Tài liệu Cardiovascular Development Methods and Protocols ppt

METHODS I N MOLECULAR BIOLOGY™

Series Editor

John M. Walker

School of Life Sciences

University of Hertfordshire

Hatfield, Hertfordshire, AL10 9AB, UK

For further volumes:

http://www.springer.com/series/7651

Cardiovascular Development

Methods and Protocols

Edited by

Xu Peng

Department of Systems Biology and Translational Medicine, College of Medicine,

Texas A&M Healthy Science Center, Temple, TX, USA

Marc Antonyak

Department of Molecular Medicine, School of Veterinary Medicine,

Cornell University, Ithaca, NY, USA

ISSN 1064-3745 e-ISSN 1940-6029

ISBN 978-1-61779-522-0 e-ISBN 978-1-61779-523-7

DOI 10.1007/978-1-61779-523-7

Springer New York Dordrecht Heidelberg London

Library of Congress Control Number: 2011944649

© Springer Science+Business Media, LLC 2012

All rights reserved. This work may not be translated or copied in whole or in part without the written permission of the

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Printed on acid-free paper

Humana Press is part of Springer Science+Business Media (www.springer.com)

Editors

Xu Peng

Department of Systems Biology

and Translational Medicine

College of Medicine

Texas A&M Healthy Science Center

Temple, TX, USA

[email protected]

Marc Antonyak

Department of Molecular Medicine

School of Veterinary Medicine

Cornell University

Ithaca, NY, USA

[email protected]

v

Preface

Congenital heart disease is the leading cause of infant death and affects approximately one

in every 100 babies born in the USA. Aberrant cardiovascular development is the reason for

congenital heart diseases and the pathogenesis of majority congenital heart disease remains

unclear. Cardiovascular system is the fi rst system to begin functioning and plays critical roles

in embryo development. From the lower invertebrate to mammalian animal, the heart mor￾phology is obviously different among Drosophila (one chamber), Zebrafi sh (two cham￾bers), Xenopus (three chambers), and rodent (four chambers), but the genetic and molecular

mechanisms in cardiovascular development are surprisingly conserved. Indeed, the knowl￾edge we get from the invertebrate and vertebrate model organisms can help us understand

and explore new strategy for the treatment of human cardiovascular disease.

The study of cardiovascular development has acquired new momentum in last 20 years

due to the advancement of modern molecular biology and new available equipments and

techniques, and we begin to understand the molecular pathways and cellular interaction in

the process of heart induction, rightward looping, chamber formation, and maturation.

Heart and vascular developments are sophisticated processes and new information expanded

very quickly. It is not diffi cult to fi nd a text book or review articles to summarize the new

advancements in the fi eld of cardiovascular development; however, it is not easy to fi nd a

book to describe the comprehensive step-by-step protocols for cardiovascular development

research. Owing to the page limitation, the current research articles cannot describe the

very detail of the experimental material and methods. The major goal of this book is to

provide the step-by-step protocols for both beginner and experience scientist in the fi eld of

cardiovascular development research.

Cardiovascular development: methods and protocols cover many new state-of-the-art

techniques in the fi eld of cardiovascular development research including in vivo imaging

and Bioinformatics. We also described many of the classical methods which are high fre￾quently used in the cardiovascular development research, such as fate mapping and immuno￾histochemistry staining. This book is divided into three parts. In part I, we summarized

using different organisms for cardiovascular developmental research. Part II focused on

using cell and molecular biology methods to study cardiovascular development. Part III

summarized the new available techniques for cardiovascular development research, such as

in vivo imaging and bioinformatics. Our primary audience of this book is for molecular

biologists and cell biologists who are working on the cardiovascular development research.

It is also a useful reference for clinician, genetic biologist, biochemists, biophysicists, or

other fi eld scientists who are interested in cardiovascular development.

Temple, TX, USA Xu Peng

Ithaca, NY, USA Marc Antonyak

vii

Contents

Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v

Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xi

PART I MODEL ORGANISMS

1 Use of Whole Embryo Culture for Studying Heart Development . . . . . . . . . . 3

Calvin T. Hang and Ching-Pin Chang

2 Quantifying Cardiac Functions in Embryonic and Adult Zebrafish . . . . . . . . . 11

Tiffany Hoage, Yonghe Ding, and Xiaolei Xu

3 Analysis of the Patterning of Cardiac Outflow Tract and Great

Arteries with Angiography and Vascular Casting . . . . . . . . . . . . . . . . . . . . . . . 21

Ching-Pin Chang

4 Morpholino Injection in Xenopus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29

Panna Tandon, Chris Showell, Kathleen Christine,

and Frank L. Conlon

5 Chicken Chorioallantoic Membrane Angiogenesis Model . . . . . . . . . . . . . . . . 47

Domenico Ribatti

6 Visualizing Vascular Networks in Zebrafish: An Introduction

to Microangiography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Christopher E. Schmitt, Melinda B. Holland, and Suk-Won Jin

7 Whole-Mount Confocal Microscopy for Vascular

Branching Morphogenesis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

Yoh-suke Mukouyama, Jennifer James, Joseph Nam,

and Yutaka Uchida

8 Visualization of Mouse Embryo Angiogenesis

by Fluorescence-Based Staining. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79

Yang Liu, Marc Antonyak, and Xu Peng

9 Miniaturized Assays of Angiogenesis In Vitro . . . . . . . . . . . . . . . . . . . . . . . . . 87

May J. Reed and Robert B. Vernon

PART II CELL AND MOLECULAR BIOLOGY METHODS

10 Analysis of the Endocardial-to-Mesenchymal Transformation

of Heart Valve Development by Collagen Gel Culture Assay . . . . . . . . . . . . . . 101

Yiqin Xiong, Bin Zhou, and Ching-Pin Chang

11 Quantification of Myocyte Chemotaxis: A Role for FAK

in Regulating Directional Motility. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111

Britni Zajac, Zeenat S. Hakim, Morgan V. Cameron,

Oliver Smithies, and Joan M. Taylor

viii Contents

12 Analysis of Neural Crest Cell Fate During Cardiovascular Development

Using Cre-Activated lacZ/ b-Galactosidase Staining . . . . . . . . . . . . . . . . . . . . 125

Yanping Zhang and L. Bruno Ruest

13 Indirect Immunostaining on Mouse Embryonic Heart

for the Detection of Proliferated Cardiomyocyte . . . . . . . . . . . . . . . . . . . . . . . 139

Jieli Li, Marc Antonyak, and Xu Peng

14 Isolation and Characterization of Vascular Endothelial Cells

from Murine Heart and Lung . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147

Yixin Jin, Yang Liu, Marc Antonyak, and Xu Peng

15 Isolation and Characterization of Embryonic and Adult Epicardium

and Epicardium-Derived Cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155

Bin Zhou and William T. Pu

16 Vascular Smooth Muscle Cells: Isolation, Culture, and Characterization . . . . . 169

Richard P. Metz, Jan L. Patterson, and Emily Wilson

17 C-kit Expression Identifies Cardiac Precursor Cells in Neonatal Mice . . . . . . . 177

Michael Craven, Michael I. Kotlikoff, and Alyson S. Nadworny

18 Cardiomyocyte Apoptosis in Heart Development:

Methods and Protocols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191

Dongfei Qi and Mingui Fu

19 Adenovirus-Mediated Gene Transfection in the Isolated

Lymphatic Vessels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199

Anatoliy A. Gashev, Jieli Li, Mariappan Muthuchamy,

and David C. Zawieja

20 Isolation of Cardiac Myocytes and Fibroblasts

from Neonatal Rat Pups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205

Honey B. Golden, Deepika Gollapudi, Fnu Gerilechaogetu,

Jieli Li, Ricardo J. Cristales, Xu Peng, and David E. Dostal

PART III NEW TECHNIQUES

21 The Application of Genome-Wide RNAi Screens in Exploring

Varieties of Signaling Transduction Pathways. . . . . . . . . . . . . . . . . . . . . . . . . . 217

Shenyuan Zhang and Hongying Zheng

22 Application of Atomic Force Microscopy Measurements

on Cardiovascular Cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229

Xin Wu, Zhe Sun, Gerald A. Meininger,

and Mariappan Muthuchamy

23 In Utero Assessment of Cardiovascular Function in the Embryonic

Mouse Heart Using High-Resolution Ultrasound Biomicroscopy . . . . . . . . . . 245

Honey B. Golden, Suraj Sunder, Yang Liu, Xu Peng,

and David E. Dostal

24 Isolation and Preparation of RNA from Rat Blood and Lymphatic

Microvessels for Use in Microarray Analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . 265

Eric A. Bridenbaugh

Contents ix

25 Visual Data Mining of Coexpression Data to Set Research Priorities

in Cardiac Development Research . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 291

Vincent VanBuren

26 High-Speed Confocal Imaging of Zebrafish Heart Development. . . . . . . . . . . 309

Jay R. Hove and Michael P. Craig

27 Measurement of Electrical Conduction Properties of Intact Embryonic

Murine Hearts by Extracellular Microelectrode Arrays. . . . . . . . . . . . . . . . . . . 329

David G. Taylor and Anupama Natarajan

Index. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 339

xi

Contributors

MARC ANTONYAK • Department of Molecular Medicine , School of Veterinary Medicine,

Cornell University , Ithaca , NY , USA

ERIC A. BRIDENBAUGH • Department of Systems Biology and Translational Medicine ,

Texas A&M Health Science Center College of Medicine , Temple , TX , USA

MORGAN V. CAMERON • Department of Pathology and McAllister Heart Institute ,

University of North Carolina , Chapel Hill , NC , USA

CHING-PIN CHANG • Department of Medicine, Division of Cardiovascular Medicine ,

Stanford Cardiovascular Institute, Stanford University School of Medicine ,

Stanford , CA , USA

KATHLEEN CHRISTINE • Department of Genetics , UNC McAllister Heart

Institute (MHI), University of North Carolina at Chapel Hill ,

Chapel Hill , NC , USA

FRANK L. CONLON • Department of Genetics , UNC McAllister Heart Institute (MHI),

University of North Carolina at Chapel Hill , Chapel Hill , NC , USA

MICHAEL P. CRAIG • Department of Molecular and Cellular Physiology ,

University of Cincinnati College of Medicine , Cincinnati , OH , USA

MICHAEL CRAVEN • Biomedical Sciences Department , College of Veterinary Medicine,

Cornell University , Ithaca , NY , USA

RICARDO J. CRISTALES • Department of Internal Medicine, Division of Molecular

Cardiology , College of Medicine, Texas A&M Health Science Center ,

Temple , TX , USA

DAVID E. DOSTAL • Department of Internal Medicine, Division of Molecular

Cardiology , College of Medicine, Texas A&M Health Science Center ,

Temple , TX , USA ; Central Texas Veterans Health Care System , Temple , TX , USA

YONGHE DING • Department of Biochemistry and Molecular Biology , Mayo Clinic ,

Rochester , MN , USA ; Department of Medicine, Division of Cardiovascular Diseases ,

Mayo Clinic , Rochester , MN , USA

MINGUI FU • Department of Basic Medical Science and Shock/Trauma Research

Center , School of Medicine, University of Missouri Kansas City ,

Kansas City , MO , USA

ANATOLIY A. GASHEV • Department of Systems Biology and Translational Medicine ,

College of Medicine, Cardiovascular Research Institute Division of Lymphatic

Biology, Texas A&M Health Science Center , Temple , TX , USA

FNU GERILECHAOGETU • Department of Internal Medicine, College of Medicine,

Division of Molecular Cardiology , Texas A&M Health Science Center ,

Temple , TX , USA

HONEY B. GOLDEN • Department of Internal Medicine, Division of Molecular

Cardiology , College of Medicine, Texas A&M Health Science Center ,

Temple , TX , USA

xii Contributors

DEEPIKA GOLLAPUDI • Department of Internal Medicine, Division of Molecular

Cardiology , College of Medicine, Texas A&M Health Science Center ,

Temple , TX , USA

ZEENAT S. HAKIM • Department of Pathology and McAllister Heart Institute ,

University of North Carolina , Chapel Hill , NC , USA

CALVIN T. HANG • Department of Medicine, Division of Cardiovascular Medicine ,

Stanford Cardiovascular Institute, Stanford University School of Medicine ,

Stanford , CA , USA

TIFFANY HOAGE • Department of Biochemistry and Molecular Biology , Mayo Clinic ,

Rochester , MN , USA ; Department of Medicine, Division of Cardiovascular Diseases ,

Mayo Clinic , Rochester , MN , USA

MELINDA B. HOLLAND • Department of Cell and Molecular Physiology,

Curriculum in Genetics and Molecular Biology , McAllister Heart Institute,

University of North Carolina at Chapel Hill , Chapel Hill , NC , USA

JAY R. HOVE • Department of Molecular and Cellular Physiology ,

University of Cincinnati College of Medicine , Cincinnati , OH , USA

JENNIFER JAMES • Laboratory of Stem Cell and Neuro-Vascular Biology,

Genetics and Developmental Biology Center , National Heart, Lung,

and Blood Institute, National Institutes of Health , Bethesda , MD , USA

SUK-WON JIN • Department of Cell and Molecular Physiology, Curriculum in Genetics

and Molecular Biology , McAllister Heart Institute, University of North Carolina

at Chapel Hill , Chapel Hill , NC , USA

YIXIN JIN • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Healthy Science Center , Temple , TX , USA

MICHAEL I. KOTLIKOFF • Biomedical Sciences Department , College of Veterinary

Medicine, Cornell University , Ithaca , NY , USA

JIELI LI • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Health Science Center , Temple , TX , USA

YANG LIU • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Healthy Science Center , Temple , TX , USA

GERALD A. MEININGER • Department of Medical Pharmacology and Physiology ,

Dalton Cardiovascular Research Center, University of Missouri-Columbia ,

Columbia , MO , USA

RICHARD P. METZ • Department of Systems Biology and Translational Medicine ,

Texas A&M Health Science Center, College of Medicine , College Station , TX , USA

YOH-SUKE MUKOUYAMA • Laboratory of Stem Cell and Neuro-Vascular Biology,

Genetics and Developmental Biology Center , National Heart, Lung,

and Blood Institute, National Institutes of Health , Bethesda , MD , USA

MARIAPPAN MUTHUCHAMY • Department of Systems Biology and Translational

Medicine , Texas A&M Health Science Center College of Medicine ,

College Station , TX , USA

ALYSON S. NADWORNY • Biomedical Sciences Department , College of Veterinary

Medicine, Cornell University , Ithaca , NY , USA

JOSEPH NAM • Laboratory of Stem Cell and Neuro-Vascular Biology,

Genetics and Developmental Biology Center , National Heart, Lung,

and Blood Institute, National Institutes of Health , Bethesda , MD , USA

Contributors xiii

ANUPAMA NATARAJAN • Department of Biology , Seminole State College of Florida ,

100 Weldon Blvd , Sanford , FL , USA

JAN L. PATTERSON • Department of Systems Biology and Translational Medicine ,

Texas A&M Health Science Center, College of Medicine , College Station , TX , USA

XU PENG • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Healthy Science Center , Temple , TX , USA

WILLIAM T. PU • Department of Cardiology, Children’s Hospital Boston ,

Boston , MA , USA ; Harvard Stem Cell Institute , Cambridge , MA , USA

DONGFEI QI • Department of Basic Medical Science and Shock/Trauma Research

Center , School of Medicine, University of Missouri Kansas City ,

Kansas City , MO , USA

MAY J. REED • Department of Medicine , University of Washington,

Harborview Medical Center , Seattle , WA , USA

DOMENICO RIBATTI • Department of Basic Medical Sciences, Section of Human

Anatomy and Histology , University of Bari Medical School , Policlinico , Bari , Italy

L. BRUNO RUEST • Department of Biomedical Sciences , Texas A&M Healthy Science

Center-Baylor College of Dentistry , Dallas , TX , USA

CHRISTOPHER E. SCHMITT • Department of Cell and Molecular Physiology,

Curriculum in Genetics and Molecular Biology , McAllister Heart Institute,

University of North Carolina at Chapel Hill , Chapel Hill , NC , USA

CHRIS SHOWELL • Department of Genetics , UNC McAllister Heart Institute (MHI),

University of North Carolina at Chapel Hill , Chapel Hill , NC , USA

OLIVER SMITHIES • Department of Pathology and McAllister Heart Institute ,

University of North Carolina , Chapel Hill , NC , USA

ZHE SUN • Department of Medical Pharmacology and Physiology,

Dalton Cardiovascular Research Center , University of Missouri-Columbia ,

Columbia , MO , USA

SURAJ SUNDER • Department of Internal Medicine, Division of Molecular Cardiology ,

College of Medicine, Texas A&M Health Science Center , Temple , TX , USA

PANNA TANDON • Department of Genetics , UNC McAllister Heart Institute (MHI),

University of North Carolina at Chapel Hill , Chapel Hill , NC , USA

DAVID G. TAYLOR • Department of Biology , Seminole State College of Florida ,

Sanford , FL , USA

JOAN M. TAYLOR • Department of Pathology and McAllister Heart Institute ,

University of North Carolina , Chapel Hill , NC , USA

YUTAKA UCHIDA • Laboratory of Stem Cell and Neuro-Vascular Biology,

Genetics and Developmental Biology Center , National Heart, Lung,

and Blood Institute, National Institutes of Health , Bethesda , MD , USA

VINCENT VANBUREN • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Healthy Science Center , Temple , TX , USA

ROBERT B. VERNON • Hope Heart Program, Benaroya Research Institute

at Virginia Mason , Seattle , WA , USA

EMILY WILSON • Department of Systems Biology and Translational Medicine ,

Texas A&M Health Science Center, College of Medicine , College Station , TX , USA

XIN WU • Department of Systems Biology and Translational Medicine ,

Texas A&M Health Science Center College of Medicine , College Station , TX , USA

xiv Contributors

YIQIN XIONG • Department of Medicine, Division of Cardiovascular Medicine ,

Stanford Cardiovascular Institute, Stanford University School of Medicine ,

Stanford , CA , USA

XIAOLEI XU • Department of Biochemistry and Molecular Biology , Mayo Clinic ,

Rochester , MN , USA ; Department of Medicine, Division of Cardiovascular Diseases ,

Mayo Clinic , Rochester , MN , USA

BRITNI ZAJAC • Department of Pathology and McAllister Heart Institute ,

University of North Carolina , Chapel Hill , NC , USA

DAVID C. ZAWIEJA • Department of Systems Biology and Translational Medicine ,

College of Medicine, Cardiovascular Research Institute Division of Lymphatic

Biology, Texas A&M Health Science Center , Temple , TX , USA

SHENYUAN ZHANG • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Healthy Science Center , Temple , TX , USA

YANPING ZHANG • Department of Biomedical Sciences , Texas A&M Healthy Science

Center-Baylor College of Dentistry , Dallas , TX , USA

HONGYING ZHENG • Department of Systems Biology and Translational Medicine ,

College of Medicine, Texas A&M Healthy Science Center , Temple , TX , USA

BIN ZHOU • Department of Cardiology, Children’s Hospital Boston, Boston, MA, USA;

Harvard Stem Cell Institute, Cambridge, MA, USA; Key Laboratory of Nutrition

and Metabolism, Institute for Nutritional Sciences, Shanghai Institutes for Biological

Sciences, Chinese Academy of Sciences, Shanghai, China; Department of Genetics,

Albert Einstein College of Medicine, Bronx, NY, USA