CARTILAGE FORM 1

 Permissive foetal development through endochondral ossification & postnatal longitudinal growth at epiphyseal growth
plate FOR Transition of cartilage to bone  Tighter management inherit stability of articular cartilage
Development General Characteristics
• CHONDROCYTE – few; synthesise extracellular matrix
• AVASCULAR – no new cells for repair; limited nutrient
supply
• ANEURAL – often no pain
• RICH IN EXTRACELLULAR MATRIX – collagen type II as
major component & type I often present; large molecular
weight proteoglycans (can hold water up to 50 times its
weight); small leucine rich proteoglycans decorin, biglycan;
cartilage oligomeric matrix protein (COMP)
CARTILAGE FORM
Classification
ARTICULAR / HYALINE
• CHONDROCYTE – 5~10% by tissue volume; arrangement varies on location within
cartilage
• WATER – 75~80% by wet weight; half in intrafibrillar space of collagen
• TYPE II COLLAGEN – 50% by dry weight of tissue; little or no type I collagen
• AGGRECAN – 30% by dry weight; large chondroitin sulphate proteoglycan; forms
aggregates of high molecular weight
MENISCUS
• HYALURONAN / FIBRO– 2% by dry weight
• CHONDROCYTE – fewer than in hyaline
cartilage
• WATER – 70~75% by wet weight
• TYPE I COLLAGEN – 90% of all
collagens in meniscus is type I; peripheral
2/3 is only type I whereas type II (60%)
dominates over type I (40%) in the 1/3
• PROTEOGLYCAN, AGGRECAN,
DECORIN, HIGLYCAN,
FIBROMODULIN – 8% of dry weight;
Aggrecan is much lower abundance than
cartilage
• HYALURONAN – 2% by dry weight
CARTILAGE STRUCTURE & FUNCTION 1
Articular/Hyaline
STRUCTURE FUNCTION
• *Synovial joint • Collagen – fibrillar organisation, mechanical resilience
• * Tissue with chondrocytes embedded within collagen matrix • Aggrecan – hydration & compressive resilience
• * Flattened in superficial zone • Aggrecan attracts ions = Attraction of water molecules into the
• * Rounded, sparsely dispersed in mid zone matrix via osmotic process
• * Interface between subchondral border and cartilage; Calcified; • Swelling pressure created by hydration supports a load (body
Column of cells due to proliferation; To resist forces in deep weight) in a similar way to cars being supported by their tyres
zone • Lubrication = Reduce
• * Proteoglycan content generally decreases as going down
friction & Preserve articular
cartilage from wear as poor
regenerative capacity
• Coefficient of friction between cartilage surfaces is very low
(0.005 ~ 0.024 – less than 2 leflon surfaces (0.04))
• Too thin to be effective shock absorber

CHRONDOCYTE
• Depends on diffusion for
nutrients = Thickness of
cartilage is limited =
Depends on diffusion of
certain signalling factors
between chondrocytes
• Responsible for metabolism of extracellular matrix
• Embedded in extracellular matrix & Don’t touch one another unlike other tissues in
body
• Normal cartilage metabolism is highly regulated balance between synthesis and
degradation = Recycling of matrix protein
• PROTEASES – MMP: Collagenases (MMP-1), Gelatinases (MMP-2), Stromolysin
CARTILAGE STRUCTURE & FUNCTION 2
Meniscal/Fibro
• 2 crescent shaped – in synovial space, knee
• Meniscus made of outer third collagen type I within matrix
= Fibro-cartilage & inner two third is type II within matrix
• Flattened cells to periphery
• FUNCTIONS – weight-bearing, shock absorption, load
distribution, articular cartilage protection, joint stability
Intervertebral Disk/Fibro
• Act as shock absorber between each of the vertebrae in the spinal
column, protect the nerves that run down the middle
• Anulus fibrousus – concentric rings of collagen type I which forms
periphery of invertebral disc – contain chrondocytes
• Nucleosus pulposus forming the centre region of intervertebral disc –
disorganised with large quantities of proteoglycan and water so swell
CARTILAGE FORMATION 1
Transient Cartilage 1
• Endochondral ossification  Appendicular skeleton comprising pectoral girdle, pelvis, limbs arise from
lateral plate mesoderm & Axial skeleton including vertebrae and ribs arise form somites of paraxial
mesoderm origin
• Regulated during embryological development to produce the wide range of skeletal forms (epigenetic)
• Apical ectodermal ridge & Zone of polarising activity control proximo-distal & anterior-posterior
patterning in growing limb bud  Regulated by signalling pathways (Indian hedgehog (IHH),
WNT/β-catenin)
• Planar cell polarity & Non-canonical WNT pathway (involving cadherins Far and Dachsous
ENDOCHONDRAL OSSIFICATION
(Fat/Dchs)) – Dchs expression regulate cell shape and directional movement during limb
• Majority of skeletonand
morphogenesis formed as cartilage anlagen  Transient nature of
growth
cartilage replacement by mechanical
• Movement-induced endochondral ossification
bone loading regulates diaphysis growth•of skeletal
from thelongitudinal Chondrocyte required to be removed to maintain steady-state
elements
• Primary ossification centre  Ends of bone  Secondary ossification centre thickness of growth plate = Chondrocyte die by apoptosis
 Cartilaginous growth plate between epiphysis & primary ossification Activation of caspases & Decreased expression of anti-apoptotic
1. centre
Initiated by embryonic mesenchymal cells migrating to form pre-cartilage factor (Bcl2) in
condensations • BUT Lack of typical apoptotic morphological changes in terminal
2. Undergo differentiation into chondrocytes hypertrophic chondrocyte = Condensation of cellular chromatin and
3. Secrete an extracellular matrix rich in collagen type II and aggrecan fragmentation of cell nucleus associated with break down of cell
4. Blood vessels attracted by hypertrophic chondrocyte & facilitated by into several phagocytotic vesicles
degradation of calcified cartilage extracellular matrix around them, MMP13 Autophagic vacuoles & Expression of autophagy-regulating
 Infiltration of bone-resorbing osteoclasts & bone-forming osteoblasts genes by growth plate chondrocytes = Cells undergo processes
occur similar to autophagy THAN apoptosis
5. Chondrocytes undergo ordered and highly regulated process of predominant • Transdifferentiation of chondrocytes of terminal hypertrophic
marginal proliferation and central maturation, hypertrophy, cell death chondrocyte  1 daughter apoptotic cell, 1 osteoblast phenoty[e
6. Latter events define the framework of endochondral bone
7. Regulated by paracrine signals (parathyroid hormone (PTH)-related peptide,
IHH from perichondrium (= create periosteum))
• Blood vessels in growth cartilage
CARTILAGE FORMATION 2
Transient Cartilage 2
MORPHOLOGY OF POSTNATAL GROWTH PLATE
• Growth plate for postnatal linear bone consist of differentiated & matured chondrocytes arranged in columns
• v parallel to axis of bone
surrounded by changing matrix
• Resting/Germinal zone = Primary zone of growth plate with undifferentiated chondrocyte progenitors
1. Chondrocytes gain a proliferative phenotype & adopt a flattened, oblate shape = Arrange into longitudinal columns
2. Production of growth plate-orientating factor
3. Endocrine/Autocrine regulation, Circadian rhythm, Age control high mitotic activity of proliferating chondrocytes
4. Chondrocytes produce collagen type II & proteoglycan-rich matrix
5. Gradually becomes hypertrophic chondrocytes (via chondrocyte membrane)
6. DISTINCT PHASES OF CHONDROCYTE VOLUME INCREASE:
3-fold increase in dry mass production & fluid uptake = True hypertrophy;
4-fold enlargement solely by cell swelling;
Insulin-like growth factor 1 (IGF1) & Regulator of longitudinal bone growth dependent dry mass & fluid volume increase = True
hypertrophy;
 Final stage dependent upon (IGF1), well-recognised regulator of longitudinal bone growth, chondrocyte hypertrophy by WNT
signalling pathway & induced secreted protein (bone growth plate, articular cartilage function)
7. Collagen type-II expression decreases during chondrocyte maturation & Hypertrophic chondrocyte initiates synthesis of collagen type-X
8. FINAL MATURATION PROCESS:
Matrix: chondrocalcin, osteonectin, osteopontin = Vascular invasion through osteoblasts and osteoclasts to gain access via migration to
replace cartilage template with bone;
*chondrocyte sense signals = express receptor activator of nuclear factor κB ligand (essential cytokine for osteoclast development,
function, survival) = initiate transformation of cartilage matrix into bony trabeculae =
Increased membrane activity of alkaline phosphatase
9. Sexual maturity reached = Mineralised tethers between epiphyseal and diaphyseal bone = Promote fusion of primary and secondary
ossification centres = Chondrocytes of growth plate reach a senescence state = Chrondocyte exhaust proliferative potential = Longitudinal
bone growth at growth plate closes
CARTILAGE FORMATION 3
Stable Cartilage
ARTICULAR CARTILAGE FORMATION
• FORMATION OF INTERZONE = Regions of undifferentiated • Combination of extrinsic mechanical factors and intrinsic factors: changes in
mesenchyme which separate developing skeletal elements in extracellular matrix composition (hyaluronan synthesis mediated by
which the joint later form – Earliest emergence of stable articular mechanical activation of MEK-ERK pathway) – Loss of tissue cohesion = Joint
cartilage during skeletal patterning cavity formation
• Densely cellular, homogenous regions with interzone • Different collagen composition:
markers (GDF5, WNT9A – inhibit chondrocyte TRANSIENT  in embryonic growth cartilage – expression of collagen
differentiation, autotoxin, chordin, MMP13) type-IIA precedes collagen type-IIB expression
• DIFFERENTIATION OF INTERZONE into 3 layers: ARTICULAR  collagen type-IIA protein doesn’t appear in interzones =
2 chondrogenic layers – cover articular surfaces of developing cartilage anlagen that form individual skeletal elements are not
opposed skeletal elements, 1 intermediate layer – separates continuous = populations of cells which will become the growth and articular
chondrogenic layers cartilage chondrocytes are discrete [EVIDENCE: GDF5-mediated tracking of
ARTICULAR CARTILAGE PHENOT YPE interzonal cells & unique expression of versican (extracellular matrix
• Expression of transcription factor ETS-related gene (ERG) is necessary toproteoglycan)
establish articular chondrocyte
in regions phenotype
of presumptive articular cartilage in joint
• Fully developed uncalcified articular cartilage divided into superficial, intermediate,
morphogenesis]deep zones with chondrocyte – chondrocyte maintain stable
phenotype characterised by small cell size and expression of tenascin-C & don’t undergo sequence of proliferation, maturation, hypertrophy, apoptosis,
ossification
• SUPERFICIAL ZONE: elongated chondrocytes orientated parallel to surface of cartilage
• DEEP ZONE: more rounded and aligned along collagen fibrils
Form interface tidemark with calcified cartilage;
Hypertrophic chondrocytes accompanied by collagen type-X and alkaline phosphatase expression
• INTERMEDIATE ZONE: collagen fibrils arranged in arches to allow transition from deep and superficial zones
• Calcified cartilage layer (~20 to 250μm) is semipermeable, act as physical barrier for vascular invasion of overlying articular cartilage, permit passage of
small molecules from underlying subchondral bone
CARTILAGE CHANGE – ageing 1
Effect Of Ageing 1
• Age-related changes in cartilage  Risk factor  Inevitable
cartilage disease
• Thinning of matrix as age
AGE RELATED CELL DEPLETION
• Cellularity varies with embryogenesis, skeletal development, ageing
19 cells per mm3 in young adults = Respond to repair;
Can decrease by 50% in femoral condyle (= cartilage tissue found at the end of femur);
Implications for superficial zone (SZ) which has high concentration of progenitor cells – top of
cartilage tissue where loss of these cells as ageing reduces regeneration
• Dead chondrocyte debris seem to enhance calcification of the ECM
• Chondrocytes from older patients proliferate less
• Adult cells rarely proliferate & with age, many cells show signs of senescence
• Related to a diminishing response of cells to growth factors
AGE RELATED
CHRONDROCYTE
•ALTERATION
Age cartilage has reduced response to
anabolic growth factors
• *marker for senescence – nitrotyrosine
CARTILAGE CHANGE – ageing 2
Effect Of Ageing 2
AGE RELATED MATRIX ALTERATION
• Reduction of proteoglycan
• Less organised
• Less aggregate of proteoglycan due to change in structure – enzymatic cleavage, reduction in size of side
chains, reduction ability to function as proteoglycan as ion and water attracting capacity = Increased
accumulation of ECM degradation products = Alteration of structure and function of cartilage tissue
• Matrix degradation increases through production of specific proteases = Proteolytic cleavage of aggrecan
from hyaluronan backbone = Increases amount of junk protein which exists within cartilage matrix
• AGEs (Advanced Glycosylation End product) = Non-enzymatic modification of proteins and lipids;
Glycated & Oxidised by reducing sugars
1. Causes shift base
2. Reorientation & Recombination = Amadori product
3. Damage to cellular structures & ECM; Alter cellular function by signalling via RAGE (= Receptor for
AGE)
4. AGE act as signalling factor, ligand, for receptors
5. Produce downstream effect to downstream effect as catabolic signalling pathway being activated
6. Degradation of cellular matrix
CARTILAGE CHANGE – exercising
Effect Of Exercising
RESPONSE TO LOADING
• Cartilage requires loading to maintain its structural properties:
Born with homogenous cartilage  18 months  Heterogencity observed  Low load areas & High load areas
• Mechanical forces modulate morphology and structure of skeletal tissues = Joints are subject to degenerate
through disuse = Require plentiful movement carried out under pressure to maintain the cartilage surface
DISUSE & EXERCISE REPAIR
• Response to cartilage disuse or unloading – due to nutrient • Cartilage
diffusion & most changes reversible if not prolonged intrinsically has
• Reduction in proteoglycan content – up to 50% a poor capacity
• Reduction in proteoglycan synthesis 30~50% for repair
• Thinning of the cartilage – up to 50% • *AUTOLOGO
• Increase in metalloproteinase (= Cause degradation US
of cartilage matrix) CHONDROC
• Loss of subchondral bone YTE
OVERUSE IMPLANTATI
• Compression of cartilage – trauma, overuse model – ON (ACI)
Disregulation of tissues
• Severe damage – surface fibrillation
• Reduced synthetic capacity of chondrocytes
• Loss of GAG &
ECM proteins
• Cell death –
apoptosis
• Impacted area has a
higher water content
= Disrupt function
of cartilage
CARTILAGE-BONE TRANSITIONS
Ectopic Initiation Of Cartilage-Bone Transitions
ARTICULAR CARTILAGE FORMATION
• Transition regulated to prevent disturbed development and/or
longitudinal bone growth
• Regulation observed in defective hypertrophic differentiation
and ossification, repair of fractured bone tissue
HETEROTOPIC

OSSIFICATION
= Common occurrence in muscle, tendon, ligaments following trauma by injury, disease, surgery
 Initiated by cartilage formation
 Endochondral ossification and ectopic bone formation in tissues can produce severe functional impairment
Fibrodysplasia Ossificans Progressiva (FOP)
• = Rare hereditary form of HO, presenting itself as painful and highly inflammatory soft tissue swellings which
progressively ossify rendering the sufferer immobile
• Endochondral ossification processes = Aberrant bone formation in patients with FOP occurs
• Increased bone morphogenetic protein (BMP) signal pathogenesis of FOP
• Mutated gene: ACVR1/ALK2 type-I receptors that mediate BMP signalling
INTERVERTEBRAL

DISC DEGENERATION (IVD)
= Major cause of back pain with expensive surgery prone to failure
 Cartilaginous IVD functions to resist compressive loads, between vertebral bodies of spine
 Consists of central nucleus pulposus contained within a fibrocartilage ring, annulus fibrosus, laterally and
cartilage end plates inferiorly and superiorly
 Nucleus pulposus similar to articular cartilage – both in its matrix composition and its cellular metabolism
Detection
• Increased alkaline phosphatase activity, levels of hypertrophic differentiation markers (collagen type-X,
osteoprotegrin (OPG), MMP13, RUNX2), levels of IVD calcification – under certain conditions of mineralisation
potential of cells of annulus fibrosus
• Regenerative properties of PTH are mediated through MAPK signalling
• PTH signal in chondrocyte hypertrophy and matrix mineralisation = PTH supplementation prevent further calcification in
degenerated discs & enhance other cell-based therapies
OSTEOARTHRITIS CARTILAGE 1
 = Degenerative joint disease – defective integrity of cartilage & changes in other tissues within the  Human mesenchymal stem cells
joints (bone, synovium, meniscus, ligament) can overcome limitations defined
 Articular cartilage subchondral trabecular loss, Subchondral bone thickening, Osteophyte formation by use of chondrocytes & gain
(through cartilage template, by endochondral ossification, involving hypertrophic chondrocytes) expression of chondrocyte
 Joint symptoms associated with loss of AC integrity hypertrophy marker collagen
• ECM degradation – collagen type II, aggrecan / PG loss type-X
• Chondrocyte death / Cluster formation
• Chondrocyte hypertrophy / Tidemark duplication
 Prevented by regulating stability of chondrocytes by limiting acquisition of their transient growth-related phenotype
ECM Degradation Risk Factors
• Aggrecan cleavage site: NVTEGE epitope BLOCKING ECM DEGRADATION
• Combination of initiation &
• MMP cleavage site: DIPEN • *Knockout ADAMTS5 from articular cartilage =
progression of osteoarthritis:
• *DMM (surgical osteoarthritis) Increase protection to remain aggrecanase in articular
GENETICS, AGEING, OBESITY
• *staining of epitope – more staining when degradation cartilage = Significant decrease in severity lesion
METABOLIC, MECHANICAL
• *MMP-13: Significant loss of articular cartilage in • *IL1 induces aggrecan degradation: Significant
TRAUMA
tribular & Significant reduction of severity of increase in TEGE (aggrecan neoepitope for
disarticulate lesion degradation)
• *Decrease in pain; Than IGG or no control treatment
OSTEOARTHRITIS CARTILAGE 2
Chondrocyte Behaviour
• Microarray profile – gene profile – change in
extracellular matrix (catabolism & anabolism)
• Gene expression profile:
Anabolic – increased ECM expression
Catabolic – increased enzymes expression
Hypertrophic markers
Growth factors & Their signalling
Cell death & Proliferation
Chondrocyte Hypertrophy Chondrocyte Apoptosis
• Hypertrophic • Increased empty
chondrocyte only seen in
lacunaes & apoptosis
terminal differential
markers – ageing & OZ
stages of chondrocytes in
severity
growth plate & calcified • Apoptosis – injurious
cartilage in articular
mechanical injury
cartilage only • Decreased OA severity –
• Increased markers of
Pan-aspase inhibition
hypertrophy in • *live cells stained green,
osteoarthritis: collagen
red cells stained in red
type X, MMP 13 (= • *Pan-Caspase inhibition
potent degradative
= apoptosis inhibition
enzyme in AC), RUNX2,
osteocalcin, alkaline
phosphatase
• Decreased collagen
type-II and aggrecan
integrity
OSTEOARTHRITIS CARTILAGE 3
Inflammatory Phenotype TGFß Signalling & OA
• IL-1 = Potent induction • TGFβ growth factor signalling
of AC degradation • SMAD2/3 – in anabolic responses in
Induction of chrondrocytes & can increase catabolism
catabolism (MMP, • BMP – in hypertrophy & differentiation of
ADAMTS4) chondrocytes
Decrease anabolism • *Other BMP pathway overactivated by
(collagen II, aggrecan) knocking out SMAD3
• Increased expression of
inflammatory markers in
OA cartilage: cytokines
(IL-1, TNFa),
chemokines, TLRs,
complements, alarmins
($100As)
• *Increase in NFkB
signalling pathway =
Significant increase in
expression of p65
TGFß Signalling & Ageing
• Chrondrocytes
change pattern
of expression =
ALK1 takes
over in response
of TGFβ =
Induce
expression of
MMP13