Robert A. Brown is Professor of Tissue Engineering and Director of the Centre for Tissue Regeneration Science at University College London, UK. He is also co-ordinator of the London Tissue Engineering Consortium (Tissue Bioreactor Science) and the British Tissue Engineering Network (BRITE Net), as well as current President of the Tissue and Cell Engineering Society (TCES). Professor Brown has published over 180 peer-reviewed publications and 18 patents/applications, collaborating across industry and academia to promote interdisciplinary research in Tissue Engineering and Regenerative Medicine

• Preface: Extreme Tissue Engineering – a User’s Guide xi1 Which Tissue Engineering Tribe Are You From? 11.1 Why do we need to engineer tissues at all? 11.1.1 Will the real tissue engineering and regenerative medicine please stand up? 21.1.2 Other people’s definitions 31.1.3 Defining our tissue engineering: fixing where we are on the scale-hierarchy 41.2 Bio-integration as a fundamental component of engineering tissues 71.2.1 Bio-scientists and physical scientists/engineers: understanding diversity in TERM 81.3 What are the ‘tribes’ of tissue engineering? 101.3.1 Special needs for special characteristics: why is networking essential for TERM? 131.4 Surprises from tissue engineering (Veselius to Vacanti) 161.5 So really is there any difference between tissue engineering and regenerative medicine? 201.5.1 Questions never really asked: repair versus regeneration? 201.5.2 Understanding the full spectrum: tissue replacement repair and regeneration 231.6 Conclusions 271.7 Summarizing definitions 28Annex 1 Other people’s definitions of tissue engineering 29Annex 2 Other people’s definitions of regenerative medicine 30Further reading 302 Checking Out the Tissue Groupings and the Small Print 332.1 Checking the small print: what did we agree to engineer? 332.2 Identifying special tissue needs problems and opportunities 372.3 When is ‘aiming high’ just ‘over the top’? 392.4 Opportunities risks and problems 412.4.1 Experimental model tissues (as distinct from spare-parts and fully regenerated tissues) 412.4.2 The pressing need for 3D model tissues 422.4.3 Tissue models can be useful spin-offs on the way to implants 422.5 Special needs for model tissues 442.5.1 Cell selection: constancy versus correctness 442.5.2 Support matrices – can synthetics fake it? 452.5.3 Tissue dimensions: when size does matter! 462.6 Opportunities and sub-divisions for engineering clinical implant tissues 462.6.1 Making physiological implants: spare parts or complete replacement? 472.6.2 Making pathological and aphysiological constructs: inventing new parts and new uses 472.6.3 Learning to use the plethora of tissue requirements as an opportunity 482.7 Overall summary 49Further reading 493 What Cells ‘Hear’ When We Say ‘3D’ 513.1 Sensing your environment in three dimensions: seeing the cues 513.2 What is this 3D cell culture thing? 543.3 Is 3D for cells more than a stack of 2Ds? 553.4 On in and between tissues: what is it like to be a cell? 583.5 Different forms of cell-space: 2D 3D pseudo-3D and 4D cell culture 623.5.1 What has ‘3D’ ever done for me? 623.5.2 Introducing extracellular matrix 633.5.3 Diffusion and mass transport 653.5.4 Oxygen mass transport and gradients in 3D engineered tissues: scaling Mount Doom 663.6 Matrix-rich cell-rich and pseudo-3D cell cultures 693.7 4D cultures – or cultures with a 4th dimension? 713.8 Building our own personal understanding of cell position in its 3D space 733.9 Conclusion 75Further reading 754 Making Support-Scaffolds Containing Living Cells 774.1 Two in one: maintaining a synergy means keeping a good duet together 774.2 Choosing cells and support-scaffolds is like matching carriers with cargo 784.3 How like the ‘real thing’ must a scaffold be to fool its resident cells? 804.4 Tissue prosthetics and cell prosthetics – what does it matter? 834.5 Types of cell support material for tissue engineering – composition or architecture? 854.5.1 Surface or bulk – what does it mean to the cells? 854.5.2 Bulk material breakdown and the local ‘cell economy’ 854.6 Three generic types of bulk composition for support materials 864.6.1 Synthetic materials for cell supports 884.6.2 Natural native polymer materials for cell supports 904.6.3 Hybrids: composite cell support materials having synthetic and natural components 984.7 Conclusions 100Further reading 1015 Making the Shapes for Cells in Support-Scaffolds 1035.1 3D shape and the size hierarchy of support materials 1045.2 What do we think ‘substrate shape’ might control? 1065.3 How we fabricate tissue structures affects what we get out in the end: bottom up or top down? 1075.4 What shall we seed into our cell-support materials? 1105.4.1 Cell loading: guiding the willing bribing the reluctant or trapping the unwary? 1115.4.2 Getting cells onto/into pre-fabricated constructs (the willing and the reluctant) 1145.4.3 Trapping the unwary: Seeding cells into self-assembling gel-forming materials 1155.5 Acquiring our cells: recruiting the enthusiastic or press-ganging the resistant 1185.5.1 From cell expansion to selection and differentiation 1215.6 Cargo crew or stowaway? 1245.6.1 Crew-type cells: helping with the journey 1245.6.2 Cargo-type cells: building the bulk tissue 1255.6.3 Stowaway or ballast-type cells 1285.7 Chapter summary 128Further reading 1296 Asymmetry: 3D Complexity and Layer Engineering – Worth the Hassle? 1316.1 Degrees of tissue asymmetry 1336.2 Making simple anisotropic/asymmetrical structures 1346.3 Thinking asymmetrically 1376.4 How do we know which scale to engineer first? 1406.5 Making a virtue of hierarchical complexity: because we have to 1446.6 Cell-layering and matrix-layering 1476.7 No such thing as too many layers: theory and practice of tissue layer engineering 1516.7.1 Examples of layer engineering 1536.8 Other forms of tissue fabrication in layers and zones 1586.8.1 Section summary 1586.9 Familiar asymmetrical construction components: everyday ‘layer engineering’ 1596.10 Summary 1607 Other Ways to Grow Tissues? 1637.1 General philosophies for repair replacement and regeneration 1637.1.1 What does reconstructive surgery have to teach us? 1657.1.2 Clues from the natural growth of tissues 1667.2 What part of grow do we not understand? 1677.2.1 Childhood growth of soft connective tissues: a good focus? 1697.2.2 Mechanically induced ‘growth’ of tissues in children 1707.2.3 Mechanically induced ‘growth’ of adult tissue 1717.2.4 Growth has a mirror image – ‘ungrowth’ or shrinkage-remodelling 1727.3 If growth and ungrowth maintain a tensional homeostasis what are its controls? 1737.3.1 Tension-driven growth and tensional homeostasis – the cell’s perspective? 1747.3.2 Mechanically reactive collagen remodelling – the ‘constant tailor’ theory 1777.4 Can we already generate tension-driven growth in in vivo tissue engineering? 1787.4.1 Mechanical loading of existing tissues 1787.5 Conclusions: what can we learn from engineered growth? 179Appendix to Chapter 7 179Further reading 1828 Bioreactors and All That Bio-Engineering Jazz 1858.1 What are ‘tissue bioreactors’ and why do we need them? 1868.1.1 Rumblings of unease in the smaller communities 1868.1.2 Hunting for special cells or special cues 1878.1.3 Farming – culture or engineered fabrication 1888.2 Bioreactors: origins of tissue bioreactor logic and its problems 1908.2.1 What have tissue engineers ever done for bioreactor technology? 1908.2.2 The 3D caveat 1918.2.3 Fundamental difference between biochemical and tissue bioreactors: 3D solid material fabrication 1938.2.4 Why should a little thing like ‘matrix’ change so much? 1948.2.5 The place of tissue bioreactors in tissue engineering logic: what happened to all the good analogies? 1958.3 Current strategies for tissue bioreactor process control: views of Christmas past and present 1998.3.1 Bioreactor enabling factors 2008.3.2 Cell and architecture control 2038.4 Extreme tissue engineering solutions to the tissue bioreactor paradox: a view of Christmas future? 2098.4.1 In vivo versus in vitro tissue bioreactors: the new ‘nature versus nurture’ question? 2098.4.2 Do we need tissue bioreactors at all? 2108.5 Overall summary – how can bioreactors help us in the future? 212Further reading 2149 Towards 4D Fabrication: Time Monitoring Function and Process Dynamics 2179.1 Controlling the dynamics of what we make: what can we control? 2189.2 Can we make tissue bioreactor processes work – another way forward? 2229.2.1 Blending the process systems: balancing the Yin and the Yang 2249.2.2 Making the most of hybrid strategies: refining the timing and sequence 2269.2.3 A real example of making tissues directly 2309.3 The 4th dimension applied to bioreactor design 2329.3.1 Change change change! 2329.3.2 For bioreactor monitoring what are we really talking about? 2339.3.3 Monitoring and processes – chickens and eggs: which come first? 2349.4 What sort of monitoring: how do we do it? 2389.4.1 Selecting parameters to be monitored 2389.4.2 What is so special about our particular ‘glass slipper’? 2419.5 The take-home message 245Further reading 24610 Epilogue: Where Can Extreme Tissue Engineering Go Next? 24710.1 So where can extreme tissue engineering go next? 247Index 249