Description
Book SynopsisIntroduces all of the essential cell biology and developmental biology background for the study of stem cells This book gives you all the important information you need to become a stem cell scientist. It covers the characterization of cells, genetic techniques for modifying cells and organisms, tissue culture technology, transplantation immunology, properties of pluripotent and tissue specific stem cells and, in particular, the relevant aspects of mammalian developmental biology. It dispels many misconceptions about stem cellsespecially that they can be miracle cells that can cure all ills. The book puts emphasis on stem cell behavior in its biological context and on how to study it. Throughout, the approach is simple, direct, and logical, and evidence is given to support conclusions. Stem cell biology has huge potential for advancing therapies for many distressing and recalcitrant diseases, and its potential will be realized most quickly when as many people as possible have a good gr
Table of ContentsPreface xi
About the Companion Website xiii
1 What is a Stem Cell? 1
Stem Cell Markers 3
Label‐ Retention 4
The Niche 5
Asymmetric Division and Differentiated Progeny 6
Clonogenicity and Transplantation 6
In Vivo Lineage Labeling 7
Conclusions 9
Further Reading 10
2 Characterizing Cells 13
Histological and Anatomical Methods 13
Histological Sections 13
Fixation 13
Sectioning 14
Staining 14
Electron Microscopy 15
Fluorescence Microscopy 16
Wholemounts 17
Immunostaining 17
In Situ Hybridization 18
Other Methods 19
RNAseq 19
Laser Capture Microdissection 19
Flow Cytometry 20
Dividing Cells 21
The Cell Cycle 21
Studying Cell Turnover 24
Reporters for the Cell Cycle 26
Identification of Very Slow Cell Turnover 26
Classification of Cell Types by Proliferative Behavior 28
Cell Death 28
Further Reading 30
3 Genetic Modification and the Labeling of Cell Lineages 31
Introducing Genes to Cells 31
Transfection and Electroporation 31
Gene Delivery Viruses 33
Controlling Gene Expression 35
Tet System 35
Cre System 35
Inhibiting Gene Activity 37
CRISPR‐Cas9 37
Transgenic Mice 38
Animal Procedures 38
Modification of Embryonic Stem Cells 40
Types of Transgenic Mice 41
Cell Lineage 42
Cell Lineage, Fate Maps, Clonal Analysis 43
Use of CreER for Lineage Analysis 44
Retroviral Barcoding 46
Clonal Analysis in Humans 47
Further Reading 47
4 Tissue Culture, Tissue Engineering and Grafting49
Simple Tissue Culture 51
Media 51
Contamination 53
Growth in Culture 54
Cryopreservation and Banking 55
GMP Cultivation 56
Complex Tissue Culture 56
Induced Differentiation 56
Three Dimensional Cell Culture 57
Artificial Organs and Organoids 59
Grafting 60
The Immune System 61
T Cells 61
The Major Histocompatibility Complex 62
T and B Cell Responses 63
Reactions to a Graft 64
Immunosuppressive Drugs 65
Animal Experiments Involving Grafting 66
Further Reading 67
5 Early Mouse and Human Development 69
Gametogenesis 70
Germ Cells 70
Mitosis and Meiosis 70
Primordial Germ Cells (PGCs) 72
Spermatogenesis 73
Oogenesis 73
Fertilization 76
Early Development 77
Preimplantation Phase 77
Implantation Period – Mouse 80
Implantation Period – Human 82
Ethical and Legal Issues Concerning the Early Human Conceptus 85
Sex Determination 86
X‐Inactivation 87
Imprinting 87
Cloning by Nuclear Transplantation (SCNT) 89
Further Reading 90
6 Pluripotent Stem Cells 93
Mouse Pluripotent Stem Cells 93
Mouse Embryonic Stem Cells 93
Differentiation of Mouse ES cells 95
Mouse iPS Cells 97
Human Pluripotent Stem Cells 101
SCNT‐Derived Embryonic Stem Cells 102
Ethical Issues Concerning Human ES Cells 102
Pluripotent Stem Cells from Postnatal Organisms 103
Applications of Pluripotent Stem Cells 104
Further Reading 105
7 Body Plan Formation 107
Embryological Concepts 107
Developmental Commitment 107
Embryonic Induction 109
Symmetry Breaking 110
Key Molecules Controlling Development 111
Genes Encoding Developmental Commitment 111
Inducing Factors 112
Wnt System 112
FGF System 113
Nodals and BMPs 114
Notch System 114
Hedgehog System 115
Growth Promoting Pathways 115
Retinoic Acid 115
Body Plan Formation 116
General Body Plan 116
Gastrulation 116
Embryo Folding 120
Further Reading 123
8 Organogenesis 125
Nervous System 125
The Brain 126
Regional Specification of the CNS 128
Rostrocaudal 128
Mediolateral 130
Dorsoventral 131
The Eye 131
The Neural Crest 132
Epidermis 134
Hair Follicles 135
Mammary Glands 136
Somitogenesis 137
The Somite Oscillator and Gradient 138
Subdivision of the Somites 139
Myogenesis 140
The Kidney 140
Blood and Blood Vessels 142
Blood 142
Blood Vessels 143
The Heart 145
The Gut 146
Regional Specification of the Endoderm 148
The Intestine 149
The Pancreas 150
The Liver 150
Further Reading 151
9 Cell Differentiation and Growth 155
Organs, Tissues and Cell Types 155
Epithelia 156
Connective Tissues 156
Cell Differentiation 158
Regulation of Gene Activity 158
Lateral Inhibition 161
Asymmetrical Cell Division 162
Neurogenesis and Gliogenesis 164
Neurons and Glia 164
Neurogenesis 166
Gliogenesis 168
Postnatal Cell Division 169
Adult Neurogenesis 169
Neurospheres 171
Skeletal and Cardiac Muscle 171
Skeletal Muscle 171
Development of Skeletal Muscle 172
Muscle Satellite Cells 173
Cardiac Muscle 175
Endodermal Tissues 176
Cell Differentiation in the Pancreas 176
Cell Differentiation in the Intestine 178
Cell Differentiation in the Liver 179
Hepatocytes and Cholangiocytes 180
Liver Growth and Regeneration 181
Transdifferentiation and Direct Reprogramming of Cell Type 183
Differentiation Protocols for Pluripotent Stem Cells 184
Further Reading 185
10 Stem Cells in the Body 189
The Intestinal Epithelium 189
Intestinal Stem Cells 191
In Vitro Culture 193
Clonality of Intestinal Crypts 193
The Epidermis 195
Hair Follicles 197
Cornea and Limbus 199
Mammary Glands 200
Mammary Stem Cells 203
The Hematopoietic System 204
Analysis by Transplantation and in Vitro Culture 204
Hematopoiesis in the Steady State 207
The Hematopoietic Niche 209
Spermatogenesis 211
Further Reading 213
11 Regeneration, Wound Healing and Cancer 217
Planarian Regeneration 217
Neoblasts 218
Amphibian Limb Regeneration 220
The Regeneration Blastema 220
Pattern Formation in Regeneration 222
Mesenchymal Stem Cells 223
Mammalian Wound Healing 225
Soft Tissue Wounds 225
Healing of Bone Fractures 225
Spinal Cord Injuries 227
Regeneration and Repair 228
Cancer 228
Genetic Heterogeneity of Cancer 230
Cancer Stem Cells 233
Further Reading 236
Index 239
