Diabetes Mellitus: Methods and Protocols

189 Pages·2003·1.36 MB·English

by  Colleen B. Kelly, Libby A. Blair, John A. Corbett, Anna L. Scarim (auth.), Sabire Özcan (eds.)

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Diabetes Mellitus
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TM
M E T H O D S I N M O L E C U L A R M E D I C I N E
John M. Walker, SERIES EDITOR
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TM
M E T H O D S I N M O L E C U L A R M E D I C I N E
Diabetes Mellitus
Methods and Protocols
Edited by
Sabire Özcan
Department of Molecular and Cellular Biochemistry
University of Kentucky Medical College, Lexington, KY
Humana Press Totowa, New Jersey
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Preface
Diabetes mellitus is the collective name for a group of diseases associated with
hyperglycemia (high levels of blood glucose) caused by defects in insulin pro-
duction, insulin action, or both. About 6.2% of the US population (17 million
people) have diabetes mellitus. It is the leading cause of kidney failure, blind-
ness, and amputations. It is also a major risk factor for heart diseases, stroke, and
birth defects.
Diabetes Mellitus: Methods and Protocols provides a state-of-the-art account
of the experimental methodology for studying the molecular defects leading to
diabetes mellitus, both at the molecular and biochemical levels. The chapters
cover a wide range of topics written by experts in their respective fields and are
organized in two sections: Insulin Production and Insulin Action. The detailed
experimental protocols presented, including the notes of interest, provide a very
useful tool for basic researchers and clinicians for investigating and treating this
disease. Each chapter starts with an introduction to a specific technique and
explains its application in the field of diabetes research. Following the list of
materials, a detailed description of the technique is presented in the methods
section in a way that enables the successful execution of the protocol. The
“Notes” section at the end discusses the pitfalls of the technique and alternative
approaches.
I am grateful to the numerous scientists who have contributed to this volume
by writing both highly detailed and understandable chapters. Special thanks also
to Prof. John M. Walker, editor of the Methods in Molecular Medicine series and
Mr. Thomas Lanigan, President of Humana Press, for bringing Diabetes Melli-
tus: Methods and Protocols to fruition.
Sabire Özcan
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Contents
Preface ................................................................................................. v
Contributors ......................................................................................... ix
PART I. INSULIN PRODUCTION
1 Isolation of Islets of Langerhans from Rodent Pancreas
Colleen B. Kelly, Libby A. Blair, John A. Corbett,
and Anna L. Scarim ............................................................ 3
2 Purification of Rat Pancreatic β-Cells
by Fluorescence-Activated Cell Sorting
Geert Stangé, Mark Van De Casteele,
and Harry Heimberg ......................................................... 15
3 Assessment of Insulin Secretion in the Mouse
Marcela Brissova, Wendell E. Nicholson,
Masakazu Shiota, and Alvin C. Powers ......................... 23
4 Detection of Insulin Production by Immunohistochemistry
Susan C. Campbell and Wendy M. Macfarlane ..................... 47
5 Quantification of the Level of Insulin Gene Expression
Susan C. Campbell and Wendy M. Macfarlane ..................... 51
6 Chromatin Immunoprecipitation Using Isolated
Islets of Langerhans
Marcelina Párrizas, Sylvia F. Boj, Reini F. Luco,
Miguel Angel Maestro, and Jorge Ferrer ....................... 61
7 Adenoviral Gene Transfer into β-Cell Lines
Amber L. Mosley and Sabire Özcan ....................................... 73
8 Utilization of NOD Mice in the Study of Type 1 Diabetes
Dennis G. Karounos and Susan E. Goes .............................. 81
PART II. INSULIN ACTION
9 Introduction of DNA into 3T3-L1 Adipocytes
by Electroporation
Shuichi Okada, Masatomo Mori, and Jeffrey E. Pessin ...... 93
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viii Contents
10 Analysis of Insulin-Stimulated Glucose Uptake
in Differentiated 3T3-L1 Adipocytes
Jaganathan Lakshmanan, Jeffrey S. Elmendorf,
and Sabire Özcan.............................................................. 97
11 Fractionation Analysis of the Subcellular Distribution
of GLUT-4 in 3T3-L1 Adipocytes
Jeffrey S. Elmendorf .............................................................. 105
12 Visualization and Quantitation of Integral Membrane
Proteins Using a Plasma Membrane Sheet Assay
John B. Knight and Ann Louise Olson ............................... 113
13 Assaying Tyrosine Phosphorylation of Insulin Receptor
and Insulin Receptor Substrates
Eileen L. Whiteman and Morris J. Birnbaum ...................... 119
14 Measuring Insulin-Stimulated Phosphatidyl-Inositol
3-Kinase Activity
Li-Ping Wang and Scott A. Summers .................................. 127
15 Assaying AKT/ Protein Kinase B Activity
Kyle L. Hoehn and Scott A. Summers ................................. 137
16 Measurements of Cellular Phosphoinositide Levels
in 3T3-L1 Adipocytes
Daryll B. DeWald ..................................................................... 145
17 Measurement of Glycogen Synthesis and Glycogen
Synthase Activity in 3T3-L1 Adipocytes
Matthew J. Brady .................................................................... 155
18 Measurement of Contraction-Stimulated GLUT-4
Translocation in Isolated Skeletal Muscle
Joseph T. Brozinick, Jr. ......................................................... 163
19 Single Embryo Measurement of Basal- and
Insulin-Stimulated Glucose Uptake
Maggie M-Y. Chi and Kelle H. Moley .................................... 171
20 Immunohistologic Staining of Muscle and Embryos to Detect
Insulin-Stimulated Translocation of Glucose Transporters
Mary O. Carayannopoulos and Kelle H. Moley ................... 179
Index ................................................................................................. 187
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Contributors
MORRIS J. BIRNBAUM • Howard Hughes Medical Institute, Cox Institute, and the University
of Pennsylvania School of Medicine, Philadelphia, PA
LIBBY A. BLAIR • Department of Biochemistry and Molecular Biology, Saint Louis
University School of Medicine, Saint Louis, MO
SYLVIA F. BOJ • Endocrinology, Hospital Clínic Universitari; Institut d’Investigacions
Biomèdiques August Pi i Sunyer, Barcelona, Spain
MATTHEW J. BRADY • Section of Endocrinology, Department of Medicine, The University
of Chicago, Chicago IL
MARCELA BRISSOVA • Department of Medicine, Vanderbilt University Medical Center,
Nashville, TN
JOSEPH T. BROZINICK, JR. • Endocrine Division, Eli Lilly and Co, Indianapolis, IN
SUSAN C. CAMPBELL • School of Biochemistry and Genetics, University of Newcastle
Upon Tyne, The Medical School, Framlington Place, UK
MARY O. CARAYANNOPOULOS • Department of Obstetrics and Gynecology, Department
of Cell Biology and Physiology, Washington University School of Medicine, St Louis,
MO
MARK VAN DE CASTEELE • Diabetes Research Center, Vrije Universiteit Brussel, Brussels,
Belgium
MAGGIE M-Y. CHI • Department of Obstetrics and Gynecology, Department of Cell
Biology and Physiology, Washington University School of Medicine, St Louis, MO
JOHN A. CORBETT • Department of Biochemistry and Molecular Biology, Saint Louis
University School of Medicine, Saint Louis, MO
DARYLL B. DEWALD • Department of Biology, Utah State University, Logan, UT
JEFFREY S. ELMENDORF • Department of Cellular and Integrative Physiology, Indiana
University School of Medicine, Center for Diabetes Research, Indianapolis, IN
JORGE FERRER • Endocrinology, Hospital Clínic Universitari; Institut d’Investigacions
Biomèdiques August Pi i Sunyer, Barcelona, Spain
SUSAN E. GOES • VA Research Service, Department of Veterans Affairs Medical Center,
Lexington, KY
HARRY HEIMBERG • Diabetes Research Center, Vrije Universiteit Brussel, Brussels,
Belgium
KYLE L. HOEHN • Department of Biochemistry and Molecular Biology, Colorado State
University, Fort Collins, CO
DENNIS G. KAROUNOS • Medical Service, Department of Veterans Affairs Medical Center,
and Department of Internal Medicine, University of Kentucky College of Medicine,
Lexington, KY
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x Contributors
COLLEEN B. KELLY • Department of Biochemistry and Molecular Biology, Saint Louis
University School of Medicine, Saint Louis, MO
JOHN B. KNIGHT • Department of Biochemistry and Molecular Biology, University of
Oklahoma Health Sciences Center, Oklahoma City, OK
JAGANATHAN LAKSHMANAN • Department of Molecular and Cellular Biochemistry,
University of Kentucky, Medical College, Lexington, KY
REINI F. LUCO • Endocrinology, Hospital Clínic Universitari; Institut d’Investigacions
Biomèdiques August Pi i Sunyer, Barcelona, Spain
WENDY M. MACFARLANE • School of Biochemistry and Genetics, University Of Newcastle
Upon Tyne, The Medical School, Framlington Place, UK
MIGUEL ANGEL MAESTRO • Endocrinology, Hospital Clínic Universitari; Institut
d’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain
KELLE H. MOLEY • Department of Obstetrics and Gynecology, Department of Cell Biology
and Physiology, Washington University School of Medicine, St Louis, MO
MASATOMO MORI • First Department of Internal Medicine, Gunma University School
of Medicine, Gunma, Japan
AMBER L. MOSLEY • Department of Molecular and Cellular Biochemistry, University of
Kentucky, Medical College, Lexington, KY
WENDELL E. NICHOLSON • Department of Medicine, Vanderbilt University Medical
Center, Nashville, TN
SHUICHI OKADA • First Department of Internal Medicine, Gunma University School of
Medicine, Gunma, Japan
ANN LOUISE OLSON • Department of Biochemistry and Molecular Biology, University
of Oklahoma Health Sciences Center, Oklahoma City, OK
SABIRE ÖZCAN • Department of Molecular and Cellular Biochemistry, University of
Kentucky, Medical College, Lexington, KY
MARCELINA PÁRRIZAS • Endocrinology, Hospital Clínic Universitari; Institut
d’Investigacions Biomèdiques August Pi i Sunyer, Barcelona, Spain
JEFFREY E. PESSIN • Department of Physiology and Biophysics, The University of Iowa,
Iowa City, IA
ALVIN C. POWERS • Department of Medicine, Vanderbilt University Medical Center,
Veterans Affairs Tennessee Valley Healthcare System, Nashville, TN
ANNA L. SCARIM • Department of Biochemistry and Molecular Biology, Saint Louis
University School of Medicine, Saint Louis, MO
MASAKAZU SHIOTA • Department of Molecular Physiology and Biophysics, Vanderbilt
University Medical Center, Nashville, TN
GEERT STANGÉ • Diabetes Research Center, Vrije Universiteit Brussel, Brussels, Belgium
SCOTT A. SUMMERS • Department of Biochemistry and Molecular Biology, Colorado
State University, Fort Collins, CO
LI-PING WANG • Department of Biochemistry and Molecular Biology, Colorado State
University, Fort Collins, CO
EILEEN L. WHITEMAN •Howard Hughes Medical Institute, Cell and Molecular Biology Graduate
Group, and the University of Pennsylvania School of Medicine, Philadelphia, PA
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INSULIN PRODUCTION