Ebook Articular cartilage (2/E): Part 1

ARTICULAR
CARTILAGE
SECOND EDITION



ARTICULAR
CARTILAGE
SECOND EDITION

Kyriacos A. Athanasiou
Eric M. Darling
Grayson D. DuRaine
Jerry C. Hu
A. Hari Reddi


CRC Press
Taylor & Francis Group
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Library of Congress Cataloging‑in‑Publication Data
Names: Athanasiou, K. A. (Kyriacos A.), author. | Darling, Eric M., author. |
Hu, Jerry C., author. | DuRaine, Grayson D., author. | Reddi, A. H., 1942author.
Title: Articular cartilage / Kyriacos A. Athanasiou, Eric M. Darling, Jerry
C. Hu, Grayson D. DuRaine, and A. Hari Reddi.
Description: Second edition. | Boca Raton : Taylor & Francis, 2016. |
Preceded by Articular cartilage / Kyriacos A. Athanasiou ... [et al.].
2013. | Includes bibliographical references and index.
Identifiers: LCCN 2016021888 | ISBN 9781498706223 (alk. paper)
Subjects: | MESH: Cartilage, Articular
Classification: LCC QM142 | NLM WE 300 | DDC 612.7/517--dc23
LC record available at />Visit the Taylor & Francis Web site at

and the CRC Press Web site at



To Thasos and Aristos, please remember that pursuit of excellence is the
virtuous objective.
Αφιερωμένο στους Θάσο και Άριστο. Αίεν αριστεύειν. —KAA
To my past and present mentors, who have contributed to my professional
success, and to my friends and family, who have contributed to my success

in everything else. —EMD
To Irene, Demitri, and Donovan, who make this all worthwhile. —GDD
To my family, my friends, and our past and current students; I think of
everyone on my treks, and wish that you could see what I see. —JCH
I dedicate this to Professor Kyriacos Athanasiou, a pioneer in articular
cartilage biomechanics and tissue engineering. —AHR


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Contents
Foreword to Second Edition............................................................... xv
Foreword to First Edition ....................................................................xix
Authors .............................................................................................xxv
List of Abbreviations.........................................................................xxix
Chapter 1 Structure and Function of Cartilage .............................. 3
1.1 Cartilage in the Body ................................................................... 4
1.1.1 Hyaline .............................................................................. 4
1.1.2 Elastic ................................................................................ 6
1.1.3 Fibrous .............................................................................. 8
1.1.4 Cartilage’s Role in the Joints ............................................. 8
1.2 Chondrocytes and Their Cellular Characteristics ....................... 11
1.2.1 Genetic, Synthetic, and Mechanical Phenotype .............. 11
1.2.2 Cytoskeletal Structure ..................................................... 15
1.2.2.1 Microfilaments..................................................... 16
1.2.2.2 IntermediateFilaments........................................ 17
1.2.2.3 Microtubules ....................................................... 17
1.2.3 MechanotransductionandHomeostasis
in Cartilage Tissue........................................................... 18

1.2.4 Pericellular Matrix Structure ............................................ 25
1.3 Cartilage Matrix Characteristics and Organization ..................... 27
1.3.1 FluidComponents ........................................................... 28
1.3.2 Zonal Variation within Cartilage ....................................... 30
1.3.2.1 Articulating Surface ............................................ 31
1.3.2.2 Superficial/TangentialZone ................................ 32
1.3.2.3 Middle/TransitionalZone ..................................... 32
1.3.2.4 Deep/RadialZone ............................................... 32
1.3.2.5 TidemarkandCalcifiedZone .............................. 33
1.3.3 Collagens in Cartilage ..................................................... 34
1.3.3.1 Tissue-Level Organization .................................. 38
1.3.3.2 CollagenCross-LinkingandBiomechanical
Properties ........................................................... 40
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Contents

1.3.4 Cartilaginous Proteoglycans ............................................ 42
1.3.4.1 CoreProteinsandGlycosaminoglycans ............. 42
1.3.4.2 Aggrecan’s Role in Articular Cartilage
CompressiveProperties ..................................... 47
1.3.4.3 Minor Proteoglycans ........................................... 47
1.3.5 Additional Molecules Critical for Articular
Cartilage Function ........................................................... 48
1.3.5.1 BoundaryLubricationbySZP/PRG4 .................. 49
1.3.5.2 Molecules for Structural Integrity ........................ 51
1.4 ArticularCartilageBiomechanicalProperties ............................ 51
1.4.1 CompressiveCharacteristics ........................................... 54
1.4.2 Tensile Characteristics .................................................... 57

1.4.3 Shear Characteristics ...................................................... 59
1.4.4 Friction and Wear Characteristics.................................... 60
1.5 Chapter Concepts ...................................................................... 63
References ........................................................................................64
Chapter 2 Articular Cartilage Development ................................. 83
2.1 CellularBehaviorduringEmbryogenesis ................................... 88
2.1.1 LimbPrecursorCellOrigin .............................................. 88
2.1.1.1 Cell Adhesion Molecules .................................... 89
2.1.1.2 InitiationofLimb .................................................. 96
2.1.2 ChondrocyteDifferentiationandJointDevelopment...... 106
2.1.2.1 SynovialJointFormation ................................... 108
2.1.3 TissueHypertrophyandOssification ............................. 109
2.2 Articular Cartilage Growth and Maturation ................................112
2.2.1 Postnatal through Childhood, Adolescence,
and Skeletal Maturity ......................................................112
2.2.2 Matrix Changes with Maturation and Aging ....................113
2.2.3 Aging and Disease .........................................................114
2.2.4 Cellular Changes ............................................................115
2.2.4.1 Aging Cells.........................................................115
2.3 SignalingPathwaysduringDevelopmentandMaintenance
of Articular Cartilage .................................................................117
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Contents

2.3.1 Signaling Factors ............................................................117
2.3.1.1 Morphogens and Growth Factors ......................117
2.3.1.2 MechanicalStimuli .............................................119
2.3.1.3 Oxygen Tension ................................................ 120

2.3.2 Signaling Cascades ....................................................... 121
2.3.2.1 Morphogen Signaling ........................................ 121
2.3.2.2 BMP/TGF-βSuperfamily ................................... 121
2.3.2.3 ReceptorsandSmads ...................................... 123
2.3.2.4 MAPK and Rho GTPase Signaling
in Articular Cartilage ......................................... 126
2.3.3 Transcription Factors ..................................................... 134
2.3.4 CartilageHomeostasis .................................................. 137
2.4 Chapter Concepts .................................................................... 139
References ...................................................................................... 141
Chapter 3 Articular Cartilage Pathology and Therapies ........... 163
3.1 Arthritis .................................................................................... 163
3.1.1 OsteoarthritisandRheumatoidArthritis ........................ 164
3.1.1.1 Osteoarthritis .................................................... 164
3.1.1.2 RheumatoidArthritis ......................................... 166
3.1.2 EtiologyandEpidemiology ............................................ 167
3.1.2.1 Genetic and Metabolic Defects ......................... 169
3.1.2.2 Crystal Pathologies ........................................... 173
3.1.2.3 Biomechanics,Loading,andInjury ................... 175
3.1.2.4 Epidemiology .................................................... 184
3.1.3 Changes in the Matrix ................................................... 186
3.1.4 Cellular Changes ........................................................... 190
3.1.5 Costs of Arthritis ............................................................ 194
3.2 CartilageInjuries ...................................................................... 195
3.2.1 Osteochondral and Chondral Defects
and Microfractures ......................................................... 195
3.2.2 CausesofCartilageInjuries .......................................... 200
3.2.3 CellularResponsestoInjury.......................................... 203
3.2.4 CostsofArticularCartilageInjuries ............................... 205
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Contents

3.3 CurrentandEmergingTherapiesforArticularCartilage
Repair and Arthritis .................................................................. 206
3.3.1 NonsurgicalTreatments ................................................. 208
3.3.2 SurgicalTreatmentsforArticularCartilageRepair .........211
3.3.2.1 Debridement ......................................................211
3.3.2.2 MarrowStimulationTechniques ........................ 212
3.3.2.3 AutologousImplants ......................................... 213
3.3.2.4 AdjunctiveTreatments....................................... 217
3.3.3 OtherTreatmentsandEmergingTechniques ................ 219
3.4 Motivation for Tissue Engineering ............................................ 222
3.5 Chapter Concepts .................................................................... 226
Appendix..........................................................................................228
References ...................................................................................... 231
Chapter 4 Tissue Engineering of Articular Cartilage ................ 257
4.1 The Need for In Vitro Tissue Engineering ................................ 260
4.1.1 Characteristics of the In VivoSystem ............................ 263
4.2 Cell Source .............................................................................. 266
4.2.1 A Need for Alternative Cell Sources .............................. 267
4.2.2 ChondrogenicDifferentiationofMesenchymal
StemCellsandProgenitorCellPopulations .................. 269
4.2.3 ChondrogenicDifferentiationofEmbryonicStemCells
andInducedPluripotentStemCells .............................. 271
4.3 BiomaterialsandScaffoldDesign ............................................ 272
4.3.1 Natural Scaffolds ........................................................... 275
4.3.2 Synthetic Scaffolds ........................................................ 281
4.3.3 CompositeScaffolds ..................................................... 289

4.3.4 Scaffoldless ................................................................... 292
4.3.5 MimickingtheZonalStructuralofArticularCartilage..... 294
4.4 Bioactive Molecules for Cartilage Engineering......................... 296
4.4.1 GrowthFactorsandCombinations ................................ 296
4.4.2 Protein Coating and Peptide Inclusion ........................... 304
4.4.3 Catabolic and Other Structure Modifying Factors .......... 313

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Contents

4.5 BioreactorsandMechanicalStimulation .................................. 316
4.5.1 Compression ..................................................................317
4.5.2 Hydrostatic Pressure ..................................................... 323
4.5.3 Shear .............................................................................329
4.5.3.1 Contact Shear ................................................... 330
4.5.3.2 Fluid Shear ....................................................... 331
4.5.3.3 Perfusion Shear ................................................ 336
4.5.3.4 Low-Shear Microgravity .................................... 341
4.5.4 Tension ..........................................................................348
4.5.5 Hybrid Bioreactors .........................................................349
4.5.6 Gas-Controlled Bioreactors ........................................... 351
4.5.7 Electrical and Magnetic Stimulation ...............................354
4.6 ConvergenceofStimuli ............................................................ 354
4.6.1 Synergismversus Additive Effects ................................354
4.6.2 Potential Mechanisms ...................................................356
4.7 Chapter Concepts .................................................................... 358
References ......................................................................................361
Chapter 5 Methods for Evaluating Articular Cartilage Quality ... 393

5.1 ImagingTechniques ................................................................. 396
5.1.1 Noninvasive Modalities ..................................................397
5.1.2 Invasive Methods ...........................................................406
5.2 Histology,Immunohistochemistry,andOtherUltrastructural
Techniques............................................................................... 407
5.2.1 Cells .............................................................................. 410
5.2.2 Collagen ........................................................................ 413
5.2.3 SulfatedGlycosaminoglycans........................................ 417
5.3 TechniquesfortheQuantitativeAssessmentofCartilage
TissueComponents ..................................................................419
5.3.1 Cells .............................................................................. 419
5.3.2 Collagen ........................................................................ 421
5.3.3 ProteoglycansandGlycosaminoglycans .......................422
5.4 BiomechanicalTechniques ...................................................... 425
5.4.1 CompressionTesting ..................................................... 426
5.4.2 Tensile Testing ............................................................... 428
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Contents

5.4.3 Shear Testing ................................................................ 430
5.4.4 Friction Testing .............................................................. 431
5.4.5 Fatigue Testing .............................................................. 432
5.4.6 MathematicalModels of Articular Cartilage Mechanics ... 434
5.4.7 Cartilage Tribology ........................................................ 437
5.5 AnimalModels ......................................................................... 438
5.5.1 SmallAnimalModels .....................................................438
5.5.2 LargeAnimalModels.....................................................443
5.5.3 Genetic and Defect Models ...........................................445

5.6 ClinicalScoringSystems ......................................................... 447
5.7 Chapter Concepts .................................................................... 450
References ...................................................................................... 451
Chapter 6 Perspectives on the Translational Aspects
of Articular Cartilage Biology .................................... 465
6.1 Challenges and Opportunities in Articular Cartilage Biology
and Regeneration .................................................................... 466
6.1.1 ComplexitiesofIntegratingNewCartilage
with Existing Cartilage ...................................................469
6.1.2 ChallengesinManufacturingComplex
Tissue- Engineered Products ......................................... 473
6.2 ImmuneResponse,Immunogenicity,andTransplants ............. 476
6.2.1 HumoralandCellularResponses .................................. 476
6.2.2 Allogeneic Transplants ..................................................480
6.2.3 Xenogeneic Transplants ................................................483
6.3 Business Aspects and Regulatory Affairs ................................ 488
6.3.1 Regulatory Bodies .........................................................489
6.3.2 Pathways to Market ....................................................... 491
6.4 U.S.StatutesandGuidelines ................................................... 497
6.4.1 Currently Available Cartilage Resurfacing Products ...... 501
6.4.2 EmergingCartilageProducts.........................................503
6.4.3 EmergingCellularTherapies .........................................503
6.5 Chapter Concepts .................................................................... 508
References ......................................................................................509
xii


Contents

Chapter 7 Experimental Protocols for Generation

and Evaluation of Articular Cartilage ........................ 521
7.1 Tissue and Cell Culture ............................................................ 522
7.1.1 Harvesting Cartilage and the Production of Explants ....523
7.1.1.1 Surgical Dissection and Cartilage Harvesting ... 526
7.1.2 Cell Isolation ..................................................................528
7.1.3 Cryopreservation of Cells ..............................................530
7.1.4 2D Cultures ...................................................................532
7.1.4.1 Monolayer ......................................................... 533
7.1.4.2 Passaging Cells ................................................ 534
7.1.5 3D Cultures ...................................................................536
7.1.5.1 Hydrogel Encapsulation .................................... 536
7.1.5.2 Cell-OnlyTechniques........................................ 541
7.2 GrossMorphology,Histology,andImmunohistochemistry....... 545
7.2.1 PhotodocumentationofGrossMorphology ...................545
7.2.2 India Ink .........................................................................546
7.2.3 Histology........................................................................547
7.2.3.1 Fixing Cryosectioned Slides ............................. 548
7.2.3.2 ClearingParaffinSlidesforStaining ................. 549
7.2.3.3 Safranin O with Fast Green Counterstain ......... 550
7.2.3.4 Picrosirius Red .................................................. 551
7.2.3.5 HematoxylinandEosin ..................................... 552
7.2.3.6 Alcian Blue ........................................................ 554
7.2.4 Immunohistochemistry ..................................................555
7.2.4.1 Antigen Retrieval............................................... 556
7.2.4.2 AntibodyDetectionforImmunohistochemistry.... 557
7.3 Protein Analysis ....................................................................... 560
7.3.1 Protein Extraction ..........................................................560
7.3.1.1 Protein Extraction for 2D Cultures .................... 561
7.3.1.2 LiquidNitrogenPulverizationandProtein
Extraction for Tissues and 3D Cultures ............ 562

7.3.2 ProteinQuantification ....................................................564
7.3.2.1 Bicinchoninic Acid ............................................. 565
7.3.2.2 CoomassieBlueAssay(Bradford) .................... 566
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Contents

7.3.3 Antibody-Based Detection Methods ..............................568
7.3.3.1 Enzyme-LinkedImmunosorbentAssay ............ 569
7.3.3.2 WesternBlot/Immunoblot.................................. 573
7.3.3.3 CellLabelingforFlowCytometry ...................... 578
7.4 InsolubleMatrixComponentExtractionandAssaying ............. 581
7.4.1 SampleDehydrationbyLyophilization ........................... 581
7.4.2 Papain Digestion ...........................................................582
7.4.3 AssayingGlycosaminoglycanContent ..........................583
7.4.3.1 SimpleDMMBAssayfortheDetection
of sGAGs .......................................................... 583
7.4.3.2 Biocolor Assay .................................................. 584
7.4.4 Assaying for Total Collagen Content .............................585
7.4.5 DNAQuantificationbyPicoGreen .................................588
7.5 RNA Extraction and Gene Expression..................................... 590
7.5.1 RNAse/DNAse-FreeDEPCWater ................................ 591
7.5.2 RNAExtractionfrom2DCultures..................................592
7.5.3 RNAExtractionfrom3DCulturesandTissue ...............595
7.5.4 Gel Electrophoresis for DNA and RNA ..........................597
7.5.5 PolymeraseChainReaction ..........................................599
7.5.6 QuantitativeReverseTranscriptionPCR .......................601
7.5.6.1 cDNA Creation .................................................. 603
7.5.6.2 QuantitativeReal-TimePCR ............................. 604

7.6 AssayingTissueBiomechanicalProperties ............................. 606
7.6.1 Compression .................................................................606
7.6.1.1 Load-Controlled Testing via the Creep
IndentationApparatusCompressionTest ......... 606
7.6.1.2 StressRelaxationCompressionTest ................ 610
7.6.2 Strain-to-Failure Tensile Test ......................................... 612
7.6.3 MeasuringtheFrictionCoefficient ................................. 613
7.7 AnimalProtocols ...................................................................... 615
References ...................................................................................... 615
Glossary ......................................................................................... 617
Index ...............................................................................................633
xiv


Foreword to Second Edition
In 1892, the American Poet Walt Whitman celebrated the remarkable design and function of synovial
joints: “the narrowest hinge in my hand puts to scorn all
machinery” (“Song of Myself,” Leaves of Grass, Walt
Whitman Complete Poetry and Collected Prose 18911892, New York, Library of America, 1982, p. 217). Over
the last half century, dramatic advances in prosthetic
joint replacement have made it possible to restore mobility and relieve pain for millions of people with advanced
joint damage. Continuing translational research, technological and procedural advances have made the current
practice of synthetic joint replacement one of the most noteworthy successes in the
annals of surgery.
Yet, Whitman’s observation stands unchallenged; no current artificial joint comes
close to replicating the function and durability of synovial joints. These complex
structures developed and progressively evolved over millions of years. Formed from
multiple self-renewing tissues, including joint capsule, ligament, in some cases
meniscus, subchondral bone, synovium, and hyaline articular cartilage, they provide
stable pain-free movement with a level of friction less than that achieved by any artificial bearing surface. The tissue central to these extraordinary functional capabilities is hyaline articular cartilage. Although it varies in thickness, cell density, and to

some extent composition and mechanical properties among joints and among mammalian species, all articular cartilages share the same general structure and perform
the same functions. These include lubricating the joint surface and minimizing peak
stresses on subchondral bone by distributing loads. Perhaps the most extraordinary
property of articular cartilage is durability; for most people, it provides normal joint
function for more than 80 years.
Deterioration of synovial joints due to multiple causes, most commonly osteoarthritis, is the leading cause of pain and impairment in middle-aged and older people.
Osteoarthritis can occur in any synovial joint and develops in every human population. Although osteoarthritis increases with aging, it is not a direct or inevitable


Foreword to Second Edition
result of aging changes alone. In addition to increasing age, excessive cumulative
joint loading and joint injury are universal risk factors for osteoarthritis in all joints
and all populations; yet, despite the recognition of these risk factors, the pathogenesis
of the joint destruction that leads to the clinical syndrome of osteoarthritis remains
obscure. No current treatments have been shown to prevent the onset or progression
of osteoarthritis, although recent findings suggest that interventions to decrease the
risk of osteoarthritis following joint injury may be possible.
In a concise cogent fashion, this second edition of Articular Cartilage summarizes
current understanding of articular cartilage structure, function, development, maintenance, and degradation. Furthermore, exciting new information included in this
volume lays the foundation for fresh approaches to preventing loss of articular cartilage and even restoring lost or diseased cartilage.
The book consists of seven chapters. Chapter 1 deals with the structure, composition,
and function of articular cartilage, including lucid explanations of how the structure
and organization of articular cartilage provide its unrivaled biomechanical properties, including lubrication of the joint surface. Chapter 2 covers articular cartilage
embryogenesis, growth and maturation, and signaling pathways that have roles in
these changes. Because articular cartilage lacks nerves and blood vessels, it was initially thought to be relatively inert; the identification of signaling pathways that control
its formation, growth, and maintenance proves that this early impression was mistaken. Progress in understanding these pathways is likely to help explain the onset
and progression of osteoarthritis and may lead to methods of detecting changes in
tissue homeostasis before the cartilage begins to deteriorate. Chapter 3, “Articular
Cartilage Pathology and Therapies,” deals with the various forms of arthritis that lead
to loss of articular cartilage and joint function, cartilage injuries and the response to

injury, and contemporary and emerging methods for cartilage repair and restoration.
Chapter 4 is devoted to tissue engineering of articular cartilage and explores the potential of in vitro tissue engineering for the restoration of articular surfaces. This chapter
then goes on to summarize the sources of cells, biomaterials, and the use of scaffolds
and bioreactors to promote formation of functional articular cartilage. As Chapter 4
clearly shows, tissue engineering has great promise for biologic restoration of synovial
joints. In advancing understanding of the disorders of articular cartilage and their
treatment or prevention, it is critical to have methods of evaluating articular cartilage
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Foreword to Second Edition
quality. Chapter 5 discusses the imaging techniques and the quantitative assessment
of cartilage components and methods of measuring mechanical properties to assess
cartilage composition, structure, and function. It also includes a summary of the uses
of large and small animal models to test the safety and efficacy of cartilage repair or
restoration therapies. Breakthroughs in treatment of damaged or degenerated joints
will depend on translating advances in basic cartilage research into clinical practice. Chapter 6 summarizes the challenges and opportunities in basic investigations
of articular cartilage biology and regeneration and covers the business and regulatory aspects of potential methods of re-creating biologic articular surfaces. Chapter 7
presents detailed explanations of experimental protocols for generating and evaluating
articular cartilage, including tissue and cell culture, tissue and matrix molecule analysis, RNA extraction, and testing mechanical properties, as well as animal protocols.
This well-organized, readable, and comprehensive second edition of Articular Cartilage
is an important milestone in the understanding of one of nature’s singular creations. It
will serve as an essential resource for those who wish to contribute new insights into
articular cartilage biology, as well as those who pursue clinically applicable technologies with the potential to reconstruct damaged or diseased joint surfaces. Although
prosthetic joint replacement for people with advanced, essentially complete, destruction of hip, knee, and shoulder joints is effective, these procedures have limitations and
in some instances devastating complications. Discovering ways to prevent the onset
or progression of synovial joint destruction and to rebuild biologic articular surfaces
would be among the most important developments in the history of medicine. This
book gives encouragement and direction to those who seek to make these discoveries.
Joseph A. Buckwalter, MS, MD

Professor and Arthur Steindler Chair
University of Iowa Department of Orthopaedics and Rehabilitation
Iowa City Veterans Administration Medical Center
Iowa City, Iowa

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Foreword to First Edition
The synovial joint is truly one of nature’s marvels, providing our skeleton with a nearly frictionless bearing surface that can withstand forces of several times body
weight for millions of loading cycles throughout life. To
date, no man-made joint has been able to approach these
capabilities. While the mammalian joint is clearly a highly
complex biological and biomechanical organ that includes
multiple structures, tissues, and cells, it is the articular
cartilage—the tissue that lines the surfaces of synovial
joints—that is fundamentally responsible for these unparalleled biomechanical properties.
Over the past century, our understanding of articular cartilage has grown exponentially. Building upon early studies that characterized the anatomy and histology of
cartilage, scientists recognized its unique mechanical properties and function. By the
mid-twentieth century, investigators had begun to develop new methods to quantify
the elastic and tribological properties of the tissue. The 1960s and the 1970s were
characterized by significant advances in the characterization of the biochemical composition of cartilage, primarily the proteoglycan and collagen components. With the
development of the biphasic theory for modeling cartilage mechanics in 1980, the
next two decades saw major breakthroughs in the understanding of the highly complex multiphasic, viscoelastic, anisotropic, inhomogeneous, and nonlinear properties
of the tissue. Simultaneously, the study of cartilage development was revolutionized

by the ongoing breakthroughs occurring in molecular biology and genetics in the
1990s. By the beginning of the twenty-first century, scientists and engineers had
made tremendous strides in understanding how the incredibly complex composition
and structure of cartilage were responsible for its load-bearing properties.
However, as with any other precision machine, even slight imbalances of the biological
or biomechanical processes responsible for maintaining the tissue can lead to cumulative and progressive changes over decades of use, ultimately causing osteoarthritic
failure of the joint. With the new depth of understanding of cartilage development,
mechanics, and biology, the fields of tissue engineering and regenerative medicine have


Foreword to First Edition
exploded in the effort to develop new therapies for preventing or treating cartilage damage by combining cells, biomaterials, bioactive molecules, and physical signals. While
there are currently no disease-modifying therapies available for treating osteoarthritis,
such tissue engineering approaches hold tremendous promise for the near future.
For the first time, the wealth of new knowledge in these areas is brought together in
a single volume. Articular Cartilage represents the most comprehensive text to date
focusing on this tissue and provides a unique and interdisciplinary approach that
encompasses the breadth of basic science, bioengineering, translational science, and
detailed methodologic approaches.
Chapter 1 broadly reviews the current state of knowledge on the structure and composition of different types of cartilage as well as the chondrocytes. In addition to
presenting the molecular components of the tissue, this chapter provides overviews
of the biomechanical function and properties of cartilage, as well as the structurefunction relationships of the primary constituents of the tissue and cells.
A critical step in understanding cartilage physiology, pathophysiology, and regeneration is an understanding of the fundamental processes involved in cartilage development, maturation, and aging. In Chapter 2, the current state of knowledge of cartilage
development is summarized, including the sequences of growth and transcription
factors necessary for proper cell-cell and cell-matrix interactions required during the
formation of the limb bud and the subsequent formation of the synovial joint. This
chapter also reviews the changes that occur in the extracellular matrix and chondrocytes with maturation and aging, under normal or pathologic conditions.
Chapter 3 focuses on the epidemiology, etiopathogenesis, and therapeutic approaches
for the major arthritides that affect cartilage and the synovial joints, namely, cartilage
injury, osteoarthritis, rheumatoid arthritis, and gout. While these represent distinct

disease processes, they are all characterized by degeneration of the articular cartilage
and, eventually, loss of joint function. In particular, significant emphasis is placed
on the role of biomechanical factors in the onset and progression of osteoarthritis.
Furthermore, a review of the (lack of) current therapeutic approaches for osteoarthritis or cartilage injury clearly reveals a substantial unmet need for disease-modifying
approaches to diseases that affect articular cartilage.
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Foreword to First Edition
With recent evidence suggesting that over 10% of osteoarthritis may arise due to joint
injury, it is clear that the development of new tissue engineering approaches for cartilage repair or regeneration can have a significant impact on this disease. Chapter 4
provides an up-to-date overview of the field of tissue engineering as applied to articular cartilage repair. Different sections provide highlights of recent advances in the
classical “three pillars” of tissue engineering: cell source, scaffold design, and external stimulation through the use of bioactive molecules and mechanical bioreactors.
The chapter also includes important discussion of the relative advantages and potential limitations of different cell types, biomaterial scaffolds, bioactive molecules, and
bioreactors.
One of the primary hindrances to the development of new therapies for joint disease
has been the lack of surrogate measures that provide valid, reliable, and responsive
readouts of disease severity or progression. Such biological markers, or “biomarkers,” may include proteins, genes, noninvasive or invasive imaging, or even biomechanical measures that reflect certain events in the disease process. In other fields
such as cardiology and infectious diseases, biomarkers such as cholesterol levels,
blood pressure, or antibody levels have served critical diagnostic and therapeutic
roles. Chapter 5 overviews a number of methods that are used to assess the structure,
composition, biology, and biomechanical function of articular cartilage. In addition
to novel imaging methods such as MRI, such assessments may include histologic or
immunohistochemical measures of joint tissues, or direct measures of tissue function through biomechanical testing. Due to the highly complex nature of cartilage,
the proper determination of tissue material-level properties often involves the use of
mathematical modeling that simulates the precise testing condition in tension, compression, shear, or contact (i.e., tribological testing). Finally, this chapter also provides a summary of different animal models and scoring systems that are often used
for modeling and assessing disease or repair processes, with a critical review of their
relative advantages and disadvantages.
With these issues in mind, Chapter 6 provides important discussion and perspectives on many of the remaining challenges and opportunities in the development and
translation of new approaches for treating diseases of articular cartilage. A variety of

issues are discussed, including some of the intrinsic characteristics of cartilage that
appear to make repair of cartilage insuperable. In this light, alternative factors are
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discussed that may influence the success of regenerative therapies for cartilage, such
as potential immunogenic responses. The ultimate success of such cell-based or biologic therapies, however, is highly dependent on practical issues such as regulatory
pathways, intellectual property concerns, the pathway to market, and potential reimbursement. This chapter provides an important snapshot of the ever-changing landscape of regulatory and commercial affairs for medical products for cartilage repair.
The final chapter of the text provides detailed working protocols for many of the
methods used to study articular cartilage. Beginning with standard cell and tissue
harvest and culture methods, the chapter also details several culture methods, such
as the use of 3D gels, that are commonly used for chondrocyte culture or cartilage
tissue engineering. Methods for cartilage assessment via histology and immunohistochemistry are also provided. Importantly, detailed methods are provided for protein
and RNA extraction from cartilage, which is generally more complex than other cells
due to the presence of significant amounts of extracellular matrix. Finally, detailed
protocols for mechanical testing of cartilage are provided.
This thorough and comprehensive text seamlessly integrates concepts of basic science, bioengineering, translational medicine, and clinical care of articular cartilage.
By revealing the wealth of knowledge we have accumulated in this area, as well as
exposing the tremendous opportunities for advancement, Articular Cartilage provides a critical template for those seeking to study one of the most complex tissues of
the human body. Only through this level of understanding will we eventually be able
to develop new methods to diagnose, prevent, or treat diseases of articular cartilage.
Farshid Guilak, PhD
Professor of Orthopaedic Surgery and Research Director
Shriners Hospital, St. Louis
Co-Director of the Washington University Center of Regenerative Medicine
Washington University
St. Louis, Missouri
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Foreword to First Edition
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