1. Bone Metabolism

2. Outline Normal calcium/phosphate metabolism
Presentation and investigation of bone metabolism disorders
Common disorders of bone metabolism

3. Bone is a specialized connective tissue that, along with cartilage, forms the skeletal system
The role of bone
Bone has a protective and supportive role.
It is metabolically active, and stores many minerals
It provide defense against acidosis
It forms a trap for some dangerous minerals such as lead

4. Since the surface to volume ratio of cancellous bone is so large it has an important role to play in calcium ion homeostasis (the importance of cancellous bone in Calcium homeostasis is exemplified by the fact that 99% of the total body Calcium is stored within the skeleton).

5. Biochemical functions of bone cells
Osteoblasts:
Synthesize webs of collagen fibers, proteoglycans and glycoproteins.
Control bone mineralization.
Osteocytes:
May contribute to the maintenance of calcium homeostasis ,
May have a role in activating bone turnover.
Osteoclasts:
They act in bone resorption and the release of calcium and phosphate into the plasma.
Osteoclasts are able to resorb bone by selectively producing an extremely low pH within the immediate micro-environment of their action by carbonic anhydrase II

6. Osteoprogenitor cells
These are functionally undifferentiated mesenchymal cells
Under the appropriate stimulation these cells may differentiate into functional osteoblasts.
Lining cells
Are former osteoblasts which have become flat and pancake-shaped, lining the entire surface of the bone.
Are responsible for immediate release of calcium from the bone if the blood calcium is too low.
Protect the bone from chemicals in the blood which dissolve crystals (such as pyrophosphate).
Have receptors for hormones and factors that initiate bone remodeling.

7. Bone consists of mineral salts (mainly calcium phosphate, approximately 60% of weight) deposited on an organic matrix.
Water comprises approximately 25% of adult bone mass.

8. Bone matrix
The matrix, which is formed before the mineral is deposited, and can be considered the scaffolding for the bone
It consists of proteins, of which collagen type I is the most important, proteoglycans, and a smaller fraction of lipids and water.

9. Collagen hydroxylation
Chapter 8, pg. 7

10. Table 1: Bone matrix composition
Water and matrix (40% of weight)
Type 1 (90% of protein content)
Other types
1. Collagen
Veriscan, Decorin, Biglycan
2. Proteoglycans
Alkaline phosphtase, Fibronectin,
Osteonectin, Osteopontin, Bone sialoprotein, Matrix extracellular protein
3. Glycoproteins
Matrix gla-protein
4. Gla-proteins
5. Serum proteins

11. Osteogenesis Imperfecta
Osteogenesis imperfecta (OI and sometimes known as Brittle Bone Disease, or "Lobstein syndrome") is a genetic bone disorder. People with OI are born without the proper protein (collagen), or the ability to make it, usually because of a deficiency of Type-I collagen.

12. Defects in Type I collagen cause:
Osteogenesis Imperfecta
Dentinogenesis Imperfecta
Blue sclera
Opalescent and cracked teeth
In support of chapter 8, pg. 6
Left panel from Langlais - Color Atlas of Common Oral Diseases. See the DVD.
Right panel from Wikipedia.
Type I: associated with OI.
Will probably need full crown coverage.

13. Genetic defects in elastin underlie
Facial features associated with Williams Syndrome
Dental features include small widely spaced teeth and malocclusion.
In support of chapter 8, pg. 10.
From Morris CA, and Mervis CB. Williams Syndrome and related disorders. Ann. Rev. Genomics Hum. Genet. 1: (2000).

14. Bone matrix: non-collagen, calcium binding proteins
They are negatively charged glycoproteins
Therefore, they have a high calcium binding potential and are implicated in bone calcification
They include Osteopontin, Osteonectin, Osteocalcin, Matrix extracellular protein, Bone sialoprotein

15. Osteopontin (MW 32 600 kD): Rich in aspartic acid + some sialic acid.
Synthesis is stimulated by vitamin D.
Bind hydroxyapatite.
Associated with the attachment of osteoclasts to the matrix.
Binds to integrins
Increases angiogenesis (makes new blood vessels) which enhances bone resorption in some situations

16. Osteonectin (MW kD):
Binds collagen and hydroxyapatite.
Also referred to as "Bone connector“, and may regulate mineralization
Its synthesis is associated with bone formation and remodeling.

17. Osteocalcin (MW 11 000): The smallest MW.
Contains γ-carboxyglutamic acid (2 or 3 residues)
Binds to hydroxyapatite (may regulate crystal size)
Its synthesis is stimulated by vitamin D.
Vitamin K is needed for the carboxylation of glutamic acid.
Used as a marker of bone metabolism, as its production and levels in the blood reflect osteoblastic activity.

18. Matrix Gla protein (MW 12 000):
Contain γ-carboxyglutamic acid.
Second smallest MW.
Synthesis is stimulated by vitamin D & require vitamin K.
Function is not clear, but it appears to inhibit mineralization

19. Bone Sialoprotein: Fibronectin Relatively abundant
Rich in sialic acid.
Binds to integrins, may assist cancer cells
No clear function.
Fibronectin
Relatively abundant
May help regulate osteoblast differentiation

20. Types of bone matrix Woven bone: Lamellar bone:
Collagen fibrils are distributed within the matrix in a haphazard arrangement.
It is rapidly formed however it is mechanically weak.
It is the first bone matrix formed during skeletal growth and development and healing.
The presence of woven bone in the context of mature skeleton is abnormal but non-specific.
Lamellar bone:
Collagen fibrils have an ordered arrangement in “curving linear arrays”.
This is a mechanically much more sound matrix.
It is the type of bone found in the mature skeleton.

21. Bone Minerals
Bone mineral is so closely associated with the organic matrix, especially with the collagen component. It consists of:
19-26% Calcium (Ca2+)
9-12% Phosphate (Po43-)
2-4 % Carbonate (Co32-)
% Magnesium (Mg2+)
Bone also contain Na and some other minerals in small amounts

22. The mineral is composed of narrow crystallites in the form of long cylinders; about 5 nm wide (60-70 nm in length), arranged parallel to the collagen fibers Some cylinders aggregate to from thicker structures approximately 20 nm wide.
The crystal structure is similar to, but not identical with naturally occurring hydroxyapatite [Ca10 (PO4)6 (OH)2 ]. Bone mineral has a higher content of carbonate and some phosphate may be replaced by sulphate or silicates.

24. BONE FORMATION AND GROWTH
Bone Formation: occurs by the coordinated activity of: chondrocytes, osteoclasts and osteoblasts
Undifferentiated mesenchymal cells
Transforming growth factors (TGFs), other growth factors,
chondrogenic stimulating activity (CSA), steroid and peptide hormones and collagens & other extracellular matrix proteins
A- Chondrogenic line
(cartilage forming)
Chondrocytes + osteoblasts
B- Osteogenic line
(bone forming)
Osteoblasts

25. Factors affecting formation & growth of bone
Pitutary gland:
Hypopituitarism → reduced rate of skeletal growth
Growth hormone:
Important until epiphesial closure.
Acts by:
influencing IGF-1 (regulate IGF-1 production)
stimulation of cell division (prechondrocytes + osteoblasts)

26. Thyroid hormone: Sex hormones: Glucocorticoids
May stimulate chondrocyte maturation.
Sex hormones:
Oestrogen help regulate the rates of bone formation and bone resorption.
It inhibits osteoclasts and their precursors.
It is also required for the process of epiphyseal closure in both sexes and testosterone additionally in males.
Glucocorticoids

27. Hormonal Abnormalities
Oversecretion of hGH during childhood produces giantism
Undersecretion of hGH or thyroid hormone during childhood produces short stature
Both men or women that lack oestrogen receptors on cells, or are unable to convert testosterone into oestrogen grow taller than normal
Oestrogen is responsible for closure of growth plate

28. Local mediators (regulators)
Bone cells produce molecules (usually proteins) that communicate with other cells.
They act on nearby cells, and thus are considered local regulators.
These factors control cell division (proliferation)

29. A mutation causing a constant activation of PTH receptor causes short stature and other signs of hyperparathyroidism, (Jansen’s chondrodysplasia)
Another mutation causing inactive receptor can be fatal in the foetus, or the bones fail to elongate (Blomstrand chondrodysplasia)

30. Blomstrand chondrodysplasia
Jansen’s chondrodysplasia

31. Examples of Local mediators: Example 1: fibroblast growth factors.
A mutation affecting FGF Receptor-3 leads to achondroplasia, the commonest cause of human short stature.

32. Achondroplasia.
This picture shows twin brothers.
The twin on the right has achondroplasia.

33. Example 2: Transcription factors, controlling the formation of mRNAs.
Mutations in TF msx1 & msx2 leads to abnormalities in craniofacial development.

34. Growth factors Bone morphogenetic proteins (BMPs):
BMPs are produced in the bone or bone marrow.
They bind to BMP receptors that are on mesenchymal stem cells within the bone marrow.
This causes the cells to produce Cbfa 1, which is a factor that activates the DNA so proteins can be made
When Cbfa 1 activates the genes, the cells differentiate into mature osteoblasts.
Without Cbfa 1, the cells would turn into fat cells instead.

35. Insulin-like growth factors (IGFs):
These growth factors are produced by osteoblastic cells in response to several bone active hormones, such as parathyroid hormone and estrogens, or BMPs.
IGFs accumulate in the bone matrix and are released during the process of bone remodeling by osteoclasts.
IGFs stimulate osteoblastic cell replication -- in other words, they cause the osteoblasts to divide, forming new cells. They may also induce differentiation.

36. Cytokines
Interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor (TNF) family of cytokines
These factors are produced by osteoblastic cells in response to systemic hormones or other cytokines.
IL-6 can cause:
Bone marrow stem cells to differentiate into pre-osteoclasts
Changes in proliferation and differentiation of osteoblasts
Inhibition of apoptosis of osteoblasts

37. Dietary Factors:
Calcium, phosphate, vitamins D, A and C are most important for endochondrial ossification.
Also important (but less) are: zinc, magnesium and vitamin K.
More recently researchers investigated the effects of moderate magnesium (Mg)-restricted diet on bone formation and bone resorption in rats Mg-restricted diet induced a decrease in bone formation and an increase in bone resorption.
Restriction of energy intake also effects bone formation.

38. MINERLIZATION OF BONE Nucleation agent theory:
Non-collagenous proteins of the matrix provide 'nucleation' sites of the correct geometry for deposition of calcium phosphate.
Note:
Calcium phosphate is relatively insoluble in water and precipitation of this salt can occur spontaneously if the concentrations of calcium, phosphate, or both, cause the 'solubility product' of calcium phosphate to be exceeded
The solubility product for calcium phosphate is: Ksp = [Ca2+]3[PO43-]2
It is pH dependent, and at pH 7.0 its value is 25.

39. ‘Matrix vesicles’ theory:
Osteoblasts and chondrocytes acquire calcium and phosphate ions, form concentrated calcium phosphate at the cell periphery and exfoliate it as vesicles (which also contain alkaline phosphate) prior to its deposition on collagen, and other proteoglycans secreted by osteoblasts

40. Both theories need enough calcium and phosphate.
Calcium and phosphate ion concentrations in the aqueous medium may be altered by binding to proteins,
Complex localized concentration changes may occur as a result of metabolic processes.
Furthermore, the 'micro-environmental' concentrations of calcium and phosphate in biological systems may not be identical to those in the bulk of the solution.

41. The structural strength (and therefore its mechanical function) of any bone is determined by the following parameters:
The volume of bone matrix
The type of bone matrix (woven or lamellar, )
The degree of mineralization of the matrix
The structural arrangement of the matrix
Alterations in any of these parameters will result in a potentially mechanically dysfunctional skeleton.

43. What is the composition of bone? The matrix
40% organic
Type 1 collagen (tensile strength)
Proteoglycans (compressive strength)
Osteocalcin/Osteonectin
Growth factors/Cytokines/Osteoid
60% inorganic
Calcium hydroxyapatite
The cells
osteo-clast/blast/cyte/progenitor

44. Calcium metabolism What is the recommended daily intake? 1000mg
What is the plasma concentration?
mmol/L
How is calcium excreted?
Kidneys mmol/24 hrs
How are calcium levels regulated?
PTH and vitamin D (+others)

45. Phosphate metabolism Normal plasma concentration? 0.9-1.3 mmol/L
Absorption and excretion?
Gut and kidneys
Regulation
Not as closely regulated as calcium but PTH most important

46. PTH Physiological role Production related to plasma calcium levels
Control of calcium levels
target organs
bone - increased Ca/PO4 release
kidneys
increased reabsorption of Ca
increased excretion of PO4
gut - indirect increase in calcium reabs by stimulting activation of vitamin D metabolism

47. Calcitonin Physiological role
Levels increased when serum Ca >2.25mmol/L
Target organs
Bone - suppresses resorption
Kidney - increases excretion

48. Vitamin D (cholecalciferol)
Sources of vit D
Diet
u.v. light on precursors in skin
Normal daily requirement
400IU/day
Target organs
bone - increased Ca release
gut - increased Ca absorption

49. Normal metabolism
Vit D
25-HCC (Liver)
Ca/PTH
1,25-DHCC ,25-DHCC
(Kidney) (Liver)

50. Factors affecting bone turnover
Other hormones
Oestrogen
gut - increased absorption
bone - decreased re-absorption
Glucocorticoids
gut - decrease absorption
bone - increased re-absorption/decreased formation
Thyroxine
stimulates formation/resorption
net resorption

51. Factors affecting bone turnover
Local factors
I-LGF 1 (somatomedin C)
increased osteoblast proliferation
TGF
increased osteoblast activity
IL-1/OAF
increased osteoclast activity (myeloma)
PG’s
increased bone turnover (#’s/inflammation)
BMP
bone formation

52. Factors affecting bone turnover
Other factors
Local stresses
Electrical stimulation
Environmental
temp
oxygen levels
acid/base balance

53. Bone metabolic disorders
Presentation?
Skeletal abnormality
osteopenia - osteomalacia/osteoporosis
osteitis fibrosa cystica - replacement of bone with fibrous tissue usually due to PTH excess
Hypercalcaemia
Underlying hormonal disorder
When to investigate?
Under 50
repeated fractures or deformity
systemic features or signs of hormonal disorder

54. Bone metabolic disorders
Assessment
History
duration of symptoms
drug reaction
causal associations
Examination
X-rays - plain and specialist
Biochemical tests
Bone biopsy

55. Biochemical tests Which investigations? Ca/PO4 - plasma/excretion
Alkaline phosphatase/osteocalcin (o’blast activity)
PTH
vit D uptake
hydroxyproline excretion

56. Osteoporosis Definition? Decrease in bone mass per unit volume
Fragility (perforation of trabecular plates)
Primary (post-menopausal/senile) Secondary

57. Primary osteoporosis Post-menopausal Aetiology?
Menopausal loss 3% vs 0.3% previously
Loss of oestrogen - incr osteoclastic activity
Risk factors?
Race
Heredity
Build
Early menopause/hysterectomy
Smoking/alcohol/drug abuse
?Calcium intake

58. Primary osteoporosis Clinical features? Prevention and treatment?
Post-menopausal
Clinical features?
Prevention and treatment?
General health measures/diet
HRT
Bisphosphonates
Calcium
Vitamin D

59. Primary osteoporosis Senile Aetiology?
7-8th decade steady loss of 0.5%
physiological manifestation of aging
Risk factors?
Prolonged uncorrected post-menopausal loss
chronic illness
urinary insuff
muscle atrophy
diet def/lack of exposure to sun/mild osteomalacia

60. Primary osteoporosis Senile Clinical features? as for post-menopausal
Treatment?
general health measures
treat fractures
as for post-menopausal (HRT not acceptable)

61. Secondary Osteoporosis
Aetiology?
Nutrition - scurvy, malnutr,malabs
Endocrine - Hyper PTH, Cush, Gonad, Thyroid
Drug induced - steroid, alcohol, smoking, phenytoin
Malignancy - carcinoma, myeloma , leukaemia
Chronic disease - RA, AS, TB, CRF
Idiopathic - juvenile, post-climacteric
Genetic -OI
Clin features?
Investigation?
Treatment?

62. Osteomalacia Definition? Aetiology?
Rickets - growth plates affected, children
Osteomalacia - incomplete mineralisation of osteoid, adults
Types - vit D def, vit-D resist (fam hypophos)
Aetiology?
Decr intake/production(sun/diet/malabs)
Decreased processing (liver/kidney)
Increased excretion (kidney)

63. Osteomalacia Clinical features? Investigation
In child
In adult
Investigation
Ca/PO4 decr, alk ph incr, Ca excr decr
Ca x PO4 <2.4
Bone biopsy

64. Osteomalacia Types Vitamin D deficient Hypophosphataemic
growth decr +++ and severe deformity with wide epiphyses
x-linked dominant
decreased tubular reabs of PO4
Ca normal but low PO4
Rx PO4 and vit D

65. Osteomalacia vs osteoporosis
Osteomal Osteopor
Ageing fem, #, decreased bone dens
Ill Not ill
General ache Asympt till #
Weak muscles normal
Loosers nil
Alk ph incr normal
PO4 decr normal
Ca x PO4 <2.4 Ca x PO4 >2.4

66. Hyperparathyroidism Excessive PTH
Due to prim (adenoma), sec (hypocalc), tert (second hyperact -> autonomous overact)
Osteitis due to fibr repl of bone
Clin feat - hypercalc
Invest - Calc incr, PO4 decr, incr PTH
Rx surg

67. Renal osteodystrophy Combination of osteomalacia secondary PTH incr
osteoporosis/sclerosis
CF - renal disorder, depends on predom pathology
Rx - vit D or 1,25-DHCC
renal disorder correction

68. Pagets Bone enlargement and thickening
Incr o-clast/blast activity -> increased tunrover
Aet - unknown but racial diff ?viral
CF - M=F, >50, ache but not severe unless fracture or tumour
Inv - x-ray app characteristic, alk ph is increased and increased hydroxyproline in urine
Rx - bisphos, calcitonin

69. Endocrine disorders Cushings
Hypopituitarism - GH def - prop dwarf or Frohlich adiposogenital syndrome
Hyperpituitarism - gigantism or acromegaly
Hypothyroidism - cretinism or myxoedema
Hyperthyroidism - o’porosis
Pregnancy - backache, CTS, rheumatoid improves SLE gets worse