Download Bone physiology

Hard tissue formation and
destruction
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Hard tissue associated with the functioning tooth:
cementum, dentin, and enamel
Specialized connective tissues (except enamel)
Collagen (esp. type I) plays a role in determining their
structure.
Hard tissue formation: cells - production of organic
matrix -capable of accepting mineral + the activity of the
enzyme alkaline phosphatase + a good blood supply
prerequisites.
Cells
 functions of synthesis and secretion
 abundant mitochondria, much rER, and Golgi
complex
Organic matrix
 type I collagen + proteoglycans +
phosphoproteins + phospholipids
 enamel: enamel proteins: amelogenin +
enamelin
 capable of accepting mineral in the form of
hydroxyapatite crystals
Mineral
 biological apatite: calcium hydroxyapatite
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[Ca10(PO4)6(OH)2]
calcium + phosphate + hydroxyl ions - ionic lattice
relationships - permit considerable variation in
composition through substitution, exchange, and
adsorption of ions
magnesium and sodium can substitute in the calcium
position
fluoride and chloride can substitute in the hydroxyl
position
carbonate can substitute in both the hydroxyl and
phosphate positions
fluoride substitution - decrease the solubility of the
crystallites
carbonate substitution - increase the solubility of the
crystallites
Mineralization
 local increase in the concentration of
inorganic ions - supersaturated tissue fluid of
calcium and phosphate ions - spontaneous
precipitation of a calcium phosphate product
- formation of ionic clusters and crystallites
(homogeneous nucleation)
 the presence of a nucleating substance but
not locally increased ionic concentration crystal formation (heterogeneous
mineralization)
factors inhibit mineralization
 tissue fluid contains other macromolecules that
could inhibit crystal formation
 the initial cluster of ions needed to form a lattice
structure is not enough or unstable
 insufficient energy for overcoming crystallization
(inhibited by inhibitors of mineralization)
Bone
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Three functions
Mechanical: support and site of muscle
attachment for locomotion
Protective: for vital organs and bone marrow
Metabolic: reserve of ions for the entire organism,
especially calcium and phosphate
Bone physiology
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Non cellular elements：
1.
33% organic：
28%collagen fiber type I,
5% ground substance : glycoproteins,
proteoglycans. These highly anionic
complexes have a high ion-binding capacity
and are thought to play an important part in
the calcification process and the fixation of
hydroxyapatite crystals to the collagen fibers.
67% inorganic：hydroxyapatite.
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Bone structure
 Lamellar structure: this organization of fibers
allows the highest density of collagen per unit
volume of tissue. These lamellae can be
parallel to each other if deposited along a flat
surface (trabecular bone and periostrum) or
concentric if deposited on the surface of a
channel centered on a blood vessel
(Haversian system)
 Woven bone: when bone is formed very
rapidly (during histogenesis, fracture healing,
tumor, or some metabolic bone disease).
They are found in more or less randomly
oriented bundles.
Cellular elements
1. Osteocytes
2. Osteblasts
3. Osteoclasts
4. Other cell types
Osteocytes
 They are found embeded deep within the bone in
small osteocytic lacunae (25000/mm3 of bone).
 They were originally bone-forming cells (osteoblasts)
that have been trapped into their own production of
bone matrix, which later became calcified.
 These cells have numerous and long cell processes
rich in microfilaments that are in contact with cell
processes from other osteocytes (gap junctions), or
with processes from the cells lining the bone surface
(osteoblasts or flat lining cell in the endosteum or
periosteum)
 These processes are organized during the formation
of the matrix and before its calcification.
Osteblasts
 The osteoblast is the bone lining cell
responsible for the production of the matrix
constituents (collagen and ground
substance).
 It originates from a local mesenchymal stem
cell (bone marrow stromal stem cell or
connective tissue mesenchymal stem cell)
 These precursors upon the right stimulation,
undergo proliferation and differentiate into
preosteoblasts and then mature osteoblasts.
Osteblasts
 Osteoblasts never appear or function individually
but are always found in clusters of cuboidal cells
along the bone surface (~100-400 cells per bone
froming site).
 They are always found lining a layer of bone
matrix that are producing and is not yet calcified
(osteoid tissue, its maturation period ~10 days)
At the ultrastructural level
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the osteoblastis characterized by
The presence of an extremely welldeveloped rough endoplasmic reticulum
and a dense granular content
The presence of a large circular Golgi
complex comprising multiple Golgi stacks
Osteblasts
 Cytoplasmic processes on the secreting side of the cell
extend deep into the osteoid matrix and are in contact with
the osteocyte processes in their canaliculi.
 Junctional complexs (gap junctions) are often found
between the osteoblasts.
 The plasma membrane of the osteoblast is
characteristically rich in alkaline phosphatase (used as an
index of bone formation) and been shown to have rceptors
for parathyroid hormone, but not for calcitonin.
Osteblasts
 OBs arise from osteoprogenitor cells of
mesenchymal origin and are the source of the
terminally differentiated osteocyte
 OBs have the ability of synthesize type I collagen
and regulates its mineralization into a specific
crystalline form
 OBs are autocrine regulatory cells. They can
synthesize and deposit growth factors in bone matrix
which when released by bone resorption processes,
restimulate further OB activity
 OBs mediate the systemic signals for the recruitment
and activity of OCs.
Osteblasts
 Osteoblasts also express receptors for
estrogens and Vit D3 in their nuclei.
 Toward the end of the secreting period the
osteoblasts will become either a flat lining
cell or an osteocyte
Bone formation
The formation of bone by mesenchymal
cells can occur by one of 2 routes
1. A direct development of bone from
mesenchymal cells, as in the calvarium,
called intramembranous ossification
2. An intervening cartilage model precedes the
formation, ie the proliferation of
mesenchymal cells is followed by their
differentiation into chondrocytes which
become hypertrophied and calcified. The
calcified cartilage is replaced by bone which
is then remodelled (endochondral
ossification)
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Bone formation
 Synthesis and intracellular processing of type I
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collagen
Secretion and extracellular processing of the
collagen
The formation of microfibrils, fibrils, and ultimately
fibers from the collagen
Maturation of collagen matrix with subsequent
nucleation and growth of HA crystals
OCs also plays an important role in development
and growth of bone by releasing polypeptide growth
factors from the extracellular mineralized matrix
(bone-derived growth factors, BDGFs)
Fig. 1-81 illustrates an osteon with osteocytes (OC) residing in
osteocyte lacunae in the lamellar bone. The osteocytes connect
via canaliculi (can) which contain cytoplasmatic projections of
the osteocytes. A Haversian canal (HC) is seen in the middle of
the osteon.
Fig. 1-82 illustrates an area of the alveolar bone in which bone
formation occurs. The osteoblasts (arrows), the bone-forming
cells, are producing bone matrix (osteoid) consisting of collagen
fi bers, glycoproteins, and proteoglycans. The bone matrix or the
osteoid undergoes mineralization by the deposition of minerals
such as calcium and phosphate, which are subsequently
transformed into hydroxyapatite.
Fig. 1-83 The drawing illustrates how osteocytes, present in the
mineralized bone, communicate with osteoblasts on the bone
surface through canaliculi.
The cellular events involved in bone
formation
 Cessation of continued OCs activity and
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disappearance of multinucleated OCs from
resorptive site
Attraction of OB precursors to the site
Proliferation of OB precursors
Differentiation of these precursors to form mature
collagen-secreting OBs
Formation of mineralized bone matrix
Cessation of OB activity
Cessation of OCs activity
Some OCs undergo a process of
programmed cell death or apoptosis at
remodeling sites, and then presumably
removed by scavenger cells
 The constituents of mineral phase of the bone
matrix could be involved in inhibition of
continued OCs activity:
1. Free ionized calcium as well as phosphate
2. TGF-b which is an inhibitor of OC activity and
is a powerful enhancer of OC apoptosis may
be produced by OCs, by OBs in the
neighborhood of OCs, or be released from the
bone matrix itself.
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Chemotaxis
 Chemotactic signals could include constituents
of the bone matrix such as fragments of
collagen or osteocalcin, or growth factors
produced in active from at the resorption site
such as TGF-b and PDGF
 include the attraction of OB precursors to the
site of resorption by a process of chemotaxis,
their stimulation is followed by differentiation to
mature cells which are capable of synthesizing
bone protein.
Proliferation
 Bone cell mitogens include insulin-like growth
factor (IGF-I, II), platelet-derived growth factor
(PDGF), and the heparin-binding fibroblast
growth factors (FGFs).
 transforming growth factor-b (TGF-b) is a
powerful bone cell mitogen
 These factors could stimulate to OB precursor
proliferation.
Differentiation
 The factors involved in stimulation of differentiation
of OB to form mature OBs.
 bone morphorgenic proteins (BMPs) are powerful
differentiation agents
 The only other agents which are known to have
similar effects are fluoride, 1,25 vitD, and retinoic
acid
 OBs synthesis and secrete the extracellular organic
matrix of bone, including type-1 collagen, osteocalcin,
osteopontin, alkaline phosphatase, proteoglycans as
well as the growth regulatory factors.
Mineralized bone matrix
 initial mineralization: involving matrix vesicle
 small, membrane-bound vesicle - providing a
microenvironment for initial mineralization
 containing alkaline phosphatase, pyrophosphatase, CaATP-ase, metalloproteinase, proteoglycans, and anionic
phospholipids - bind to calcium and inorganic phosphate form calcium-inorganic phosphate phospholipid complexes
(must maintain)
 deposition of apatite crystallites in relation to the collagen
fibrils and noncollagenous proteins
Alkaline phosphatase
 may or may not associated with matrix vesicles
 function: hydrolyzing phosphate ions from organic
radicals at an alkaline pH
 nonspecific; may have more than one distinct
function in mineralization
 breaking down pyrophosphate -removing its
inhibition of hydroxyapatite crystal growth permitting crystal growth to proceed
Regulatory factors involved in
normal bone formation
 TGF-b is one of the most abundant of the growth regulatory
factors in bone matrix.
 TGF-b exerts its major effect on bone formation by
stimulating proliferation of OB precursors and increasing
the pool of committed OBs, and it also increases OB
chemotaxis.
 BMPs are growth regulatory factors in extended TGF-b
family, and in particular BMPs 2,3, and 4 are expressed by
OBs as they differentiate.
 BMP2 and BMP4 seem to have effects on bone cell
differentiation
 heparin-binding fibroblast growth factors (FGFs)
are powerful bone forming factors. Both acidic and
basic FGFs stimulate bone formation in vivo which
are probably mediated predominately by
stimulation and proliferation of OB precursors.
 PDGF is expressed by bone cells and there are
also receptors for PDGF on bone cells
Cascade theory of bone
formation
 In the early phases, this probably involves a number
of chemotactic and proliferative factors such as
PDGF, TGF-b and IGF.
 As these cells multiply, they express mitogenic
growth factors such as TGF-b, PDGF and the FGFs.
 As the cells starts to differentiate, they express a
number of genes involved in the bone formation
process such as osteoclacin, alkaline phosphatase,
and type I collagen.
 They also express the BMPs, which are
autostimulatory. As a consequence, expression of
BMPs may be responsible for the final stages of
bone cell differentiation at the formation of normal
mineralized bone.
Osteoclasts and bone resorption
 The osteoclast is the bone lining cell
responsible for bone resorption
 It is a giant multinucleated cell (4-20 nuclei)
usually found in contact with a calcified bone
surface and with a lacuna (Howship’s
lacunae), which is the result of its own
resorptive activity.
 Possible 4 or 5 osteoclasts in the same
resorptive site, but usually only 1 or 2 per site.
 The cytoplasm is “foamy” with many
vacuoles
 The contact zone with bone is
characterized by the presence of a
ruffled border and dense patches on
each side of it known as the sealing
zone.
At the ultrastructural level
 The abundance of Golgi complexs
characterically disposed around each nuclei,
mitochondria, and transport vesicles loaded
with lysosomal enzymes.
 The attachment of the cell to the matrix is
performed via integrin receptors, binding to
specific sequences in matrix proteins.
Origin of the osteoclast
 Mononuclear-phagocyte
lineage are the most likely
candidates to differentiate
into osteoclasts.
 May occur at the
promonocyte stage, but
monocytes and
macrophages, already
commited to their own
lineage, might still be able
to form osteoclasts under
the right circumstances.
Resorptive events
 attachment of osteoclasts to the mineralized surface of
bone
 creation of a sealed acidic environment through action of
the proton pump, which demineralizes bone and exposes
the organic matrix
 degradation of this exposed organic matrix to its
constituent amino acids by the action of released enzymes
such as acid phosphatase and cathepsin B
 uptake of mineral ions and amino acids by the cell
Mechanisms of bone resorption
 Lysosomal enzymes: in the endopalsmic
reticulum, Golgi, and many transport vesicle;
these lysosomal enzymes are secreted, via
the ruffled border, into the extracellular bone
resorbing compartment.
 Nonlysosomal enzymes: metalloproteinase
such as collagenase, stromelysin and
elastase. All these proteinases would not
retain much activity in the low pH of the ruffle
border.
Mechanisms of bone resorption
 Proton pumps: the presence of an electrogenic
proton pump ATPase.
 The protons are provided to the pumps by the
enzyme carbonic anhydrase, highly concentrated in
the cytosol of this cell, and ATP and CO2 are
provided by mitochondria.
 The basolateral membrane activity exchanges
bicarbonate for chloride (HCO3 /Cl ), thereby
avoiding and alkalinization of cytosol.
 The basolateral sodium pumps might be involved is
secondary active transport of calcium and/or protons
in association with Na+, Ca++ exchanger and/ or Na+,
H+ antiport
Extracellular bone resorbing
compartment
 A low pH: dissolves the crystals, exposing the matrix
 Lysosomal enzymes: now at optimal pH, degrade the
matrix components.
 Substrate: the residues from this extracellular
digestion are either internalized, or transported
across the cell (transcytosis) and releasesed at the
basolateral domain, or else released during periods
of relapse of the sealing zone, possibly induced by a
calcium sensor in response to the rise via
extracellular calcium in the bone resorbing
compartment.
Bone remodeling
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A balance between synthesis and breakdown is
now widely called “coupling” of bone resorption
and formation.
Resorption involves three steps:
The bone resorbing agent, PTH, induces a change
of shape of the osteoblast, thus facilitating the
access of osteoclasts to the bone surface.
The osteoblasts systhesis the collagenase and
plasiminogen activator which digests the osteoid,
exposing the mineralized matrix, which may be
chemotactic to the OC.
The osteoblasts release the short-range soluble
activator to the OC.
Remodeling of the trabecular bone starts with resorption of the bone surface by
osteoclasts (OCL) as seen in Fig. 1-90a. After a short period, osteoblasts (OB) start
depositing new bone (Fig. 1- 90b) and fi nally a new bone multicellular unit is formed,
clearly delineated by a reversal line (arrows) as seen in Fig. 1-90c.
The osteoblast-osteoclast
relationship
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Osteoblast contains the receptors for the major bone
resorbing agents such as, PTH, the eicosanoids (PGE2,
PGI2, LTB4), 1,25 dihydroxyvitamin D3, and cytokines
(IL-1, TNF), and transmit resorptive signal to the
osteoclast.
OBs secrete a short-range souble activator
(lipoxygenase metabolites of arachidonic acid) for OCs
OBs appear to play a pivotal role in the regulation of
bone resorption.
Biology of bone healing
（granulation tissue）
Initial injury - Caused
extravasation and cell signaling
 Proteolytic degradation of ECM produces
chemotactic remnants, luring monocytes and
macrophages to wound bed
 Activated macrophages release FGF,
stimulating endothelial cells to express
plasminogen activator and procollagenase.
 Growth factors released from the a granules
of degranulating platelets are beacons for
PMNs, lymphocytes, monocytes, and
macrophages.
Proliferation
 Granulation tissue develops consisting of new blood
vessels, collagen isotypes and cells.
 At least 18 isotypes of collagens: type I is associated with
bone, type II with cartilage, types III and V with granulation
tissue, types IV and VI with the endothelial matrix, and type
X with hypertrophic cartilage.
 Collagenous substratum which is a key instructional
substratum of the repairing wound functions as a
provisional solid-state matrix for cellular attachment and
seletive binding of growth factors.
 The functional role of a callus is to stabilize the bone
fragments
Proliferation
 At the injury locus, the lure for monocytes
that will convert into osteoclasts appears to
be fragments of fibronectin and degradation
products from extracellular matrix.
 Macrophages at the wound site express FGF
and VEGF, prompting neoangiogenesis.
Remodeling
 The processes associated with hemostatic
remodeling are known as activation-resorptionformation.The processes will take between 3 to
6 months
 Osteoblasts are activated by signaling factors
（PTH）.
 Osteoclasts home in to the osteoblast vacant
zone, attach, resorb an in response to an as yet
unidentified signal, cease resorbing and
abandon their attachment.
 Osteoclastic resorptive pits become repopulated
by a contigent of osteoblasts that express
osteoid, which calcifies, retsoring bone.
The mediators of bone remodeling
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Parathyroid hormone
Parathyroid hormone related peptide (PTHrP)
Vitamin D metabolites
The cytokines
Eicosanoids
Growth factors
Bacterial products
Mechanical stress
Parathyroid hormone
 An 84 amino acid
 Physiological conc. appears to promote
bone formation, incrased conc. promote
osteroclastic bone resorption
 In high conc. PTH inhibits osteoblastic
collagen systhesis.
Parathyroid hormone related peptide
(PTHrP)
 Tumors factors produce hypercalcium and
reduce serum phosphate, and elevate
circulating levels of 1,25 dihydroxyvitamin
D3 and increase the excretion of cAMP and
phosphate.
 It can stimulate bone resorption
Vitamin D metabolites
 It is steroid-like compounds
 The major active metabolite of Vit D is
1,25 dihdroxycholecalciferol which
affects bone formation and also cause
bone resorption
 Its effect on bone resorption appears to
be by the differentiation of committed
progenitor cells into mature cells.
The cytokines
 Short range soluble mediators, which have
effects on bone remodeling and stimulate
bone resorption either directly or by
enhancing the recruitment of OCs.
 IL-1 exists in 2 forms: IL-1a and IL-1b. IL-1
has complex effects on bone remodeling and
it stimulates bone resorption and bone cell
replication.
 IL-1 does not have a direct action on the OCs,
but like PTH acts via the OB
 TNFa and TNFb can stimulate bone resorption
through OBs. It is of similar potency to IL-1 that
can be inhibited by IFNg.
 TNF also inhibits collagen and non-collagenous
protein systhesis, and this effect can be inhibited
by indomethcin and enhanced IFNg.
 IL-6 with sIL-6R can induce OCs formation
and bone resorption.
 IL-1 increase the synthesis of IL-6, which
increases bone resorption probably through
the recruitment of cells of the OC lineage.
The synthesis of IL-6 is diminished by
estrogens.
 Colony-stumilating factors play a role in the
maturation of OCs, and
granulocyte/macrophage-colony stumilating
factor secretion is stimulated by PTH.
Eicosanoids
 The PGs are local pathological mediators of bone
destruction, in particular where there is
inflammation.
 HETEs (hydroxyeicosatetraenoic acids) and LTs
(leukotrienes) are active important bone resorption
factors in inflammatory tissue.
Growth factors
 Transforming growth factors (TGF, TGF-a and
TGF-b) : are a family of polypeptides with
biological propeties similar to those of epidermal
growth factors (EGF).
 They have been shown to be potent bone
resorbing factors
 TGFs, EGF, and PDGF has also been reported to
be mediated entirely by PGs
Bacterial products
 Lipopolysaccharide (LPS) lipoteichoic acids, and
peptidoglycans have been shown to stimulate
bone resorption.
 These products may also act as foreign antigens
and induce monocyte macrophages and then
bone cells to produce PGs and cytokines such as
IL-1, leading to bone resorption.
Inhibitors of bone resorption
 Calcitonin(CT): is synthesized by C cells in the
thyroid gland and acts directly on OCs. It inhibits
OCs bone resorption.
 IFNg inhibits both proliferation and differentiation of
OCs progenitors.
 Bisphosphonates inhibits OCs bone resorption
probably by making the mineralized surface
inaccessible to the cell by binding to the HA crystals.
 IL-1 receptor antagonist (IL-1ra) which is produced
by monocytes, binds to IL-1 receptor. It is a very
effective inhibitor of OC bone resorption stimulated
by IL-1 and TNF.