Human Cell Culture: Volume VI: Embryonic Stem Cells PDF
Preview Human Cell Culture: Volume VI: Embryonic Stem Cells
John R. Masters Bernhard O. Palsson James A. Thomson Editors Hum a n C e l l C u l t u r e 6 Embryonic Stem Cells M a s t e r s · P a l s s o n · T h oEmdss. o n Embryonic Stem Cells FM.qxd 18/4/07 5:02 PM Page i HUMAN CELL CULTURE Volume VI: Embryonic Stem Cells FM.qxd 18/4/07 5:02 PM Page ii Human Cell Culture Volume 6 The titles published in this series are listed at the end of this volume. FM.qxd 18/4/07 5:02 PM Page iii Human Cell Culture Volume VI Embryonic Stem Cells edited by John R. Masters University College London, London, UK Bernhard O. Palsson University of California, San Diego, CA, USA and James A. Thomson University of Wisconsin, Madison, WI, USA FM.qxd 18/4/07 5:02 PM Page iv A C.I.P. Catalogue record for this book is available from the Library of Congress. ISBN-13 978-1-4020-5982-7 (HB) ISBN-13 978-1-4020-5983-4 (e-book) Published by Springer, P.O. Box 17, 3300 AA Dordrecht, The Netherlands. www.springer.com Printed on acid-free paper All Rights Reserved © 2007 Springer No part of this work 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, with the exception of any material supplied specifically for the purpose of being entered and executed on a computer system, for exclusive use by the purchaser of the work. FM.qxd 18/4/07 5:02 PM Page v CONTENTS Preface vii 1. Defined Culture Media for Human Embryonic Stem Cells 1 Tenneille Ludwig and James A. Thomson 2. Generation of Disease-specific Human Embryonic Stem Cell Lines 17 Stephen Minger 3. Characterization and Differentiation of Human Embryonic Stem Cells 27 Andrew L. Laslett, Adelia Lin, and Martin F. Pera 4. Genetic Modification of Human Embryonic Stem Cells 41 Thomas P. Zwaka 5. Hematopoietic Differentiation 53 Chantal Cerdan, Veronica Ramos-Mejia, and Mickie Bhatia 6. Neural Differentiation 85 Zhi-Jian Zhang, Jason S. Meyer, and Su-Chun Zhang 7. Germ Cell Differentiation 109 Vanessa T. Angeles and Renee A. Reijo Pera 8. Mesodermal Differentiation 129 Nadav Sharon and Nissim Benvenisty v FM.qxd 18/4/07 5:02 PM Page vi vi Contents 9. Three-dimensional Culture of Human Embryonic Stem Cells 149 Sharon Gerecht, Jason A. Burdick, Christopher Cannizzaro, and Gordana Vunjak-Novakovic 10. Extraembryonic Cell Differentiation 173 Lyle Armstrong and Majlinda Lako 11. Pancreatic Cell Differentiation 189 Bettina Fishman, Hanna Segev, and Joseph Itskovitz-Eldor 12. Cardiomyocyte Differentiation 211 Dinender K. Singla, Shreeya Jayaraman, Jianhua Zhang, and Timothy J. Kamp 13. Human Embryonal Carcinoma (EC) Cells: Complementary Tools for Embryonic Stem Cell Research 235 Peter D. Tonge and Peter W. Andrews 14. Quality Control of Human Stem Cell Lines 255 Glyn N. Stacey Index 277 FM.qxd 18/4/07 5:02 PM Page vii PREFACE The aim of this volume is to describe methods for culturing human embryonic stem cells and the culture conditions needed to direct these cells to differentiate into specialized cell types. Human embryonic stem cells are potentially capable of differentiation into any other cell type, including endoderm, mesoderm, and ectoderm. Consequently there is a great deal of academic and commercial interest in uti- lizing these cells in the treatment of a wide variety of medical conditions, as well as certain ethical considerations. The maintenance and differentiation of human embryonic stem cells is the focus of many large programs in cell biology and of many groups wishing to translate their research to the clinic. However, human embryonic stem cells are difficult first to establish in culture and second to maintain in an undifferentiated state. The development and optimization of techniques for growing and maintaining stem cells and for directing them to dif- ferentiate along specific cell lineages are crucial to the clinical application of these cells and are the focus of this book. Transplanted organs and tissues are often used to replace those that are diseased or destroyed, but the number of people needing a transplant far exceeds the number of organs available. Pluripotent stem cells offer the possibility of a renewable source of replacement cells and tissues to treat conditions such as Parkinson’s and Alzheimer’s diseases, spinal cord injury, stroke, burns, heart disease, diabetes, osteoarthritis, and rheumatoid arthritis. This book deals with human embryonic stem cells and the derivation, maintenance, and differentiation of human adult stem cells will be the subject of the next volume. vii Ch01.qxd 18/4/07 4:42 PM Page 1 CHAPTER 1 DEFINED CULTURE MEDIA FOR HUMAN EMBRYONIC STEM CELLS 1,2,3 1,2,3,4 TENNEILLE LUDWIG AND JAMES A. THOMSON 1 WiCell Research Institute, P.O. Box 7365, Madison, WI 53707 2 National Primate Research Center, University of Wisconsin Graduate School, 1220 Capitol Court, Madison, WI 53705 3 Genome Center of Wisconsin, University of Wisconsin-Madison, 425 Henry Mall, Madison, WI 53706 4 Department of Anatomy, University of Wisconsin Medical School, 470 N. Charter Street, Madison, WI 53706-1509 Human embryonic stem (ES) cells can differentiate into any tissue type within the body, and thus offer tremendous potential for use in regenerative medicine. However, traditional tissue culture methods employed in their derivation and proliferation jeopardize that potential because of the inclusion of animal- derived products. The improved safety facilitated by the development of defined culture systems that eliminate the need for animal products are key in the trans- fer of human ES cell technology from the bench to the bedside. Here we describe recent advances in the culture of human ES cells in defined culture medium. Human ES cells were first isolated using mouse embryonic fibroblast (MEF) feeder layers and bovine serum-containing medium (Thomson et al. 1998). This technique, using serum and feeder layers to support difficult-to-culture cells, was first developed more than half a century ago when Puck used mitotically inac- tivated feeder cells along with serum to maintain delicate HeLa cells in culture (Puck and Marcus 1955). Years later, the technique was adapted for both murine embryonic carcinoma (EC) (Evans 1972) and ES cell culture (Evans and Kaufman 1981; Martin 1981), and has been used in the derivation of all human ES cell lines that are currently available for federal funding in the USA (see http://stemcells.nih.gov/research/registry). However, use of feeder layers and animal serum products in human ES cell culture carries the risk of cross con- tamination of xenogenic or allogenic pathogens or cell byproducts. This risk was confirmed when it was found that human ES cells incorporate a nonhuman sialic acid during culture with animal products (Martin et al. 2005). The incor- poration of this immunogenic molecule during routine culture raises the specter 1 J.R. Masters et al (eds.), Embryonic Stem Cells, 1–16. © 2007 Springer. Ch01.qxd 18/4/07 4:42 PM Page 2 2 Ludwig and Thomson of immune rejection and renders the resulting cells inappropriate for transplan- tation therapies unless the antigen is removed. The inherent risks of culture with animal feeders and serum products were recognized early, and almost as soon as the cell lines were isolated, improved culture systems were being developed. Because serum is highly variable lot to lot, and is known to mask media deficiencies, research to improve the quality of the medium required that it be eliminated from the culture system. Amit and colleagues first discovered that human ES cells could be maintained and clon- ally propagated on MEFs in the absence of serum using a combination of basic fibroblast growth factor (bFGF) and a commercially available serum replacer (Knockout Serum Replacer, KOSR: Gibco) (Amit et al. 2000). While a substan- tial improvement over serum products in terms of variability, KOSR contains “Albumax,” a poorly defined lipid-rich bovine albumin, making it problematic for culturing cells for clinical use. The same medium supports human ES cells in the absence of direct contact with feeder layers if it is conditioned on MEFs prior to use (Xu et al. 2001). This method became known as “feeder-free” culture, despite the fact that it is not free from potential feeder-layer contaminants, merely free of direct human ES cell contact with MEFs. In an effort to remove animal products from the culture system, many inves- tigators replaced MEFs with human-sourced feeder cells. Fetal muscle and fetal epithelial (Richards et al. 2002), adult epithelial (Richards et al. 2003), fallopian tube (Richards et al. 2002), marrow (Cheng et al. 2003), foreskin (Amit et al. 2003; Choo et al. 2004; Hovatta et al. 2003), and placental cells (Genbacev et al. 2005) were all used with success. Feeder layers were also derived from differentiated human ES cells (Stojkovic et al. 2004), addressing the limited availability and tremendous variability often seen with primary cell culture mate- rial. All but one (Genbacev et al. 2005) of these studies, however, utilized bovine albumin containing KOSR or calf serum, and therefore, did not eliminate all potential sources of human ES cell contamination. Additionally, it is important to note that feeder layers, regardless of the source, are extremely labor-intensive to prepare, and may be the primary factor limiting large-scale culture of human ES cells necessary for therapeutic use. The variability between lots is also a cause of inconsistency in human ES cell culture, and the use of primary human tissues has the potential to introduce human pathogens to the culture. Clearly, a culture system completely independent of feeders of any kind is more desirable. The role that feeder layers play in maintaining self-renewal in human ES cell culture, however, remains poorly understood. While leukemia inhibitory factor (LIF) activation of the LIF/Stat pathway is sufficient to sustain pluripotency in the absence of feeder layers in murine ES cells, it does not have the same effect in human ES cells (Thomson et al. 1998; Reubinoff et al. 2000; Daheron et al. 2004; Sato et al. 2004), suggesting a role for other pathways in maintaining human ES cells self-renewal. Amit and associates demonstrated a role for trans- forming growth factor beta (TGFβ), which when combined with bFGF allows feeder-independent growth of human ES cells, but with about a 20% background
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