bone healing and factors affecting it

1. Speaker-Raghavendra MS
Moderator-Dr Marulasiddappa G

2.  Fracture is a break in the continuity of bone or
periosteum
 The healing of fracture is in many ways similar
to soft tissue healing except that the end result
is mineralized mesenchymal tissue i.e BONE
 Fracture healing starts as soon as bone breaks
and continues for many months

3.  In 17th century Albrecht Haller, observed invading
capillary buds in fracture callus and thought that
blood vessels are responsible for callus formation
 John Hunter, a pupil of Haller, described the
morphologic sequence of fracture healing.
 In 1873, Kolliker observed the role of
multinucleated giant cells, osteoclast to be
responsible for bone resorption.
 In1939, Gluksman suggested pressure and
shearing stresses are possible stimuli for fracture
healing.
 In 1961, Tonna and Cronkie demonstrated the role
of local mesenchymal cells in fracture repair.

4.  Bone marrow
 Periosteum
 Cortex
 Soft tissue

5. Is a membrane that lines the outer surface of
all bones except at the joints of long bones.
Is made up of :
 Outer FIBROUS layer : made up of white
connective and elastic tissue.
 Inner CAMBIUM layer : which has a looser
composition, is more vascular and contains
cells with osteogenic potency.

6. 1. Anchors tendons and ligaments to bone.
2. Acts as a limiting membrane.
3. Participates in growth (appositional) and repair
through the activities of the osteoprogenitor cells .
4. Periosteum helps in fracture healing by forming
periosteal callus.
5. It also lessen the displacement of the # and helps in
reduction.
6. Allows passage of blood vessels, lymphatics and
nerves into and out of the bone.

8.  Are basophilic, cuboidal to pyramidal in shape ,
associated with bone formation, these cells are
located where new bone is forming, eg: in the
periosteum.
 Osteoblasts often appear stratified as in an
epithelium.
 The nucleus is large with a single prominent
nucleolus.
 Osteoblasts contain the enzyme alkaline
phosphatase used to calcify the osseous matrix.
 They synthesize type 1 collagen, osteocalcin (bone
Gla protein) and osteonectin

10.  Giant, multinuclear cells which vary greatly in
shape.
 They are found on the surfaces of osseous
tissue usually in shallow depressions called
Howship’s lacunae.
 The cytoplasm is slightly basophilic and
contains lysosomal vacuoles.
 Under E.M. the cell surface facing the osseous
matrix shows numerous cytoplasmic
projections and microvilli described as a
ruffled border.

11. RUFFLED BORDER
MICROSCOPIC PICTURE ELECTRON MICROSCOPY
OSTEOCLASTS
11

12.  Basically, an osteoblast that has been enclosed
within the bony matrix in a space called the lacuna.
 The cytoplasm of the osteocyte is faintly basophilic
containing fat droplets and granules of glycogen
with single dark stained nucleus.
 In developing bone, the cytoplasmic processes from
one osteocyte make contact with the processes (i.e.:
cannaliculi) from adjoining osteocytes. In mature
bone, the processes are withdrawn almost
completely.
 In mature bone the empty canaliculi remain as
passage ways for the diffusion of nutrients and
wastes between bone and blood.
12

13. 13

14.  Are flattened epithelium in adult skeleton
found on resting surfaces.
 Plays active role in differentiation of
progenitor cells
 Controls osteoclasts, mineral hemostasis
and may secrete collagenase.
 Lines – endosteal surface of marrow cavity
- periosteal surface
- vascular channels within osteons.
14

15.  Endoochondral
 Intramembranous ossification
 Oppositional new bone formation
 Osteonal migration
(creeping substitution)

16.  Mechanism by which a long bone grows in
width.
 Osteoblasts differentiate directly from pre
osteoblasts and lay down seams of osteoid.
 Does NOT involve cartilage anlage.
16

17. 17

18.  Mechanism by which a long bone grows in
length.
 Osteoblasts line a cartilage precursor.
 The chondrocytes hypertrophy, degenerate and
calcify (area of low oxygen tension).
 Vascular invasion of the cartilage occurs
followed by ossification (increasing oxygen
tension).
18

19. Picture courtesy Gwen Childs,
PhD.

20.  The process of bone remodelling by osteoclastic
resorption and creation of new vascular
channals with osteoblastic bone formation
resulting in new haversian systems
 Seen in bone healing after bone grafting

21.  Primarily a
mechanism to
remodel bone.
 Osteoclasts at the
front of the cutting
cone remove bone.
 Trailing osteoblasts
lay down new bone.
21

22. 22

23. • Stage of hematoma formation
• Stage of inflammation and cellular
proliferation
Reactive
phase
• Stage of callus formation
• stage of consolidation
Reparative
phase
• Stage of remodeling
Remodeling
phase
1975, Cruess and Dumont 1989, FROST

24.  Torn vessels will
bleed and form
hematoma
 A mass of clotted
blood(hematoma)
formed around the
fracture site

25.  Hematoma is invaded
inflammatory cells &
they degrade necrotic
tissue
 Release of TGF-
β,PDGF by
macrophages
 Procuring
osteoprogenitor cells

26.  new capillaries organize fracture hematoma
into granulation tissue – procallus
 Fibroblasts and osteogenic cells invade
procallus.
 Make collagen fibers which connect ends
together
 Chondroblasts begin to produce fibrocatilage
 This is called soft callus(7days-6wks)

28.  Osteoblasts lay more
boney trabacule
 Fibrocartilagenous
frame work is
calcified to form
boney callus(6-12wks)
 Now fracture is
painless and allows
weight bearing

29.  Globular callus is
slowly remodeled
over years
 More trabacule are
layed in the line of
stress and non
stressed area is
resorbed
 Hence bone will take
original shape

30.  A bone will adapt to mechanical stress and
strain by changing size, shape and structure

32.  Direct (primary) bone healing
 Indirect (secondary) bone healing

33.  Mechanism of bone healing seen when there
is no motion at the fracture site (i.e. absolute
stability)
 Does not involve formation of fracture callus
 Osteoblasts originate from endothelial and
perivascular cells

34.  Contact Healing
 Direct contact between the fracture ends allows healing
to be with lamellar bone immediately
 Gap Healing
 Gaps less than 200-500 microns are primarily filled with
woven bone that is subsequently remodeled into
lamellar bone
 Larger gaps are healed by indirect bone healing
(partially filled with fibrous tissue that undergoes
secondary ossification)

36.  Mechanism for healing in
fractures that have some
motion, but not enough to
disrupt the healing process.
 Bridging periosteal (soft)
callus and medullary (hard)
callus re-establish structural
continuity
 Callus subsequently
undergoes endochondral
ossification
 Process fairly rapid - weeks

37.  Growth factors
Transforming growth factor
Bone morphogenetic proteins
Fibroblast growth factors
Platelet-derived growth factors
Insulin-like growth factors
 Cytokines
Interleukin-1,-4,-6,-11, macrophage and
granulocyte/macrophage (GM) colony-stimulating factors (CSFs)
and Tumor Necrosis Factor
 Prostaglandins/Leukotrienes
 Hormones
 Growth factor antagonists

38.  Super-family of growth factors (~34 members)
 Acts on serine/threonine kinase cell wall
receptors
 Promotes proliferation and differentiation of
mesenchymal precursors for osteoblasts,
osteoclasts and chondrocytes
 Stimulates both enchondral and
intramembranous bone formation
 Induces synthesis of cartilage-specific proteoglycans
and type II collagen
 Stimulates collagen synthesis by osteoblasts

39.  These are included in the TGF-β family
 Except BMP-1
 Sixteen different BMP’s have been identified
 BMP2-7,9 are osteoinductive
 BMP2,6, & 9 may be the most potent in
osteoblastic differentiation
 Involved in progenitor cell transformation to pre-
osteoblasts
 Work through the intracellular Smad pathway
 Follow a dose/response ratio

40.  Osteoinductive proteins initially isolated from
demineralized bone matrix
 Induce cell differentiation
 BMP-3 (osteogenin) is an extremely potent inducer of
mesenchymal tissue differentiation into bone
 Promote endochondral ossification
 BMP-2 and BMP-7 induce endochondral bone
formation in segmental defects
 Regulate extracellular matrix production
 BMP-1 is an enzyme that cleaves the carboxy termini of
procollagens I, II and III

41. Table from Dimitriou, et al.,
Injury, 2005

42.  Used at doses between 10x & 1000x native
levels
 Negligible risk of excessive bone formation
 rhBMP-2 used in “fresh” open fractures to
enhance healing and reduce need for secondary
procedures after unreamed IM nailing
 It is found that application of rhBMP-2 decreases
infection rate in Type IIIA & B open fractures.
 BMP-7 approved for use in recalcitrant
nonunions in patients for whom autografting is
not a good option (i.e. medically unstable,
previous harvesting of all iliac crest sites, etc.)

43.  BMP-2
 Increased fusion rate in spinal fusion
 BMP-7 equally effective as ICBG in nonunions
(small series: need larger studies)
 Must be applied locally because of rapid
systemic clearance

44.  May have important role in bone formation
 Noggin
 Extra-cellular inhibitor
 Competes with BMP-2 for receptors
 BMP-13 found to limit differentiation of
mesenchymal stromal cells
 Inhibits osteogenic differentiation

45.  Both acidic (FGF-1) and basic (FGF-2)
forms
 Increase proliferation of chondrocytes and
osteoblasts
 Enhance callus formation
 FGF-2 stimulates angiogenesis

46.  A dimer of the products of two genes, PDGF-A
and PDGF-B
 PDGF-BB and PDGF-AB are the predominant forms
found in the circulation
 Stimulates bone cell growth
 Mitogen for cells of mesenchymal origin
 Increases type I collagen synthesis by
increasing the number of osteoblasts
 PDGF-BB stimulates bone resorption by
increasing the number of osteoclasts

47.  Two types: IGF-I and IGF-II
 Synthesized by multiple tissues
 IGF-I production in the liver is stimulated by
Growth Hormone
 Stimulates bone collagen and matrix synthesis
 Stimulates replication of osteoblasts
 Inhibits bone collagen degradation

48.  Interleukin-1,-4,-6,-11, macrophage and
granulocyte/macrophage (GM) colony-
stimulating factors (CSFs) and Tumor Necrosis
Factor
 Stimulate bone resorption
 IL-1 is the most potent
 IL-1 and IL-6 synthesis is decreased by estrogen
 May be mechanism for post-menopausal bone
resorption
 Peak during 1st 24 hours then again during
remodeling
 Regulate endochondral bone formation

49. Cytokine Bone Formation Bone Resorption
IL-1 + +++
TNF-α +
+++
TNF-β +
+++
TGF-α --
+++
TGF-β ++
++
PDGF ++ ++
IGF-1 +++ 0
IGF-2 +++ 0
FGF +++ 0

50.  Effect on bone resorption is species dependent
and their overall effects in humans unknown
 Prostaglandins of the E series
 Stimulate osteoblastic bone formation
 Inhibit activity of isolated osteoclasts
 Leukotrienes
 Stimulate osteoblastic bone formation
 Enhance the capacity of isolated osteoclasts to form
resorption pits

51.  Estrogen
 Stimulates fracture healing through receptor
mediated mechanism
 Modulates release of a specific inhibitor of IL-1
 Thyroid hormones
 Thyroxine and triiodothyronine stimulate
osteoclastic bone resorption
 Glucocorticoids
 Inhibit calcium absorption from the gut causing
increased PTH and therefore increased osteoclastic
bone resorption

52.  Parathyroid Hormone
 Intermittent exposure stimulates
 Osteoblasts
 Increased bone formation
 Growth Hormone
 Mediated through IGF-1 (Somatomedin-C)
 Increases callus formation and fracture
strength

53.  Metalloproteinases
 Degrade cartilage and bones to allow
invasion of vessels
 Angiogenic factors
 Vascular-endothelial growth factors
 Mediate neo-angiogenesis & endothelial-
cell specific mitogens
 Angiopoietin (1&2)
 Regulate formation of larger vessels and
branches

54. Systemic Factors
 A. Age
 B. Activity level including
 1. General immobilization
 2. Space flight
 C. Nutritional status
 D. Hormonal factors
 1. Growth hormone
 2. Corticosteroids (microvascular
osteonecrosis)
 3. Others (thyroid, estrogen,
androgen, calcitonin,
 parathyroid hormone,
prostaglandins)
 E. Diseases: diabetes, anemia,
neuropathies, tabes
 F. Vitamin deficiencies, A, C, D, K
 G. Drugs: nonsteroidal
antiinflammatory drugs (NSAIDs),
 anticoagulants, factor XIII, calcium
channel blockers
 (verapamil), cytotoxins,
diphosphonates, phenytoin,
 sodium fluoride, tetracycline
 H. Other substances (nicotine,
alcohol)
 I. Hyperoxia
 J. Systemic growth factors
 K. Environmental temperature
 L. Central nervous system trauma

55.  Local Factors
A. Factors independent of injury, treatment, or
 complications
 1. Type of bone
 2. Abnormal bone
 a. Radiation necrosis
 b. Infection
 c. Tumors and other pathological conditions
 3. Denervation
B. Factors depending on injury
 1. Degree of local damage
 a. Compound fracture
 b. Comminution of fracture
 c. Velocity of injury
 d. Low circulatory levels of vitamin K1
 2. Extent of disruption of vascular supply to bone, its
 fragments (macrovascular osteonecrosis), or soft
 tissues; severity of injury
 3. Type and location of fracture (one or two bones,
 e.g., tibia and fibula or tibia alone)
 4. Loss of bone
 5. Soft tissue interposition
 6. Local growth factors

56.  C. Factors depending on treatment
 1. Extent of surgical trauma (blood supply, heat)
 2. Implant-induced altered blood flow
 3. Degree and kind of rigidity of internal or external
 fixation and the influence of timing
 4. Degree, duration, and direction of load-induced
 deformation of bone and soft tissues
 5. Extent of contact between fragments (gap, displacement,
 overdistraction)
 6. Factors stimulating posttraumatic osteogenesis
 (bone grafts, bone morphogenetic protein, electrical
 stimulation, surgical technique, intermittent
 venous stasis [Bier])
 D. Factors associated with complications
 1. Infection
 2. Venous stasis
 3. Metal allergy

57.  Electromagnetic (EM) devices are based on
Wolff’s Law that bone responds to mechanical
stress: In vitro bone deformation produces
piezoelectric currents and streaming potentials.
 Exogenous EM fields may stimulate bone
growth and repair by the same mechanism
 Clinical efficacy very controversial
 No studies have shown PEMF to be effective in “gap
healing” or pseudarthrosis

58.  Microamperes
 Direct electrical current
 Capacitively coupled electric fields
 Pulsed electromagnetic fields (PEMF)

59.  Approved by the FDA for the treatment of non-
unions
 Efficacy of bone stimulation appears to be
frequency dependant
 Extremely low frequency (ELF) sinusoidal electric
fields in the physiologic range are most effective (15
to 30 Hz range)
 Specifically, PEMF signals in the 20 to 30 Hz range
(postural muscle activity) appear more effective than
those below 10 Hz (walking)

60.  Low-intensity ultrasound is approved by the
FDA for stimulating healing of fresh fractures
 Modulates signal transduction, increases gene
expression, increases blood flow, enhances
bone remodeling and increases callus torsional
strength in animal models

61.  Human clinical trials show a decreased time
of healing in fresh fractures treated
nonoperatively
 Four level 1 studies show a decrease in healing
time up to 38%
 Has also been shown to decrease the healing
time in smokers potentially reversing the ill
effects of smoking