Periodontal Ligament Turn Over & Cell

Matrix Interactions

Dr Harshavardhan Patwal

Introduction

How unique is periodontal ligament?

Periodontal ligament cells & extracellular matrix constituents.

Ground substances.

Periodontal ligament fibers.Collagen

Periodontal ligament homeostasis and adaptationto functional demand.

Degradation And Remodelling Of Connective Tissue Matrix.

Mechanisms of degradation and remodelling of connective tissue matrix.

Fibroblast To Matrix Adhesions And Tractions

The periodontal ligament is composed of a complex vascular & highly cellular connective tissue that surrounds the tooth root & connects it to the inner wall of the alveolar bone (McCulloch CA et al).

It is continuous with the connective tissue of the gingiva & communicates with the marrow spaces through vascular channels in the bone.

Average width of periodontal ligament space is about 0.2mm

Space is diminished around teeth Not in function Unerupted teeth.

Introduction:

Increased in teeth subjected to hyperfunction.

The periodontal ligament is unique among the various ligament and tendon systems of the body in that it is the only ligament to span two distinct hard tissues – namely, tooth cementum and bone.

Function: Supporting the teeth in their sockets. Withstand considerable forces of mastication. It has the capacity to act as a sensory receptor necessary for the

proper positioning of the jaws during mastication. It is a cell reservoir for tissue homeostasis and repair ⁄

regeneration.

When periodontal ligament cells are removed from the cementum by mechanical means or displaced or altered under the influence of certain compounds-ankylosis may occur.

Bone tissue invade the periodontal ligament space & establish a direct connection between the tooth & the wall of the alveolar socket.

How Unique Is The Periodonal Ligament?

When periodontal ligament fibroblasts or their progenitors are allowed to gain access to the root & repopulate the area.

These cells may come from the adjacent periodontal ligament.

Invasion of periodontal ligament fibroblasts in an ankylosed site must be preceded by cells that have the capacity to resorb bone / cementum.

A new periodontal ligament space may be created at the cost of the cementum & also of the alveolar process.

Shortly, this space is colonized by periodontal ligament cells that form new periodonal ligament fibers,new cementum & a new alveolar wall in which the fibers insert (Wasserlink et al 1994).

Moreover,masticatory function can accelerate the resolution of ankylotic areas & restoration of normal periodontal ligament width (Andersson et al 1985).

Boyko et al 1981- seeded fibroblasts from either periodonal ligament or gingiva onto the surface of tooth roots, which were subsequently inserted in artificial sockets.

Apart from ankylotic areas, - areas with newely formed periodonal ligament fibers inserted in newely formed cementum.

Observed only when periodontal ligament fibroblasts were used as seeding cells.

Gingival cells could not induce regeneration of the periodontal ligament (Lang et al).

Indicating that the periodontal ligament cells have a specialized properties.

Cellular elements.

Periodontal fiber groups.

Ground substances.

Periodontal Ligament Composed Of

The periodontal ligament consists of cells and an extracellular compartment comprising collagenous and noncollagenous matrix constituents.

The cells include osteoblasts and osteoclasts, fibroblasts, epithelial cell rests of Malassez, monocytes and macrophages, undifferentiated mesenchymal cells, and cementoblasts and odontoclasts.

The extracellular compartment consists mainly of well-defined collagen fiber bundles embedded in an amorphous background material, known as ground substance.

Periodontal Ligament Cells And Extracellular Matrix Constituents

The predominant cell type is the fibroblast, which occupies about 30% of the volume of the periodontal ligament space in rodents (Beertsen W,).

FIBROBLASTS

The fibroblasts of the periodontal ligament originate in part from the ectomesenchyme of the investing layer of the dental papilla and from the dental follicle (Ten Cate AR et al) and are different from cells in other connective tissues in a number of respects.

Freeman & Ten Cate demonstrated that periodontal ligament fibroblasts near the cementum are derived from the ectomesenchymal cells of the investing layer of the dental papilla, while fibroblasts near the alveolar bone are derived from perivascular mesenchyme.

Although periodontal ligament cells are frequently considered as a homogeneous population, there are some data indicating that the periodontal ligament contains a variety of fibroblast populations with different functional characteristics (McCulloch CAG et al).

Whether these subsets are derived from a single type of progenitor cell is unknown.

The fibroblasts on the bone side of the periodontal ligament exhibit more abundant alkaline phosphatase activity than those on the tooth side (Groeneveld MC et al).

This enzyme plays a key role in phosphate metabolism probably in the mineralization process (Beertsen et al) & perhaps also in acellular (afibrillar) cementum formation (Groeneveld et al)

Alkaline phosphatase activity in rat molar PDL shows a high corelation with acellular cementum thickness- patient suffering from hyphophosphatasia.

Cementum formation is greatly impaired.

The enzyme is not restricted to cementoblasts & osteoblasts but is found in all periodontal ligament fibroblasts, especially along their outer plasma-membrane.

The rapid degradation of collagen by fibroblast phagocytosis is the basis for the very fast turnover of collagen in the periodontal ligament (Everts V,et al).

The generation of highly specialized cell populations that can remodel and heal damaged tissues in a temporally and spatially appropriate manner is thought to be essential for the repopulation and differentiation responses in healing periodontium.

The signals that regulate these processes include cell matrix and direct cell-cell interactions which are known to control cell proliferation, differentiation and cell function (Gumbiner BM et al).

For example, periodontal ligament cells produce cell adhesion proteins like vitronectin, tenascin and undulin as well as several integrin subunits (Steffensen B et al).

The fibroblasts of the periodontal ligament are connected by specialized junctional complexes which include as gap junctions (Shore RC et al).

Notably, mechanical stimulation of the periodontal ligament stimulates the expression of connexin, an important protein in the formation of gap junctions in periodontal ligament cells (Lewis JE et al).

The PDL fibroblast contains a prominent nucleus, which has a single distinct nucleolus & clearly defined nuclear pores.

It has been suggested that periodontal fibroblasts have a role in generating the force of eruption by contraction, in a way similar to myofibroblasts that are responsible for wound contraction.

Sodek 1977 – as fibroblasts produce the extracellular matrix of the PDL which demostrates a very high rate of turn over, the cells contain significant amounts of the organelles involved in protein synthesis & degradation.

Epithelial cells:

The epithelial cells in the periodontal ligament are remnants of HERS and known as the epithelial cell rests of Malassez.

They occur close to the cementum as a cluster of cells that form an epithelial network, and seem to be more evident or abundant in furcation areas.

Undifferentiated mesenchymal cells:

An important cellular constituent of the periodontal ligament is the undifferentiated mesenchymal cell, or progenitor cell.

The fact that new cells are being produced for the periodontal ligament whereas cells of the ligament are in a steady state means that selective deletion of cells by apoptosis must balance the production of new cells.

In periodontal wound healing, the periodontal ligament contributes cells not only for its own repair but also to restore lost bone and cementum ( Anusaksathien O,et al).

Recently, cells with stem cell characteristics have been isolated from the human periodontal ligament (Hollenbach E,et al).

Filling The Space Between The Fibers & Cells.

GlycosaminoglycansHyaluronic AcidProteoglycans

Glycoproteins

Fibronectin

Laminin

Ground Substances

The cell surface proteoglycans participate in several biological functions, including:

Cell adhesion,

Cell-cell & cell matrix interactions,

Binding to various growth factors as co-receptors

Cell repair (Worapamorn W et al)

Ground substance: The periodontal ligament ground substance has been

estimated to be 70% water and is thought to have a significant effect on the tooth’s ability to withstand stress loads.

There is an increase in tissue fluids within the amorphous matrix of the ground substance in areas of injury and inflammation.

Periodontal Ligament Fibers

Periodontal ligament Fibers:

The predominant collagens of the periodontal ligament are type I, III, and XII, with individual fibrils having a relatively smaller average diameter than tendon collagen fibrils.

The vast majority of collagen fibrils in the periodontal ligament are arranged in definite and distinct fibre bundles, and these are termed principal fibers.

Elastic fibers: There are three types of elastic fibers:

Elastin, Oxytalan, and Elaunin.

Only oxytalan fibers are present within the periodontal ligament; however, elaunin fibers may also be found in association with fiber bundles in the gingival ligament.

Oxytalan fibers:

These are bundles of microfibrils that run more or less vertically from the cementum surface, forming a three-dimensional branching meshwork that surrounds the root and terminates in the apical complex of arteries, veins, and lymphatics.

They are also associated with neural elements.

They are thought to regulate vascular flow in relation to tooth function.

Because they are elastic, they can expand in response to tensional variations, with such variations then registered on the walls of the vascular structures.

Principal fibers- which are collagenous & arranged in bundles & follow a wavy course.

The terminal portions of the principle fibers that are inserted into cementum & bone – sharpeys fibers.

They are associated with abundant noncollagenous proteins typically found in bone & cementum (Mc Kee MD et al)OsteopontinBone sialoprotein

These proteins are thought to contribute to the regulation of mineralization & to tissue cohesion at sites of increased biomechanical strain.

The principal fibers of the periodontal ligament are arranged in six groups:(Sloan & Carter)TransseptalAlveolar crestHorizontalObliqueApical Interradicular

Principal Fibers

Components of the extracellular matrix:Collagen: Collagens are a large family of triple helical proteins that are

widespread throughout the body and are important for a broad range of functions, including tissue scaffolding, cell adhesion, cell migration, cancer, angiogenesis, tissue morphogenesis and tissue repair.

It is a protein composed of different amino acids, the most important of which are glycine, proline, hydroxylysine & hydroxyproline (Carneiro J et al).

Each polypeptide chain has a repeating Gly-XY triplet in

which glycyl residues occupy every third position and the

X and Y positions are frequently occupied by proline and 4-

hydroxyproline, respectively.

 The three α chains are held together by interchain

hydrogen bonds. Highly ordered hydration networks

surround the triple helices.

The amount of collagen in a tissue can be determined by

its hydroxyproline content.

Collagen is responsible for maintenance of the frame work

& the tone of the tissues & it exhibits a wide range of

diversity.

Vertebrate collagens are classified by function and domain homology:

Fibril-forming collagens: (I,II,III,V,XI)

  Fibril-associated collagens with interrupted triple helices (FACITs): (IX,XII,XIV,XVI)

 Network-forming collagens :( IV,VIII,X)

 Transmembrane collagens:(XIII,XXV,XVII)

 Endostatin-producing collagens:(XV,XVIII)

 Anchoring fibrils:(VII)

Beaded-filament-forming collagen:(VI)

Collagen is synthesized by fibroblasts, chondroblasts, osteoblasts, odontoblasts.

The principle fibers are composed mainly of collagen type I (Bosshardt DD et al)

Reticular fibers are composed of type III.

Collagen type IV is found in basal lamina (Romanos GE et al)

Type VI collagen has been immunolocalized in periodontal ligament & gingiva (Everts V et al)

Recent in vitro studies have shown that type VI collagen

simulates fibroblasts proliferation in a non-integrin-mediated pathway(Atkinson JC et al)-

Type V collagen – associated with the cell surface & to coat larger type III & type I fibrils.

Immunocytochemical studies have also localized collagen types XII & XIV in the PDL (Karimbux NY et al).

Type XII, a member of the fibril associated collagens, forms small fibrils that have a role in the organization of the network of larger collagen fibrils.

N.Y. Karimbux and I. Nishimura- Temporal and Spatial Expressions of Type XII Collagen in the Remodeling Periodontal Ligament during Experimental Tooth Movement.

The type XII collagen expression may be closely associated with the functional regeneration of the PDL.

Collagen biosynthesis occurs inside the fibroblasts to form tropocollagen molecules.

In collagen type I & III the fibrils associate to form fibers, & in collagen type I the fibers associate to form bundles.

In comparision to collagens,the non-collagenous proteins occur in small amounts in the PDL.

The adhesion molecules, fibronectin, tenascin, & vironectin, are among the glycoproteins found in the PDL (Pitaru et al).

Fibronectin is widely distributed in the PDL.

Vitronectin is an attachment factor associated with elastic fibers in loose connective tissue.

Non-collagenous Proteins

It has been localized throughout the PDL, including in cells lining cementum & bone surfaces.

Vitronectin participates in the regulation of blood coagulation, plasminogen activation, & fibrinolysis.

The fibroblasts interact with the extracellular matrix through receptor-ligand interactions.

Many of the matrix ligands are noncollagenous proteins, such as extracellular adhesion factors (Hynes et al).

Eg,binding of a fibronectin fragment to the a5b1 fibronectin receptor of rabbit synovial fibroblasts leads to secretion of collagenase.

In contrast,when a5b1 receptors are occupied by inact fibronectin molecules,no collagenolytic response is observed.

PDL fibroblasts react in a similar way to fibronectin fragments by increasing the expression of collagenase & stromelysin.

It is increasingly apparent that receptor-matrix informational exchange is involved in regulating fibroblast connective tissue remodelling.

Such a unique and dynamic connective tissue system involving multiple tissues requires exquisite regulation at the cellular level.

Maintenance and remodeling of periodontal ligament collagen fibers (Deporter DA, Ten Cate AR), together with the embedding and calcification of their extremities to form Sharpey’s fibers (Johnson RB et al), requires the concerted action of numerous cell types (Garant PR et al)

Central to these integrated activities is the periodontal ligament fibroblast, whose responsibilities include the formation and remodeling of the periodontal ligament fibers, and presumably a signaling system to maintain periodontal ligament width and thickness across the soft tissue boundary

In the periodontal ligament, cellular signals are, in part, mediated by the forces transmitted to the fibroblasts via collagen fibrils with which they are in direct contact( Garant PR et al).

At the tissue level, periodontal ligament fibroblasts are rather regularly dispersed throughout the ligament and are generally oriented with their long axis parallel to the direction of the collagen fibrils.

During development and the initial formation of the periodontal ligament, the cytoplasm-to-nucleus ratio is high, and fibroblasts appear very active in terms of having an extensive network of rough endoplasmic reticulum, a well-developed Golgi apparatus and abundant secretory granules containing predominantly type I collagen

MATRIX REMODELING & ADAPTABILITY.

Several studies have indicated that the extracellular matrix collagens of the periodontal ligament have an extremely high turnover and remodeling rate, much higher than in gingiva, skin and bone (Sodek J.).

Turnover and remodelling in the periodontal ligament imply synthesis and breakdown of matrix components, particularly the collagenous fiber meshwork that extends between cementum and bone.

Turnover describes a process in which the structural organization of the tissue remains unchanged.

During remodelling the three-dimensional organization of the fiber meshwork is adapted to accommodate for positional changes of the tooth in its socket or changes in functional state (such as hypofunction) (Beertsen et al).

Both processes can occur simultaneously & may therefore be indistinguishable.

Sodek et al rapid remodeling is a unique characteristic of the periodontal ligament that relates to the adaptability of the periodontal tissues.

(Sicher H.et al) suggested that remodeling of the ligament is confined largely to the mid-region of the periodontal ligament where fibers from the bone and fibers from the tooth interdigitate in an ‘‘intermediate plexus’’.

Turnover and remodeling activity in teeth of limited eruption, like the molars of rodents, are found throughout the width of the periodontal ligament from cementum to bone (Rippin JW.).

To adapt to changes of tooth position, the fiber systems in the periodontal ligament must be degraded and new fibers synthesized.

Since the periodontal ligament is not made up of single strands of straight collagen fibers but consists instead of a complex meshwork, remodeling does not necessarily occur at all sites synchronously.

There is apparently some flexibility in the system to permit adaptational changes by breaking down short stretches of collagen fiber bundles or single fibrils while leaving others intact.

This highly localized remodeling process Is facilitated by the phagocytosis of collagen.

Unlike the bulk removal of collagen that is effected by extracellular matrix metalloproteinases, collagen phagocytosis enables periodontal ligament fibroblasts to very precisely remove collagen fibrils at specific sites (Everts V).

Svoboda et al- relationship between turnover rate of collagen in the various tissues constituting the periodontium & the amount of collagen ingested by fibroblasts, the highest amount being found in the tissues with the highest turnover, most notably the periodontal ligament.

A remarkable capacity of the periodontal ligament is that it maintains its width more or less over time, despite the fact that it is squeezed in between two hard tissues.

Compelling evidence exists indicating that populations of cells within the periodontal ligament, both during development and during regeneration, secrete molecules that can regulate the extent of mineralization and prevent the fusion of tooth root with surrounding bone, e.g. ankylosis.

Periodontal Ligament Homeostasis And Adaptation

To Functional Demand

Balance between the activities of bone sialoprotein and osteopontin may contribute to establishing and maintaining an unmineralized periodontal ligament region.

Matrix Gla protein is also present in periodontal tissues; based on its role as an inhibitor of mineralization, it may also act to preserve the periodontal ligament width.

At the cell level, Msx2 prevents the osteogenic differentiation of periodontal ligament fibroblasts by repressing Runx2 ⁄ Osf2 transcriptional activity (Kanda-Nakamura C et al).

The periodontal ligament has also the capacity to adapt to functional changes.

When the functional demand increases, the width of the periodontal ligament can increase by as much as 50%, and the fiber bundles also increase markedly in thickness.

Conversely, a reduction in function leads to narrowing of the ligament and a decrease in number and thickness of the fiber bundles.

These functional modifications of the periodontal ligament also implicate corresponding adaptive changes in the bordering cementum and alveolar bone.

Breakdown of the collagenous matrix is a normal event in tissues undergoing morphogenesis, morphostasis and growth.

However, it is vital that this process is subject to rigid control, as failure to maintain an appropriate balance between degradation and synthesis can lead to net destruction or net gain, resulting,

Degradation And Remodelling Of Connective Tissue Matrix

The earliest manifestations of vitamin C deficiency (scurvy) were seen as intraoral lesions and tooth loosening (Hunt and Paynter, 1959), which indicated that the periodontal tissues were subject to rapid turnover of collagen.

The earliest attempts to determine collagen turnover in the PDL used autoradiographic techniques (Stallard, 1963 et al).

These studies (mostly using 3H-labelled proline) suggested that turnover of collagen in the periodontal tissues was rapid.

However, the technique fails to discriminate between specific radiolabelling of collagen and incorporation of the tracer into other proteins, many of which (particularly intracellular proteins) turn over at a much higher rate than extracellular proteins.

Orlowski (1976, 1978), overcame these difficulties in part bY measuring the specific activity of hydroxyproline 24 hours after injection with 3H-proline.

Turnover in the PDL was found to be higher than that of the gingival, with a half life of around 9.5 days and a turnover time of 13.5 days for the rat incisor PDL.

Sodek (1976, 1977) utilized a novel short-term approach to measure incorporation of 3H-labelled proline into newly synthesized (and therefore salt-extractable) and mature collagen in the rat molar PDL, gingival, alveolar bone and skin.

Hydroxyproline-specific activities for theses tissues revealed that the rate of collagen synthesis in the PDL was twice that in gingival, four time that in skin, and six times that in alveolar bone.

This study also showed that the conversion of newly synthesized collagen into mature insoluble collagen was highly efficient in the PDL compared with the other tissues (which showed up to 50 per cent degradation of newly synthesized collagen).

This supported the Suggestion of Guis and Slootweg (1973) based upon collagen extractability, that newly synthesized tropocollagen matured rapidly in bovine PDL.

The half life of mature collagen in rat molar PDL was calculated by Sodek to be 1 day, compared with 5 days in gingival, 6 days in alveolar bone and 15 days in skin.

(Sodek, 1978) comparing molar and incisor PDLs calculated a longer half life of 3 days for mature collagen in the incisor ligament.

Imberman et al. (1986) applied the so-called ‘pool expansion’ technique in order to determine the half life of collagen in the rat incisor and molar PDLs; they then compared these with those of the skin, gingival, and palatal mucosa.

These authors calculated the half lives of collagen to be: 7.8 days for incisor PDL, 8.8 days for molar PDL, 150 days for incisor gingival tissue, 8.8 days for molar gingival, 50 days for skin and 21 days for palatal mucosa.

Finally, using Poole’s approach (1971), Sodek and Ferrier (1988) attempted to validate the apparently rapid rate of collagen turnover in the periodontal tissues of the rat.

This study yielded half life values for mature, insoluble collagen as 3 days for molar PDL, 6 days for incisor PDL, and 10 days for gingiva.

The discrepancies between values reported by different

workers using a variety of techniques clearly indicate the

difficulties involved in obtaining definitive data for

collagen turnover rates in the PDL and other connective

tissues in general.

However, the general consensus is that the turnover of

collagen in PDL is unusually high.

This may have important implications in the aetiology of

chronic inflammatory periodontal disease , such that an

imbalance in the synthesis and degradation process may

result in net collagen loss (Page and Ammons, 1974).

Sodek (1989) suggested that the differences in turnover rate with tooth type might be due to direction of the functional loading on the tissue, i.e tensional as opposed to compressional.

Finally, perhaps related to the constraints turnover of collagen in the PDL also appears to be in some way linked with rate of eruption, (Berkovitz,et al).

Rippin (1976, 1978) reported an increase in collagen turnover with reactivated eruption following extraction of opposing teeth, but no such increase was observed when eruption was accelerated in the rat incisor (Van den Bos and Tonino, 1984).

The precise relationship between rate of eruption and PDL collagen turnover remains obscure.

Degradation of the extracellular matrix can occur through a number of different pathways, including activation of matrix metalloproteinases (MMPs), release of reactive oxygen species and phagocytosis of matrix components,release of a wide range of cytokines & other inflammatory mediators that affect the enzyme release & fibroblasts function.

Mechanisms Of Degradation And Remodeling Of Connective Tissue Matrix

Matrix metalloproteinases:

Matrix metalloproteinase gene family encodes a total of 24 homologous proteinases, classified into collagenases, gelatinases, stromelysins, membrane type matrix metalloproteinases and other matrix metalloproteinases depending on their substrate specificity and molecular structure (Uitto VJ et al).

MMPs are secreted by connective tissue cells (predominantly fibroblasts) but are also produced by some leucocytes (polymorphonuclear neutrophil leucocytes and macrophages.)

The secreted MMPs are subsequently activated by proteinases such as plasmin, which in turn are regulated by tissue protein factors such as plasminogen activators.

The plasminogen activators are serine proteinases.

Matrix metalloproteinases play a major role in connective tissue breakdown (Birkedal-Hansen H,et al).

To date, MMP-1, -2, -3, -8, -9, and -13 have been identified in inflamed periodontal tissues.

The enzymes synthesized by fibroblasts & epithelial cells are belived to be mosetly involved in normal tissue remodeling (sodek et al)

LPS & TNF-alpha activate MMP synthesis by keratinocytes.

IL-I enhances MMP expression by fibroblasts, whereas LPS stimulate PGE2 production by these cells.

The activities of PGE2 include suppression of cell proliferation, inhibition of collagen synthesis, & with IL-6, stimulation of bone resorption (page et al).

Bacterial plaque products can also stimulate MMP production by a variety of cells (Birkedal-Hansen et al).

Tissues also contain another group of matrix metalloproteinase inhibitors known as tissue inhibitor of metalloproteinases (TIMPs) (Brew K,et al).

At least four tissue inhibitor of metalloproteinases (TIMP-1, TIMP-2, TIMP-3, TIMP-4) are expressed by vertebrates and these act by preventing the conversion of precursor forms of matrix metalloproteinase to their active forms.

TGF-b, steroids, & inteferon –y induce the expression of TIMP-1 & TIMP-2.

Tetracycline & their analogues –inhibit (ingman et al).

Tetracycline, which also inhibit the expression of 72-kd gelatinase (MMP-2), does not affect fibroblasts collagenase, indicating that the MMP in gingival crevicular fluid are derived from PMN (Ingman et al).

Phagocytosis

Phagocytosis is also a significant pathway of collagen degradation in the physiological turnover and remodeling of periodontal connective tissues (Ten Cate AR et al).

During normal tissue homeostasis, collagen degradation can take place within the lysosomal apparatus of phagocytic cells.

Phagocytosis of collagen has been proposed to involve recognition of collagen fibrils through specific membrane-bound receptors (possibly integrins), followed by partial digestion of the fibrils by proteinases such as gelatinases (MMP- 2, -9) and final degradation by lysosomal enzymes cysteine proteinases such as cathepsin B or L (Everts V,et al).

Cells defective in phagocytosis may contribute to gingival overgrowth and fibrosis as well as compromise normal wound repair and tissue regeneration (McCulloch CA et al).

Reactive oxygen species:

Oxygen-derived free radicals such as the superoxide radical and the hydroxyl radical are integral reaction products of normal cellular metabolism, but these are elevated in cells undergoing active respiratory bursts (Baboir BM).

Tissues and cells may be exposed to oxygen-derived free radicals during inflammatory reactions, particularly where polymorphonuclear leukocytes and macrophages are in abundance.

Oxygen-derived free radicals are highly reactive molecular species which can disrupt cellular proteins, nucleic acids and membrane lipids as well as cause depolymerization of matrix components such as collagen, hyaluronan and proteoglycans (Freeman BA).

What advantage could there be in degrading collagen intracellularly instead of extracellularly?

Phagocytosis allows a more precise & selective control for the collagen fibers to be degraded.

Whereas the release of extracellular collagenolytic enzyme affords a more rapid, extensive degradation around the cells,observed during inflammation.

Together all these studies indicate that the PDL is characterized by a rapid turnover & a high remodeling capacity.

Under steady-state conditions, collagen degradation in this tissue is carried out via a process of collagen phagocytosis without the involvement of collagenase.

When cells come into contact with one another, & sometimes with the

extracellular matrix, specialized junctions may form at specific sites on the

contacting cell membranes.

The specialized junctions may be classified as

Occluding (tight) junctions (zonula occludens)

Adhesive junctions Cell-cell

Zonula adherens

Macula adherens

Cell-matrix Focal adhesions

Hemidesmosomes

Communicating (gap) junctions.

Cell Matrix Interactions

Cell-matrix junctions have a structural organization similar to that of cell-cell adhesive junctions, but they use different molecular components & attach the cell to the extracellular matrix.

In focal adhesions the transmembrane component is a member of the integrin family of adhesion molecules.

Integrins are heterodimers of different alpha & beta subunits that occur in different combinations with specificity for various extracellular matrix molecules.

The cytoplasmic adapter proteins, which include the actin-binding proteins a-actinin,vinculin, & talin, link the transmembrane integrins to the actin cytoskeleton.

Binding of the integrin to collagen, laminin, fibronectin, & other extracellular matrix proteins results in recruitement & remodeling of actin cytoskeleton.

Ligand binding by integrins also leads to the recruitment & activation of various intrcellular signaling molecules, including guanine nucleotide-binding proteins & several protein kinases.

Fibroblast To Matrix Adhesions And Tractions• Fibroblast attached to the substratum of the extra cellular

matrix via surface receptors for collagen and fibronectin.

Attachment to the substratum is essential for cellular migration and for organization for the extra cellular fibrillar matrix.

In the formation of these adherent contacts, the cell membrane integrin a5 b1 attaches to the arginine- glycine-aspartic acid sequences of fibronectin.

Assembly is initiated by binding of soluble fibro nectin molecules to the cell of its integrin receptors A5b1 and avb3

Specific Cell-matrix Interaction Through Integrins

The cytoplasmic domain of the integrin receptor attaches to the peripheral cytoplasmic protein talin, which in term interacts with the protein called vinculin.

With the binding of fibronectin to collagen fibrils , the molecular linkage extends from the cytoplasmic contractile apparatus to an extracellular fibrin network-exerting traction on collagen fibers.

In fibroblast under tension, filamentous actin and smooth muscle myocin associate to from contractive stress fiber that terminated plasma membrane in fibro nexus junctions.

Through such cell to matrix contact the extra cellular matrix can exert and effect on cells shape and behaviour.

Through such cell to matrix contact the extra cellular matrix can exert and effect on cells shape and behaviour.

Tension in the ECM is transmitted to fibroblasts integrin receptors, leading to signaling event that alters the activity of te cell.

Human PDL fibroblast respond to increased tension by upregulating the

expressions of IL-1b103 and by secreting prostaglandin E2.

In this “outside –in” type of signalling, tension transmitted to the fibroblast

causes a rise in the activity of several small guanosine triphosphatases, which

regulate the enzyme cascades that lead to changes in cell shape and function.

Cells can also alter the binding strength of their integrin receptors through

cytoplasmic signaling pathways.

This represents a form of “inside-out” signal transduction.

The linkage between the cell surface and the immediate extracellular matrix

serves as a node whereby the cell can receive regulatory information from the

outside and provides a mechanism for exerting an organizing influence on the

adjacent matrix.

When fibroblasts move through a collagen gel matrix, they tend to align collagen fibrils parallel to the long axis of migration.

If the substratum on which cells are placed is sufficiently anchored-cells move over the substratum.

Not anchored-pulled towards the cells.

In vitro studies of matrix contraction have shown that fibroblasts are able to generate the same degree of tension-contracting wound.

PDGF & IGF-1 & TGF-b promote fibroblasts contraction of type I collagens gels.

PDGF promotes gel contraction by stimulating actin cytosketetal polymerization & increase the expression of integrins.

Both gingival & PDL fibroblasts exhibit high gel-contracting abilities.

Each fibroblasts may have a minimum of 105 fibronectin receptors.

On highly mobile fibroblasts-receptors are diffusely distributed.

Stationary cells-arranged in linear arrays.

Immunocytochemical localization of fibronectin in the periodontal ligament has shown that it forms aggregates about 90nm thick.(Cho Mi et al)

The fibronexus is a terminal for anchorage of stress fibers to the cell surface and the ECM .

The stress fibers comprise well defined bundles of actin and non sarcomeric myosin oriented parallel to the long axis of the fibroblast .

Stress fibers are found in fibroblasts involved in transferring tension to an extracellular fibre network that is firmly attached to stable structures in their immediate vicinity and PDL fibroblasts can form robust stress fibers similar to those observed in myofibroblasts of healing wounds.

Well developed stress fibers and fibronexus contact have been observed in fibroblasts of the transseptal fiber group between molar teeth of monkeys.(Garant PR)

V.-J. Uitto- Expression of Fibronectin and Integrins in Cultured Periodontal Ligament Epithelial Cells.

During the different stages of cell growth and spreading, the integrins appeared to become reorganized, which facilitated cell-cell contact and allowed for migration of the cell colonies.

PDL has a high potential for regeneration & repair.

Regeneration of a functional ligament requires corelated development of new cementum & bone for the attachment of sharpeys fibers.

Repair of the PDL involves the replacement of small areas of damaged ligament.

In repair, new fibroblasts are derived from perivascular progenitor cells in the adjacent normal PDL.

Clinical Correlations

Migration of fibroblasts into the area to be repired is facilitated by the presence of fibrin & fibronectin networks.

New collagen fibers are laid down raoidly & often without functional orientation or attachment to the adjacent hard tissues.

Studies of potential progenitor-cell pools have shown that the marrow spaces of the alveolar bone,particularly along lateral communications between the PDL & the marrow, are sites of cell proliferation.

After extensive damage to the PDL connective tissue, the PDL compartment is populated by an increased number of bone-forming cells, & ankylosis of the tooth to the alveolar bone results.

Surgical attempts to regenerate new PDL attachment have revealed that success depends on:

After removal of inflamed tissue, the root surface must be debrided of contaminants,such as bacterial endotoxins.

Gingival epithelial cells must be prevented from gaining access to the root surface.

The geometric nature of the lesion to be repired is a factor in the prognosis for success.

Defects with intact lateral bone surfaces & with an ample amount of normal PDL adjacent to the area to be repaired are more likely to undergo satisfactory regeneration than are lesions that have horizontal loss of attachment.

Application of growth factors:

Single application of PDGF & IGF to the root surface produced new cementum with functionally oriented PDL fibers & new crestal alveolar bone 4wks postsurgery in monkeys.

Regeneration of PDL is accelerated by the application of PDGF-BB to acid – demineralized root surfaces in membrane-guided repair of furcation defects in beagle dogs.

BMP-7-significant new bone,cementum,& PDL within 8weeks following itts application to furcation defects in beagle dogs.

Recent studies have shown that amelogenin acts as a cell adhesion factor.

Clinical periodontology-carranza.

Oral cells & tissues-garant.

Biology of the periodontal connective tissues

Oral histology-tencate.

Perio 2000-2000

Perio2000-1997.

Perio2000-2006.

The periodontal ligament-berkovitz.

REFERENCES