rolander
Chilo of the Nutrition Foundation of Italy for her invaluable
advice and for the organization of the Institute as well as
for the editorial work.
Giovanni Gall,i
Enrica Bosisio
v
CONTENTS
Coordinate Regulation of the Three Enzymes in
Cholesterol Biosynthesis • •••••• 1
Ajit Sanghvi
Microsomal Cholesterol 7a-Hydroxylase Phosphatases. 15
Ajit Sanghvi, Warren Diven, and Joyce Sweeney
Bile Acids and Cholesterol Absorption • 19
George V. Vahouny and Linda L. Gallo
Regulation of Cholesterol Absorption in Humans. • • • • • • 35
Maurizio Ponz de Leon
Biochemical Methods for Serum Bile Acid Analysis. • • • • • 65
A. Roda, S. Girotti, P. Filippetti,
A. Piacentini, and P. Simoni
Diagnostic Usefulness of Serum Bile Acid
Determination in Liver Diseases • 81
L. Barbara in collaboration with D. Festi,
A. Roda, A.M. Morselli Labate,
B. Grigolo, R. Frabboni, and S. Vicari
Quantitation and Metabolism of Bile Acids in
Hyperlipoproteinemias 87
Tatu A. Miettinen
Effects of Lipid Lowering Diets and of Bile Acid
Sequestrants on Plasma Lipoproteins and
Biliary Metabolism. • • • • • • • • 101
Cesare R. Sirtori
Faecal Bile Acids in Health and Disease • • • • • • • • • • 113
Michael H. Thompson
Bile Acids and Intestinal Pathophysiology 131
George V. Vahouny and Marie M. Cassidy
The Therapy of Cholesterol Gallstones
with Bile Acids •••••••••••• 143
E. Roda in collaboration with F. Bazzoli,
G. Mazzella, M. Malavolti, and F. Luzza
Dissolution of Cholesterol Gallstones in Bile • 151
Gianfranco Salvioli
Bile Acids, Fiber and Cancer. 161
David Kritchevsky
Intestinal Operations and Bile Acids: Their
Effects on Adaptation and Carcinogenesis
in the Gut. . . . . . .. . . . . . . . 171
J.B. Bristol, J.B. Rainey, and R.C.N. Williamson
Similarities and Differences between Primary
Biliary Cirrhosis and Primary
Sclerosing Cholangitis •••••••• 185
D.H. Van Thiel, S. Lasky, J.S. Gavaler,
T. Whiteside, and R.R. Schade
Liver Transplantation: After 20 years of
Experience the Procedure Has Come of Age. • • • • • 195
David H. Van Thiel, Robert R. Schade, and
Thomas E. Starzl
Effects of Inhibitors of Cholesterol Biosynthesis
in Isolated Rat Hepatocytes • • • • • 205
Giuliana Cighetti, Giovanni Galli,
Ri ta Paroni, and ~1arzia Galli Kienle
Bile Acid Synthesis by Cultured Hepatocytes
Regulation by Cholesterol Availability. 209
Haya Herscovitz and Alisa Tietz
Maturation of Bile Acid Uptake in Freshly Isolated
Hepatocytes from Fetal and Newborn
Rabbi ts . . . . . . . . . . . . . 219
M.A. Evans, R. Bhat, M.S. Bernstein,
R.J. Anderson, and D. Vidyasagar
Sulfated Lithocholic Acid Conjugates and Cholestasis:
Clinical Implications and Protecting Factors. 225
Folkert Kuipers, Charles M.A. Bijleveld,
C.M. Frank Kneepkens, John Fernandes, and
Roel J. Vonk
viii
Abnormal Fecal Bile Acid Excretion in Cystic
Fibrosis: Physiopathological Mechanism
and Clinical Implications • • 235
Carla Colombo and Annamaria Giunta
Effect of Chenodeoxycholic and Ursodeoxycholic
Acid Treatment on Intestinal Microflora
of Patients with Cholesterol Gallstones 241
Annamaria Ferrari, Novella Pacini, and
Enrica Canzi
The Influence of a Lactovegetarian, a Vegan
and a Mixed Diet on Colonic Function
of Man: Preliminary Results •• 249
A. van Faassen, W. van Dokkum,
N.A. Pikaar, and R.J.J. Hermus
INDEX • 253
COORDINATE REGULATION OF THE THREE ENYMES IN
CHOLESTEROL BIOSYNTHESIS
Ajit Sanghvi
Department of Pathology
University of Pittsburgh School of Medicine
Pittsburgh, PA 15261
It has been observed by several investigators, including Kandutsch
et al. (1), and Spector et al. (2), Myant and Mitropoulos (3), and others
that it is crucial that the concentration of unesterified, or free
cholesterol, within the cell be regulated between very narrow limits. It is
known that the unesterified cholesterol which is present in cell
membranes functions to modulate fluidity of the phospholipid bilayer. A
change in the membrane content of free cholesterol, therefore, may
affect fluidity and thereby bring about changes in permeability, which in
turn may affect the properties of membrane-bound enzymes and transport
systems. It is for this reason that an excess of free cholesterol within the
cell may be considered to be toxic to vital cell functions. The storage of
cholesterol in the form of cholesterol esters, and in the case of liver cell,
the utilization of cholesterol via the bile acid synthesis, provides an
elegant means for the control of the amount of unesterified cholesterol
present within the cell.
In this article, I will review the evidence which is consistent with
the concept that the activities of three enzymes--namely, HMG-CoA
reductase, acyl-CoA cholesterol acyl transferase or ACAT, and cholesterol
7 a-hydroxylase--which are intimately concerned with the metabolism of
cholesterol are regulated in a coordinate manner by a process of
phosphorylation/dephosphorylation of the enzymes. Based on this
evidence, Dr. Scallen and our group have formulated the proposal that
such regulation provides a short-term mechanism for intracellular
unesterified cholesterol homeostasis.
As shown in Fig. 1, the reduction of HMG-CoA to mevalonic acid,
which is the commitment step in cholesterol biosynthesis, is catalyzed by
HMG-CoA reductase. The first indication that the activity of this
enzyme may be regulated by phosphorylation/dephosphorylation was
G. Galli et al. (eds.), Liver, Nutrition, and Bile Acids
© Plenum Press, New York 1985
Acetyl-CoA Cholesterol
t t
Reductase
HMG-CoA Mevalonate
~
2 NAOPH
Fig. 1 General outline of formation of cholesterol from acetyl-CoA.
provided by the studies of Beg et al. in 1973 (4). These studies were
prompted by an earlier observation of Bricker and Levey in 1972 (5) that
cholesterol synthesis was lowered by cAMP. Beg and Gibson showed that
reductase activity in rat liver microsomes was severely diminished when
they were preincubated in the presence of AT P and magnesium chloride.
They also observed that the diminished reductase activity, as a result of
ATP magnesi!lm treatment, was restored towards almost normal levels if
these treated microsomes were centrifuged and resuspended in a buffer
containing a fraction of the cytosol. In turn, this reactivation was
blocked by 100 mM sodium fluoride. This was evidence suggesting a
reversible modulation of reductase activity possibly brought about by the
action of a protein kinase and a phosphoprotein phosphatase. Further
studies by Begg, Stonik and Brewer (6) showed that there was progressive
inactivation of 'reductase activity with two different concentrations of
ATP and reactivation of the enzyme in the presence of EOTA and a
phosphotase. These observations by Beg and others were further
confirmed by Brown et al. (7) when they showed a progressive increase in
HMG-CoA reductase activity with increasing amounts of alkaline
phosphatase in the incubations. These data also indicated a greater
activation of reductase activity if the time of preincubation with alkaline
phosphatase was increased from 30 minutes to 60 minutes. Their own
studies led Brown and Goldstein to speculate that in the live animal,
HMG-CoA reductase was almost fully active--that is, dephosphorylated-
to the extent of 85-90%, and that the inactive--that is, phosphorylated-
form of the enzyme observed in vitro may simply represent an artifact at
the time the animals are being killed and the liver is being removed or a
result of procedural manipulations during microsome preparation. They
concluded, correctly, that the long-term alteration in cholesterol
synthesis does not involve changes in the state of phosphorylation of
reductase, but rather it requires changes in the total amount of enzyme
protein.
In contrast, recent studies in Dr. Scallen's laboratory (8, 9) and from
Beg and Brewer (10) support the view that short-term changes in
reductase activity in the intact animal most likely involve a change in the
2
