MSS26 Molecular Mechanism Of Bone Development Flashcards
Forelimb development
- Lateral plate mesoderm
- -> skeletal tissues of limbs - Somites (Paraxial mesoderm):
- -> nerves, blood vessels, muscles
Limb patterning
- Stylopod:
- Humerus - Zeugopod
- Radius
- Ulner - Autopod
- Carpals
- Metacarpal
- Phalanges
***Limb outgrowth
Proximal-distal: ***FGF signaling
- **AER (apical ectodermal ridge) produce **FGF8
- FGF8 diffuse from ***distal to proximal
Anterior-posterior: ***Hedgehog signaling
- **ZPA (zone of polarising activity) produce **Shh (sonic hedgehog)
- Shh diffuse from ***posterior to anterior
- produce ***gradient of morphogen
Dorsal-ventral signaling: WNT signaling
- influence dorsal tissue
ALL these signals do not act alone
4 phases of skeletal morphogenesis
- Migration of mesenchymal cells
- Cell interaction: between epithelial cells and mesenchymal cells
- Condensation –> condensed mesenchymal cells
- Differentiation
mesenchymal cells –> chondrocytes / osteoblasts
2 processes of bone formation
- Intramembranous
Mesenchymal cells directly differentiate
–> osteoblasts
–> Clavicles, Craniofacial skeleton
2. Endochondral Mesenchymal cells --> cartilage cells --> cartilage template of future bone --> - ***Cartilage replacement - ***Blood vessel invasion - ***Mineral deposition - ***Bone synthesis --> Axial and appendicular bones
Control of osteoblast differentiation
- **RUNX2
- a transcription factor (small family)
- -> Gene activator
- master gene for osteoblast differentiation
- regulates osteoblast specific genes
- RUNX2 deficient mice
- -> normal cartilage skeleton develops
- -> but NO bone tissue formed
Cleidocranial dysplasia (CCD)
- Mutation in RUNX2 –> haploinsufficiency
Symptoms:
- Delayed closure of ***cranial suture and fontanelles
- Hypoplastic / aplastic ***clavicles
- ***Dental abnormalities
- delayed eruption of deciduous and permanent teeth
- supernumerary teeth of permanent dentition
Control of chondrocyte differentiation
- **SOX9
- a transcription factor (large family)
- -> Gene activator
- SRY-related (sex-determining region Y gene) HMG (high-mobility-group) box (DNA binding region)
- required for expression of cartilage-specific ***ECM
- -> Collagen II, IX, XI, Aggrecan
Campomelic dysplasia
- Mutation in SOX9
Symptoms:
- Bowing and angulation of ***long bone
- ***Scapular and pelvic hypoplasia
- ***Vertebral column abnormalities
- Decreased no. of ribs
- Small cranium resulting from severe craniofacial anomalies
- XY sex reversal (male with normal female external genitalia)
***Transcriptional regulation of Osteochondroprogenitor
Mesenchymal cells
-(SOX9)-> Osteochondroprogenitor –>
- (RUNX2 on, SOX9 off) –> osteoblast –> functional osteoblast
- (SOX9 keeps on) –> chondrocyte –> hypertrophic chondrocyte
Developing limbs are formed in proximodistal sequence
FGF signaling
- FGF8 produced by ***AER (apical ectodermal ridge)
- FGF8 diffuse from ***distal to proximal
Cartilages are formed as continuous rods
- -> Apoptosis
- -> Bifurcations and segmentations
- -> joints
Anterior-posterior patterning
- ***Sonic hedgehog (Shh), Gli proteins
- determine sequence of fingers
Absence of Gli protein
- -> Greig cephalopolysyndactyly syndrome (fused fingers/toes, widely spaced eyes, macrocephaly)
- -> polydactyly / abnormally wide thumb or big toe
Hedgehog processing and signaling
- Hh sending cell
- -> send signals to Hh receiving cell
- -> with Ptc receptor, Smo receptor (controlled by Ptc)
- -> Hh binds to Ptc receptor
- -> Stop Ptc control on Smo
- -> ***Gli complex stabilized
- -> Gene activator
Overall: Hh —> Ptc —X—> Smo —> stabilise Gli complex —> gene activator
(No Hh signaling
- -> Ptc inhibit Smo
- -> Gli complex degraded
- -> Gene repressor)
Shh-controlled processing (e.g. cleavage) regulates ***transcriptional activity of Gli3
Gene activator / repressor –> determine cell behaviour
Morphogens function as gradient
ZPA produce Shh from ***posterior to anterior
- -> produce gradient of morphogen across different region
- -> different regions have ***different positional values / threshold
- -> different cell fates
Endochondral bone formation
- Cartilage model
- ***Chondrocyte proliferation then hypertrophy
- Cartilage mineralization
- Vascular invasion
- Cartilage replacement with bone
- Growth plate cartilage
- Linear growth of long bone
Balance between Proliferating cells and Hypertrophic cells
controlled by:
- FGF and receptors
- Ihh
- PTH/PTHrP and receptors
FGF and receptors
- key roles in Chondrogenesis
- Negative regulators of chondrocytes:
- -> when FGF bind to FGFR3
- -> ***Suppress proliferation (in contrast to usual role of FGF: ↑ cell proliferation)
- Heparan sulphate proteoglycan (HSPG) required for FGF activity
- > = 18 FGFs, 4 FGFR
- FGFR: tyosine kinase receptor
Mechanism: FGF bind to FGFR --> dimerization of FGFR --> phosphorylation of STAT1 to STAT1-P --> transcription activation of p21 --> bind cdk4,2 complex --> inhibit cyclin D, E dependent kinase activity --> ***inhibit cell cycle progression from G1 (cell growth) to S (DNA synthesis) --> ***block chondrocyte proliferation --> retard bone growth
Defects caused by mutations in FGF3 receptors
Too much FGF signaling
Osteochondrodysplasia:
- Achondroplasia
- Hypochondroplasia
- Thanatophoric dysplasia
Proposed effects of FGFR3 mutation
- Mutations in **transmembrane domain
- -> **stabilization of dimer
- -> too much phosphorylation
- -> too much inhibition of cell cycle - Formation of disulphide bonds between receptors
- -> ***ligand independent activation of receptor - Mutation in tyrosine kinase domain
- -> ***ligand independent activation of receptor
ALL these are Gain-of-function mutations
PTH, PTHrP and PTH1R
PTH1R: Receptor for PTH and PTHrP
PTHrP, PTH1R: major roles in normal endochondral bone formation and elongation
PTH and 1,25-dihydroxyvitamin D3
- regulators of Ca and PO4 homeostasis
PTHrP (PTH related protein):
–> tumour-associated humoral hypercalcemia
Inactivation and Activation of PTHrP-R and PTHrP
PTHrP-R Inactivation: (齋differentiate唔proliferate)
No suppression of differentiation of proliferative chondrocyte into pre-hypertrophic chondrocyte
–> ↑ hypertrophic differentiation + ↓ proliferation
Differentiation speed faster than proliferation speed
–> Retard growth
PTHrP-R Activation: (齋proliferate唔differentiate)
↓ hypertrophic differentiation + ↑ proliferation
–> still Retard growth because hypertrophic differentiation essential for bone growth
Conclusion: must strike balance
Indian hedgehog (Ihh)
- Morphogen
- Stimulate chondrocyte proliferation (form cartilage)
- Prevent chondrocyte hypertrophy (form bone)
Mechanism:
- Ihh expressed in pre-hypertrophic cells (between proliferative and hypertrophic cells)
- 2 effects:
1. Short range effect: stimulate proliferating chondrocytes to proliferate
- Long range effect: diffuse to perichondrium
- -> activate PTHrP
- -> PTHrP diffuses back to pre-hypertrophic layer to bind to PTHrP-R
- -> suppress differentiation of proliferative chondrocyte to pre-hypertrophic chondrocyte
- -> ↓ no. of pre-hypertrophic chondrocyte
- -> ↓ Ihh sent to PTHrP in perichondrium
- -> -ve feedback to regulate proportion of proliferating and hypertrophic chondrocytes in growth plates
***Summary
RUNX2 mutation (唔夠direct differentiation into osteoblasts —> form唔到flat bone): - Cleidocranial dysplasia (CCD)
SOX9 mutation (唔夠chondrocyte differentiation —> form唔到long bone): - Campomelic dysplasia
Proximodistal sequence:
- FGF signaling
Anterior-posterior patterning:
- Sonic hedgehog (Shh), Gli proteins
Balance between proliferating vs hypertrophic chondrocyte controlled by:
- FGF and receptors (suppress proliferation)
- Ihh (stimulate proliferation, activate PTHrP)
- PTH/PTHrP and receptors (stimulate proliferation, suppress hypertrophy)
