Bone development and growth

Question 1

What are the tissues and cell types in the development of the mammalian skeleton?

Answer 1

Two tissues = cartilage + bone

Three cell types = chondrocytes, osteoblasts and osteoclasts

Question 2

What is the osteochondral progenitor?

Answer 2

A common mesenchymal progenitor cell

Question 3

What is the embryonic origin of bone?

Answer 3

Mesenchymal cells in different parts of the embryo come from different cell lineages
1 = neural crest (neuroectoderm) - craniofacial
2 = paraxial mesoderm (somites) - axial skeleton
3 = lateral plate mesoderm - limb skeleton

Question 4

What is intramembranous ossification?

Answer 4

Flat bone formation - mesenchymal condensation - differentiate into osteoblasts

Question 5

What is endochondral ossification?

Answer 5

Long bone formation - mesenchymal condensation - differentiate into chondrocytes producing cartilage. Bone is formed on the cartilage template

Question 6

Why is angiogenesis important?

Answer 6

Brings the vascular endothelial growth factor where it is needed for vascularisation

Question 7

What is mesenchymal condensation?

Answer 7

Aggregation of mesenchymal cells at future skeletal locations
- positional identity = locations of intital skeletal formation determine which of the three mesenchymal cell lineages contribute to the future skeleton

Question 8

What is axial patterning and what controls it?

Answer 8

Somites (segmented mesenchymal structures located in either side of the neural tube) bud off from the anterior tip of the presomitic mesoderm
- Notch, Wnt, FGF pathways control patterning of the axial skeleton

Question 9

What is a limb bud and how does it form?

Answer 9

Small buds of mesodermal cells covered by an ectodermal cap

- requires three signalling molecules and three signalling centres that interact with each other

Question 10

What is proximal - distal patterning (P-D)?

Answer 10

Shoulder → digit tip
3 limb segments
- stylopod, zeugopod, autopod (proximal - distal)

Question 11

What happens if you remove the AER early?

Answer 11

Only the stylopod region forms

Question 12

What happens if the AER is removed late?

Answer 12

The autopod region is not formed but the stylopod and zeugopod regions are formed

Question 13

What is the AER?

Answer 13

Apical Ectodermal Ridge

• thickened epithelium at the most distal end of the limb bud
• signalling centre that direct P-D patterning
• FGF-4, FGF-8, maintain FGF10, important for the proliferation of the mesoderm

Question 14

What is the zone of polarising activity (A/P patterning)?

Answer 14

Thumb to little finger

• area of mesenchyme that contains signals which instruct the developing limb bud to form along the A-P axis
• Shh expressed in ZPA and mediates ZPA activity
• other signals include: bHLH transcription factors (twist 1)

Question 15

What does a mutation in the twist 1 gene cause?

Answer 15

Premature fusion of the caluarial bones and limb abnormalities

Question 16

What is dorsal/ventral patterning?

Answer 16

Back of the hand - palm

• non AER limb ectoderm that covers the limb bud - this tissue controls D-V polarity of the ectoderm and underlying mesoderm
• Signals such as Wnt and BMP activate LIM homeobox transcription factors that determine dorsal identity

Question 17

How is embryonic cartilage and bone formed?

Answer 17

Endochondral progenitor in the mesenchyme condensation expresses both Runx2 and Sox9

Mesenchyme forms either cartilage (endochondral, controlled by Sox9 with low levels of 𝛃-catenin) or bone (membranous, controlled by Runx2 with high levels of 𝛃-catenin)
- once cartilage is formed Ihh signalling mediated by Runx2 forms bone

Question 18

Why is embryonic bone and cartilage formed dependant on canonical Wnt signalling?

Answer 18

1. induces expression of genes required for osteoblastic differentiation
2. inhibits the expression of genes required for chondrocyte differentiation

Question 19

How does intramembranous ossification occur?

Answer 19

Occurs within mesenchymal condensations that are rich in blood vessels

• flat bones
• Wnt signalling in condensations is high - promoting osteoblastic differentiation rather than chondrocyte differentiation
• Mesenchymal cells = osteogenic cells = osteoblasts

Question 20

How does bone within a limb form?

Answer 20

Endochondral ossification

Question 21

What is endochondral ossification?

Answer 21

• Mesenchymal cells proliferate, differentiate in prechondrocytes → chondrocytes forming a cartilage matrix
• This causes the cells at the centre to become hypertrophic (stop proliferating)
• Perichondrial cells at the periphery differentiate into osteoblasts forming a bone collar
• Hydrotrophic chondrocytes at the centre direct formation of bone by attracting blood vessels bringing in osteoblast precursors and then the hydrotrophic chondrocytes will die by apoptosis
• Chondrocytes still proliferate and lengthen the bone
• Once chondrocytes become hypertrophic and attract blood vessels bringing more osteoblasts to form bone creating a secondary ossification centre
• In the middle, bone replaces the cartilage matrix that disintergrates because chondrocytes die by apoptosis
• New bone and woven bone will be replaced by lamellar bone

Question 22

What is secondary ossification in the epiphysis?

Answer 22

1. Occurs at time of birth in humans
2. Epiphysis initially lacks perichondrium (no osteoblasts)
3. Blood vessels invade the centre of the epiphysis
4. Transport osteoclasts and osteoblasts
5. Osteoblasts initiate bone growth and formation of secondary ossification centres

Question 23

Which regions do not have their cartilage replaced by bone?

Answer 23

1. End of epiphysis - joint cartilage

2. Region between diaphysis and epiphysis - epiphyseal growth plate (EGP)

Question 24

What is the importance of VEGF in endochodral ossification?

Answer 24

Immunohistochemical analysis of mammal embryo for presence of VEGFR-2
- resting and proliferating chondrocytes have strong anti-angiogenic effects - not labelled with the antibody against VEGF receptors with hypertrophic chondrocytes which strongly express VEGF receptors and elicit revascularisation

Question 25

What are the important signalling pathways in chondrocyte differentiation in developing cartilage?

Answer 25

BMP - (Bone Morphogen Protein)
Ihh - (Indian hedgehog - synthesised by chondrocytes leaving the proliferative pool)
PTHrp - (PTH - related peptide - keeps proliferative chondrocytes in proliferative pool and delays production of Ihh, no PTHrp chondrocytes become hypertrophic)
Wnt

Question 26

How does bone grow in diameter?

Answer 26

Periosteal apposition exceeds endocortical reabsorption (thicker cortex)
- bone diameter increases during growth

Question 27

What are the requirements for reabsorption during bone growth?

Answer 27

Cortex in diaphysis must be reabsorbed inside and reformed outside

• increase space for bone marrow whilst limiting weight of bone
• in response to muscle activity and mechanical loading

Question 28

What are factors affecting bone growth?

Answer 28

• Vitamin C: collagen synthesis
• Vitamin D: absorption of Ca2+ in the intestine
• Hormones: (thyroid, estrogen)
• Growth hormone: secreted by pituitary - stimulates division of cartilage cells
• Insufficient GH = dwarfism
• Excessive GH = gigantism or acromegaly in adults
• IGF-1: expansion of proliferating chondrocytes

Question 29

What effect do thyroid hormones have on bone growth (length)?

Answer 29

Growth arrest in hyperthyroidism (replacement of cartilage by bone)

Question 30

What effects does oestrogen have on bone growth?

Answer 30

Expression is induced at the end of puberty which causes epiphyseal growth plate cells to hypertrophy and die
- growth plate closure and a lack of oestrogen function

Question 31

What effect do thyroid hormones have on bone growth (width)?

Answer 31

+ =

• mechanical forces
• Androgen
• PTH
• = estrogens