Advances in Metabolic Disorders: Somatomedins and Some Other Growth Factors Proceedings of the Twenty-Eighth Nobel Symposium Held at Hässelby, Sweden, September 4–7, 1974

Advances in Metabolic disorders

Somatomedins and Some Other Growth Factors

Rolf Luft

Department of Endocrinology and Metabolism, Karolinska Hospital, Stockholm, Sweden

Kerstin Hall

Deportment of Endocrinology and Metabolism, Karolinska Hospital, Stockholm, Sweden

ISSN  0065-2903

Volume 8 • Number suppl (C) • 1975

Table of Contents

Cover image

Title page

Advances in Metabolic Disorders

Copyright page

Contributors

Contents of Previous Volumes

Volume 1

Volume 2

Volume 3

Volume 4

Volume 5

Volume 6

Volume 7

Introductory Speech

Publisher Summary

Regulation of Cellular Growth

Publisher Summary

I Introduction

II How Do Growth-Regulating Factors Work?

III Cyclic AMP and Cell Growth

IV How Does Insulin Act to Lower Cyclic AMP Levels?

V How Might Cyclic AMP and Steroids Work to Control Cell Growth?

VI Cyclic GMP

VII Insulin Receptors, Cyclic AMP, and Cyclic GMP

VIII Malignant Transformation

IX Putting It All Together

Somatomedins

Publisher Summary

I Introduction

II Bioassay of Somatomedin

III Somatomedin in Man

IV Purification of Somatomedin

V Mechanism of Production of Somatomedin

VI Biological Action of Purified Somatomedins in Vitro

VII Conclusions and Discussion

Acknowledgment

Isolation and Chemistry of Human Somatomedins A and B

Publisher Summary

I Introduction

II Procedure for Isolation of Somatomedins

III Somatomedin A

IV Somatomedin B

V Discussion

Acknowledgment

The Measurement of Somatomedin A by Radioreceptor Assay

Publisher Summary

I Introduction

II Radioreceptor Assay Methodology

III Specificity of Somatomedin A Radioreceptor Assay

IV Somatomedin A in Human Serum

V Conclusions

Acknowledgment

Radioimmunoassay of Somatomedin B

Publisher Summary

I Introduction

II Radioimmunoassay Methodology

III Somatomedin B in Plasma

IV State of Somatomedin B in Plasma and Plasma Fractions

V Summary and Conclusions

Acknowledgments

The Response of Cultured Human Normal Glial Cells to Growth Factors

Publisher Summary

I Introduction

II The Glial Cell System

III The Glial Cell Bioassay of Growth-Promoting Substances

IV Somatomedins

V Fibroblast Growth Factor

VI Growth Factors Released by Cultured Cells

VII Platelets and Serum Factors

VIII Discussion

IX Summary

Note Added in Proof

Acknowledgments

Somatomedin A and B: Demonstration of Two Different Somatomedinlike Components in Human Plasma

Publisher Summary

I Introduction

II 35SO4-Incorporation into Cell Cultures

III Stimulation of Sulfate Incorporation

IV Separation of Two Somatomedinlike Polypeptides from Human Plasma

V Summary

Acknowledgment

Preliminary Studies of Somatomedin in Vitro and in Vivo in Rats

Publisher Summary

I Introduction

II Materials and Methods

III Effect of Somatomedin in Vitro

IV Effect of Somatomedin in the Rat

Explorations of the Insulinlike and Growth-Promoting Properties of Somatomedin by Membrane Receptor Assays

Publisher Summary

I Introduction

II Studies of Insulinlike Properties of Somatomedin

III Comparisons between Insulinlike Activity (ILA) and Somatomedin Activity in Human Plasma Fractions

IV Studies with Purified Somatomedin C

V Measurement of Plasma Somatomedin Levels with the Specific Somatomedin C Receptor Assay

VI Conclusion

VII Summary

Acknowledgment

Somatomedin Generation by Perfused Livers

Publisher Summary

I Introduction

II Methods

III Results

IV Discussion

Acknowledgments

Somatomedin Levels in Human Beings

Publisher Summary

I Introduction

II Somatomedin Levels in Man

III Relation of Growth Hormone to Somatomedin

Acknowledgments

Further Observations on Plasma Somatomedin Activity in Children

Publisher Summary

I Introduction

II Plasma Somatomedin in a Nonselect Population of Children of Similar Age

III Further Observations on a Somatomedin Inhibitor in Malnutrition

Conclusions

IV Plasma Somatomedin Activity in Children with Emotional-Deprivation Short Stature

Conclusions

V Summary

Acknowledgments

Interaction of Somatomedin and a Peptide Inhibitor in Serum of Hypophysectomized and Starved, Pituitary-Intact Rats

Publisher Summary

I Introduction

II Methods

III Results

IV Discussion

Purification Procedures for NSILA-S

Publisher Summary

I Large-Scale Purification of NSILA-S from Serum

II Partial Purification of NSILA-S from Individual Sera

Acknowledgments

Biological Properties of NSILA-S

Publisher Summary

I Insulinlike Effects of NSILA-S in Vitro

A Adipose Tissue

B Muscle

II Effects of NSILA-S in Vivo

III Growth Effects of NSILA-S

IV Membrane Receptors for NSILA-S

V Binding Protein for NSILA-S in Plasma

Acknowledgment

NSILA-S, Physiologic and Pharmacologic Significance as an Insulinlike Hormone and as a Growth-Promoting Hormone

Publisher Summary

I Introduction

II NSILA-S in Normal Subjects, Acromegalics, and Pituitary Dwarfs

III Multifactorial Endocrine Regulation of NSILA-S

Acknowledgment

In Vitro Effects of Growth Hormone on Cyclic AMP Metabolism in the Isolated Rat Diaphragm

Publisher Summary

I Introduction

II Materials and Methods

III Results

IV Discussion

V Summary

Acknowledgments

Interaction of Epidermal Growth Factor (EGF) with Cultured Fibroblasts

Publisher Summary

I Introduction

II Chemical and Physical Properties of EGF

III Effects of EGF in Vivo and in Organ Culture Systems

IV Stimulation of Fibroblast Proliferation by EGF

V Interactions of 125I-EGF with Human Fibroblasts

VI Conclusions

Acknowledgments

Nerve Growth Factor: Structure and Mechanism of Action

Publisher Summary

I Introduction

II Structure of Nerve Growth Factor (NGF)

III Biological Function of NGF

IV Mechanism of Action of NGF

V Conclusions and Perspectives

Fibroblast Growth Factor: Its Localisation, Purification, Mode of Action, and Physiological Significance

Publisher Summary

I Introduction

II Localisation and Purification of FGF

III Control of DNA Synthesis in 3T3 Cells by FGF and Glucocorticoids

IV Stimulation of Division and Morphological Changes in Sparse and Confluent 3T3 Cell Populations by FGF, Dexamethasone, and Insulin

V Mitogenic Effect of FGF on Other Cell Lines

VI Action of FGF upon Cell Transformation Mutants in Culture: Loss of Growth-Initiating Ability at Permissive Temperatures

VII Induction of the Pleiotypic Responses by FGF in Quiescent Cultures of BALB/c 3T3 Cells

VIII Changes in Intracellular Cyclic Nucleotides Induced by FGF in Quiescent Cultures of BALB/c 3T3 Cells

IX In Vivo Effect of FGF

Acknowledgments

Growth Regulation in Cultures of Embryonic Rat Fibroblasts by the Serum Factors S1 and S2

Publisher Summary

I Introduction

II Isolation and Properties of the Serum Factors S1 and S2

III Effects of S1 and S2 on Embryonic Rat Fibroblasts in Tissue Culture

IV Effects of S1 and S2 on Permanent Rat Cell Lines

V Concluding Remarks

Acknowledgments

The Endocrine Role of the Thymus and Its Hormone, Thymosin, in the Regulation of the Growth and Maturation of Host Immunological Competence

Publisher Summary

I Introduction

II Background of Present Concepts of the Nature and Role of the Thymus Gland and Lymphoid Cell Growth and Maturation in Host Immunological Competence

III Chemical Nature of Thymosin and Other Recently Characterized Thymic Factors

IV Biological Roles of Thymosin and Other Thymic Factors as Revealed by in Vitro and in Vivo Studies

V Possible Mechanisms of Action of Thymic Humoral Factors

VI Suggested Interrelationships of the Endocrine Roles of the Thymus and of Growth-Promoting Factors

VII Potential Practical Applications of Thymic Humoral Factors

VIII Summary

Acknowledgments

Cyclic AMP and the Malignant Transformation of Cells

Publisher Summary

I Introduction

II cAMP Levels in Normal and Transformed Cells

III cAMP Treatment of Cells

IV Enzymes of cAMP Metabolism

V NRK Cells

VI KNRK Cells

VII Summary

Postsession Discussion of Papers*

Publisher Summary

I Purification, Structure, and Synthesis of Growth Factors

II Quantitative Measurements of Growth Factors

III In Vitro Actions of Growth Factors

IV Biological and Physiological Actions of Growth Factors

V Clinical Aspects of Growth Factors

General Discussion

I Summary by Hogue-Angeletti

II Summary by Cohen

III Summary by White

IV Summary by Gospodarowicz

V Summary by Van Wyk

Concluding Remarks

Publisher Summary

Subject Index

Advances in Metabolic Disorders

Edited by

Rachmiel Levine

City of Hope Medical Center

Duarte, California

Rolf Luft

Department of Endocrinology and Metabolism

Karolinska Hospital

Stockholm, Sweden

Supplementary Volumes

1. Early Diabetes

Edited by Rafael A. Camerini-Dávalos and Harold S. Cole

2. Vascular and Neurological Changes in Early Diabetes

Edited by Rafael A. Camerini-Dávalos and Harold S. Cole

Copyright page

COPYRIGHT © 1975, by Academic Press, Inc.

ALL RIGHTS RESERVED.

NO PART OF THIS PUBLICATION MAY BE REPRODUCED OR TRANSMITTED IN ANY FORM OR BY ANY MEANS, ELECTRONIC OR MECHANICAL, INCLUDING PHOTOCOPY, RECORDING, OR ANY INFORMATION STORAGE AND RETRIEVAL SYSTEM, WITHOUT PERMISSION IN WRITING FROM THE PUBLISHER.

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United Kingdom Edition published by

ACADEMIC PRESS, INC.(LONDON) LTD.

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LIBRARY OF CONGRESS CATALOG CARD NUMBER: 64-14568

ISBN 0-12-027308-X

PRINTED IN THE UNITED STATES OF AMERICA

Contributors

J.B. Baseman,     Department of Bacteriology, University of North Carolina, School of Medicine, Chapel Hill, North Carolina (127)

K. Benirschke,     Department of Obstetrics and Gynecology, University of California, San Diego, California (301)

Ralph A. Bradshaw,     Department of Biological Chemistry, Division of Biology and Biomedical Sciences, Washington University, St. Louis, Missouri (285)

Graham Carpenter,     Department of Biochemistry, Vanderbilt University, School of Medicine, Nashville, Tennessee (265)

D.R. Clemmons,     Department of Pediatrics, University of North Carolina, School of Medicine, Chapel Hill, North Carolina (127)

Stanley Cohen,     Department of Biochemistry, Vanderbilt University, School of Medicine, Nashville, Tennessee (265)

William H. Daughaday,     Metabolism Division, Department of Medicine, Washington University School of Medicine, St. Louis, Missouri (151, 159)

E. Eigenmann,     Metabolic Unit, Department of Medicine, University of Zürich, Zürich, Switzerland (237)

Werner Frank,     Max-Planck-Institut für Virusforschung, Abteilung für Physikalische Biologie, Tübingen, West Germany (337)

William A. Frazier,     Department of Psychiatry, University of California, San Diego, School of Medicine, La Jolla, California (285)

E.R. Froesch,     Metabolic Unit, Department of Medicine, University of Zürich, Kantonsspital, Zürich, Switzerland (203, 211, 237)

Linda Fryklund,     AB KABI, The Recip Polypeptide Laboratory, Stockholm, Sweden (19, 47, 61)

Lynn P. Gimpel,     Department of Physiology, Emory University, Atlanta, Georgia (249)

Allan L. Goldstein,     Division of Biochemistry, Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, Texas (359)

Denis Gospodarowicz,     The Salk Institute for Biological Studies, San Diego, California (301)

Johanna Grimm,     Max-Planck-Institut für Virusforschung, Abteilung für Physikalische Biologie, Tübingen, West Germany (337)

Kerstin Hall,     Department of Endocrinology and Metabolism, Karolinska Hospital, Stockholm, Sweden (19, 47, 61, 73)

R. Heimann,     Departments of Medicine and Biochemistry, University of Zürich, Zürich, Switzerland (203, 237)

U. Heinrich,     Department of Pediatrics, Erasmus University, Rotterdam, The Netherlands (171)

A.C. Herington*,     Metabolism Division, Department of Medicine, Washington University School of Medicine, St. Louis, Missouri (151)

R.L. Hintz,     Department of Pediatrics, University of Connecticut, School of Medicine, Farmington, Connecticut (127)

Rainer Hoffman,     Max-Planck-Institut für Virusforschung, Abteilung für Physikalische Biologie, Tübingen, West Germany (337)

Ruth A. Hogue-Angeletti,     The Institute for Cancer Research, Philadelphia, Pennsylvania (285)

R.E. Humbel,     Departments of Medicine and Biochemistry, University of Zürich, Zürich, Switzerland (203)

Olle Isaksson,     Department of Physiology, University of Göteborg, Göteborg, Sweden (249)

U. Kaufmann,     Metabolic Unit, Department of Medicine, University of Zürich, Zürich, Switzerland (211)

Jack L. Kostyo,     Department of Physiology, Emory University, Atlanta, Georgia (249)

Kenneth J. Lembach,     Department of Biochemistry, Vanderbilt University, School of Medicine, Nashville, Tennessee (265)

Jon Lindstrom,     The Salk Institute for Biological Studies, San Diego, California (301)

Rolf Luft,     Department of Endocrinology, Karolinska Hospital, Stockholm, Sweden (3, 73, 431)

M. Mäder,     Metabolic Unit, Department of Medicine, University of Zürich, Zürich, Switzerland (211)

R.N. Marshall*,     Department of Pediatrics, University of North Carolina, School of Medicine, Chapel Hill, North Carolina (127)

C. Meuli,     Metabolic Unit, Department of Medicine, University of Zürich, Zürich, Switzerland (211)

B. Morell,     Metabolic Unit, Department of Medicine, University of Zürich, Zürich, Switzerland (211)

Ira Pastan,     Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland (7, 377)

L.S. Phillips†,     Metabolism Division, Department of Medicine, Washington University School of Medicine, St. Louis, Missouri (151)

Hans-Jurgen Ristow,     Max-Planck-Institut für Virusforschung, Abteilung für Physikalische Biologie, Tübingen, West Germany (337)

W.J. Ritschard,     Diagnostic Research Department, F. Hoffmann-La Roche & Co., Basel, Switzerland (203)

Phillip Rudland,     Imperial Cancer Research Fund, London, England (301)

William D. Salmon, Jr.,     Veterans Administration Hospital and Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee (183)

U. Schlumpf,     Departments of Medicine and Biochemistry, University of Zürich, Zürich, Switzerland (203, 237)

Hans Sievertsson,     AB KAB1, The Recip Polypeptide Laboratory, Stockholm, Sweden (19, 47, 61)

Kazue Takano,     Department of Endocrinology and Metabolism, Karolinska Hospital, Stockholm, Sweden (19, 61)

L.E. Underwood,     Department of Pediatrics, University of North Carolina, School of Medicine, Chapel Hill, North Carolina (127)

Knut Uthne,     AB KABI, The Recip Polypeptide Laboratory, Stockholm, Sweden (47, 101, 115)

Sylvia Van Buul,     Department of Pediatrics, Erasmus University, Rotterdam, The Netherlands (171)

J.L. Van Den Brande,     Department of Pediatrics, Erasmus University, Rotterdam, The Netherlands (171)

F. Van Roon,     The Bergweg Hospital, Rotterdam, The Netherlands (171)

A.C. Van Steirtegem

The Free University of Brussels, Brussels, Belgium

The Medical Mission of CEMUBAC to IRSAC, Kuvu, Zaire (171)

J.J. Van Wyk,     Department of Pediatrics, University of North Carolina, School of Medicine, Chapel Hill, North Carolina (127)

Josef Veser,     Max-Planck-Institut für Virusforschung, Abteiling für Physikalische Biologie, Tübingen, West Germany (337)

M. Waldvogel,     Metabolic Unit, Department of Medicine, University of Zürich, Zürich, Switzerland (211)

Åke Wasteson,     Institute of Medical and Physiological Chemistry, Uppsala University, Uppsala, Sweden (85, 101)

Bengt Westermark,     The Wallenberg Laboratory, Uppsala University, Uppsala, Sweden (47, 85, 101)

Abraham White

Syntex Research, Palo Alto, California

Department of Biochemistry, Stanford University School of Medicine, Stanford, California (359)

Rosalyn S. Yalow

Solomon A. Berson Research Laboratory, Veterans Administration Hospital, Bronx, New York

Department of Medicine, Mt. Sinai School of Medicine, The City University of New York, New York (73)

J. Zapf,     Departments of Medicine and Biochemistry, University of Zürich, Zürich, Switzerland (203, 211, 237)

T. Zurcher,     Department of Pediatrics, Erasmus University, Rotterdam, The Netherlands (171)

Numbers in parentheses indicate the pages on which the authors’ contributions begin.

*Present address: Medical Research Centre, Prince Henry’s Hospital, St. Kilda Road, Melbourne, Australia.

*Present address: Department of Pediatrics, University of Texas Medical School, Houston, Texas.

†Present address: Center for Endocrinology, Metabolism, and Nutrition, Northwestern University School of Medicine, Chicago, Illinois.

Contents of Previous Volumes

Volume 1

GLYCOGEN STORAGE DISEASE

H. G. Hers

THE PARATHYROIDS

G. D. Aubach and John T. Potts, Jr.

MITOCHONDRIAL RESPIRATORY CONTROLS: BIOCHEMICAL, PHYSIOLOGICAL, AND PATHOLOGICAL ASPECTS

Lars Ernster and Rolf Luft

OSTEOPOROSIS

B. E. C. Nordin

BASAL METABOLIC RATE AND THYROID HORMONES

J. R. Tata

INSULIN ANTAGONISTS AND INHIBITORS

J. Vallance-Owen

ALDOSTERONE: ITS BIOCHEMISTRY AND PHYSIOLOGY

John H. Laragh and William G. Kelley

FOLIC ACID DEFICIENCY IN MAN AND ITS INTERRELATIONSHIP WITH VITAMIN B12 METABOLISM

A. Leonard Luhby and Jack M. Cooperman

AUTHOR INDEX—SUBJECT INDEX

Volume 2

GOUT

James B. Wyngaarden

NITROGEN-RETAINING STEROIDS AND THEIR APPLICATION IN DISEASE

A. Querido and A. A. H. Kassenaar

MACROGLOBULINEMIA

Jan Waldenstrom

TESTING THE FUNCTIONAL CAPACITY OF THE TRYPTOPHAN-NIACIN PATHWAY IN MAN BY ANALYSIS OF URINARY METABOLITES

J. M. Price, R. R. Brown, and Norma Y ess

THE SYNDROME OF TESTICULAR FEMINIZATION

A. Louis Southren

AUTHOR INDEX—SUBJECT INDEX

Volume 3

KETOGENESIS AND ITS REGULATION

O. Wieland

SPONTANEOUS DIABETES AND/OR OBESITY IN LABORATORY RODENTS

Albert E. Renold

ETHANOL-INDUCED HYPOGLYCEMIA

Leonard L. Madison

HORMONAL SYNDROMES ASSOCIATED WITH NEOPLASIA

Mortimer B. Lipsett

METABOLIC DISORDERS OF THE OVARY

Bruno Lunenfeld, Aliza Eshkol, Vaclav Insler, and Zaki Kraiem

THE NATURE AND SIGNIFICANCE OF THE LONG-ACTING THYROID STIMULATOR

Joseph P. Kriss

THYROID AUTOANTIBODIES IN THYROID DISEASES

Noel R. Rose and Ernest Witebsky

ERYTHROPOIETIN

Clifford W. Gurney

AUTHOR INDEX—SUBJECT INDEX

Volume 4

INTESTINAL FACTORS IN THE REGULATION OF INSULIN SECRETION

Vincent Marks and Ellis Samols

ANTIDIURETIC HORMONE SYNTHESIS, RELEASE, AND ACTION UNDER NORMAL AND PATHOLOGICAL CIRCUMSTANCES

Niels A. Thorn

DISACCHARIDASE DEFICIENCY

David H. Alpers and Kurt J. Isselbacher

HORMONAL CONTROL OF FETAL DEVELOPMENT AND METABOLISM

Alfred Jost and Luc Picon

THE METABOLISM OF THE PLACENTA

Andrew J. Szabo and Richard D. Grimaldi

METABOLIC ASPECTS OF OBESITY

E. S. Gordon

THE USE OF LIQUID FORMULA DIETS IN METABOLIC STUDIES: 15 YEARS′ EXPERIENCE

E. H. Ahrens, Jr.

AUTHOR INDEX—SUBJECT INDEX

Volume 5

HYPOTHALAMIC CONTROL OF THE SECRETION OF ADENOHYPOPHYSIAL

HORMONES

Roger Guillemin

ADRENOCORTICAL CONTROL OF EPINEPHRINE SYNTHESIS IN HEALTH AND DISEASE

R. J. Wurtman and L. A. Pohorecky

THE RELATIONSHIP BETWEEN ANGIOTENSIN AND ALDOSTERONE

G. W. Boyd and W. S. Peart

THE METABOLIC INFLUENCE OF PROGESTINS

Richard L. Landau and James T. Poulos

THE THYMUS GLAND: EXPERIMENTAL AND CLINICAL STUDIES OF ITS ROLE IN THE DEVELOPMENT AND EXPRESSION OF IMMUNE FUNCTION

Allan L. Goldstein and Abraham White

CONTROL OF GLUCOSE METABOLISM IN THE HUMAN FETUS AND NEWBORN INFANT

Peter A. J. Adam

HUMAN ADIPOSE TISSUE DYNAMICS AND REGULATION

Per Björntorp and Jan Östman

CORONARY DISEASE AND CARBOHYDRATE AND FAT ABNORMALITIES

Michael V. Herman and Richard Gorlin

AUTHOR INDEX—SUBJECT INDEX

Volume 6

THE ROLE OF THE SKIN IN CARBOHYDRATE METABOLISM

John A. Johnson and Ramon M. Fusaro

ON THE TRANSMISSION OF ALLOXAN DIABETES AND OTHER DIABETOGENIC INFLUENCES

Martin G. Goldner and Gabriel Spergel

GLUCAGON AND DIABETES MELLITUS

Roger H. Unger

RECENT CONTRIBUTIONS TO THE STUDY OF DIABETIC ANGIOPATHY AND NEUROPATHY

Knud Lundbaek

DISORDERS DUE TO ENZYME DEFECTS IN THE RED BLOOD CELL

Ernest Beutler

THE PINEAL GLAND—A NEUROENDOCRINE TRANSDUCER

R. Collu and F. Fraschini

CALCITONIN IN METABOLIC DISORDERS

Asher Haymovits and John F. Rosen

AUTHOR INDEX—SUBJECT INDEX

Volume 7

GROWTH HORMONE AND SOMATOMEDIN

Kerstin Hall and Rolf Luft

NATRIURETIC HORMONES

Norman G. Levinsky

METHODOLOGICAL ASPECTS ON THE ESTIMATION OF GENETIC EFFECTS OF ENVIRONMENTAL AGENTS IN MAN

N. Ryman and J. Lindsten

METABOLIC ASPECTS OF DESERT ADAPTATION (MAN)

Shlomo Samueloff

Proceedings of a Symposium on Insulin, City of Hope Medical Center Duarte, California, February 13, 1972

PHILOSOPHY AND OUTLOOK

Charles H. Best

SYNTHESIS AND SECRETION OF INSULIN IN DYNAMIC PERFUSION SYSTEMS

G. M. Grodsky, H. Sando, S. Levin, J. Gerich, and J. Karam

STRUCTURE AND FUNCTION OF THE ENDOCRINE CELL TYPES OF THE ISLETS

Paul E. Lacy

MECHANISMS OF INSULIN ACTION

Rachmiel Levine

DIABETES MELLITUS⇔A DISEASE OF PANCREATIC AND EXTRAPANCREATIC ORIGIN

E. Cerasi and R. Luft

THE PATHOGENESIS OF PANCREATIC ISLET CELL HYPERPLASIA AND INSULIN INSENSITIVITY IN OBESITY

Richard J. Mahler

OBESITY AND DIABETES MELLITUS

E. F. Pfeiffer and H. Laube

SUBJECT INDEX

Introductory Speech

Rolf Luft*,     *Department of Endocrinology, Karolinska Hospital, Stockholm, Sweden

Publisher Summary

This chapter presents an introduction to growth hormone from the hypophysis. In the past half century, periodically there have been a series of major breakthroughs related to growth hormone: 1) the early observation that there was a growth-promoting factor in the pituitary from which the name arose, 2) the establishment of its diabetogenic action in animals and man, 3) the demonstration of the species specificity of its biological action, 4) the introduction of radioimmunoassay of growth hormone, which led to an appreciation of the acute responsiveness of growth hormone to a variety of stimuli, and 5) an appreciation that the growth-promoting properties of this hormone appear to be mediated by a peripherally produced factor that is growth-hormone dependent. The growth process may be considered to comprise all the metabolic steps in the cells involved in the transition from a resting to a dynamic state, termed positive pleiotypic responses: uridine uptake, RNA synthesis, polyribosome formation, protein synthesis, lipid synthesis, and glucose utilization. The chapter also presents a discussion on the identification and the role of factors that are growth hormone dependent or promote growth, or both. The somatomedins have been defined as factors that meet both the criteria.

This introduction is intended to establish the framework for Nobel Symposia in general, and this one in particular. Nobel Symposia are distinguishable from the usual congresses in that they are devoted more to the exchange of ideas and concepts than to presentation of data. By tradition there are certain prerequisites to be met in planning a Nobel Symposium: the area of research to be discussed should be of immediate interest; the state of the art in the particular area should have developed to a level that makes it appropriate and necessary to have mutual exchange among the scientists involved: the organizers should have special expertise in the field. We believe this Symposium meets these criteria.

The starting point for the development of the field we are here to discuss is the growth hormone from the hypophysis. In the past half century, periodically there have been a series of major breakthroughs related to growth hormone: the early observation that there was a growth-promoting factor in the pituitary from which the name arose; the establishment of its diabetogenic action in animals and man; the demonstration of the species specificity of its biological action; the introduction of radioimmunoassay of growth hormone, which led to an appreciation of the acute responsiveness of growth hormone to a variety of stimuli; and, last, an appreciation that the growth-promoting properties of this hormone appear to be mediated by a peripherally produced factor that is growth-hormone dependent. In this Symposium our efforts will be concentrated on the last observation.

The term growth hormone was employed originally because it was known that its action was to promote bodily growth—i.e., linear growth or increase in size. It has since become obvious that the hormone has a wide variety of actions. Thus, the growth process may be considered to comprise all the metabolic steps in the cells involved in the transition from a resting to a dynamic state, what we term positive pleiotypic responses: uridine uptake, RNA synthesis, polyribosome formation, protein synthesis, lipid synthesis, and glucose utilization.

Our Symposium is developed around the identification and role of factors that are growth hormone dependent or promote growth, or both. The somatomedins have been defined as factors that meet both criteria. Three such factors have been characterized—somatomedins A, B, and C—although it is not yet established whether A and C are essentially identical. There exist other growth factors, including the fibroblast growth factor, the nerve growth factor, the epithelial growth factor, a substance produced by cultured liver cells, and others, whose growth-hormone dependence has yet to be established. Some of our deliberations during these days will be concerned with whether these growth factors are to be classified among the somatomedins.

It is of a special interest that a well characterized, very familiar, anabolic substance known to be present in the blood, insulin, meets both criteria for the designation as a somatomedin. Its concentration is in part growth-hormone dependent, and it is necessary for growth. In this context it is noteworthy that some, if not all, of the other growth-promoting factors to be discussed here appear to exert an insulinlike action on their specific target tissue. The similarity of one or more of the somatomedins and nonsuppressible insulinlike activity, NSILA-S, will occupy a part of our discussion. Furthermore, one of the panelists will discuss his finding of structural similarities between the nerve growth factor and proinsulin.

Another similarity between insulin and the other growth-promoting factors might reside in their mode of action. Most of our current knowledge in this field is derived from information obtained from studies on how insulin works. One early and prominent action of insulin is depression of the level of cyclic AMP, from which a number of metabolic events ensue. It has now been amply demonstrated that growth is accompanied by such a depression of the cyclic AMP level, while growth inhibition follows elevation of cyclic AMP. It is of obvious interest that data are presented at this Symposium showing that the actions of the growth-promoting factors are accompanied by diminution of the available cyclic AMP pool. The participants will also discuss the significance of cyclic GMP in the action of growth-promoting factors. We are here entering another field of immediate interest, that dealing with the control of cellular processes: whether the growth processes are controlled by a bidirectional system of the yin-yang type whereby stimulation or inhibition is achieved by changes in opposite directions in the concentrations of cyclic AMP and cyclic GMP.

Another area to be discussed during these days deals with the transfer of information by growth-promoting factors from the outside of the cell to its macromolecular machinery. Cell surfaces contain receptors that are specific for polypeptide hormones. A number of reports deal with such receptors, specific for one or more of the growth-promoting factors. Specific assays, based on the receptor affinity for such factors, have been developed for the quantitative measurement of these substances in tissues and fluids. Again, insulin is of interest since, in some tissues, it seems to share its receptor sites with somatomedin A and C and NSILA-S. In this connection, I think it is justified to pay tribute to those who, by developing the competitive radioassays, have contributed so much to the whole field of the action of growth-promoting factors—and the action of other hormones too.

Like many breakthroughs in biomedical investigation, the impetus to the study of the somatomedins was derived from clinical and physiological observations. The finding that the effect of growth hormone on tissues required in vivo administration of the hormone and could not be fully reproduced by its in vitro addition, prompted the search for a serum factor generated by administration of growth hormone. At least one such factor was found, and over the past 20 years much effort has been directed to improving methods of measurement of the factor(s). In recent years the research has developed along more fundamental approaches. First, investigations were directed toward the interaction of these factors at the level of the cell membrane and to the specific receptors residing thereon. As we will learn at this Symposium, one must probe even deeper and consider the molecular action within the cell generated by the signal from without. Even as the totality of action of these factors requires the complete interaction between events outside and within the cells, so too does our understanding of the factors require interaction between scientists with research interests involving extracellular phenomena and those with interests involving intracellular ones.

This Symposium takes cognizance of the broad spectrum of studies relating to growth and to the diversity of scientific talents required. At this meeting we hope to attain cross-fertilization between the multiplicity of scientific disciplines concerned with different aspects of this problem. This interchange of ideas may well lead to the synthesis of concepts and to providing a framework to contain pieces of new information even as pieces fit together in a complex jigsaw puzzle.

Regulation of Cellular Growth

Ira Pastan*,     *Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland

Publisher Summary

This chapter discusses the mechanisms regulating cellular growth. Many of the hormones that control the activity of one or another endocrine gland also promote the growth of that target gland. Hormones, such as insulin and growth hormone, affect many different tissues. Generally, when untransformed fibroblasts are grown in culture under standard conditions—a physiological salt solution fortified with serum and other nutrients—they display an interesting type of growth control. The cells multiply until a confluent monolayer is reached, and then their growth slows markedly or ceases. This type of growth control bears a number of names, including density-dependent inhibition of growth and contact inhibition of growth. One well-characterized anabolic substance is insulin, and insulin stimulates cell growth. Besides insulin, other large molecular weight factors have also been found to stimulate cell growth. These include insulinlike factors called somatomedins a substance produced by transformed cells, a substance produced by cultured liver cells, and a pituitary factor present in semipurified preparations of luteinizing hormone.

I Introduction

II How Do Growth-Regulating Factors Work?

III Cyclic AMP and Cell Growth

IV How Does Insulin Act to Lower Cyclic AMP Levels?

V How Might Cyclic AMP and Steroids Work to Control Cell Growth?.

VI Cyclic GMP

VII Insulin Receptors, Cyclic AMP, and Cyclic GMP

VIII Malignant Transformation

IX Putting It All Together

References

I Introduction

I have been asked by the organizers of this meeting to summarize the current state of knowledge of the mechanisms regulating cellular growth. Although to understand growth regulation has never been a direct goal of any of my research endeavors, it has always been closely associated with those things I have tried to study. My comments will reflect this point of view.

I first became interested in growth control while trying to understand the mechanism of hormone action. Many of the hormones that control the activity of one or another endocrine gland also promote the growth of that target gland. Indeed many contain the suffix -tropin or -tropic as part of their name. These include TSH, LH, FSH, and ACTH. Hormones such as insulin and growth hormone affect many different tissues. The mechanism of action of these hormones was quite obscure until the discovery of cAMP (cyclic adenosine monophosphate) by Earl Sutherland and his co-workers. Then it became clear, over a period of perhaps 10 years, that many hormones exerted at least some of their effects on target tissues by raising cAMP levels (Robison et al, 1971). In the rush to identify the precise role of cAMP in mediating the actions of hormones, rather little attention was given to investigating how the tropic effects of these hormones were mediated. Indeed most research in this area focused on studying events that occurred immediately after the addition of a hormone to a tissue.

Because of the difficulty of investigating hormone action in vivo, many attempts have been made to culture endocrine cells in vitro. Gordon Sato has had the most success in this regard. The endocrine cells that grow well in culture are mainly derived from animal tumors (Sato et al, 1969). Unfortunately for the investigator, these tumors no longer need specific tropic hormones for growth, although such cells still respond to these hormones in other ways. For example, there is an adrenal cell line that synthesizes and secretes steroids only when ACTH is present, but grows as well in the absence of ACTH as in its presence (Sato et al, 1969). These cells do, however, require factors present in serum to grow in culture.

While endocrinologists were following this line of investigation, others interested in cell culture for its own sake, or virologists interested in growing viruses in vitro, had developed methods of growing fibroblastic cells and tumor cells in culture. The fibroblastic cells were usually prepared from embryos or from very young animals. When an embryo is dispersed by trypsin treatment, the cells that grow out in culture have the appearance of fibroblasts. Actually these cells do not make much collagen.

Generally, when untransformed fibroblasts are grown in culture under standard conditions (a physiological salt solution fortified with serum and other nutrients), they display an interesting type of growth control. The cells multiply until a confluent monolayer is reached, and then their growth slows markedly or ceases. This type of growth control bears a number of names including density-dependent inhibition of growth and contact inhibition of growth (Stoker and Rubin, 1967). Todaro and Green (1963) have developed permanent lines of mouse cells (3T3 cells) in which growth ceases abruptly at a rather low cell density, about 50,000 cells/cm². The density to which cells grow, and in some cases the rate at which they achieve this density, is affected by many factors, but especially the quantity of serum present, the frequency of feeding, and the pH of the medium (Ceccarini and Eagle, 1971; Rubin, 1971).

Serum, even after dialysis to remove small molecular weight factors, contains a number of substances that enhance growth. Various workers have had a go at trying to purify and isolate these factors, but their high biological potency and the small quantities present in serum have made this endeavor difficult. Nevertheless, it is apparent that there are a number of factors present that affect cell growth, cell survival, and cell motility (Holley and Kiernan, 1971).

One well-characterized anabolic substance is insulin, and insulin will stimulate cell growth. But whether or not the amount of insulin in serum is responsible for the growth-promoting effects of serum is not completely clear to me. Besides insulin, other large molecular weight factors have also been found to stimulate cell growth. These include insulinlike factors now called somatomedins (Daughaday et al., 1972; Salmon and Hosse, 1971; Scher et al., 1974) a substance produced by transformed cells (Rubin, 1970), a substance produced by cultured liver cells (Dulak and Temin, 1973), and a pituitary factor present in semipurified preparations of luteinizing hormone (Gospodarowicz, 1974). Although I list these as distinct factors, it seems entirely possible that some of these are closely related, if not identical, molecules. Such factors do not stimulate growth under all conditions. Usually it is necessary to put the test cells in a resting state, for example, in a low serum medium, to obtain a growth-promoting response.

Besides large molecular weight factors a number of smaller molecules have been shown to stimulate growth. One of these is the adrenal steroid hydrocortisone (Gospodarowicz, 1974). Another is the cation Zn²+ (Rubin, 1972).

II How Do Growth-Regulating Factors Work?

As best as I can make out, our current ideas of how growth-promoting factors work are derived from information generated by studies on how insulin works and especially how insulin affects the metabolism of muscle, liver, and fat tissue. Some very prominent early actions of insulin on these tissues are (1) increased transport processes and particularly increased glucose uptake and (2) decreased levels of cAMP and changes in glycogen and fat metabolism resulting from low levels of cAMP (Cahill, 1971). Since cAMP analogs as well as agents that raise cAMP levels (some prostaglandins and some hormones) antagonize many of the metabolic effects of insulin, it is thought that at least some of the metabolic responses to insulin are due to its ability to lower cAMP levels. It does not appear that the fall in cAMP levels either causes or is a consequence of enhanced hexose uptake, but rather that both actions of insulin are a consequence of some other initial event. I shall return to this point but emphasize it here because there is coupling of glucose transport and cAMP metabolism in the bacterium Escherichia coli (Pastan and Perlman, 1969). When E. coli are placed in a glucose-containing medium and rapidly take up glucose, cAMP levels rapidly fall. This fall is at least in part due to a rapid loss of cAMP from the cells into the medium (Makman and Sutherland, 1965).

In various cultured fibroblastic cells, the addition of insulin and other serum factors rapidly promotes increased glucose uptake, uridine uptake, phosphate uptake, and a fall in cAMP levels (Shaw and Amos, 1973; Rozengurt and De Asua, 1973; Pastan and Johnson, 1974). Subsequently cellular growth is increased. This is particularly striking in serum-deprived cells. Since treatment of cells with agents that raise levels of cAMP blocks the increase in uridine uptake, it appears that cAMP may be regulating uridine uptake (Rozengurt and De Asua, 1973). However, the changes in phosphate uptake do not appear to be secondary to changes in cAMP, and it has been suggested that the amount of phosphate in the medium may govern cAMP levels (Rozengurt and De Asua, 1973). Treatment of growing cells with cAMP analogs, or with agents that raise cAMP levels, slows growth (Pastan and Johnson, 1974). Thus an antagonism between the effects of insulin (or serum) and cAMP is evident. I shall not discuss the actions of the various serum factors any further, for this is a major topic of this conference.

III Cyclic AMP and Cell Growth

What is the evidence that cAMP regulates cell growth? There are at least four lines of evidence: (1) as mentioned above, agents that raise cAMP levels slow growth; (2) agents that lower cAMP levels (insulin, trypsin, serum) promote growth; (3) cAMP levels vary directly with the doubling time of logarithmically growing mouse and rat cells; and (4) the highest levels of cAMP are observed in density inhibited, non-growing 3T3 or NRK cells (reviewed in Pastan and Johnson, 1974).

The cell membrane has been considered to be a site at which growth-controlling factors act. Indeed many workers have proposed that alterations in the cell membrane result in the abnormal growth of transformed cells. One of the attractive features of implicating cAMP in growth control is that the enzyme that makes cAMP (adenylate cyclase) is exclusively located in the plasma membrane. In addition, one of the two enzymes that degrade cAMP (cAMP phosphodiesterases) is also located in the plasma membrane (Russell and Pastan, 1973).

There are at least two stages of the cell cycle, at which cAMP acts to inhibit growth. One is early in G1; the other is G2 (Fig. 1). A variety of experiments have led to these conclusions. In these experiments the cAMP analog N⁶−2’-0-dibutyryl cAMP (Bt2cAMP) has been employed.

Fig. 1 Cyclic AMP and the cell cycle.

Cells arrested in G1 can be stimulated to grow by trypsinization and replating at a low cell density, or by the addition of serum. (Both these treatments lower cAMP levels.) Bt2cAMP added at the time of trypsinization (Willingham et al., 1972) or of serum addition (Froelich and Rachmeler, 1972) blocks the usual burst of DNA synthesis occurring 12–20 hours later. If the time of addition of Bt2cAMP is delayed for 6–8 hours, DNA synthesis is not inhibited, but mitosis is still prevented. Such cells have traversed G1 and S and are arrested somewhere in G2. Other evidence that cAMP arrests growth in the G2 phase of the cell cycle comes from analyses of time-lapse movies of L cells treated with Bt2cAMP for 3 days (Pastan et al., 1972). When the Bt2cAMP is removed from these cells, there is a burst of mitosis; about 25% of the cells divide within 2 hours.

In addition to its inhibitory effects on cell growth, cAMP probably also has a stimulatory role at certain stages of the cell cycle (Willingham et al., 1972; Zeilig et al., 1974). The sites of these positive and negative actions are illustrated in Fig. 1.

IV How Does Insulin Act to Lower Cyclic AMP Levels?

Despite the fact that it is relatively easy to show that insulin lowers cAMP levels in intact cells, it has been difficult to elucidate the mechanism of this response. Two types of response to insulin have been observed. Some workers have reported that insulin decreases adenylate cyclase activity when added to cellular homogenates (Hepp, 1971; Illiano and Cuatrecasas, 1972). It is of particular interest that somatomedin has also been found to inhibit adenylate cyclase (Tell et al., 1973). However, many laboratories have failed to demonstrate an effect of insulin on adenylate cyclase in cell-free extracts. The basis of this conflict is as yet unresolved.

Another effect observed with insulin is an increase in the activity of cAMP phosphodiesterase (Loten and Sneyd, 1970). This action of insulin has not been observed in broken cell preparations, but only when intact cells were treated with the hormone, the cells homogenized, and the enzyme activity then measured (Loten and