Periodontal ligament in health part 1

1

PERIODONTAL LIGAMENT IN HEALTH AND

DISEASE

Dr Heenal AdhyaruDepartment of Periodontology

2

INTRODUCTION PERIODONTIUM: The

tissues that invest and support the teeth including the gingiva, alveolar mucosa, cementum, periodontal ligament, and alveolar and supporting bone. (GTP 2001)

PERIODONTIUM:1. Gingiva2. Periodontal ligament3. Cementum4. Alveolar bone

Glossary of periodontal terms, 2001 4th edition

3

The periodontal ligament is composed of a complex vascular and highly cellular connective tissue that surrounds the tooth root and connects it to the inner wall of the alveolar bone.

Carranza's Clinical Periodontology 10th edition

4

Periodontal ligament: The connective tissue that surrounds and attaches roots of teeth to the alveolar bone. (GTP 2001)

Glossary of periodontal terms, 2001 4th edition

5

Terms: Desmodont, gomphosis, pericementum,

dental periosteum, alveolodental ligament, periodontal membrane.

Because it is a complex soft connective tissue providing continuity between two mineralized connective tissues, the term periodontal ligament appears to be the more appropriate.

Orban’s Oral Histology and Embryology, 12th edition

6

COMPOSITIONPDL

Cellular element

Extra cellular substance

Fibers

Ground substance


7

CELLULAR ELEMENTS The principal cells of the healthy,

functioning periodontal ligament are concerned with the synthesis and resorption of alveolar bone, the fibrous connective tissue of the ligament and cementum.

Compared with most connective tissues, the periodontal ligament is highly cellular.


8B K B Berkovit’s The periodontoal ligament in

health and disease, 2nd edition.

9

The cells of the PDL may be divided as:

1. Synthetic cells Fibroblasts Osteoblasts cementoblasts

2. Resorptive cells Osteoclasts Fibroblasts cementoclasts

3. Progenitor cells4. Epithelial rests

of Malassez5. Defense cells

Mast cells Macrophages Eosinophils


10

FIBROBLASTS The fibroblasts lie between the collagen

fibers and although various shapes have been described it is likely that their appearance is governed by the surrounding matrix (Ross 1968).

If the PDL is sectioned both transversely and longitudinally, it can be deduced that the cells take the form of a flattened irregular disc, approx. 30 ɥm in diameter. (Berkovitz, 1988).

However, a defined 3D reconstruction of PDL fibroblasts has yet to be accomplished.B K B Berkovit’s The periodontoal ligament in


11

Nucleus: Flatterned disc

shape Diameter approx

10ɥm (Shore and Berkovitz, 1979)

Occupy upto 30% of cell volume. (Beertsen and Everts, 1977; Yamasaki et al, 1987a)B K B Berkovit’s The periodontoal ligament in


12

When stained with colloidal silver (Crocker and Nar, 1987) it demonstrates either one or two regions of acidic proteins which are associated with the nucleolar organizer regions (Shore et al., 1991)

B K B Berkovit’s The periodontoal ligament in health and disease, 2nd edition.

13

In the aged PDL, multinucleated fibroblasts may appear, arising either from fusion of mononuclear cells or perhaps by faulty division. (Sasaki and Grant, 1993)


14

As fibroblasts produce the ECM of the PDL, which demonstrates a very high rate of turnover (Sodek, 1977), the cells contain significant amounts of the organelles involved in protein synthesis and degradation.

Cho and Garant (1981a) have demonstrated, using tritiated proline in a pulse-chase experiment, that the synthetic pathway is from rough endoplasmic reticulum (RER) to Golgi complex and via secretory vesicles to the cell membrane.


15

Rough Endoplasmic Reticulum:


16

Golgi complex:


17

Mitochondria: Distributed throughout the cell, except

for the finest cell processes. Their profiles vary from elongated to

round, reflect either an inherent variability of shape or simply the plane of section.

They occupy approx. 3 – 3.5% of the cell volume in humans (Yamasaki et al., 1987a)

This value not seem to altered by different occlusal loading levels or eruption in rates (Shore et al., 1982, 1985)B K B Berkovit’s The periodontoal ligament in


18

Lysosomes: They are in the form of large

membrane-bound vesicles containing a homogeneous matrix that is more electron-dense than the surrounding cytoplasm.


19

PDL fibroblasts contain small fragments of collagen fibrils within membrane bound vesicles (Ten Cate, 1972; Listgarten, 1973; Beertsen et al., 1974; Eley and Harrison, 1975; Frank et al., 1976; Shore and Berkovitz, 1979)


20

Intracellular collagen profiles: This constitute the temporal sequence

of the intracellular degradation of collagen (Ten Cate et al., 1976)


21

Cytoskeleton: Cell possess a cytoskeleton that

provides a structural framework, facilitates intracellular transport, supports cell junctions and transmits signals about cell contact and adhesion, and permit motility.

The three structural elements of the cytoskeleton are Microfilaments Intermediate filamentsMicrotubules

Ten Cate’s Oral Histology 8th edition

22

Microfilaments: Microfilaments are of 5-7 nm in

diameter and are composed predominantly of polymerized actin (F actin), although other proteins are present as well (Brinkley, 1982)

B K B Berkovit’s The periodontoal ligament in health and disease, 2nd edition

23

The microfilaments are present in the cytoplasm of the cell either as a network that fills the cell processes (Beertsen et al., 1974) or as bundles beneath the cell membrane that resembles stress fibers seen in fibroblasts in vitro (Beersten et at., 1974; Shore and Berkovitz, 1979)B K B Berkovit’s The periodontoal ligament in

health and disease, 2nd edition

24

It is possible that the cells undergo continual short term localized movements in order to maintain the matrix around them.

As the PDL has an extremely high rate of turnover and therefore a constant need for fibril orientation, this local motility may be of considerable significance in maintaining PDL integrity.

That the presence of stress fibers may be linked to fibril orientation.


25

Microtubules: Microtubules are tubular or cylindrical

structures with an average diameter of 25 nm and are composed of the protein tubulin.

They are randomly arranged in cytoplasm (Shore and Berkovitz, 1979)


26

Microtubules are linked to fibroblast motility and in facilitating protein export (Ehrlich and Bornstein, 1972).

The distruption of microtubules leads to internal accumulation of procollagen (Cho and Garant, 1981b)

They are often seen to radiate from centrioles, structures which themselves consist of a hollow tube of microtubules.

A structure frequently associated with the centriole of PDL fibroblasts is a solitary cilium.


27

Solitary cilia lie within invaginations of the cell membrane with their distal ends protruding into the surrounding matrix.


28

Cilium lacks the central doublet of microtubules and dynein side arms associated with the outer doublets.


29

Degree of polarization of PDL fibroblasts and their organelles in relation to collagen secretion and /or motility can be considered on two levels:Whether individual cells are polarized in

terms of shape and organelle content andWhether the population of cells within the

tissue as a whole is polarized in a particular orientation.


30

Beertsen et al. (1979) and Garant and Cho (1979, 1989) have suggested that both the Golgi complex and the centriolar region of PDL fibroblasts may be situated between the leading edge of the cell and the posteriorly located nucleus.

Significant number of cells appeared to be polarized in opposing direction i.e. some cells may be moving apically while neighboring cells are moving incisally.

Microtubules have also been considered as indicators of polarity.


31

Intermediate filaments (IF): They are approximately 10 nm in diameter

and have a diverse protein composition. They are important in the maintenance of

cell shape and contact between adjacent cells and extra cellular matrix.

In cells of mesenchymal origin they are polymers of the protein Vimentin while in epithelial cell, they consists of cytokeratins.

The filaments form bundles, called tonofilaments which anchor onto desmosomes.


32

In humans, PDL may possess significant accumulations of IFs particularly within cell processes (Yamasaki et al., 1987b; Berkovitz, 1988)


33

Webb et al (1994) suggests that periodontal fibroblasts and cementoblasts co express vimentin and cytokeratin immediately before and during the active phase of eruption.

Once eruption has ceased, the expression of cytokeratin ceases also.


34

Intracellular contact:1. Tight junctions (Zonula occludents)2. Adhesive junctions

1. Cell to cell1. Zonula adherens2. Macula adherens (desmosomes)

2. Cell to matrix1. Focal adhesions2. hemidesmosomes

3. Communicating (gap) junctions


35

On molecular level, intercellular junctions typically consists of three components:1. A transmembrane

adhesive protein2. A cytoplasmic adapter

protein3. A cytoskeletal filament

These three components differ depending on the type of junctions


36

Contacts not normally being found in significant numbers between fibroblasts of adult connective tissues (Gabbiani, 1979; Moxham et al., 1984)

In the PDL, two major types of contact are seen:Gap junction Simplified desmosomes


37

Gap junction Simplified desmosomes


38

Cell surface receptors: Specific cell surface receptors shown to

be present on PDL fibroblasts are those for EGF-Epidermal growth factor (Thesleff et al.,

1987; Topham et al., 1987; Cho et al., 1991) IL-1ß: Interleukin-1ß (Saito et al., 1991)

In addition, in vitro studies suggest the presence of receptors for I-LGF (insuline like growth factor), PDGF (platelet derived growth factor), growth hormone (Blom et al., 1992; Matsuda et al., 1992) and parathyroid hormone (Ngan et al., 1988)B K B Berkovit’s The periodontoal ligament in


39

CEMENTOBLASTS Cementoblasts are the cells responsible

for secreting the organic matrix of cementum.


40

When active they may appear as a distinct layer of cells on the root surface, somewhat similar to the osteoblastic layer but usually not as regular in arrangement.

Some of the cementoblast cell processes do not approach cementum and their cytoplasmic contents are not polorized, the cells may contribute matrix to the PDL.

Synthetic pathway similar to fibroblasts and they appear to have less RER but more mitochondria than PDL fibroblasts (Yamasaki et al., 1987b)B K B Berkovit’s The periodontoal ligament in


41

One prominent feature is the accumulation of numerous glycogen granules, the number decreasing the further the distance from the cementum surface (Yamasaki et al., 1986)

They also appear to contain significant quantities of both intermediate and actin filaments.

Intercellular contacts: gap and simplified desmosome (Yamasaki et al., 1987b)

Receptors for growth hormone (Zhang et al., 1993) and EGF (Cho et al., 1991)


42

OSTEOBLASTS Osteoblasts within the PDL are found on

the surface of the alveolar bone.


43

The cells do not appear to possess receptors for EGF (Martineau- Doize et al., 1987)

Intercellular contacts: gap junction and simplified desmosome.

They also form contacts via gap junctions with osteocytes lying within lacunae in the adjacent bone, thus forming a coordinated system throughout the bone tissue (Holtrop and Weinger, 1972)

As bone deposition proceeds, osteoblasts become incorporated in the matrix as osteocytes (in which the organelle content is reduced).B K B Berkovit’s The periodontoal ligament in


44

Cells that may be osteoblast precursors are often seen beneath the osteoblast layer in the vicinity of adjacent blood capillaries.

As they proceed through the stages of differentiation from precursor via committed osteoprogenitor to preosteoblast, they first migrate away from the bone surface into the body of the PDL before eventually taking up their functional position (Roberts et al., 1987)

Once in the functional state, the cells may remain active for a period of up to 20 days.


45

When osteogensis is not occurring, a distinct layer of osteoblasts is absent.

Osteoblasts (and osteoclasts) are present over only approximately 10-15 % of bone surfaces (Jowsey et al., 1965); the remaining 85-90% of the bone surface is covered by flattened cells with scanty cytoplasm, the so called bone-lining cells.


46

OSTEOCLASTS Although it has been claimed that bone

resorption may be mediated via osteocytes (Belanger, 1971), resorption of bone surface is accomplished via a distinct cell type, the osteoclast.

They are found on the surface of the alveolar bone: Found within resorption lacunae They are large and multinucleated They have a ‘ruffled border’ adjacent to the

resorbing surface, enclosed by a smooth ‘clear’ zone




48

They do not cover the whole of the resorbing surface at any one time (Owen and Shetlar, 1968); rather they ‘service’ a much larger area by demonstrating considerable motility (Hancox, 1972; Jones and Boyde, 1977)


49

The multinucleated cells associated with the resorption of cementum and dentine have sometimes been referred to as cementoclasts and odontoclasts.

However the evidence suggests that all multinucleated resorptive cells involved in the removal of mineralized tissues are morphologically and functionally similar (Yaeger and Kraucunas, 1969; Freilich, 1971; Addision, 1979)


50

EPITHELIAL CELLS Epithelial cells represent the remains of

the developmental epithelial root sheath of Hertwig, which is involved in mapping out the shape of the roots and in the differentiation of root odontoblasts (Thomas and Kollar, 1988)

The epithelial cell rests (ECR) can be distinguished from the fibroblasts: Close packing of their cuboidal cells Stain more deeply Completely surrounded by connective tissue

cells (Brunette et al., 1979)B K B Berkovit’s The periodontoal ligament in


51

Immediately after disruption of the root sheath, the ECR are found in groups of one or two cells, with only a partial basal lamina.

Subsequently, the epithelial rests become more cellular and are contained within an almost complete basal lamina with narrowed intercellular spaces.

As laminin is chemotactic to epithelial cells, the basal lamina may therefore play a role in the formation, differentiation and maintenance of the ECR (Hamamoto et al., 1991).


52

ECR a high nuclear-cytoplasmic ratio. They exhibit basal cell-like,

undifferentiated and hyperproliferative characteristics, as indicated by expression of cytokeratins 5, 6, 14, 16 and 19 (Salonen et al., 1991)




54

ECR are located closer to the cementum.

Average distance 27ɥm in apical region and 41ɥm in cervical region. (Valderhaug and Zander, 1967)

Distribution changes with age: more numerous in children and less numerous in older individuals (Reitan, 1961; Simpson, 1965; Wesselink and Beertsen, 1993)


55

Up to the second decade of life, ECR are found most commonly in the apical region of the PDL, whereas later in life the majority of cell rests are located cervically in the gingiva of the alveolar crest (Reeve and Wentz, 1962)

In this cervical region, some of the ECR are presumably derived from the gingival epithelium and the junctional epithelium (Wentz et al., 1950)


56

As non functional cells usually disappear, the persistence of ECR suggests that they are not totally inactive and may sub-serve some function (Spouge, 1980).

They take up tritated thymidine, indicating some degree of turnover (Trowbridge and Shibata, 1967; McCilloch and Melcher, 1983c)

Show intense binding of EGF indicating ECR are activated by a local rise in tissue level of this GF. (Thesleff, 1987)


57

Functions: ECR secrete enamel like proteins onto

the root surface (Slavkin et al., 1988; Luo et al., 1991)

Mediating repair cementogensis (Brice et al., 1991)

Factor liminting the resorption and maintenance of the periodontal space. (Loe and Waerhaug, 1961; Lindskog et al, 1983)

Cells have been implicated in the aetiology of periodontal cysts




59

CELL KINETICS WITHIN PDL The question arises as to whether

periodontal fibroblasts, cementoblasts and osteoblasts all arises from a common precursor or whether each cell type has its own specific precursor cell.

Nuclear size can help distinguish some cell type (Roberts et al., 1981), while receptors to EGF are present during root development on preosteoblasts and periodontal fibroblasts, but not on precementoblasts, cementoblasts and osteoblasts. (Cho et al., 1991)B K B Berkovit’s The periodontoal ligament in


60

Dividing progenitor cells within the PDL are located predominantly paravascularly (Gould et al., 1982; McCulloch and Melcher, 1983b, c; Roberts et al., 1987) and give rise to cells that can migrate towards the bone and cement surfaces, where they differentiate into osteoblasts and cementoblasts (McCulloch and Melcher, 1983b)

This population may represent stem cells that are not terminally differentiated but that continue to divide at a slow rate.B K B Berkovit’s The periodontoal ligament in


61

McCulloch and Melcher (1983c) investigated the relationship between cell density and cell generation within the PDL.

Labelling indices are highest in zones adjacent to blood vessels.

Also in the middle of the ligament where cell density was lower, compared with zones adjacent to bone and cementum, where cell density was higher.


62

Yee (1979), Gould et al. (1980) and Gould (1983) have studied the morphology of progenitor cells in the PDL.

They have relatively undifferentiated appearance (e.g. small size, scarcity of intracellular organelles, and a high nuclear-cytoplasmic ratio), some cells have been shown to be relatively well differentiated, containing much RER and even intracollagen profiles.


63

The cell cycle time for roughly one half of the cells in the normal PDL has been calculated to be less then 48hrs. (Roberts, 1975a; Roberts et al., 1981)

For PDL fibroblasts, Gould et al. (1983) have calculated a turnover time of 45 days, there being a slight increase with age.


64

Using orthodontically induced osteogenesis as the model system, Roberts and Chase (1981) found that the initial layer of osteoblasts was unlabelled.

This implies that among the cells of normal PDL are preosteoblasts that are sufficiently differentiated to become osteoblasts without synthesizing DNA.

All cells across the ligament synthesize collagen using tritiated proline (Beertsen and Everts, 1977; Rippin, 1978) indicating that such preosteoblasts contribute to the formation of PDL collagen before finally differentiating into osteoblasts (Roberts et al., 1982)B K B Berkovit’s The periodontoal ligament in


65

Roberts et al. (1981, 1982 and 1987) and Roberts and Ferguson (1989) used nuclear size (Nuclear diameter) to classify cells in the PDL after orthodontically induced osteogensis.

A. 40-79 ɥm3 : small precursor cellsB. 80-119 ɥm3 : typical PDL fibroblastsC. 120-169 ɥm3 : G1 preosteoblastsD. >170 ɥm3 : G2 preosteoblasts


66

McCulloch et al. (1987) have provided some evidence that raises the possibility that cells may migrate out from the endosteal spaces in the alveolar bone and into the ligament, thereby augmenting the populations of fibroblasts, osteoblasts and cementoblasts.


67

Osteoclasts are derived from haemopoietic stem cells.

This cells enter the ligament as mononuclear cells from the haemopoietic system as required and then fuse to form the typical large, multinucleated giant cell.

1. Fusion of monocytes or macrophages or both.

2. Share a common progenitor with cells of the monocyte-macrophage line

3. From pluripotent haemopoietic stem cell entirely separate from that of the monocyte-macrophage line.


68

DEFENCE CELLS The PDL contains defence cells: Macrophages, Mast cells and Eosinophils.


69

Macrophages: MuCulloch et al. (1989) described the

detailed distribution of the macrophage in the healthy PDL using electron microscopy.


70

Macrophages comprise of 4% of the cells of the PDL

They are located close to blood vessels. As the labelling index of perivascular

cells is high compared with other regions of the ligament, it is possible that lymphokines released from macrophages may be involved in cell kinetics.

Dual role:Phagocytosing dead cellsSecreting growth factors


71

Mast cells: Granules released by mast cells may be

phagocytosed by fibroblasts, suggesting that other interactions may occur between mast cells and fibroblasts (Atkins et al., 1985)

B K B Berkovit’s The periodontoal ligament in health and disease, 2nd editio

72

REFERENCES: Carranza's Clinical Periodontology,

10th edition. Glossary of periodontal terms, 2001

4th edition Orban’s Oral Histology and

Embryology, 12th edition. Ten Cate’s Oral Histology 8th

edition. B K B Berkovit’s The periodontoal

ligament in health and disease, 2nd edition.

Health & Medicine

Periodontal ligament in health part 1