Principles of Tissue Engineering, Fifth Edition
Edited by Robert Lanza, Robert Langer, Joseph P. Vacanti and Anthony Atala
Edited by Robert Lanza, Robert Langer, Joseph P. Vacanti and Anthony Atala
Contents
List of contributors xxix
Preface xli
1. Tissue engineering: current status and future perspectives 1
Prafulla K. Chandra, Shay Soker and Anthony Atala
Clinical need 1
Current state of the field 1
Smart biomaterials 2
Cell sources 4
Whole organ engineering 8
Biofabrication technologies 9
Electrospinning 9
Inkjet three-dimensional bioprinting 12
Extrusion three-dimensional bioprinting 12
Spheroids and organoids 13
Imaging technologies 16
Tissue neovascularization 16
Bioreactors 16
Organ-on-a-chip and body-on-a-chip 17
Integration of nanotechnology 18
Current challenges 19
Future directions 21
Smart biomaterials 21
Cell sources 22
Whole organ engineering 24
Biofabrication technologies 24
Tissue neovasculatization 25
Bioreactors 25
Integration of nanotechnology 25
Conclusions and future challenges 26
References 26
Further reading 35
2. From mathematical modeling and
machine learning to clinical reality 37
Ben D. MacArthur, Patrick S. Stumpf and
Richard O.C. Oreffo
Introduction 37
Modeling stem cell dynamics 37
Positive feedback_based molecular switches 38
Variability in stem cell populations 40
Modeling tissue growth and development 41
Monolayer tissue growth in vitro 42
Tissue growth on complex surfaces in vitro 42
Three-dimensional tissue growth in vitro 43
Pattern formation 44
Machine learning in tissue engineering 45
Supervised methods 46
Unsupervised methods 46
Machine learning of cellular dynamics 47
Regulatory network inference 47
From mathematical models to clinical reality 47
References 48
3. Moving into the clinic 53
Chi Lo, Darren Hickerson, James J. Yoo,
Anthony Atala and Julie Allickson
Introduction 53
Current state of tissue engineering 53
Pathway for clinical translation 54
Regulatory considerations for tissue
engineering 58
Conclusion 60
Acknowledgment 60
References 60
Further reading 60
Preface xli
1. Tissue engineering: current status and future perspectives 1
Prafulla K. Chandra, Shay Soker and Anthony Atala
Clinical need 1
Current state of the field 1
Smart biomaterials 2
Cell sources 4
Whole organ engineering 8
Biofabrication technologies 9
Electrospinning 9
Inkjet three-dimensional bioprinting 12
Extrusion three-dimensional bioprinting 12
Spheroids and organoids 13
Imaging technologies 16
Tissue neovascularization 16
Bioreactors 16
Organ-on-a-chip and body-on-a-chip 17
Integration of nanotechnology 18
Current challenges 19
Future directions 21
Smart biomaterials 21
Cell sources 22
Whole organ engineering 24
Biofabrication technologies 24
Tissue neovasculatization 25
Bioreactors 25
Integration of nanotechnology 25
Conclusions and future challenges 26
References 26
Further reading 35
2. From mathematical modeling and
machine learning to clinical reality 37
Ben D. MacArthur, Patrick S. Stumpf and
Richard O.C. Oreffo
Introduction 37
Modeling stem cell dynamics 37
Positive feedback_based molecular switches 38
Variability in stem cell populations 40
Modeling tissue growth and development 41
Monolayer tissue growth in vitro 42
Tissue growth on complex surfaces in vitro 42
Three-dimensional tissue growth in vitro 43
Pattern formation 44
Machine learning in tissue engineering 45
Supervised methods 46
Unsupervised methods 46
Machine learning of cellular dynamics 47
Regulatory network inference 47
From mathematical models to clinical reality 47
References 48
3. Moving into the clinic 53
Chi Lo, Darren Hickerson, James J. Yoo,
Anthony Atala and Julie Allickson
Introduction 53
Current state of tissue engineering 53
Pathway for clinical translation 54
Regulatory considerations for tissue
engineering 58
Conclusion 60
Acknowledgment 60
References 60
Further reading 60
Part One
The basis of growth and
differentiation 63
4. Molecular biology of the cell 65
J.M.W. Slack
The cell nucleus 65
Control of gene expression 66
Transcription factors 67
Other controls of gene activity 67
The cytoplasm 68
The cytoskeleton 69
The cell surface 71
Cell adhesion molecules 71
Extracellular matrix 72
Signal transduction 73
Growth and death 74
Culture media 75
Cells in tissues and organs 76
Cell types 76
Tissues 77
Organs 77
Reference 78
Further reading 78
5. Molecular organization of cells 79
Jon D. Ahlstrom
Introduction 79
Molecules that organize cells 79
Changes in cell_cell adhesion 80
Changes in celleextracellular matrix adhesion 80
Changes in cell polarity and stimulation of cell
motility 81
Invasion of the basal lamina 81
The epithelial_mesenchymal transition
transcriptional program 82
Transcription factors that regulate
epithelial_mesenchymal transition 82
Regulation at the promoter level 82
Posttranscriptional regulation of
epithelial_mesenchymal transition
transcription factors 83
Molecular control of the
epithelial_mesenchymal transition 83
Ligand-receptor signaling 83
Additional signaling pathways 85
A model for epithelial_mesenchymal transition
induction 85
Conclusion 86
List of acronyms and abbreviations 86
Glossary 86
References 87
6. The dynamics of cell_extracellular
matrix interactions, with implications
for tissue engineering 93
M. Petreaca and M. Martins-Green
Introduction 93
Historical background 93
Extracellular matrix composition 93
Receptors for extracellular matrix molecules 94
Cell_extracellular matrix interactions 96
Development 96
Wound healing 100
Signal transduction events during
cell_extracellular matrix interactions 104
Relevance for tissue engineering 111
Avoiding a strong immune response that
can cause chronic inflammation and/or
rejection 111
Creating the proper substrate for cell survival
and differentiation 111
Providing the appropriate environmental
conditions for tissue maintenance 112
References 113
7. Matrix molecules and their ligands 119
Allison P. Drain and Valerie M. Weaver
Introduction 119
Collagens 120
Fibrillar collagens 121
Fibril-associated collagens with interrupted
triple helices (FACIT) 122
Basement membrane_associated collagens 123
Other collagens 123
Major adhesive glycoproteins 123
Fibronectin 123
Laminin 125
Elastic fibers and microfibrils 126
Other adhesive glycoproteins and
multifunctional matricellular proteins 126
Vitronectin 126
Thrombospondins 126
Tenascins 126
Proteoglycans 127
Hyaluronan and lecticans 127
Perlecan 128
Small leucine-rich repeat proteoglycans and
syndecans 128
Conclusion 128
References 128
8. Morphogenesis and tissue
engineering 133
Priscilla S. Briquez and Jeffrey A. Hubbell
Introduction to tissue morphogenesis 133
Biology of tissue morphogenesis 133
Morphogens as bioactive signaling molecules
during morphogenesis 134
The extracellular matrix as a key regulator
of tissue morphogenesis 135
Cell_cell interactions during tissue
morphogenesis 136
Tissues as integrated systems in the body 136
Engineering tissue morphogenesis 138
Cells as building units in tissue engineering 138
Biomaterial scaffolds as artificial extracellular
matrices 139
Morphogens as signaling cues in tissue
engineering 140
Tissue remodeling in healthy and diseased
environments 140
Current focuses and future challenges 141
References 141
9. Gene expression, cell determination,
differentiation, and regeneration 145
Frank E. Stockdale
Introduction 145
Determination and differentiation 145
MyoD and the myogenic regulatory factors 147
Negative regulators of development 148
MicroRNAs—regulators of differentiation 148
Pax in development 149
Satellite cells in skeletal muscle differentiation and
repair 149
Tissue engineering—repairing muscle and fostering
regeneration by controlling determination and
differentiation 150
Conclusion 152
References 152
Part Two
In vitro control of tissue
development 155
10. Engineering functional tissues:
in vitro culture parameters 157
Jennifer J. Bara and Farshid Guilak
Introduction 157
Key concepts for engineering functional
tissues 158
Fundamental parameters for engineering
functional tissues 158
Fundamental criteria for engineering
functional tissues 159
Importance of in vitro studies for engineering
functional tissues 159
In vitro studies relevant to tissue engineering
and regenerative medicine 159
In vitro platforms relevant for high throughput
screening of drugs and other agents 160
Influence of selected in vitro culture
parameters on the development and
performance of engineered tissues 161
Culture duration 161
Biomaterials 162
Bioreactors and growth factors 166
Bioreactors and mechanical forces 169
Conclusion 171
Acknowledgments 172
References 172
Further reading 177
11. Principles of bioreactor design for
tissue engineering 179
Hanry Yu, Seow Khoon Chong,
Ammar Mansoor Hassanbhai, Yao Teng,
Gowri Balachander, Padmalosini Muthukumaran,
Feng Wen and Swee Hin Teoh
Introduction 179
Macrobioreactors 180
Design principles 181
Sustainable bioreactors 188
Cell manufacturing quality attributes and
process analytics technology 189
Future outlook 189
Microbioreactors 191
Design principles 191
Types of microreactors 194
Components and integration into
microreactors 194
Applications 195
Summary 197
Acknowledgments 197
References 197
12. Regulation of cell behavior by
extracellular proteins 205
Amy D. Bradshaw
Introduction 205
Thrombospondin-1 205
Thrombospondin-2 207
Tenascin-C 208
Osteopontin 209
Secreted protein acidic and rich in cysteine 210
Conclusion 212
References 212
13. Cell and matrix dynamics in
branching morphogenesis 217
Shaimar R. Gonza´lez Morales and
Kenneth M. Yamada
Introduction 217
The basis of branching morphogenesis 217
Branching morphogenesis in the lung 218
Branching morphogenesis in the
salivary gland 220
Branching morphogenesis in the kidney 222
Contributions of other cell types 224
MicroRNAs in branching morphogenesis 225
Extracellular matrix components in
branching morphogenesis 226
Laminin 226
Collagen 226
Heparan sulfate proteoglycan 227
Fibronectin and integrins 228
Basement membrane microperforations 228
Mathematical and computational
models 230
Geometry 230
Mechanical forces 230
Signaling mechanisms 230
Conclusion 231
Acknowledgments 232
References 232
14. Mechanobiology, tissue
development, and tissue
engineering 237
David Li and Yu-li Wang
Introduction 237
Mechanical forces in biological systems 237
Tension 237
Compression 238
Fluid shear 238
Cellular mechanosensing 238
The cytoskeleton 239
Stretch-activated ion channels 239
Cell_cell adhesions 240
Cell_substrate adhesions 240
The extracellular matrix 241
Cellular effects of mechanotransduction 243
Substrate adhesion, spreading, and
migration 243
Cell_cell interactions in collectives 243
Proliferation and differentiation 244
Mechanotransduction in biological
phenomena 245
Wound healing 245
Tissue morphogenesis 247
Cancer metastasis 248
Mechanobiology in tissue engineering 248
Bone-implant design 248
Organs-on-a-chip 250
References 252
Part Three
In Vivo Synthesis of Tissues and
Organs 257
15. In vivo engineering of organs 259
V. Prasad Shastri
Introduction 259
Historical context 259
Nature’s approach to cellular differentiation
and organization 260
Conceptual framework of the in vivo
bioreactor 261
In vivo bone engineering—the bone
bioreactor 261
In vivo cartilage engineering 264
Induction of angiogenesis using biophysical
cues—organotypic vasculature engineering 265
De novo liver engineering 267
Repairing brain tissue through controlled
induction of reactive astrocytes 269
Conclusions and outlook 269
References 270
Part Four
Biomaterials in tissue
engineering 273
16. Cell interactions with polymers 275
W. Mark Saltzman and Themis R. Kyriakides
Methods for characterizing cell interactions
with polymers 275
In vitro cell culture methods 275
In vivo methods 278
Cell interactions with polymers 280
Protein adsorption to polymers 280
Effect of polymer chemistry on cell behavior 280
Electrically charged or electrically conducting
polymers 284
Influence of surface morphology on cell
behavior 284
Use of patterned surfaces to control cell
behavior 285
Cell interactions with polymers in suspension 286
Cell interactions with three-dimensional
polymer scaffolds and gels 287
Cell interactions unique to the in vivo setting 287
Inflammation 287
Fibrosis and angiogenesis 288
References 289
17. Polymer scaffold fabrication 295
Matthew L. Bedell, Jason L. Guo,
Virginia Y. Xie, Adam M. Navara and
Antonios G. Mikos
Introduction 295
Design inputs: materials, processing, and cell
types 297
Materials and inks 297
Processing and cell viability 299
Cell types and biological interactions 300
Assessment of cell viability and activity 301
3D printing systems and printer types 302
Inkjet printing 303
Extrusion printing 304
Laser-assisted bioprinting 305
Stereolithography 305
Open source and commercial 3D printing
systems 306
Print outputs: patterning, resolution, and
porous architecture 307
Printing/patterning of multiple inks 308
Print resolution 308
Porous architecture 309
Assessment of scaffold fidelity 309
Printing applications: vascularized and
complex, heterogeneous tissues 310
Conclusion 310
Acknowledgments 311
Abbreviations 311
References 311
18. Biodegradable polymers 317
Julian Chesterman, Zheng Zhang,
Ophir Ortiz, Ritu Goyal and
Joachim Kohn
Introduction 317
Biodegradable polymer selection criteria 317
Biologically derived polymers 318
Peptides and proteins 318
Biomimetic materials 322
Polysaccharides 322
Polyhydroxyalkanoates 325
Polynucleotides 326
Synthetic polymers 326
Aliphatic polyesters 326
Aliphatic polycarbonates 330
Biodegradable polyurethanes 330
Polyanhydrides 331
Polyphosphazenes 331
Poly(amino acids) and pseudo-poly
(amino acids) 332
Combinations (hybrids) of synthetic and
biologically derived polymers 333
Using polymers to create tissue-engineered
products 333
Barriers: membranes and tubes 334
Gels 334
Matrices 334
Conclusion 335
References 335
19. Three-dimensional scaffolds 343
Ying Luo
Introduction 343
Three-dimensional scaffold design and
engineering 343
Mass transport and pore architectures 344
Mechanics 346
Electrical conductivity 348
Surface properties 349
Temporal control 352
Spatial control 354
Conclusion 355
References 355
Part Five
Transplantation of engineered
cells and tissues 361
20. Targeting the host immune
response for tissue engineering and
regenerative medicine applications 363
Jenna L. Dziki and Stephen F Badylak
Introduction 363
Immune cells and their roles in building
tissues after injury 363
Neutrophils 364
Eosinophils 364
Macrophages 364
Dendritic cells 364
T and B cells 365
Specialized immune cell functions beyond
host defense 365
Tissue engineering/regenerative medicine
strategies as immunotherapy 365
Future considerations for immune cell targeting
tissue engineering/regenerative medicine
therapies 366
References 366
Further reading 368
21. Tissue engineering and
transplantation in the fetus 369
Christopher D. Porada, Anthony Atala and
Grac¸ a Almeida-Porada
Introduction 369
Rationale for in utero therapies 370
In utero transplantation 371
Early murine experiments with in utero
transplantation 372
In utero transplantation experiments in large
preclinical animal models 372
Barriers to in utero transplantation success 373
Clinical experience with in utero
transplantation 376
Rationale for in utero gene therapy 376
Hemophilia A as a model genetic disease for
correction by in utero gene therapy 377
The need for better hemophilia A
treatments 378
Preclinical animal models for hemophilia A
and recent clinical successes 378
Sheep as a preclinical model of hemophilia A 379
Feasibility and justification for treating
hemophilia A prior to birth 380
Mesenchymal stromal cells as hemophilia A
therapeutics 383
Preclinical success with mesenchymal stromal
cell_based hemophilia A treatment 384
Risks of in utero gene therapy 385
Genomic integration_associated insertional
mutagenesis 385
Potential risk to fetal germline 386
Conclusion and future directions 387
References 388
22. Challenges in the development of
immunoisolation devices 403
Matthew A. Bochenek, Derfogail Delcassian
and Daniel G. Anderson
Introduction 403
Rejection and protection of transplanted
cells and materials 403
Rejection pathways 404
Cellular nutrition 404
Therapeutic cells 405
Primary cells 405
Immortalized cell lines 406
Stem cells 407
Device architecture and mass transport 407
Transplantation site 408
Improving oxygenation of immunoprotected
cells 409
Controlling immune responses to implanted
materials 410
Steps in the foreign body reaction 411
The role of geometry in the foreign body
reaction 411
Tuning chemical composition to prevent
attachment 412
Directing immune cell behavior in the
transplant niche 412
References 412
Part Six
Stem cells 419
23. Embryonic stem cells 421
Irina Klimanskaya, Erin A. Kimbrel and Robert
Lanza
Introduction 421
Approaches to human embryonic stem cell
derivation 421
Maintenance of human embryonic stem cell 425
Subculture of human embryonic stem cell 425
Nuances of human embryonic stem cell
culture 426
Directed differentiation 426
Safety concerns 430
Conclusion 431
Acknowledgment 431
References 431
24. Induced pluripotent stem cell
technology: venturing into the
second decade 435
Yanhong Shi, Haruhisa Inoue,
Jun Takahashi and Shinya Yamanaka
Disease modeling 435
Drug discovery 436
Stem cell_based therapeutic development 438
Concluding remarks 440
Acknowledgements 440
References 440
25. Applications for stem cells 445
Andres M. Bratt-Leal, Ai Zhang, Yanling Wang
and Jeanne F. Loring
Introduction 445
Reprogramming of somatic cells into induced
pluripotent stem cells 445
Epigenetic remodeling 446
Reprogramming techniques 446
Induced transdifferentiation 448
Genomic stability 448
Applications of induced pluripotent stem cells 448
Disease modeling 448
Challenges and future possibilities in disease
modeling 450
Disease-modifying potential of induced
pluripotent stem cells 451
Other applications for induced pluripotent
stem cells 452
Conclusion 452
List of acronyms and abbreviations 453
References 453
26. Neonatal stem cells in tissue
engineering 457
Joseph Davidson and Paolo De Coppi
Introduction 457
Stem cells 457
Embryonic stem cells 457
Induced pluripotent stem cells 458
Perinatal stem cells 458
Scaffolding specifics in fetal and neonatal
tissue engineering 459
Synthetic materials 459
Natural materials 459
Relevance to prenatal therapy 460
Immunology 460
Physiology 460
Conditions of interest 461
Spina bifida 461
Gastroschisis 461
Congenital diaphragmatic hernia 461
Esophageal atresia 461
Congenital heart disease 462
Congenital airway anomalies 462
Bladder 463
Bone and bone marrow 463
Conclusion 463
References 463
27. Embryonic stem cells as a cell
source for tissue engineering 467
Ali Khademhosseini, Nureddin Ashammakhi,
Jeffrey M. Karp, Sharon Gerecht, Lino Ferreira,
Nasim Annabi, Mohammad Ali Darabi, Dario
Sirabella, Gordana Vunjak-Novakovic and
Robert Langer
Introduction 467
Maintenance of embryonic stem cells 468
Directed differentiation 471
Genetic reprogramming 471
Microenvironmental cues 472
Three-dimensional versus two-dimensional
cell culture systems 475
High-throughput assays for directing stem cell
differentiation 475
Physical signals 477
Isolation of specific progenitor cells from
embryonic stem cells 479
Transplantation 480
Transplantation and immune response 481
Future prospects 482
Conclusion 483
Acknowledgments 483
Conflicts of interest 483
References 483
Further reading 490
Part Seven
Gene therapy 491
28. Gene therapy 493
Stefan Worgall and Ronald G. Crystal
Strategies of gene therapy 493
Ex vivo versus in vivo gene therapy 494
Ex vivo 494
In vivo 495
Chromosomal versus extrachromosomal
placement of the transferred gene 495
Gene transfer vectors 495
Nonviral vectors 497
Adenovirus 497
Adeno-associated virus 499
Retrovirus 500
Lentivirus 501
Cell-specific targeting strategies 502
Targeting of Ad vectors 502
Targeting of adeno-associated virus vectors 505
Targeting of retroviral and lentiviral vectors 505
Regulated expression of the transferred gene 505
Using gene transfer vectors for gene editing 507
Combining gene transfer with stem-cell
strategies 508
Gene transfer to stem cells 508
Gene transfer to control uncontrolled
stem-cell growth 508
Gene transfer to instruct stem-cell
differentiation 508
Gene transfer to regulate gene expression 509
Challenges to gene therapy for tissue
engineering 509
Acknowledgments 510
References 510
29. Gene delivery into cells and tissues 519
Christopher E. Nelson, Craig L. Duvall, Aleˇs
Prokop, Charles A. Gersbach and Jeffrey M.
Davidson
Introduction 519
Fundamentals of gene delivery 519
Biodistribution, targeting, uptake, and
trafficking 521
Tissue biodistribution/targeting 521
Cellular uptake and intracellular trafficking 523
Viral nucleic acid delivery 526
Introduction to viral gene therapy 526
Types of viral vectors 527
Engineering viral vectors 528
Nonviral nucleic acid delivery 530
Introduction to nonviral nucleic acid delivery 530
Oligonucleotide modifications 531
Conjugates 531
Synthetic polymers 531
Polymers derived from natural sources or
monomers 534
Lipid-based delivery systems 536
Inorganic nanoparticles 537
High-throughput screening 537
Engineering tissues with gene delivery 538
Introduction to engineering tissue with gene
delivery 538
Viral delivery to engineer tissues 538
Nonviral delivery from scaffolds 540
Nucleic acid delivery for tissue engineering
advances into the clinic 541
Future challenges 541
Outlook 542
Acknowledgments 543
References 543
Part Eight
Breast 555
30. Breast tissue engineering:
implantation and three-dimensional
tissue test system applications 557
Karen J.L. Burg and Timothy C. Burg
Introduction 557
Breast anatomy and development 557
Breast cancer diagnosis and treatments 558
Breast reconstruction 558
Synthetic implants 559
Tissue flaps 559
Cell transplants 559
Cellular scaffolds 560
Special considerations 565
Breast cancer modeling 565
Animal models 565
Breast tissue test systems 566
In silico breast cancer models 570
Concluding remarks 571
Acknowledgement 571
References 571
Part Nine
Cardiovascular system 577
31. Cardiac progenitor cells, tissue
homeostasis, and regeneration 579
Wayne Balkan, Simran Gidwani, Konstantinos
Hatzistergos and Joshua M. Hare
Origin of cardiac stem/progenitor cells 579
Modeling cardiac development with
pluripotent stem cells 581
In vivo fate mapping of cardiac progenitors 582
Neonatal cardiac repair 582
Reprogramming cardiac fibroblasts 584
Cardiac resident mesenchymal stem cells 584
Cardiomyocytes and cardiac repair/
regeneration 585
Cell-based therapy 585
Cardiac progenitor/stem cell therapy 586
Combination stem cell therapy 586
Pluripotent stem cells 586
Future directions 588
References 588
32. Cardiac tissue engineering 593
Yimu Zhao, George Eng, Benjamin W. Lee,
Milica Radisic and Gordana Vunjak-Novakovic
Introduction 593
Clinical problem 593
Engineering cardiac tissue: design principles
and key components 594
Cell source 594
Scaffold 598
Biophysical stimulation 599
Directed cardiac differentiation of human
stem cells 599
Derivation of cardiomyocytes from human
pluripotent stem cells 599
Purification and scalable production of stem
cell_derived cardiomyocytes 601
Scaffolds 601
Decellularization approach 601
Artificial scaffolds 602
Biophysical cues 604
Electrical stimulation 604
Mechanical stimulation 604
Perfusion 606
In vivo applications of cardiac tissue
engineering 606
Engineered heart issue 606
Vascularized cardiac patches 608
Electrical coupling of cardiomyocytes on the
heart 608
Modeling of disease 609
Generation of patient-specific
cardiomyocytes 609
Engineered heart tissue models 609
Cardiac fibrosis 609
Titin mutation_related dilated
cardiomyopathy 611
Diabetes-related cardiomyopathy 611
Chronic hypertension induced left ventricle
hypertrophy 611
Barth syndrome 611
Tissue engineering as a platform for
pharmacologic studies 611
Summary and challenges 612
Acknowledgments 612
References 612
33. Blood vessels 617
Luke Brewster, Eric M. Brey and
Howard P. Greisler
Introduction 617
Normal and pathologic composition of
the vessel wall 617
Developmental biology cues important in
vascular tissue engineering 618
Conduits 618
Arteries 618
Veins 618
Current status of grafts in patients 618
Conduit patency and failure 618
Venous reconstruction 619
Hemodialysis vascular access 619
Inflammation and the host response to
interventions and grafts 620
Host environment and the critical role of the
endothelium 621
Prevalent grafts in clinical use 622
Vascular tissue engineering 623
Early efforts—in vitro tissue-engineered
vascular grafts 623
Endothelial cell seeding 623
In vitro approaches to tissue-engineered
vascular grafts 624
In vivo tissue-engineered vascular grafts 625
Bioresorbable grafts 625
The living bioreactor 626
Cellular and molecular mediators of graft
outcome 626
Conclusion and predictions for the future 630
References 630
34. Heart valve tissue engineering 635
Kevin M. Blum, Jason Zakko, Peter Fong,
Mark W. Maxfield, Muriel A. Cleary and
Christopher K. Breuer
Introduction 635
Heart valve function and structure 635
Cellular biology of the heart valve 636
Heart valve dysfunction and valvular repair
and remodeling 637
Heart valve replacement 638
The application of tissue engineering
toward the construction of a replacement
heart valve 640
Tissue engineering theory 640
Biomaterials and scaffolds 640
The search for appropriate cell sources 643
Cell seeding techniques 644
Bioreactors 645
Neotissue development in tissue engineered
heart valves 645
Clinical applications of the tissue engineered
heart valve 647
Conclusion and future directions 648
References 649
Part Ten
Endocrinology and metabolism 655
35. Generation of pancreatic islets
from stem cells 657
Ba´rbara Soria-Juan, Javier Lo´ pez-Beas,
Bernat Soria and Abdelkrim Hmadcha
Introduction 657
State-of-the-art 657
The challenge of making a β-cell 658
Recent achievements (first generation of
pancreatic progenitors used in the clinic) 658
Need of late maturation: cabimer protocol 659
Strategies to maintain cell viability 659
Encapsulation and tolerogenic strategies 661
The concept of cellular medicament 661
Conclusion 662
Acknowledgments 662
References 662
36. Bioartificial pancreas: challenges
and progress 665
Paul de Vos
Introduction 665
History of the bioartificial pancreas 666
Replenishable cell sources and encapsulation 666
Macro- or microedevices 667
Factors contributing to biocompatibility of
encapsulation systems 669
Avoiding pathogen-associated molecular
patterns in polymers 670
Natural and synthetic polymers 670
Multilayer capsule approaches 670
Antibiofouling approaches 671
Formation of polymer brushes 671
Immunomodulatory materials 672
Intracapsular environment and longevity
of the encapsulated islet graft 672
Concluding remarks and future
considerations 673
Acknowledgments 674
References 674
37. Thymus and parathyroid
organogenesis 681
Craig Scott Nowell, Kathy E. O’Neill, Paul Rouse,
Timothy Henderson, Ellen Rothman Richie,
Nancy Ruth Manley and Catherine Clare Blackburn
Structure and morphology of the thymus 681
Thymic epithelial cells 682
Complexity of the thymic epithelium
compartment 682
Functional diversity 683
In vitro T cell differentiation 683
Thymus organogenesis 685
Cellular regulation of early thymus
organogenesis 685
Origin of thymic epithelial cells 686
Thymic epithelial progenitor cells 686
Human thymus development 688
Cervical thymus in mouse and human 688
Molecular regulation of thymus and
parathyroid organogenesis 689
Molecular control of early organogenesis 689
Transcription factors and regulation of third
pharyngeal pouch outgrowth 691
Specification of the thymus and
parathyroid 692
Foxn1 and regulation of thymic epithelial cell
differentiation 695
Medullary development and expansion 696
Maintenance and regeneration of thymic
epithelial cells: Progenitor/stem cells in
the adult thymus 696
Strategies for thymus reconstitution 697
Summary 698
Acknowledgments 699
References 699
Part Eleven
Gastrointestinal system 707
38. Stem and progenitor cells of the
gastrointestinal tract: applications for
tissue engineering the intestine 709
Kathryn M. Maselli, Christopher R. Schlieve,
Mark R. Frey and Tracy C. Grikscheit
Introduction 709
Stem cells of the intestine 709
Cell types of the epithelial layer 709
Stem and progenitor cell types 710
Signaling pathways in the intestinal
epithelium 712
The Wnt pathway 712
The Notch pathway 713
Epidermal growth factor receptor/ErbB
signaling 713
The Hedgehog pathway 714
The BMP pathway 714
Tissue engineering the intestine with stem/
progenitor cells 714
Organ-specific stem cell progenitors versus
pluripotent stem cells 714
Synthetic and biological scaffolds 715
Primary intestinal-derived organoid units 716
Pluripotent stem cell approaches—human
intestinal organoids 717
Remaining barriers to the generation of
tissue-engineered intestine 718
Conclusion 718
Acknowledgment 718
References 718
39. Liver stem cells 723
Dagmara Szkolnicka and David C. Hay
Introduction 723
Liver architecture and function 723
Liver development 723
Fetal liver stem cells 724
Hepatocytes and liver progenitors in organ
regeneration 724
Molecular signaling and processes involved in
liver regeneration 724
Hepatocytes’ role in liver regeneration 725
Cholangiocytes and liver stem cells in liver
regeneration 725
Pluripotent stem cell_derived hepatoblasts
and hepatocytes 726
3D liver organoids and expansion 727
Pluripotent stem cell_derived liver
organoids 728
Bile duct_derived organoids 728
Hepatocyte-derived organoids 728
Novel scaffolds for liver organoids 729
Organoids as a model to study liver cancer
disease 730
Reprogramming of human hepatocytes to liver
progenitors using different culture
conditions 730
Conclusion 731
References 731
Further reading 736
40. Hepatic tissue engineering 737
Amanda X. Chen, Arnav Chhabra,
Heather E. Fleming and Sangeeta N. Bhatia
Liver disease burden 737
Current state of liver therapies 738
Extracorporeal liver support devices 738
Biopharmaceuticals 738
Liver transplantation 738
Hepatocyte transplantation 740
Current clinical trials 740
In vitro models 740
Two-dimensional liver culture 741
Three-dimensional liver constructs 741
Physiological microfluidic models of liver 742
Controlling three-dimensional architecture
and cellular organization 742
In vivo models 743
Cell sourcing 743
Cell number requirements 743
Immortalized cell lines 744
Primary cells 744
Fetal and adult progenitors 744
Reprogrammed hepatocytes 744
Extracellular matrix for cell therapies 744
Natural scaffold chemistry and modifications 745
Synthetic scaffold chemistry 745
Modifications in scaffold chemistry 745
Porosity 746
Vascular and biliary tissue engineering 746
Vascular engineering 746
Host integration 747
Biliary network engineering 747
Conclusion and outlook 747
References 748
Part Twelve
Hematopoietic system 755
41. Hematopoietic stem cells 757
Qiwei Wang, Yingli Han, Linheng Li and
Pengxu Qian
Introduction 757
Hematopoietic stem cells and hematopoietic
stem cells niche 757
Effects of biomaterials on hematopoietic stem
cells 758
Applications 759
Engineering hematopoietic stem cells niche
for in vitro expansion 759
Manipulation of the multilineage
differentiation of hematopoietic stem cells 760
In vivo tracking hematopoietic stem cells 761
Future perspectives 761
Acknowledgments 761
References 761
42. Blood components from
pluripotent stem cells 765
Erin A. Kimbrel and Robert Lanza
Introduction and history of modern
hematology 765
Red blood cells 765
Megakaryocytes/platelets 769
White blood cells 770
Lymphocytes—T cells 770
Lymphocytes—NK cells 773
Lymphocytes—NKT cells 775
Monocyte-derived dendritic cells 776
Monocyte-derived macrophages 777
Granulocytes—neutrophils 778
Perspectives 779
References 779
43. Red blood cell substitutes 785
Andre Francis Palmer and Donald Andrew Belcher
Introduction 785
Replicating red blood cell functions 785
Hemoglobin-based oxygen carriers 785
Hemoglobin toxicity 787
Oxygen delivery 789
Viscosity and colloid osmotic pressure 789
Cross-linked and polymeric hemoglobin 790
Surface conjugated hemoglobin 790
Encapsulated hemoglobin 791
Sources of hemoglobin 791
Recombinant hemoglobin 792
Erythrocruorins 792
Perfluorocarbons 793
Perspectives 794
Organ transplant preservation 794
Cancer treatment 795
Tissue-engineered construct oxygenation 795
References 795
Part Thirteen
Kidney and genitourinary system 803
44. Stem cells in kidney development
and regeneration 805
Kyle W. McCracken and Joseph V. Bonventre
Kidney development 805
Early embryonic origins of nephrogenic
tissues 806
Development of the nephric duct and
ureteric bud 808
Maintenance and differentiation of the
nephron progenitor cell 809
Role of stromal lineages in kidney
organogenesis 811
Nephron endowment 812
Kidney repair and regeneration 813
Stem cells in kidney repair 813
Sources of nephrogenic cells 814
Differentiation of renal tissue from
pluripotent stem cells (organoids) 815
Conclusion 817
Disclosures 818
Acknowledgements 818
References 818
45. Tissue engineering of the kidney 825
Ji Hyun Kim, Anthony Atala and James J. Yoo
Introduction 825
Cell-based tissue engineering of the kidney 826
Cell sources 826
Tissue-engineered cellular three-dimensional
renal constructs 830
Cell-free tissue engineering of the kidney 835
In situ kidney regeneration 835
Granulocyte-colony stimulating factor 835
Stromal cell_derived factor-1 837
Conclusion and future perspectives 837
Acknowledgment 838
References 838
46. Tissue engineering: bladder and
urethra 845
Yuanyuan Zhang, James J. Yoo and
Anthony Atala
Introduction 845
Cell sources 846
Bladder and ureter cells 846
Stem cell sources 846
Mechanism of cell therapy 848
Biodegradable biomaterials 850
Synthetic scaffolds 850
Natural collagen matrix 851
Preclinical models 854
Tissue regeneration models 854
Fibrotic bladder model 854
Clinical trials 856
Clinical translation 856
Clinical studies 857
Conclusion 858
References 858
47. Tissue engineering for female
reproductive organs 863
Renata S. Magalhaes, James K. Williams and
Anthony Atala
Introduction 863
Uterus 863
Acellular tissue engineering approaches
for uterine tissue repair 864
Cell-seeded scaffolds for partial uterine repair 864
Scaffold-free approaches for partial uterine
repair 865
Uterine cervix tissue engineering 865
Ovary 865
Tissue engineering ovarian follicles 866
Vagina 866
Tissue engineering approaches for neovagina
reconstruction 866
Conclusion and future perspectives 867
References 867
48. Male reproductive organs 871
Hooman Sadri-Ardekani, John Jackson and
Anthony Atala
Introduction 871
Testes 871
Spermatogonial stem cell technology 871
Androgen-replacement therapy 873
Ejaculatory system 874
Engineering vas deferens 874
Spinal ejaculation generator 875
Penis 875
Penile reconstruction 875
Penile transplantation 876
Stem cell therapy for erectile dysfunction 876
Conclusion 877
References 877
Part fourteen
Musculoskeletal system 881
49. Mesenchymal stem cells in
musculoskeletal tissue engineering 883
Yangzi Jiang, Dan Wang, Anna Blocki and
Rocky S. Tuan
Introduction 883
Mesenchymal stem cell biology relevant to
musculoskeletal tissue engineering 883
Mesenchymal stem cell identification 883
Tissue sources of mesenchymal stem cells 885
Mesenchymal stem cell isolation and in vitro
culture 886
Mesenchymal stem cell self-renewal and
proliferation capacity 887
Skeletogenic differentiation of mesenchymal
stem cells 888
Plasticity of mesenchymal stem cells 888
Mesenchymal stem cell heterogeneity 889
Mesenchymal stem cell effect on host
immunobiology 889
Safety of using mesenchymal stem cells for
transplantation 891
Mesenchymal stem cells in musculoskeletal
tissue engineering 891
Cartilage tissue engineering 891
General properties of articular cartilage 892
Cells for cartilage tissue engineering 892
Bone tissue engineering 897
Osteochondral tissue engineering 898
Engineering other skeletal tissues with
mesenchymal stem cells 899
Tendon/ligament 899
Meniscus 900
Gene therapy in musculoskeletal tissue
engineering 901
Conclusion and future perspectives 901
Acknowledgments 902
References 902
50. Bone tissue engineering and bone
regeneration 917
J.M. Kanczler, J.A. Wells, D.M.R. Gibbs, K.M.
Marshall, D.K.O. Tang and Richard O.C. Oreffo
Introduction 917
Skeletal stem cells 917
Fracture repair—the (limited) self-reparative
capacity of bone 919
A framework for bone repair:
biomaterial-driven strategies for bone
regeneration 922
Growth factors: biomimetic-driven strategies
for bone regeneration 923
Bone biofabrication 924
Development of vascular bone 925
Preclinical development—ex vivo/in vivo
small and large animal preclinical models 926
Clinical translation 929
Summary and future perspectives 931
Acknowledgments 931
References 931
51. Tissue engineering for regeneration
and replacement of the intervertebral
disk 937
Stephen R. Sloan Jr., Niloofar Farhang, Josh Stover,
Jake Weston, Robby D. Bowles and Lawrence J.
Bonassar
Introduction 937
Intervertebral disk structure and function 938
Cell-biomaterial constructs for intervertebral
disk regeneration 940
Nucleus pulposus cell-biomaterial implants 940
Annulus fibrosus repair and regeneration 942
Composite cell-biomaterial intervertebral disk
implants 944
Cellular engineering for intervertebral disk
regeneration 945
Cell therapy preclinical studies 946
Cell therapy clinical studies 947
Growth factors and other biologics for
intervertebral disk regeneration 948
In vitro studies 948
In vivo studies: growth factors 952
In vivo studies: other biologics 953
Gene therapy for intervertebral disk
regeneration 953
Gene transfer studies: viral 954
Gene transfer studies: nonviral 954
Endogenous gene regulation 955
Gene therapy in summary 955
In vivo preclinical models for intervertebral
disk regeneration and replacement 955
Concluding remarks 957
Acknowledgment 957
References 957
52. Articular cartilage injury 967
J.A. Martin, M. Coleman and J.A. Buckwalter
Introduction 967
Articular cartilage injury and joint
degeneration 968
Mechanisms of articular cartilage injuries 968
Response of articular cartilage to injury 970
Matrix and cell injuries 970
Chondral injuries 971
Osteochondral injuries 971
Preventing joint degeneration following injury 972
Promoting articular surface repair 972
Penetration of subchondral bone 972
Periosteal and perichondrial grafts 973
Cell transplantation 973
Artificial matrices 973
Growth factors 973
Antiinflammatories 974
Conclusion 974
Acknowledgments 974
References 974
Further reading 977
53. Engineering cartilage and other
structural tissues: principals of bone
and cartilage reconstruction 979
Batzaya Byambaa and Joseph P. Vacanti
Introduction 979
Biomaterials for cartilage tissue engineering 979
Cell sources for cartilage tissue engineering 980
Biofabrication of cartilage tissue 981
Magnetic resonance imaging and
computerized tomography scans 981
Scaffolds for cartilage tissue engineering 981
Bioprinting techniques for fabrication of
cartilage constructs 982
Bioinks for cartilage tissue printing 982
Osteochondral tissue engineering 985
References 985
54. Tendon and ligament tissue
engineering 989
Spencer P. Lake, Qian Liu, Malcolm Xing,
Leanne E. Iannucci, Zhanwen Wang and
Chunfeng Zhao
Introduction 989
Tendon and ligament composition, structure,
and function 990
Composition 990
Structure 990
Function 990
Requirements for a tissue-engineered
tendon/ligament 991
Scaffold 992
Cell 994
Bioactive factors 995
Three-dimensional bioprinting and bioink 996
Bioink inspired from ligament and tendon
structures 997
Tissue engineering tendon and ligament in
clinical application 998
Summary 999
References 1000
55. Skeletal tissue engineering 1007
Matthew P. Murphy, Mimi R. Borrelli,
Daniel T. Montoro, Michael T. Longaker and
Derrick C. Wan
Introduction 1007
Distraction osteogenesis 1008
Critical-sized defects 1010
Cellular therapy 1010
Cytokines 1013
Scaffolds 1014
Tissue engineering in practice 1016
Conclusion 1017
References 1017
Part Fifteen
Nervous system 1023
56. Brain implants 1025
Lars U. Wahlberg
Introduction 1025
Cell replacement implants 1025
Primary tissue implants 1025
Cell line implants 1027
Cell protection and regeneration implants 1028
Cell implants secreting endogenous factors 1028
Cell implants secreting engineered factors
(ex vivo gene therapy) 1029
Encapsulated cell brain implants 1029
Controlled-release implants 1030
Combined replacement and regeneration
implants 1030
Disease targets for brain implants 1031
Surgical considerations 1032
Conclusion 1032
References 1032
57. Brain_machine interfaces 1037
Jose´ del R. Milla´n and Serafeim Perdikis
Introduction 1037
Brain_machine interface signals 1037
Voluntary activity versus evoked potentials 1038
Mutual learning 1040
Context-aware brain_machine interface 1040
Future directions 1041
References 1042
58. Spinal cord injury 1047
Nicolas N. Madigan and Anthony J. Windebank
Introduction 1047
Epidemiology 1047
Spinal cord organization 1047
Spinal cord injury 1048
Available clinical interventions 1049
The continuum of physical, cellular, and
molecular barriers to spinal cord
regeneration 1049
The role of tissue engineering in spinal cord
injury repair 1051
Bioengineering for integrated spinal cord
biocompatibility 1052
Animal models of spinal cord injury 1052
Principles of biomaterial fabrication for
spinal cord injury repair 1054
Biomaterials for spinal cord tissue
engineering: natural polymers 1058
Extracellular matrix polymers 1058
Polymers from marine or insect life 1065
Polymers derived from the blood 1071
Biomaterials for spinal cord tissue
engineering: synthetic polymers 1072
Poly α-hydroxy acid polymers 1073
Nonbiodegradable hydrogels 1077
Conclusion and future directions:
the promise of clinical translation 1080
References 1080
59. Protection and repair of hearing 1093
Su-Hua Sha, Karl Grosh and Richard A. Altschuler
Introduction 1093
Protection from “acquired” sensory hair cell
loss 1093
Oxidative stress and stress-related
mitochondrial pathways 1094
Calcium influx 1094
Endoplasmic reticulum stress 1094
Prevention of ototoxicity 1094
Prevention of acoustic trauma 1096
Antiinflammatory agents 1097
Heat shock proteins 1097
Neurotrophic factors 1098
Protection from excitotoxicity: “acquired”
loss of auditory nerve connections to hair
cells 1098
Gene transfer for the prevention and
treatment of genetic deafness 1099
Interventions for hair cell repair: gene
therapy for transdifferentiation 1099
Interventions for repair: hair cell and
auditory nerve replacement—exogenous
stem cells 1101
Interventions for repair/replacement:
cochlear prostheses 1101
Fully implantable cochlear prostheses 1101
Interventions for repair/replacement: central
auditory prostheses 1102
Local delivery to cochlear fluids 1103
Conclusion 1103
Acknowledgments 1103
References 1104
Further reading 1112
Part Sixteen
Ophthalmic 1113
60. Stem cells in the eye 1115
Chao Huang, Julie Albon, Alexander Ljubimov
and Maria B. Grant
Introduction 1115
Endogenous ocular stem cells 1115
Corneal stem cells 1115
Stromal stem cells 1119
Endothelial stem cells 1119
Conjunctival epithelial stem cells 1120
The bioengineered cornea 1120
Retinal progenitor cells 1120
Mu¨ ller stem cells 1121
Retinal pigment epithelium stem cells 1121
Nonocular stem cells 1121
Induced pluripotent stem cells (iPSCs) 1121
Embryonic stem cells/iPSCs in retinal
regeneration 1121
Bone marrow stem cells 1124
References 1126
61. Corneal replacement tissue 1135
Maria Mirotsou, Masashi Abe and Robert Lanza
Introduction 1135
Corneal anatomy and structure 1135
Epithelium 1136
Stroma 1138
Endothelium 1139
Conclusion 1140
References 1141
62. Retinal degeneration 1145
Erin A. Kimbrel and Robert Lanza
Epidemiology of visual impairment and
blindness 1145
Structure/function of the retina and cell types
affected in retinal degenerative diseases 1145
Age-related macular degeneration 1147
History of retinal pigment epithelium as a
cellular therapy for age-related macular
degeneration 1147
Retinal pigment epithelium from pluripotent
stem cells 1149
Retinitis pigmentosa 1150
Photoreceptors from pluripotent stem cells 1151
Glaucoma 1153
Stem cell_based therapies to treat glaucoma 1154
Diabetic retinopathy 1155
Stem cell_based therapies to treat diabetic
retinopathy 1155
Future directions and competing therapies 1156
References 1157
63. Vision enhancement systems 1163
Gislin Dagnelie, H. Christiaan Stronks and
Michael P. Barry
Introduction 1163
Visual system, architecture, and (dys)function 1163
Current- and near-term approaches to vision
restoration 1166
Enhancing the stimulus through
optoelectronic and optical means 1166
Visual prostheses based on electrical tissue
stimulation 1167
Retinal cell transplantation 1170
Optic nerve protection and regeneration 1171
Drug delivery 1172
Genetic interventions 1172
Emerging application areas for engineered
cells and tissues 1173
Photosensitive structures 1174
Optogenetics 1174
Outer retinal cell transplantation 1177
Cell matrices supporting axonal regrowth 1177
Repopulating ischemic or diabetic retina 1178
Assessing the functional outcomes of novel
retinal therapies 1178
Conclusion: toward 2020 vision 1179
Acknowledgment 1179
References 1179
Further reading 1183
Part Seventeen
Oral/Dental applications 1185
64. Biological tooth replacement and
repair 1187
Anthony J. (Tony) Smith and Paul T. Sharpe
Introduction 1187
Tooth development 1187
Whole tooth-tissue engineering 1189
Stem cell-based tissue engineering of teeth 1189
Bioteeth from cell-seeded scaffolds 1189
Root formation 1190
Cell sources 1191
Dental-tissue regeneration 1191
Natural tissue regeneration 1191
Importance of the injury-regeneration
balance 1192
Signaling events in dental regeneration 1193
Control of specificity of dental-tissue
regeneration 1193
Dental postnatal stem cells 1194
Directed tissue regeneration 1195
Signaling-based strategies 1195
Cell- and gene-based strategies 1196
Conclusion 1197
References 1197
65. Tissue engineering in oral and
maxillofacial surgery 1201
Simon Young, F. Kurtis Kasper, James Melville,
Ryan Donahue, Kyriacos A. Athanasiou,
Antonios G. Mikos and Mark Eu-Kien Wong
Introduction 1201
Special challenges in oral and maxillofacial
reconstruction 1201
Current methods of oral and maxillofacial
reconstruction 1204
Mandibular defects 1205
Maxillary defects 1207
Relevant strategies in oral and maxillofacial
tissue engineering 1208
Bone applications 1208
Cartilage applications 1212
Oral mucosa applications 1214
Composite tissue applications 1215
Animal models 1215
The future of oral and maxillofacial tissue
engineering 1216
References 1216
66. Periodontal tissue engineering and
regeneration 1221
Xiao-Tao He, Rui-Xin Wu and Fa-Ming Chen
Introduction 1221
Stem cells for periodontal bioengineering 1222
Intraoral mysenchymal stem cells 1222
Periodontal tissue_derived stem cells 1223
Stem cells from apical papilla 1224
Dental follicle stem cells 1224
Hertwig’s epithelial root sheath 1225
Stem cells from dental pulp or exfoliated
deciduous teeth 1225
Extraoral mysenchymal stem cells 1225
Bone marrow_derived mysenchymal stem
cells 1225
Adipose-derived stem cells 1226
Selection of cell types 1226
Signaling molecules 1227
Types of signals 1228
Crucial delivery barriers to progress 1230
Gene delivery as an alternative to growth
factor delivery 1231
Scaffolding and biomaterials science 1232
Requirements of cell scaffolds 1232
Biomaterial-based immune modulation 1233
Classes of biomaterials 1233
Biomaterial redesign for periodontal
application 1235
Periodontal bioengineering strategies 1236
Cell-free approaches 1237
Cell-based approaches 1239
Challenges and future directions 1242
Closing remarks 1243
Acknowledgments 1243
References 1243
Part Eighteen
Respiratory system 1251
67. Cell- and tissue-based therapies
for lung disease 1253
Jeffrey A. Whitsett, William Zacharias,
Daniel Swarr and Vladimir V. Kalinichenko
Introduction: challenges facing cell and
tissue-based therapy for the treatment of
lung disease 1253
Lung morphogenesis informs the process of
regeneration 1254
Integration and refinement of signaling and
transcriptional pathways during lung
formation 1256
The mature lung consists of diverse
epithelial and mesenchymal cell types 1256
Structure and function of pulmonary
vasculature 1257
Embryonic development of alveolar
capillaries 1258
Evidence supporting lung regeneration 1259
A diversity of lung epithelial progenitor/stem
cells is active during regeneration 1260
Role of lung microvasculature in lung repair 1262
Endothelial progenitor cells in lung repair 1262
Pulmonary cell-replacement strategies for
lung regeneration 1263
Induced pluripotent stem cells for study of
treatment of pulmonary disease 1263
Differentiation of induced pluripotent stem
and embryonic stem cells to pulmonary
epithelial cell lineages 1264
Bioengineering of lung tissues 1265
Mesenchymal stromal cells and mesenchymal
stromal cell products for the treatment of
lung disease 1265
Important role of the extracellular matrix in
lung structure and repair 1265
Tissue engineering for conducting airways 1266
Pulmonary macrophage transplantation for the
treatment of interstitial lung disease 1266
Conclusion 1266
Acknowledgments 1266
References 1266
68. Lung tissue engineering 1273
Micha Sam Brickman Raredon, Yifan Yuan
and Laura E. Niklason
Introduction 1273
Design criteria for pulmonary engineering 1273
Decellularized scaffolds and biofabrication
approaches 1274
Pulmonary epithelial engineering 1276
Proximal airway engineering 1276
Distal airway engineering 1276
Mesenchymal support of pulmonary
epithelium 1277
Pulmonary endothelial engineering 1277
Endothelial cell sources for lung tissue
engineering 1278
Endothelial seeding into lung scaffolds 1278
Organomimetic endothelial culture 1279
Mesenchymal support of pulmonary
microvasculature 1280
Bioreactor technologies for pulmonary
engineering 1280
Conclusion 1281
References 1281
Part Nineteen
Skin 1287
69. Cutaneous epithelial stem cells 1289
Denise Gay, Maksim V. Plikus, Iris Lee, Elsa
Treffeisen, Anne Wang and George Cotsarelis
Introduction 1289
Interfollicular epidermal stem cells 1289
Models for skin renewal: epidermal
proliferative unit versus committed
progenitor 1290
Hair follicle stem cells 1291
The bulge as stem cell source 1291
Defining characteristics of the bulge as a
stem cell source 1292
Multiple hair follicle stem cell subpopulations
by marker expression 1294
Stem cells of other ectodermal appendages 1295
Sebaceous glands 1295
Sweat glands 1296
Nails 1296
Hair follicle stem cells in skin homeostasis,
wound healing, and hair regeneration 1297
Homeostasis 1297
Wound healing 1297
Wound-induced hair follicle neogenesis and
regeneration 1298
Epithelial stem cells in aging 1298
Role of stem cells in alopecia 1299
Skin as an active immune organ 1300
Cross talk between hair follicles and the
immune system 1300
The inflammatory memory of skin cells 1301
Tissue engineering with epidermal stem cells 1301
Epidermal stem cells as a therapy: the future 1302
Conclusion 1302
References 1302
70. Wound repair: basic biology to
tissue engineering 1309
Richard A.F. Clark, Michael Musillo and
Thomas Stransky
Introduction 1309
Basic biology of wound repair 1310
Inflammation 1310
Transition from inflammation to
repair 1310
Reepithelialization 1310
Granulation tissue 1312
Wound contraction and extracellular matrix
organization 1316
Chronic wounds 1317
Scarring 1318
Pathological scars 1318
Scarless healing 1319
Tissue engineered therapy with skin cells 1320
Engineered epidermal constructs 1320
Engineered dermal constructs 1321
Engineered skin substitutes 1321
Skin autograft harvesting without scarring 1322
Tissue-engineered therapy with stem cells,
bioactives, and biomaterials 1322
References 1324
71. Bioengineered skin constructs 1331
Vincent Falanga
Introduction 1331
Skin structure and function 1331
The epidermis 1331
The dermis 1332
The process of wound healing 1333
Impaired healing and its mechanisms 1333
Acute versus chronic wound healing 1333
Bacterial colonization 1333
Growth factor imbalances 1334
Matrix metalloproteinase activity 1334
Moist wound healing in chronic wounds 1334
Ischemia 1334
Abnormalities at the cellular level 1335
Engineering skin tissue 1335
Design considerations 1335
Commercial considerations 1336
Process considerations 1337
Regulatory considerations 1337
Immunological considerations 1338
Summary: engineering skin tissue 1338
Epidermal regeneration 1338
Dermal replacement 1339
Bioengineered living skin equivalents 1339
Bioengineered skin: FDA-approved
indications 1340
Cutaneous indications 1340
Oral indications 1341
Apligraf and Dermagraft: off-label uses 1341
The importance of wound bed preparation 1344
Proposed mechanisms of action of
bioengineered skin 1345
Construct priming and a new didactic
paradigm for constructs 1347
Other considerations 1348
Conclusion 1348
References 1349
Further reading 1352
Part Twenty
Tissue-engineered food 1353
72. Principles of tissue engineering for
food 1355
Mark Post and Cor van der Weele
Introduction 1355
Why tissue engineering of food? 1355
Specifics of tissue engineering for medical
application 1356
Uniqueness 1356
Function 1356
Skeletal muscle and fat tissue
engineering 1357
Tissue engineering of skeletal muscle 1357
Tissue engineering of fat 1359
Specifics of food tissue engineering 1361
Scale 1361
Efficiency 1362
Taste, texture, juiciness 1362
Enhanced meat 1363
Other foods 1363
Consumer acceptance 1364
Regulatory pathway 1365
Conclusion 1365
References 1365
73. Cultured meat—a humane meat
production system 1369
Zuhaib F. Bhat, Hina Bhat and Sunil Kumar
Introduction 1369
Need and advantages of cultured meat 1370
Cultured meat 1372
Scaffolding techniques 1372
Self-organizing tissue culture 1373
Organ printing 1375
Biophotonics 1375
Nanotechnology 1375
Challenges and requirements for industrial
production 1375
Generation of suitable stem cell lines from
farm-animal species 1376
Safe media for culturing of stem cells 1377
Safe differentiation media to produce muscle
cells 1377
Tissue engineering of muscle fibers 1378
Scaffolds 1378
Industrial bioreactors 1379
Fields 1380
Atrophy and exercise 1380
Senescence 1381
Meat processing technology 1381
Associated dangers and risks 1381
Regulatory issues 1381
Consumer acceptance and perception 1382
Role of media in publicity of cultured meat 1382
Market for cultured meat 1382
Conclusion 1383
References 1384
Part Twentyone
Emerging technologies 1389
74. Three-dimensional bioprinting for
tissue engineering 1391
Jun Tae Huh, James J. Yoo, Anthony Atala and
Sang Jin Lee
Introduction 1391
3D Bioprinting strategy: from medical
image to printed bioengineered tissue 1391
Three-dimensional bioprinting techniques 1392
Jetting-based bioprinting 1392
Extrusion-based bioprinting 1394
Laser-assisted bioprinting 1394
Laser-based stereolithography 1395
Digital light processing 1395
Hybrid and other techniques 1396
Biomaterials as bioinks for three-dimensional
bioprinting 1396
Hydrogel-based bioinks for cell-based
three-dimensional bioprinting 1396
Biodegradable synthetic polymers for
structure-based three-dimensional
bioprinting 1399
Scaffold-free cell printing 1399
Three-dimensional bioprinting in tissue
engineering applications 1400
Three-dimensional bioprinted vascular
structures 1400
In vitro tissue models 1400
Three-dimensional bioprinted implantable
tissue constructs 1403
Conclusion and future
perspectives 1409
Abbreviations 1410
Glossary 1410
References 1411
75. Biofabricated three-dimensional
tissue models 1417
David B. Berry, Claire Yu and Shaochen Chen
Introduction 1417
Current methods of three-dimensional
biofabrication 1418
Biomaterials for three-dimensional
fabrication 1421
Three-dimensional tissue models for drug
screening, disease modeling, therapeutics,
and toxicology 1425
Conclusion and future directions 1435
Acknowledgments 1435
References 1435
76. Body-on-a-chip: three-dimensional
engineered tissue models 1443
Thomas Shupe, Aleksander Skardal and Anthony
Atala
Introduction 1443
Advanced in vitro modeling
systems—progression from two-dimensional
to three-dimensional models 1444
Organ-on-a-chip technologies and their
applications 1445
Microengineering and biofabrication 1446
Liver-on-a-chip 1447
Vessel-on-a-chip 1447
Lung-on-a-chip 1448
Heart-on-a-chip 1448
Cancer-on-a-chip 1448
Body-on-a-chip: integrated multiorgan
systems and future applications 1449
The importance of multiorganoid integration 1449
Cutting edge body-on-a-chip: the first highly
functional multiorganoid systems 1452
Conclusion and perspectives 1455
References 1456
77. Monitoring and real-time control of
tissue engineering systems 1459
Jean F. Welter and Harihara Baskaran
Introduction 1459
Current state-of-the-art 1460
General environmental monitoring and
real-time control 1460
Tissue-level monitoring 1462
Mechanical properties 1462
Cell-level monitoring 1463
Reporter-based gene expression imaging 1463
Tissue-specific 1463
Cartilage monitoring and real-time control 1463
Skin 1464
Concluding remarks 1464
Acknowledgments 1465
References 1465
78. Biomanufacturing for regenerative
medicine 1469
Joshua G. Hunsberger and Darren H.M. Hickerson
Current landscape of biomanufacturing 1469
Highlighting current workflows for
biomanufacturing 1470
Current challenges in biomanufacturing for
regenerative medicine 1470
Current platform technologies enabling
biomanufacturing 1472
Regulatory challenges for biomanufacturing 1473
Food and Drug Administration guidance
documents 1474
Creating standards 1475
The future: envisioned advanced
biomanufacturing 1476
Closed-modular biomanufacturing systems 1476
Off-the-shelf products 1477
Preservation advances 1477
Synthetic biology advances 1477
Cell banking advances 1477
Medical applications for biomanufacturing in
regenerative medicine 1477
Space exploration 1478
References 1479
Part Twentytwo
Clinical experience 1481
79. Tissue-engineered skin products 1483
Jonathan Mansbridge
Introduction 1483
Types of therapeutic tissue-engineered skin
products 1484
Components of tissue-engineered skin
grafts as related to function 1484
Scaffold 1484
Keratinocytes 1485
Fibroblasts 1485
Extracellular matrix 1485
Subcutaneous fat 1485
Components of the immune system 1486
Melanocytes 1486
Adnexal structures 1487
Commercial production of tissue-engineered
skin products 1487
Regulation 1487
Product development 1487
Overall concept 1487
Allogeneic cell source 1488
Viability of product and avoidance of a final
sterile fill 1488
Shelf life 1488
Size, user convenience 1489
The manufacture of Dermagraft and
TransCyte 1489
Cells 1489
Medium 1489
Bioreactor design 1490
The Dermagraft and TransCyte production
processes 1490
Release specifications 1491
Distribution and cryopreservation 1491
Problems with commercial culture for tissue
engineering 1492
Clinical trials 1492
Immunological properties of
tissue-engineered skin 1493
Commercial success 1494
Mechanism of action 1494
Future developments 1495
Conclusion 1496
References 1496
80. Tissue-engineered cartilage
products 1499
Henning Madry
Introduction 1499
Cartilage defects, osteoarthritis, and
reconstructive surgical options 1499
Cartilage defects pathophysiology 1499
Surgical treatment options for articular
cartilage defects 1500
Tissue-engineered cartilage products for
orthopedic reconstruction 1500
Cells for tissue-engineered cartilage repair 1500
Scaffolds for clinical tissue-engineered
cartilage repair 1501
Collagen scaffolds 1501
Hyaluronan 1502
Synthetic polymers 1502
Agarose and alginate 1502
Scaffold-free three-dimensional systems 1502
Bioreactors for tissue-engineered cartilage
repair 1502
Clinical nomenclature of scaffold-based
techniques 1503
Clinical generations of autologous
chondrocyte implantation 1503
Acellular, scaffold-based products 1503
Particulated autologous or allogenic articular
cartilage 1503
Commercial autologous chondrocyte
implantation products 1503
MACI (Vericel, Cambridge, MA,
United States) 1503
ChondroCelect (TiGenix, Leuven, Belgium) 1504
Spherox (Co.don, Berlin, Germany) 1504
Novocart 3D (Tetec, Reutlingen, Germany) 1504
BioSeed C (Biotissue, Geneva, Switzerland) 1504
Novocart Inject (Tetec, Reutlingen,
Germany) 1504
Chondron (Sewon Cellontech, Seoul, Korea) 1505
Cartipatch (Tissue Bank of France, Ge´nie
Tissulaire, Lyon, France) 1505
CARTISTEM (Medipost, Seongnam, Korea) 1505
Clinical application of autologous chondrocyte
implantation in reconstructive articular
cartilage surgery 1505
Indications for autologous chondrocyte
implantation 1505
Contraindications 1505
Surgical steps 1506
Clinical results of autologous chondrocyte
implantation 1506
Overview 1506
Data from prospective randomized clinical
trials 1507
Long-term results of autologous chondrocyte
implantation 1508
Clinical factors affecting the clinical
outcomes of autologous chondrocyte
implantation 1508
Conflict of interest 1509
References 1509
81. Bone tissue engineering 1511
Hani A. Awad, Regis J. O’Keefe and Jeremy J. Mao
Introduction 1511
Conventional bone tissue engineering
strategies: cells, scaffolds, and biofactors 1511
Delivery of molecules and/or scaffolds to
augment endogenous bone regeneration 1512
Biomaterials development and
three-dimensional printing 1513
Clinical successes and opportunities in
regenerative repair of craniofacial defects 1516
Conclusion 1517
Acknowledgments 1517
References 1517
82. Tissue-engineered cardiovascular
products 1521
Doris A. Taylor, Camila Hochman-Mendez, Joern
Huelsmann, Abdelmotagaly Elgalad and
Luiz C. Sampaio
Clinical situation/reality 1521
Considerations for tissue-engineered
cardiovascular constructs 1521
Components for tissue-engineered
cardiovascular constructs 1521
Cell sources 1521
Scaffolds 1524
Tissue-engineered cardiovascular constructs 1525
Vascular grafts 1525
Valves 1526
Cardiac patches 1527
Building the next level of complexity: whole
heart 1529
Pathway to approval and commercialization 1530
Future perspectives 1532
References 1532
83. Tissue organoid models and
applications 1537
Timothy S. Leach, Anthony Dominijanni,
Sean V. Murphy and Anthony Atala
Introduction 1537
Cell sources 1537
Types of organoid models 1538
Cardiac organoid 1539
Liver organoid 1540
Brain organoid 1540
Lung organoid 1541
Gastrointestinal tract organoid 1541
Other organoid models 1542
Applications 1542
Tumor and disease models 1542
Drug analysis 1543
Organ-on-a-chip 1544
Developmental biology 1544
Conclusion 1545
References 1545
Part Twenty three
Regulation, commercialization
and ethics 1551
84. The regulatory process from concept to
market 1553
Kyung Eun Sung, Judith Arcidiacono, Donald W.
Fink Jr., Andrea Gray, Johnny Lam, Winson Tang,
Iwen Wu and Raj K. Puri
Introduction 1553
Regulatory background 1553
Overview of development and approval
process 1554
Early-stage development 1554
Chemistry, manufacturing, and controls 1555
Pharmacology and toxicology 1555
Clinical 1556
US Food and Drug Administration/sponsor
meetings 1557
Submitting an investigational new drug
application 1557
Required US Food and Drug Administration
forms 1557
Investigational new drug application
contents 1558
US Food and Drug Administration review
of an original investigational new drug
application submission 1559
Later-stage development topics 1559
Compliance with current good
manufacturing practice 1559
Product readiness for Phase 3 1559
Potency assay 1560
Pharmacology and toxicology 1560
Phase 3 clinical development 1560
Combination products 1561
Tissue-engineered and regenerative medicine
products 1562
3D bio-printed tissue-engineered/
regenerative-medicine products 1563
Medical devices 1563
Least burdensome principles 1563
Breakthrough device program 1563
Evaluation of devices used with regenerative
medicine advanced therapy 1564
Expedited review programs 1564
Other regulatory topics 1565
Minimal manipulation and homologous
use of human cells, tissues, and cellular and
tissue-based products 1565
Clinical research involving children 1566
Expanded access to investigational drugs for
treatment use 1566
Charging for investigational drugs under an
investigational new drug application 1566
Responsibilities of sponsors and investigators 1566
Clinical research conducted outside of the
United States 1568
Use of standards 1568
US Food and Drug Administration
international regulatory activities 1568
The role of cell-based products in medical
product testing 1568
Conclusion 1568
Acknowledgments 1568
Appendix I: Code of Federal Regulations
citations relevant to cellular product
development 1569
Appendix II: The list of acronyms 1569
References 1570
85. Business issues 1573
Matthew Vincent
Introduction 1573
The aging population 1573
Rise of regenerative medicine 1575
Product development 1577
Embryonic stem cells 1578
Induced pluripotent stem cells 1579
Direct reprogramming of differentiated cells 1580
Small molecule-induced differentiation 1580
Reimbursement 1580
Conclusion 1582
References 1582
86. Ethical issues 1585
Laurie Zoloth
Introduction 1585
Duty and healing: natural makers in a broken
world 1587
To make is to know: notes on an old problem
about knowledge 1587
What is a thing? The perils of deconstruction 1588
What contextual factors should be taken into
account, and do any of these prevent the
development and use of the technology? 1588
What purposes, techniques, or applications
would be permissible and under what
circumstances? 1589
On what procedures and structures, involving
what policies, should decisions on
appropriate techniques and uses be based? 1590
Conclusion 1590
References 1590
Index 1593