I AM A STRONG TOOTH

MOLECULAR BIOLOGY OFCEMENTUM

ANDALVEOLAR BONE

Dr. KRITIKA JANGIDMDS- Periodontics

Cementum

• Its name is derived from a Latin word, “Caementum”, meaning quarried stone or chips of stone used in making mortar.

• It was demonstrated in 1835.

Cementum

• It is a hard, mineralised, avascular connective tissue found on the anatomic roots of the teeth.

Uniqueness of Cementum• Medium of attachment

• It is avascular and not innervated

• Does not undergo continuous remodelling like bone, but continues to grow in thickness throughout life.

Physical Characteristics

• Softer than Dentin (Selvig and Selvig 1962)• It is light yellow in color.• It is different from enamel by its lack of luster

and its darker hue.• It is lighter than dentin.

Thickness of Cementum

Coronal half 16 - 60µm

Apical 20- 150µm

Distal surface is thicker than mesial, probably because of functional stimulation like mesial drift.

Composition

• Its content is– 45–50% Inorganic:

• Ca, PO4 as Hydroxy-apatite.• Numerous trace elements• Highest fluoride content

– 50-55% Organic and Water:• Collagen Type I• Proteoglycans

Minerals:

• The mineral component is the same as in other calcified

tissues i.e. hydroxyapatite Ca10(PO4)6(OH)2

• with small amounts of amorphous calcium phosphates.

Minerals:• Due to its lower crystallinity, cementum has a greater capacity

for adsorption of fluoride and other elements.

• Cementum has a high fluoride content compared to other

mineralized tissues (up to 0.9%), shows a general increase with

age and vary with fluoride supply to the individual.

• Cementum contains 0.5% magnesium, about half that in

dentin, and it is lower at the surface than in deeper layers of

cementum.

– Significance - composition of cementum is more similar to

bone tissue than to dentin.

• Cementum also contains 0.1-0.3% sulfur as a constituent of

the organic matrix.

• Trace elements may be present in concentrations detectable

by electron microprobe analysis, in particular Cu, Zn and Na.

Organic Components:

• The organic matrix of cementum is composed primarily of

collagen.

• Predominantly type I and type III (Birkedal and Hansen et al

1977)

• Type I collagen plays structural as well as morphogenic role and

provides scaffolding for mineral crystals

• It is the major component accounting for 90% of organic

components in cellular cementum.

• The type III collagen, which coats type I collagen fibrils, accounts for only 5%. (Rao et al 1979, Wang et al 1980)

• It is a less cross-linked collagen found in high concentrations during development, repair and regeneration of mineralized tissues

• Type XII collagen- A fibril associated collagen with interrupted

triple helices that binds to type 1 collagen and also to non

collagenous proteins

• Trace amounts of Type XIV are also found in extracts of mature

cementum, however these may be contaminants from periodontal

ligament.

• Based on immunostaining, the cementum does not appear to have

either Type V or Type VI collagen (Becker and Romanos et al 1991)

Non collagenous proteins:

– Bone sialoprotein (BSP)– Osteopontin(OPN)– Alkaline phosphatase– Fibronectin– Osteocalcin– Osteonectin– Proteoglycans– Proteolipids– Vitronectin – Growth factors

• BSP and OPN are expressed in AEFC and AAFC.

• Both are phosphorylated and sulfated glycoproteins.

• They bind tightly to the collagenous matrices and

hydroxyapatite, and possess cell attachment properties

through the Arg-Gly-Asp sequence, that binds to integrins.

Bone Sialoprotein(BSP)

– Root surface cells express the BSP, and it is also present in mature teeth.

– BSP is believed to have adhesion function to root surface cells and participate in initiating mineralization.

– It is chemotactic to pre-cementoblasts and promotes their adhesion and differentiation.

Osteopontin• OPN is present within the periodontal ligament region of the

mature teeth. • Many cells express the OPN during periods of cementogenic

activity. • It regulates cell migration, differentiation, and survival through

the interaction with integrins.• Participates in inflammation by regulating monocyte-

macrophage activation, phagocytosis, and nitric oxide production.

• It may regulate biomineralization by regulating bone cell differentiation and matrix mineralization.

• Fibronectin• Believed to bind cells to the extracellular matrix

• Tenascin• Present in HERS during odontoblast differentiation.

• And later at the attachment site of periodontal ligament with the root

surface.

• Osteonectin• Expressed by cementoblasts producing cellular extrinsic fiber

cementum and cellular intrinsic fiber cementum

• Osteocalcin:– Appears to be involved in the mineralization process.

• Biochemical analysis of extracts of human cementum have identified – Chondroitin sulfate, – Dermatan sulfate and – Hyaluronic acid.

• Enzymes:

– Alkaline phosphatase is believed to participate in

cementum mineralization.

– The enzyme activity adjacent to cellular intrinsic fiber

cementum is higher than that to acellular extrinsic fiber

cementum.

• Growth factors

– BMP-2, 3 and 4, PDGF, α and β-FGFs, TGFβ, PTH and IGF-1.

• Molecules unique to cementum have also been described.

• One of these are an IGF-1, referred early as Cementum Growth Factor (CGF).

• The second molecule is a collagenous protein referred to as Cementum Attachment Protein (CAP). CAP promotes the adhesion and spreading of mesenchymal cells.

.

Cemento-Enamel Junction60 – 65% Cementum overlaps the enamel.

Overlapping of cementum on enamel is due to local degeneration of the reduced enamel epithelium

With the result that, the connective tissue elements of the dental follicle enter and effect cementogenic activity.

30% - 35% there is an edge-to-edge butt joint.

5 – 10 % the cementum and enamel fail to meet.

Cemento-Dentinal Junction

• It is relatively smooth in permanent teeth.

• Sometimes scalloped in deciduous teeth.

• Contains large quantities of collagen associated with GAGs like chondroitin sulfate and dermatin sulfate resulting in increases water content and contributes to the stiffness.

• In histological sections, the cementum usually

stains more intensely than dentin.

• The cementum is more electron dense than

dentin.

• The collagen fibrils are in distinct bundles in

cementum where as

• The collagen fibrils are haphazard in dentin.

CEMENTOGENESIS

Current theories suggest

1. Infiltrating dental follicle cells recieve a reciprocal inductive signal from the dentin or the surrounding HERS and differentiate into cementoblasts.

2. HERS cells transform into cementoblasts.

• Evidence is increasing that ECRM are not simply residual cells but may also participate in maintenance and regeneration of periodontal tissues.

• Some HERS cells remain attached to the forming root surface, they can produce focal deposits of enamel like material called enamel pearls.

Cementoblasts

• Arise from the undifferentiated mesenchymal cells

• Synthesise collagen and protein polysaccharides- organic matrix of cementum.

• Mitochondrea, golgi, RER

• Inner cells of dental follicle: CIFC• HERS- AEFC

Cementocytes• In the apical 1/3rd,cementoblasts trapped in rapidly

calcifying cemental matrix, later, differentiate into cementocytes.

• These locate in spaces termed lacunae & have numerous cytoplasmic processes coursing in canaliculi, that are preferentially directed towards the periodontal ligament.

• This is how cementocytes derive their nutrition from periodontal ligament & contribute to the vitality of this mineralized tissue.

• While adjacent canaliculi of neighboring cells communicate frequently, the processes remain independent.

• Thus, the metabolites progress mostly by diffusion through the canaliculi of cellular cementum.

Cementoclasts:

• They are multinucleated giant cells, which are indistinguishable from osteoclasts.

• Responsible for root resorption that leads to primary teeth exfoliation & also in the permanent dentition in mesial surfaces in compliance with mesial migration & may occur due to occlusal trauma & orthodontic therapy.

MOLECULAR INSIGHT TOCEMENTOGENSIS

GROWTH FACTORS

• BMPs- Members of TGFβ superfamily that act through transmembrane serine/threonine protein kinase receptors.

• BMP-2, BMP-4, BMP-7• Known to promote differentiation of

preosteoblasts and cementoblasts precursor cells• PDGF± IGF- Promote cementum formation by

altering cell cycle activities.• FGF: Cell proliferation, migration and

vasculogenesis.

• BMPs have been used successfully to induce periodontal regeneration in a number of experimental models and certain clinical situations.

EPITHELIAL FACTORS

• Dental epithlial • Ectomesenchymal cells

• Enamel proteins• PTH related protein• Basal lamina constituents

ADHESION MOLECULES

• Contain cell adhesion motifs arginine-glycene-aspartic acid

• Bone sialoprotein- Promotes mineralisation

• Osteopontin- Regulate the extent of mineral growth.

COLLAGENS

• I• III• XII: Assist in maintaining the PDL space versus

continuous formation of cementum

Gla proteins

• Matrix/ Bone Gla proteins• Contain γ- carboxyglutamic acid• Matrix Gla protein: Inhibitor of mineralisation.

Significant role in preventing abnormal ectopic calcification

• Osteocalcin: Marker for cells associated with mineralisation (osteoblasts, cementoblasts, odontoblasts) hence considered to be a rgulator of crystal growth

TRANSCRIPTION FACTORS

• Runt-related transcription factor 2 (Core binding factor alpha 1)

• Osterix

• Involved in cementoblast differentiation

• BMPs have been identified as factors promoting Runx-2

SIGNALLING MOLCULES

• RANK• RANKL• OPG

Types of CementumEmbryologically

Primary & Secondary

According to location on teeth ( Kronfield 1928).

- Radicular cementum- found on root surfaces.- Coronal Cementum to Cementum that forms on the

enamel covering the crown.

On the basis of cellularity (Gottlieb 1942).

- Acellular / Primary Cementum.- Cellular / Secondary Cementum.

Schroder(1986) classified cementum x 5 subtypes based on cellularity & organisation of collagen fibres into- Acellular afibrillar cementum.

- Acelluar extrinsic fiber cementum.- Acellular intrinsic fiber cementum.- Cellular intrinsic fiber cementum.

- Cellular mixed stratified cementumBased on the origin of the collagen matrix

- Extrinsic.- Intrinsic.- Mixed.

Depending on the location & patterning

- Intermediate.- Mixed stratified cementum.

Differences Between Acellular & Cellular Cementum…..

Acellular Cementum Cellular Cementum

Formation Forms before tooth reaches occlusal plane

After tooth reaches occlusal plane

Cells Does not contain any cells Contains cementocytes

Location Coronal portion of root Apical portion of root

Rate of formation

Slow Rapid

Incremental lines

More Sparse

Cont…..Acellular Cementum Cellular

CementumFunction Forms after regenerative

periodontal surgical procedureContributes to the

length of the root during growth

Calcification More calcified Less calcified

Sharpey’s fibers More Less

Regularity of fibers

Regular Irregular

Thickness 20 – 50µm near the cervical region &150 – 200µm near the

apex.

Thickness of 1 – several mm.

Acellular Afibrillar Cementum (AFC):

• Contains neither cells, nor extrinsic / intrinsic fibers, apart from a mineralized ground substance.

• It is a product of cementoblasts, found deposited on the enamel over small areas of the dental crown just coronal to the CEJ.

• Thickness is about 1 - 15 µm.

Acellular Extrinsic Fiber Cementum (AEFC):-

Composed almost entirely of densely packed bundle of Sharpey's fiber and no cells.

• A product of fibroblasts and cementoblasts,• found on the cervical ⅓ of roots, but may

extend further apically..• Cementoblasts that produce AEFC

differentiate in close proximity to the advancing root edge.

• During root development, the first formed cementoblasts align along the newly formed, but not yet mineralized, mantle dentin surface & exhibit fibroblastic characteristics.

• Deposit collagen fibrils within it so that dentin & cementum fibers intermingle.

• Initially AEFC consists of mineralized layer with a short fringe of collagen fibers implanted perpendicular to the root surface.

• Cementoblasts then migrate away from the surface but continue to deposit collagen so that a fine fiber bundle lengthens & thickens.

• These cells also secrete non – collagenous matrix proteins that fill in the spaces between the collagen fibers.

• AEFC has the potential to adapt to functionally dictated alterations such as mesial tooth drift.

Cellular Mixed Stratified Cementum (CMSC):

• Contains both collagen fibers & calcified matrix.

• It is the co – product of cementoblasts & fibroblasts and consists of both extrinsic & intrinsic fibers.

• Appears primarily in the apical third of the roots & in furcation areas.

• Consists of AEFC and CIFC that alternate & appear to be deposited in irregular sequence upon one another.

- Schroeder, (1993).

• Deposited @ 0.1 – 0.5 µm / year.

Cellular Intrinsic Fiber Cementum (CIFC):

• Contains cells but no extrinsic (Sharpey's) fibers.

• Once the tooth is in occlusion, a more rapidly formed & less mineralized variety of cementum, (CIFC) is deposited on unmineralized dentin surface near the advancing root edge.

• Formed by cementoblasts & fills resorption lacunae (resorptive cementum.)

• Can easily repair a resorptive defect of the root due to its capacity to grow faster than any other form of cementum.

Acellular Intrinsic Fiber Cementum (AIFC):

• An acellular variant of cellular intrinsic fiber cementum that is also deposited during adaptive responses to external forces (i.e.,) slow deposition rate so that cells are not engulfed in their matrix & that forms without leaving cells behind.

- Bosshardt & Schroder, (1990) .

• In the light microscope, CIFC is identified easily because of the inclusion of cementocytes within lacunae with processes directed towards the tooth surface, laminated structure & presence of cementoid on its surface.

• Fine, densely packed intrinsic fibers running parallel to the root surface & larger, haphazardly incorporated extrinsic fibers running at right angles to the root surface.

• Cellular intrinsic fiber cementum is initially deposited on root surface areas where no acellular extrinsic fiber cementum has been laid down on the dentin (furcation and on the apical root portions).

Clinical Considerations

EXPOSURE TO ORAL ENVIRONMENT:

• Cementum becomes exposed to oral environment in gingival recession & as a result of loss of attachment in pocket formation.

• Cementum is sufficiently permeable to be penetrated by organic substances, inorganic ions & bacteria, leading to hypersensitivity to thermal changes / tactile stimulation, root caries, etc... sometimes resulting in pulpal pathology.

Changes in the Periodontium During Pocket Formation……….

• The changes may be grouped as

• Structural• Chemical • Cytotoxic

Structural Changes:

• Presence of pathologic granules, representing areas of collagen degeneration / areas not fully mineralized initially.

- Bass, (1951).

• Areas of Increased Mineralization: -

Selvig, (1966) as a result of exchange on exposure to the oral cavity, of minerals & organic component at the cementum saliva interface.

• Areas of Demineralization:-

Exposure to oral fluids & bacterial plaque results in proteolysis of the embedded Sharpey’s fibres, leading to softening of the cementum, which undergoes fragmentation & cavitation (root caries) → Pulpal sensitivity / severe pain.

Chemical Changes……

• Exposed cementum has an increased mineral content (Selvig 1966)- Ca, Mg, P, F. & may be resistant to decay.

Cytotoxic Changes:

• These include bacterial penetration into cementum as deep as the CDJ.

• In addition, bacterial products such as endotoxins are also found deep in the cemental wall of  the periodontal pocket.

Anomalies

Scurvy (Vit.C deficiency)

• It affects the deposition of bone, dentin and cementum.

• It also produces atrophy of formative cells.• This can lead to increase in bone resorption

leading to tooth loss.

Rickets • The cementum is hypomineralized.• The bone matrix also exhibits

hypomineralization.

Hypophospatasia• There is a total absence of cementum.• It is a rare hereditary disease with loosening

and premature loss of deciduous teeth.

Hypercementosis

• It is abnormal thickening of cementum.• It may be diffuse or circumscribed.• It may affect all teeth/ one tooth/ few

portions of a single tooth.• It could be cemental hypertrophy or cemental

hyperplasia.

Cementicles

• These are calcified bodies appearing on/in cementum and PDL.

• It is usually ovoid or round.• It is classified as,

– Free– Attached– Embedded

• It forms as a response to local trauma or hyperactivity.

• Its number increases with age.

The nidus for the calcifying process are dead cells associated with epithelial rests of malassez, mineralized sharpeys fibres and phleboliths.

A wall of Cementum of varying thickness form around this with diameter ranging between 0.2- 0.3mm

• They may fuse into interstitial cementicles.

Resorption and Repair• Cementum resists resorption under normal

conditions.• But it can resorb after trauma or excessive occlusal

loads.• If the original contour of the root surface is re-

established, it is called Anatomical repair.• In Functional repair, only a thin layer is deposited

and a bay like recess remains. The outline is followed by the alveolar bone.

Other Anomalies

• Cemental tears can occur.• Transverse fractures of root may occur and it

heals by formation of new cementum.

BONE

Circumferential lamellae

Concentric lamellae

OSTEON

Interstitial lamellae

•outer "fibrous layer" and• inner "cambium layer" (or "osteogenic layer").

PERIOSTEUM

Are responsible for formation, resorption and maintenance of osteoarchitecture

• Osteogenic cells

• Osteoprogenitors• Preosteoblasts• Osteoblasts• Osteocytes• Bone lining cells

• Osteoclast

BONE

CELLS

OSTEOBLASTS

• Mononucleated cell that synthesize collagenous and non collagenous proteins of the bone matrix proteins

• Exhibit high levels of alkaline phosphatase.

• Cleave organically bound phosphate.

• Liberated phosphate contributes to the initiation and progressive growth of bone mineral crystals.

• Plump• Cuboidal (when active) or slightly flattened• Primary role: Production of organic matrix of

bone• Abundant RER, Golgi complexes, rich in

secretory granules• Type 1 collagen dominant• Type III collagen in small amounts• Non- Collagenous proteins and PG also present

• Cytokines and growth factors• BMP-2• BMP-7• Transforming growth factor β• IGF-1• IGF-2• PDGF

• IGF-1 + PDGF: Increase the rapidity of bone formation and bone repair

• Used in bone healing and bone growth after periodontal surgical procedures.

• Enhance osseous integration after placement of dental implants

Hormones

• PTH and Vit D: Enhance resorption at high conc., bone formation at low conc.

• Calcitonin and estrogen: Inhibit resorption• Glucocorticoids: Inhibit resorption and

formation

Leptin

• A circulating hormone• Produced by adipocytes• Regulates food intake and body weight;

control body mass• Acts on hypothalamus• Inhibit differentiation of osteoclasts• Promote differentiation of osteoprogenitor

cells

Bone lining cells

• Last phenotype of osteoblast lineage prior to activation of bone remodeling cycle

• Form sheets over bone. Estimated 80% of bone surface is covered by lining cells

• Scarce organelles with less protein synthesizing machinery

• Flattened cells that line the quiescent bone surfaces• These quiescent surface is the primary site of

mineral ion exchange between blood and bone.

OSTEOCYTES

• As osteoblasts produce bone, some cells get entrapped in matrix they secrete , whether mineralised or unmineralised.

• Smaller than osteoblasts

OSTEOCLASTS

• Large multinucleated cell

MORPHOLOGY(EM)

An active OCL occupies hollowed out depressions called HOWSHIP’S LACUNAE

Secretory functions

• Cathepsins B,D,K have been localised in vacoules near the ruffled border

• These degrade collagen at acidic Ph

• Express MMP1, MMP2,MMP3,MMP9,• Digest collagen I,IV &V

• Cathepsins act first at low pH, followed by MMP’s

• Carbonic Anhydrase II in preosteoclasts & osteoclasts

• This enzyme generates hydrogen ions, which by proton pump is extruded in to sealed pit via the ruffled border, creates an acidic environment

Sequence of events

• Attachment of OCL to mineralised surfaces of bone

• Creation of acidic environment, which demineralises bone to expose matrix

• Degradation of exposed matrix by acid phosphatase & Cathepsins

• Endocytosis of degradative products by ruffled border

• Translocation & release of degradative products at opposite border

MATRIX COMPONENTS

COLLAGEN

80- 90% of organic component

TYPE III & XII

TYPE I

TYPE V(5%)

• Type I provides structural integrity

• Type III is present as mixed fibers with type I in sharpey’s fibers

• Type XII is related to mechanical strain

I, V, XII

III & Some

XII

OSTEOBLAST FIBROBLAST

NONCOLLAGENOUS PROTEINS

• Osteocalcin

• Osteonectin

• Osteopontin

• Bone Sialoprotein

OSTEOCALCIN

• Bone gla protein,15% of NC proteins• Small/molecular mass of 6Kd• 3 γ carboxy glutamic acid residues(gla groups)• Dual role in bone formation & resorption

• Expressed by osteoblasts and odontoblasts

• Secreted in to bone matrix at the time of mineralisation (Boskey et al 1992)

• Carboxy terminal segment act as chemottractant to OCL precursors

OSTEONECTIN

• SPARC(Specific Protein Acidic Rich in Cysteine

• 40Kd protein• 20% of NC proteins

• Also known as endothelial ”culture shock” protein, basement membrane protein (BM 40)

Four domains

• 1 - Calcium binding• 2 - Cysteine rich domain• 3 - α helical domain• 4 - EF hand domain

OSTEOPONTIN

• Phosphoprotein with high content of serine, asparagine & glutamate

• 55Kd low phosphorylated species produced by preosteoblasts

• 44Kd highly phosphorylated species that binds to HA(Sodek et al 1995)

• 8 α helices with RGD sequence in the middle flanked by 2 segments of β sheet structures

• Amino terminus binds to calcium & HA

DUAL ROLE IN REMODELING

• Highest expression is seen in preosteoblasts during bone formation and mature osteoblasts during bone remodeling

• Through the RGD sequence , binds to α5β3 of OCL, plays a role in remodeling

BONE SIALOPROTEIN

• Highly glycosated , acidic phosphoprotein with high sialic acid content

• Highly expressed only in mineralised tissues• RGD sequence, helps in attachment of Ob tomineralised tissues • Glutamic acid domainare sites for HA binding

• Implicated in nucleation of HA during de novo bone formation & in initial mineralisation of newly formed bone

PROTEOGLYCANS• DECORIN• 46Kd protein core• Single c.sulfate chains at

amino terminus• Binds to the gap region

of collagen fibrils• Binds to TGF β• Regulates fibrillogenesis

• BIGLYCAN• 46Kd protein core• Two c.sulfate chains at

amino terminus• Binds to TGF β, collagen• Regulates fibrillogenesis

Both comprise <10% of non collagenous proteins & decreases with maturation of bone

OTHER PROTEINS

• Procollagen Peptides• Thrombospondin • Fibronectin• Vironectin,

Modulate cell attachment

BONE COUPLING

REFERENCES

• Biology of the periodontal connective tissues by Bartold and Sampath Narayan

• Molecular biology of periodontium by KV Arun• Molecular and cellular biology of alveolar bone. (Perio

2000)• Targeting osteoclast-osteoblast communication

(nature.com)• Ten cate’s oral histology• Orbans oral histology and embryology• Carranza 10th ed