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What is the periodontal ligament and what is its function?

1. Berkovitz BK. Periodontal ligament: structural and clinical correlates. Dent Update. 2004 Jan-Feb;31(1):46-50, 52, 54. Review.

2. Cho MI, Garant PR. Development and general structure of the periodontium. Periodontol 2000. 2000 Oct;24:9-27. Review.

3. Cohn SA: Transalveolar fibres in the human periodontium. Arch Oral Biol 20:257-259, 1975.

4. Selliseth NJ, Selvig KA. The vasculature of the periodontal ligament: A scanning electron microscopic study using corrosion casts in the rat. J Periodontol65:1079-1087, 1994.

What are the dimensions of the PDL? Do these change with age and/or function?

1. Oehmke MJ, Schramm CR, Knolle E, Frickey N, Bernhart T, Oehmke HJ. Age-dependent changes of the periodontal ligament in rats. Microsc Res Tech. 2004 Mar 1;63(4):198-202.

2. McCulloch, C.A., Lekic, P., McKee, M.D., Role of physical forces in regulating the form and function of the periodontal ligament. Periodontol 2000, 24: 56 – 72, 2000

3. Coolidge ED: The thickness of human periodontal membrane. JADA 24:1260-1270, 1937.

For all periodontal tissues, what are the various types of collagen and where they are found? How are these arranged and what are their functions?

1. Chavrier C, Couble MC, Maglorie H, Grimaud JA : Connective tissue organization of healthy human gingiva. Ultrastructural localization of collagen types I, II, III, and IV. J. Periodontal Res. 19:221-229, 1984.

2. Page RC, Ammons WF. Collagen turnover in gingiva and other mature connective tissues of the marmoset. Arch. Oral Biol. 19:651-658, 1974.

3. Zwarych PD, Quigley MB: The intermediate plexus of the periodontal ligament: History and further observations. J. Dent. Res., 44:383-391, 1965.

What cell type(s) is(are) responsible for collagen production in the periodontal tissues? What are some of the characteristics of these cells?

1. Giannopoulou C, Cimasoni G. Functional characteristics of gingival and periodontal ligament fibroblasts. J Dent Res 1996; 75: 895-902.

2. Mariotti AJ, Cochran DL : Characterization of fibroblasts derived from human periodontal ligament and gingiva. J. Periodontol. 61:103-111, 1990.

3. Palaiologou AA, Yukna RA, Moses R, Lallier TE. Gingival, dermal, and periodontal ligament fibroblasts express different extracellular matrix receptors. J Periodontol. 2001 Jun;72(6):798-807.

4. Kramer PR, Nares S, Kramer SF, Grogan D, Kaiser M. Mesenchymal stem cells acquire characteristics of cells in the periodontal ligament in vitro. J Dent Res. 2004 Jan;83(1):27-34

5. McCulloch C, Melcher A: Cell migration in the periodontal ligament of mice. J Periodontal Res.18:339-352, 1983

6. Lallier TE, Miner QW Jr, Sonnier J, Spencer A. A simple cell motility assay demonstrates differential motility of human periodontal ligament fibroblasts, gingival fibroblasts, and pre-osteoblasts. Cell Tissue Res. 2007 May;328(2):339-54. Epub 2007 Jan 31

What changes occur in collagen in periodontal disease? How is collagen degraded? What cells and agents are involved?

1. Chavrier C, et al. Immunohistochemical study of types I, II, III, and IV collagen in fibrosis of diseased gingiva during chronic periodontitis: A light and electron microscopic study. J. Periodontal Res. 22:29-36, 1987.

2. Buduneli N, Atilla G, Guner G, Oktay G. Biochemical analysis of total collagen content and collagen types I, III, IV, V and VI in gingiva of various periodontitis categories. J Int Acad Periodontol. 2001 Jan;3(1):1-6.

3. Christner P: Collagenase in the human periodontal ligament. J Periodontol 51:455-461, 1980

4. Bildt MM, Bloemen M, Kuijpers-Jagtman AM, Von den Hoff JW. Collagenolytic fragments and active gelatinase complexes in periodontitis. J Periodontol. 2008 Sep;79(9):1704-11.

5. Chang YC, Lai CC, Yang SF, Chan Y, Hsieh YS. Stimulation of matrix metalloproteinases by black-pigmented Bacteroides in human pulp and periodontal ligament cell cultures. J Endod. 2002 Feb;28(2):90-3.

How does smoking affect collagen and collagen production?

1. Zhou J, Olson BL, Windsor LJ. Nicotine increases the collagen-degrading ability of human gingival fibroblasts. J Periodontal Res. 2007 Jun;42(3):228-35.

2. Takeuchi H, Kubota S, Murakashi E, Zhou Y, Endo K, Ng PS, Takigawa M, Numabe Y. JNicotine-induced CCN2:from smoking to periodontal fibrosis. Dent Res. 2010 Jan;89(1):34-9. Epub .

What is the relationship between the periodontal ligament and periodontal regeneration?

1. MacNeil RL, Somerman MJ. Development and regeneration of the periodontium: parallels and contrasts. Perio 2000 19:8-20, 1999.

2. Sculean A, Donos N, et al. Presence of oxytalan fibers in human regenerated periodontal ligament. J Clin Periodontol 26:318-321, 1998.

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What is the periodontal ligament and what is its function?

Berkovitz 2004                              No Article

PURPOSE: To review certain structural aspects of the periodontal ligament (PDL) including: collagen, ground substance, cells, nerves, and blood vessels.

DISCUSSION:

Periodontal Ligament main functions:

• 1)tissue attachment between tooth and alveolar bone and is responsible for displacing forces

• 2)is responsible for the mechanisms whereby a tooth attains, and the maintains, its functional position

• 3)maintains and repairs alveolar bone and cementum

• 4)mechanoreceptors are involved in the neurological control of mastication

Extracellular matrix is made up of:

• 1)collagen fibers: mostly types I and III (3:1 ratio), but types V, VI, VII, and XII also exist. PDL collagen has a high rate of turnover but the significance of this has not been determined.  The nature of the collagen may change in periodontal disease (increase in type V collagen).

• 2)oxytalan fibers: pre-elastin type fibers make up about 3% of the PDL fibers, are attached to the cementum of the tooth and the function is unknown but may have some role in tooth support.

• 3)ground substance: turnover rate faster than collagen. Its functions are ion and water binding and exchange, control of collagen fibrillogenesis and fiber orientation. May play a role in tooth support, eruptive mechanisms and prevention of PDL mineralization.  Content changes in periodontal disease (dermatan sulphateàchondroitin sulphate).

Cells: PDL consists of heterogenous cell population including:connective tissue cells-fibroblasts (most numerous);  formative cells- cementoblasts, osteoblasts;resorbing cells- osteoclasts and odontoclasts/cementoclasts;  stem cells/precursors; defense cells and epithelial cells (rests of Malassez)

Fibroblasts:The typical fibroblast shows a well-developed rough endoplasmic reticulum, Golgi complex and many mitochondria and secretory vesicles. They synthesize and secrete collagen (and ground substance) as well as degrade collagen (through intracellular collagen vacuoles). Cellular activities of PDL fibroblasts can be modulated by bioactive molecules made by themselves, by local inflammatory cells, or be present within the extracellular matrix of the PDL or bone/cementum. PDL fibroblasts produce numerous growth factors and cytokines such as IGFI, BMPs, PDGF, IL-1, and TGFβ. PDL fibroblasts also release prostaglandins which may influence bone cell activity. They are also rich in alkaline phosphatase, cellular retinoic acid-binding protein, and in receptors to epidermal growth factor.

Cell Kinetics and Cell Phenotype:Cell formation and cell differentiation increases markedly with wounding or after the application of orthodontic loads, while different stimuli may recruit progenitors giving rise to different cell types. It is not clear whether periodontal fibroblasts, cementoblasts, and osteoblasts all arise from a common precursor, or whether each cell type has its own specific precursor cell.

Cementogenesis and Periodontal Regeneration:Cementogenesis is a key component of periodontal regeneration but there are still gaps in our knowledge concerning this process in the normal state. Recent studies have shown that enamel matrix protein (EMP) can be applied to a cleaned root surface and can result in periodontal regeneration. The underlying mechanism of this is unknown though it is postulated that EMP may interact with PL fibroblasts via integrins. Cementum attachment proteins (CAP) may play a role during cementogenesis and during periodontal regeneration.

Nerves:The sensory nerves of the PDL show endings of the Ruffini type that plan an important part in the reflex control of mastication. Sensory nerve endings also release neuropeptides, such as substance P, that can have widespread effects on both blood vessels and cells, though their exact role in PDL biology is yet unknown.

Vessels:Major blood vessels of the PDL lie between the principal fiber bundles, close to the wall of the alveolus. The majority of vessels appear to be postcapillary venules. The presence of fenestrated capillaries are related to the high metabolic requirements of the PDL. The number of fenestrations are not fixed and vary according to stage of eruption.

CONCLUSION/BLBasic knowledge concerning the structure and development of the PDL has relevance in understanding and achieving periodontal regeneration.

Cho 2000                             No Article

B: The periodontal tissues develop as the root forms, mostly arising from the dental follicle that is of neural crest origin. These tissues both develop and function as a unit.

P: Review of the development of periodontal tissues by emphasizing the origin and lineage of the cells responsible for formation of their structural components

D: Review of cell and tissue structure of periodontium using TEM, histochemistry, cytochemistry and radioautography to develop periodontal regeneration techniques.

I – Tooth bud formation

Tooth bud is capable of giving rise to all components of a mature tooth. 2 major components are the dental papilla (odontoblasts and dental pulp) and the dental follicle (cementum, PDL and alveolar bone)

The dental follicle develops as neural crest cells migrate to developing branchial arches then interact with early oral epithelium to form tooth primordial. These ectomesenchymal cells aggregate to form dental papilla and dental follicle.

II – Hertwig’s epithelial root sheath

Double layer (inner and outer enamel epithelial cells): separates dental papilla from dental follicle. It is continuous with the apical rim of the enamel organ, but continuity is lost at onset of root formation

The inner epithelial layer induces odontoblast differentiation. (the developing root gives rise to fibroblasts, preodontoblasts and precementoblasts). It also produces proteins such as bone sialoprotein, osteopontin and amelin along with components of the basement membrane. It has been hypothesized, but not proven, that these secreted elements cause differentiation of cementoblasts (stimulates cementogenesis), but this has not been proven

The outer layer of HERS breaks up at the onset of cementogenesis. More recent studies: possible that some of these epithelial cells undergo mesenchymal transition into fibroblasts (secrete acellular cementum) and cementoblasts (secrete cellular cementum), whereas the rest retain an epithelial phenotype and survive in the PDL as rests of Mallasez (traditional thinking)

III.- Cementum

Avascular mineralized tissue covering the entire root surface. Forms the interface between root dentin and PDL.

2 types: cellular cementum has cementocytes within, acellular does not. Further grouping is based on presence of collagen fibers; intrinsic has collagen fibers formed by cementoblasts and extrinsic has collagen fibers formed from fibroblasts.

Acellular afibrillar cemenum: over cervical enamel at CEJ, major component is glycosaminoglycans, its functional significance is unknown.

Cellular intrinsic fiber cememtum: contains cementocytes embedded in a collagenous matrix of intrinsic collagen fibers. Found in old resorption lacunae and root fracture sites

Cellular mixed stratified cementum: located primarily on the apical one third of the root and in the furcation area of multirooted teeth. It is composed of alternating layers of acellular extrinsic fiber cementum and cellular intrinsic fiber cementum/acellular intrinsic fiber cementum, and is covered by a thin layer of acellular extrinsic fiber cementum for attachment to the periodontal ligament. Serves to reshape the root surface in order to compensate for physiological drift and nonphysiological shifting of teeth in their alveolar sockets.

Acellular extrinsic fiber cementum covers 40% to 70% of the root sur- face and is comprised of collagen fibers and glycosa- minoglycans. It serves the exclusive function of an- choring the root to the periodontal ligament.

Cementogenesis of acellular extrinsic fiber cementum: In humans, cementoblast differentiation and cementogenesis are closely related with root formation. HERS detaches from dentin at the apical edge of the developing root. Fibroblasts of the dental follicle lay down “fringe fibers” which will be incorporated into the layer of acellular extrinsic fiber cementum once mineralization begins. These fringe fibers will become continuous with the principal fibers of the PDL once they develop.

When root development is ~ 2/3 complete, shifts to formation of the cellular varieties. Rapid deposition from multiple sites leads to entrapment of cementoblasts in the matrix as cementocytes.

IV. PDL

Develops prior to tooth eruption at the time of root formation. Perifollicular mesenchymal cells have increased cellular volume and synthetic activity, become elongated, then actively synthesize and deposit collagen fibrils and glycoproteins. These fiber bundles will eventually merge with the fringe fibers and be embedded in bone/cementum as Sharpey’s fibers.

Distinct groups of principle fibers histologically: dentogingival, alveolar crest, transseptal, interradicular, horizontal, oblique and apical fiber bundles.

Mature PDL has 3 distinct regions: 1. bone-related with lot of cells/blood vessels, 2. cementum-related with dense well organized collagen bundles, and 3. middle zone: fewer cells and thinner collagen

Contain undifferentitated stem cells that retain the potential to differentiate into fibroblasts, cementoblasts and osteoblasts (possible that damage to PDL causes differentiation to osteoblasts which then causes ankylosis)

Fibroblasts: most abundant cell in PDL. 1. needed to maintain normal width of PDL, 2. can give rise to osteoblasts and cementoblasts, 3. produce acellular extrinsic fiber cementum in mature PDL, 4. responsible for collagen fiber formation and removal, and 5. express numberous epidermal growth factor receptors

V. Gingiva

Comprised of gingival epithelium and CT. covers the tooth-bearing part of alveolar bone and cervical neck of tooth. Regional morphological variations ( oral GE, oral SE, JE)

At eruption: a thick reduced enamel epithelium overlying the enamel fuses with oral epithelium, transforms, then establishes dentogingival junction (JE directly attached to tooth). 10-20 cells wide at coronal end, 1-2 cells at apical end. In health, the apical aspect of JE is at CEJ.

CT attachment: densely packed collagen bundled anchored to the acellular extrinsic fiber cementum just below terminal part of JE. Stability of CT is key factor in limiting apical migration of JE. Gingival CT fibroblasts secrete collagen matrix organized into fiber bundles

Gingival supra-alveolar fiber apparatus: transseptal, circular, semicircular, transgingival & intergingival fibers: connect/link adjacent teeth in the arch. Secure against rotation and maintain linkage during drift.

VI. Alveolar Bone

Maxilla and mandible have 2 components: alveolar process (houses the tooth roots) and basal body.

Alveolar process consists of thin alveolar bone proper (socket wall), inner and outer cortical plates, and spongy bone in between. Size, shape, location and function of the teeth determines the morphology of alveolar process.

Begins formation during late bell stage of development, separating individual tooth germs. Changes occur as the root forms: osteoblasts differentiate from dental follicle and form alveolar bone proper.

Major function is to anchor roots of teeth and to absorb/distribute occlusal pressures. This is achieved by insertion of Sharpey’s fibers into alveolar bone proper.

Cohn 1975                             Article

BG: It has been established that numerous cemento-alveolar fibers in the mouse, marmoset, and macaque monkeys do not terminate in bone but pass without interruption through the wall of the alveolus. These unusual fibers were termed transalveolar fibers.

P:To examine the transalveolar fibers (the cemento-alveolar fibers that traverse the entire thickness of the alveolus) in humans.

M+M:Jaws of 16 adult cadavers and a few fresh specimens were fixed and cut into blocks of 2 adjacent teeth. Blocks then cut into 10um sections in a mesio-distal plane and stained by a modified Mallory (trichrome stain using aniline blue, acid fuchsin, and orange G reveals collagen of CT) method.

R:Many cemento-alveolar fibers traversed the entire thickness of the alveolus instead of being anchored in bone as conventional Sharpey’s fibers. Observed only in regions of lamellar bone, lacking Haversian systems. The fibers could join roots of adjacent teeth, the roots of the same tooth, the periosteum of the alveolar process, or to the lamina propria on gingival surfaces.

BL:The author proposes orientation and distribution of transalveolar fibers likely represents a functional adaptation to occlusal and muscular forces, permitting maximum support of the tooth in its alveolus.

CR:How many is many? This study does not specify.

Selliseth 1994                             No Article

P: To examine the 3-dimensional architecture of the microvascular system of the rat periodontal ligament (PDL).

M&M: 6 rats had liquid acrylic resin perfused through each of carotid arteries. Vascular corrosion casts were prepared and examined by SEM.

D: The results show that the microvasculature forms a highly organized system presumably related to the specialized functions of the periodontium. Cervically, arterioles and venules communicated with the profuse capillary network of the gingiva. The mid-root segment of the PDL contained arterioles and venules that coursed occluso-apically near the alveolar wall, as well as capillary loops located closer to the surface. Arterioles entered PDL through vascular canals from the bone marrow when proceeded coronally and branched into an interconnected capillary network. The capillaries formed hairpin loops pointing coronally. At the apical portion, capillary loops were larger in diameter, coursed apically, and anastomosed freely until entering a venule.

BL: Cervically, a dense capillary system may be required for antimicrobial defenses and rapid tissue turnover. Mid root vasculature supports the suspensory structures, while the apical region has a venous cap designed maybe for cushioning of masticatory forces. The large vessel diameter combined with an irregular lumina surface at the tip of capillary loops indicates reduced blood velocity and turbulence in the functional part of the PDL vasculature where exchange of metabolites mainly occur.

What are the dimensions of the PDL? Do these change with age and/or function?

Oehmke 2004                             Article

Purpose: To analyze the normal age-dependent changes and regional differences of the collagen renewal rate of the PDL in rats.

Materials and methods: Nine male rats were used and divided in 3 groups: Group A: 1 month old, Group B: 8 months old and Group C: 18 months old. Animals received an injection of 3H-proline for he labeling of newly formed collagen and were killed 8 hours after that. Parts from the mandible, muscles and blood were sampled. The mesial roof o the 1^stand 2^nd lower right molars of each animal were evaluated. Autoradiography was used to visualize the activity of fibroblasts as collagen-forming cells.

Results: Autoradiograms demonstrated age-related alterations and regional differences of the collagen renewal rate of the PDL. In the cervical third of the PDL of Group A, the density of silver grains (labeled molecules) was significantly higher than in the adjacent bone or dentine. In 8-month-old rats a lower density of silver grains was observed and was further diminished at 18-month-old specimen. In the middle root third, there was a marked decrease in the number of silver grains with increasing age and an irregular shaped PDL comparing to the cervical third. In the apical the situation in younger rats is similar to that in the cervical third. The density of silver grains was slightly reduced in the 8-month-old animals but was stull markedly higher in the apical zone. IN the 18-month-old rats there were even fewer silver grains. The labeling was reduced with age, it was always lowest in the middle root third and highest in the apical third, with the values of the cervical third in between.

Conclusion:In all age groups, the formation of collagen occurred mainly in the apical and cervical root thirds, presumably subject to functional demand.

McCulloch 2000                             Article

P: Review of how cytoskeletal proteins mediate protective responses to applied force which may enable the cells to survive in a mechanically active environment.

PDL: bundles of fibers arranged in a meshlike net stretching between the cementum and bone. It is the only ligament to span two distinct hard tissues (cementum and bone). It has a dynamic relationship to external forces, with specific metabolic requirements and architectural tissue design.

Maintenance and remodeling of collagen as well as calcification of the extremities to form Sharpey’s fibers require numerous cell types with multiple signaling mechanisms. The PDL fibroblast appears to be responsible for formation and remodeling of PDL fibers.

PDL fibroblast dispersed throughout the ligament, generally organized with their long axis parallel to the direction of the collagen fiber. There appear to be multiple organized contact points for cell signaling cascades to occur quickly in response to external stimuli. Other cells existing within the PDL including endothelial cells, epithelial rests of Malassez, sensory cells, osteogenic and osteoclastic cells and cementoblasts.

The ECM of the PDL appears to have a much higher turnover rate than other dental tissues. It is possible that only certain portions of the fibrils are broken down, and not the entire fiber, in response to certain stresses. This is important as portions of the fibers are embedded into bone which is remodeled under different circumstances than cementum.

B/c the PDL maintains a generally constant width throughout life, several mechanisms must maintain homeostasis. Cytokines and growth factors are important for acting locally, and there are several signaling signals in place to respond to mechanical forces in order to maintain width. The PDL secretes and expresses proteins for regulating PDL growth, but also appears to be able to influence bone metabolism as well.

If PDL cells are removed from the root or disturbed by medications, bone generally grows into the PDL space and ankylosis occurs.

PDL and alveolar bone cells are exposed to physical forces in vivo in response to mastication, parafunction, speech and ortho movement. The actual process of remodeling is, at this time, poorly understood. It also relatively unknown how force is specifically transferred from the ligament into the bone. Models being studied generally revolve around the stress-strain relationship.

BL: The fibroblasts and osteoblasts within the PDL have the necessary signaling and effector mechanisms to both sense forces and produce an applied response to maintain the PDL width and cell viability.

Coolidge 1937                             Article

P: To examine the thickness of the human periodontal ligament at different ages and in teeth showing different types of occlusion.

M&M:1145 measurements from 172 teeth of 15 human jaws were used. PDL thickness was measured at the alveolar crest, at mid-root, at the apex of the tooth, and at the bifurcation of multi-rooted teeth. Measurements were made at M/D, B/L, or all 4 surfaces. Plaster casts of the models were mounted on articulators to reproduce occlusal relationships and determine the amount of function: if there were fewer teeth in the arch they were labeled as heavy function while teeth w/o antagonists were labeled as no function. Unerupted or embedded teeth were labeled as a separate group.

R: The thickness of the PDL decreased with age (except around teeth in heavy function). PDL thickness generally increases with function, but can vary with the type of stress. PDL was thinner on the pressure side of drifting teeth, and thicker on the tension side. Drifting and malaposed teeth were found to have a relatively thick periodontal membrane. The following are averages of the findings:

11 – 16 years: 0.21mm heavy function: 0.18mm

32 – 50 years: 0.18mm no function: 0.13mm

51 – 67 years: 0.15mm embedded teeth: 0.08 mm

Malposed/drifted 0.19 mm

BL: The thickness of the PDL is a variable amount and is affected by age, intensity of functional forces, and the tooth position (drifting, malposition). Avg thickness is 0.10-0.20 mm.

For all periodontal tissues, what are the various types of collagen and where they are found? How are these arranged and what are their functions?

Chavrier 1984                             Article

BG: Type I and III collagen are the main collagenous components of the healthy human gingival connective tissue (99% of the total extractable collagen) with a predominance of type III collagen in the gingival papillae underlying gingival basement membrane, and around the blood vessel walls, while Type IV collagen was the main collagenous component of basement membrane (accounts for less than 1%.)

P: To examine the morphological pattern of organization of the gingival CT and its collagen

components, by using the indirect immunoperoxidase labeling procedure.

M&M: 7 healthy dental students had 6 mm³ of attached gingiva biopsied. 2 mm³ sample block

were subsequently cut and divided into two groups: a) Standard EM, and b) Indirect

Immunolabeling, c) controls.

R: Standard EM – 2 Patterns observed:

Dense tissue (Predominant), with large dense bundles of long, thick, striated collagen fibers (60-70nm). Often in close contact with mature fibroblasts

Loose CT, underlying gingival basement membrane or blood vessels walls. Short thin (40-60 nm) striated collagen fibers, mixed with non-striated material (mast and plasma cells).

Immunoperoxide labeling – 2 patterns observed:

Type I collagen arranged in thick bundles (60-70nm), with 64nm space between the fibers.

Mixed I and III collagen, with type III, to be the predominant and organized either in fibrous (short, thin, striated fibers) or fibrillar (wide-spread, thread-like material) form. Type IV, was limited to the lamina densa of the basement membrane.

D: Healthy gingival CT has heterogenicity of collagen that can be divided into 2 types of

organization and composition. One type of collagen predominates in each of the 2 patterns of

gingival CT. Fibroblasts in 1 region produce more of its respective collagen. Type 1 = stability

and Type III = early regeneration

BL: Collagen types are reflected in the tissue’s function. Immuno-typing is an advance in the attempt to distinguish different patterns of organization in healthy gingival connective tissue.

Page and Ammons 1974                             Article

P: To study the normal biologic properties of CT turnover in healthy tissues.

M&M:  12 marmosets were used in this study.  Each animal was given (U)¹⁴C-L-proline on 2 consecutive days.  24 hours after the second dose the animals were killed and blood and tissue was collected.  Several CT (tendons, skin, gingiva, palate) expected to exhibit varying rates of collagen turnover, were selected for analysis and comparison with the gingiva. Any areas that displayed gingival inflammation were excised and not included.   The extent of incorporation of  (U)¹⁴C-L-proline into collagen hydroxyproline and the subsequent loss of the label from the tissue was evaluated over 17 weeks.

R: Initially there was a very high incorporation of proline and conversion into hydroxyproline in all of the tissues.  During subsequent periods, significant numbers of counts remained only in the gingiva, but after 17 weeks activity began to approach the values seen in other CT.

D: The rate of conversion of ¹⁴C-proline into ¹⁴C-L-hydroxyproline by CT reflects the rate of collagen production, and the rate of loss of the labeled hydroxyproline from the tissue indicates the levels of collagen degradation. The gingiva differs from other CT in that a far greater portion of the newly synthesized molecules is required for incorporation into the insoluble collagen. The data shows that the turnover rate of mature insoluble collagen in normal gingiva is rapid (5X) when compared to other connective tissues.

In inflammation, the observed net loss of collagen may result from interference with collagen production and turnover, rather than from destruction of previously existing collagen.

Zwarych 1965                             Article

PURPOSE: To determine if an intermediate plexus exists in the mammalian periodontal ligament in teeth of limited growth.

METHODS: 8 adult white mice were sacrificed and decapitated.  Heads were fixed, bisected sagitally, and embedded in paraffin and sectioned so that mesio, distal, occlusal, and oblique plane sections could be studied histologically.

RESULTS: The sections of mouse molars support the concept of continuity of the principal fibers across the periodontal space.  No “intermediate plexus” was found.  Shortly after passing from the alveolar septum, the closely packed fibers became more loosely arranged and the individual fibers took separate courses joining a few adjacent fibers to become attached to the cementum.  A greater number of bundles containing fewer fibers were attached to the cementum in contrast to a lesser number of bundles containing more fibers attached to the alveolar septum. 

DISCUSSION:  The evidence indicates what the PDL fibers are continuous across the periodontal space.  More numerous bundles with fewer fibers are attached to cementum while fewer bundles with more fibers are attached to the alveolar bone of the socket, allowing displacement forces applied to a given area of cementum to be transmitted to a greater area of the alveolar septum.

CONCLUSION/BLThere is no “intermediate plexus” in mice molar teeth with limited eruption. Principle fibers are continuous from the cementum to alveolar bone, leading to enhanced compressibility and strength of the periodontal ligament in normal tooth movement.

What cell type(s) is(are) responsible for collagen production in the periodontal tissues? What are some of the characteristics of these cells?

Giannopoulou 1996                             Article

P:To study and compare the functional characteristics of gingival and PDL fibroblasts.

M&M:Gingival and PDL fibroblasts (GF & PDLF) from 5 healthy Caucasian males 25-30 years old were isolated and compared in vitro. Patients were undergoing extractions for orthodontic reasons. The CT cells were taken from PDL of premolar teeth and adjacent healthy gingiva or interdental papilla. The cells were prepared and observed under SEM. The effect of extracellular matrix components (ECM) on attachment, proliferation and protein synthesis were examined. The agents used were: collagen type I, IV, gelatin, fibronectin, laminin and vitronectin. Muscle differentiation markers and the effects of epithelial cells were also examined.

R:GF and PDLF appeared similar under SEM. They appeared rounded, with a spherical nucleus in the center and typical prolongations. Generally, in primary cultures, the proliferation of gingival fibroblasts was faster than that of PDL fibroblasts but the differences were not SS. All ECM components enhanced attachment; however, while collagen types I and IV were more effective in promoting the attachment of GF, gelatin, laminin, and vitronectin promoted attachment of PDLF. Both cell types demonstrated same degree of enhanced attachment with fibronectin. Most ECM components increased the proliferation rate of GF and the biosynthetic activity (protein synthesis) of PDLF.The biochemical markers were similarly distributed between the 2 cell types, except for alkaline phosphatase, which was detected only in the cellular extract of PDLF. Both GF & PDLF strongly expressed alpha-smooth-muscle actin, but only PDLF were positive for smooth-muscle myosin. Epithelial cells significantly stimulated the proliferation of both GF and PDLF but had no effect on their biosynthetic activity.

BL:This in vitro investigation confirmed that GF and PDLF have a similar morphology, but physiological and chemical differences may better explain their in vivo functional differences.

Mariotti 1990                             Article

P:To determine the characteristics of fibroblasts derived from human PDL and gingiva.

M+M: PDL fibroblasts (PDLF) were isolated from impacted 3^rdmolars (21-35year old) healthy adults. Human gingival fibroblast (hGF) were isolated from interproximal papilla of premolar or molar gingiva. PDLF and hGF were incubated then replanted on 500,000-cells/100mm-culture dish. Fibroblasts were used between the 3^rd and 5^th passages only.

R:

GF grew faster than PDLF, in a 500,000 cells / 100 mm culture; a total confluence occurred in 4 days for GF and 6 days for PDLF.

DNA content of growing cells was greater in GF than PDLF.

Total protein content in GF was slightly greater than PDLF at day 7 but NSSD.

Greater trend on non-collagen protein synthesis in GF, and more collagen synthesis in PDLF.

GF had greater amounts of hyaluronic acid and heparin and lesser amounts of chondroitin sulfates A and C.

The growth characteristics of PDLF and GF was similar but did exhibit specific differences in proliferate rates and macromolecular synthesis.

D: An explanation for GF cells reaching confluence earlier than PDLF may be because of the gingival cells being larger than PDLF cells. This is supported by the findings that the DNA and protein content of GF cultures are initially greater than PDLF.

BL: GF cells reached total confluence faster than PDLF cells and had different productions of macromolecules.

Palaiologou 2001                             Article

B: Fibroblasts are the main cell of the periodontal ligament and gingiva and play important roles in function/regeneration of periodontal tissues. Glycoproteins are important for cell-cell and cell-matrix interactions. Fibronectin is the main glycoprotein in connective tissue and serves to orient fibroblasts to collagen and provide protein attachment for cell-matrix adhesions linking collagen and fibrin to the cell surface and underlying actin cytoskeleton. Fibronectin contains Arg-Gly-Asp (RGD) sequence that is part of the cell binding site and plays a crutial role in migration of fibroblasts and maintaining structural integrity of connective tissue.

P: To evaluate any differences in binding of fibroblasts (human gingival (GF), periodontal ligament (PDLF, and dermal (DF))to various ECM proteins (fibronectin, laminin, vitronectin, other peptides) and collagen (Type I and IV).  Differences in integrin expression was also looked at for the different types of fibroblasts.

M&M: GF were taken from a 13-year-old boy that was systemically healthy and underwent surgery for hereditary gingival hyperplasia. Dermal fibroblasts were ordered and came from a 12 week old female embryo. Stock cultures of polyclonal human periodontal ligament fibroblasts were used. ECM proteins were commercially obtained. Cells were allowed to adhere to the substratum for 45 min to 2 hours. Non adherent cells were removed by the 3^rd wash of PBS and lysed by freezing. ECM proteins were prepared according to manufacturer’s instructions and added to each sample and incubated for 30 min. Adherent cells were quantified fluorometrically using a fluorescent die. Guanidine thiocyanate was used to extract the RNA from the 3 cell types. RT-PCR was used to assess transcript expression. The primers used, specific for the integrin subunits to quantify ECM receptor transcript expression, were derived from the published DNA sequence for human integrins.

R: GF and PDLF adhered to vitronectin and collagen type 1 and IV more than DF. PDLF adhered more to laminin than, whereas GF and DF did not. All adhered well to fibronectin and RGD peptide. There were found to be innate differences between the fibroblast types and the integrin transcripts they expressed.

BL: GF and PDLF are more similar in the ECM proteins that they adhere to and the integrins they express when compared to DF. Moving forward, experiments using dermal fibroblasts would not necessarily be useful in studies looking at oral tissues.

Cr: Could have tested several fibroblasts from other patients in addition to the 13 y/o boy with hereditary gingival hyperplasia to confirm results.

Kramer 2004                             Article                   mesenchymal stem cells

Purpose: To determine if mesenchymal cells differentiate into a specific cell type.

Materials and methods: Samples of extracted teeth were obtained from female subjects needing extractions. Proliferative cells having various morphologies were produced from the section in culture between 7 and 10 days. Cell with PDL morphology were diluted and cultured and processed for immunohistochemistry. Human male mesenchymal cells were obtained and cultured according to the manufacturer’s directions. Mesenchymal stem cells were mixed with periodontal cells isolated from the tooth explants at rations of 1:1, 2:1 and 10:1. Co-cultures of cells were isolated after 0, 3, 7, 14 and 21 days and processed for immunohistochemistry and in situ hybridization.

Results: Collagen III staining was restricted to PDL cells and not in the cementum and dentin layers. Osteopontin was present in the PDL, osteocalcin was heterogeneous in the PDL and observed in the bone, dentin and cementum and BMP-2/4 could be detected in the PDL, cementum and bone tissue. PDL did not stain for bone sialoprotein. These indicate that PDL can be differentiated from other periodontal tissues within the explant. The staining pattern for mesenchymal cells was different form that seen for PDL and have different morphology. Co-culture for 7 days led to an overall change in the mesenchymal stem cells structure, to a more fibroblast – like morphology. Osteocalcin and osteopontin expression was up-regulated in mesenchymal stem cells following co-culture for 7 and 21 days and expression of bone sialoprotein was reduced.

Conclusion: Data demonstrate mesenchymal stem cells’ potency to develop periodontal ligament characteristics and suggest that the cells may have the potential to form other periodontal tissues.

McCulloch 1983                             Article

Purpose: First to confirm that cells in a normally functioning PDL do migrate; second to analyze the rate at which the cells cycle; third to analyze the relationship between proliferating cells within the PDL in order to assess further their capacity for migration and for clonal proliferation within the PDL.

Materials and methods:

• 135 mice were injected ³H-Tdr dilated with PBS then sacrificed at 1hr, 1 day, 3days, 7days, 14days, 60days and three control mice were injected with PBS.

• Then mice mandibles were block sectioned in the molar area and analysis of radioautographs using labeling index and grain counts.

Results:

• Analysis of radioautographs using labeling index and grain counts demonstrated that the majority of labeled cells divided within 3days, but a measurable population of cells had not divided after 14days.

• The labeling index of cells adjacent to alveolar bone increased 8 times within 1 day, indicating that labeled cells had migrated to the bone surface.

• Migration of cells to the vicinity of the cementum was observed 3 days after labeling.

• The percentage of labeled cells located within 20 m of one another increase to 40% within 3 days, suggesting clonal proliferation of PDL cells.

BL: Cells of PDL migrate under physiological conditions. This conclusion is supported by first, the evidence that progeny of labeled cells can migrate from their paravascular location; second, that members of clones migrate from the site of their birth; and third, that cells migrate to the surfaces of bone and cementum.

Lallier 2007                             Article

P:To investigate the motility of the cells of the periodontium, since this may influence their ability to aid in tissue regeneration. Moreover, to determine whether different ECM proteins (collagen I, collagen III, collagen V,fibronectin, and laminin) can be used to promote differential cell (PDL and HGF and osteoblasts) motility.

M&M:Periodontal and gingival fibroblasts were established from patients who had healthy gingiva but who underwent oral surgery at the Louisiana State University School of Dentistry for the purpose of removing impacted wisdom teeth. Cell lines of pre-osteoblasts (ATCC-CRL-11372) were obtained from the American Type Culture Collection. Mature osteoblasts were obtained by growing pre-osteoblasts at 37°C for 5 days prior to use in subsequent assays. Cell motility assay, cell movement evaluation, cell adhesion assay and cell proliferation assay were carried out for different cell types and different ECM proteins. ELIZA was done to detect integrin β1, α1, and α2 subunit expression, using different Integrin subunit antibodies.

R:Gingival fibroblasts are twice as motile as PDL fibroblasts, whereas osteoblasts are essentially non-motile. Collagens promote the greatest motility of gingival fibroblasts in the following order: collagen III>collagen V>collagen I. Differences in motility do not correlate with cell proliferation or integrin expression. Osteoblasts display greater attachment to collagens than does either fibroblast population, but lower motility. Gingival fibroblast motility on collagen I is generally mediated by α2 integrins, whereas motility on collagen III involves α1 integrins.

BL:ECM proteins, differentially promote the cell motility of periodontal cells. Because of their greater motility, gingival fibroblasts have more of a potential to invade periodontal wound sites, which may explain the formation of disorganized connective tissue masses rather than the occurrence of the true regeneration of the periodontium.

What changes occur in collagen in periodontal disease? How is collagen degraded? What cells and agents are involved?

Chavrier 1987                             Article

P: To study the distribution, ultrastructure and organization of type I, III, IV collagen in fibrotic gingival CT of pts with long standing cases of chronic periodontitis.

M&M: 5 pts with progressive, long standing periodontitis provided tissue samples that were evaluated by immunofluorescence (IF), Standard electrom microscope (SEM) , and immunoperoxidase electron labeling (IPEL).

R: IF: The diseased CT was made up of both type I and type III collagen. Type I collagen was strongly fluorescent and appeared to be the main gingival collagenous comp

HOME PERIO TOPICS 

Discussion Topics

A. What are the various modes of intercellular attachment? Draw and label a hemidesmosome.

B. Is a gingival sulcus necessary and/or desirable?

C. What is the “biologic width” and what is its significance?

D. What is the turnover rate of the oral epithelia? Draw and label the exfoliation of the cells of the junctional epithelium.

E. What is the embryogenesis of the junctional epithelium?

F. Define primary and secondary junctional epithelium.

G. How does the junctional epithelium heal after excision? After incision?

H. Can and/or should sulcular epithelium be keratinized? Why?

I. Describe the changes that occur in the junctional epithelium at the light and electron microscope level during gingivitis and periodontitis development.

How does a periodontal pocket form?

Anatomy and Development

1. Kobayashi K, et al. Ultrastructure of the dento-epithelial junction. J. Periodontal Res. 11:313-330, 1976

2. Stern IB: Current concepts of the dentogingival junction: the epithelial and connective tissue attachments to the tooth. J. Periodontol. 52:465-476, 1981.

3.Ten Cate AR:  The dento-gingival junction.  An interpretation of the literature.  J. Periodontol.  46:475-477, 1975.  (Review)

4. Pollanen MT., Salonen JI, Uitto VJ. Structure and function of the tooth –epithelial interface in health and disease. Periodontl 2000. 2003; 31:12-31

5. Hujoel PP, White BA, Garcia RI, Listgarten MA. The dentogingival epithelial surface area revisited. J Perio Res 36:48-55, 2001.

6. Messer RL, Davis CM, Lewis JB, Adams Y, Wataha JC. Attachment of human epithelial cells and periodontal ligament fibroblasts to tooth dentin. J Biomed Mater Res A. 2006 Oct;79(1):16-22.

Can you quantify inflamed periodontal tissues?

7. Nesse W, Abbas F, van der Ploeg I, Spijkervet FK, Dijkstra PU, Vissink A. Periodontal inflamed surface area: quantifying inflammatory burden. J Clin Periodontol. 2008 Aug;35(8):668-73. Epub 2008 Jun 28.

Describe the dimensions of the DGJ and the significance of the “biologic width” in dentistry.

8. Garguilo, A., et al: Dimensions and relations of the detogingival junction in humans. J Periodontl 32: 261-267, 1961

9. Vacek, J.S., et al: The dimensions of the human dentogingival junction. Int J Perio Rest Dent 14: 155 – 165, 1994

10. Perez J, Smukler H, Nunn M. Clinical dimensions of the supraosseous gingivae in healthy periodontium. J Periodontol 2008; 79: 2267 – 2272

11. Novak MJ, Albather HM, Close JM. Redefining the biologic width in severe generalized, chronic periodontitis: Implications for therapy. J Periodontol 2008; 79: 1864

Describe the pocket epithelium

12.Muller-Glauzer W, Schroeder HE. The pocket epithelium: A light and electron microscopic study. J. Periodontol. 53:133-144, 1982.

What are the characteristics of the junctional epithelium, including its structure, function, turnover rate?

13. Hatakeyama S, Yaegashi T, Oikawa Y, Fujiwara H, Mikami T, Takeda Y, Satoh M. Expression pattern of adhesion molecules in junctional epithelium differs from that in other gingival epithelia. J Periodontal Res. 2006 Aug;41(4):322-8.

14. Bosshardt DD, Lang NP. The junctional epithelium: from health to disease. J Dent Res. 2005 Jan;84(1):9-20. Review.

Wound Healing

15. Taylor AC, Campbell MM. Reattachment of gingival epithelium to the tooth. J Periodontol. 43:281-293, 1972.

16. Listgarten MA. Ultrastructure of the dento-gingival junction after gingivectomy. J. Periodontal Res. 7:151-160, 1972.

17. Braga AM, Squier CA : Ultrastructure of regenerating junctional epithelium in the monkey. J. Periodontol. 51:386-392, 1980.

18. Tomofuji T, Sakamoto T, Ekuni D, Yamamoto T, Watanabe T. Location of proliferating gingival cells following toothbrushing stimulation. Oral Dis. 2007 Jan;13(1):77-81

What is the significance of keratinization of sulcular epithelium?

19. Caffesse RG et al: The effect of mechanical stimulation on the keratinization of sulcular epithelium. J. Periodontol. 53:89, 1982.

20. Squier CA: Keratinization of the sulcular epithelium – a pointless pursuit? J. Periodontol. 52:426-429, 1981.

Epithelial Rests

21. Grant DA, Bernick S. A possible continuity between epithelial rests and epithelial attachment in miniature swine. J. Periodontol. 40:87-95, 1969.

22. Spouge JD. The rests of Malassez and chronic marginal periodontitis. J Clin Periodontol 11: 340-347, 1984.

23. MacNeil RL, Thomas HF. Development of the murine periodontium. II. Role of the epithelial root sheath in formation of the periodontal attachment. J Periodontol 1993;64:285-291.

24. Rincon JC, Young WG, Bartold PM. The epithelial cell rests of Malassez–a role in periodontal regeneration? J Periodontal Res. 2006 Aug;41(4):245-   52. Review.

25. Shimonishi M, Hatakeyama J, Sasano Y, Takahashi N, Uchida T, Kikuchi M, Komatsu M. In vitro differentiation of epithelial cells cultured from human periodontal ligament. J Periodontal Res. 2007 Oct;42(5):456-65.

26. Becktor KB, Nolting D, Becktor JP, Kjaer I. Immunohistochemical localization of epithelial rests of Malassez in human periodontal membrane. Eur J Orthod. 2007 Aug;29(4):350-3. Epub 2007 Jul 2.

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Topic Overview

Anatomy and Development

Kobayashi 1976                    Article

:to present new data on the dento-epithelial junction.

The dento-epithelial junctions of 5 Rhesus monkeys were examined by SEM. After making gingival incisions teeth with intact gingiva including the margin of the alveolar bone were excised and examined with electron microscope.

The junctional epithelium contacted the enamel, the afibrillar cementum covering the enamel surface and the root cementum (fibrillar cementum). Multiple hemidesmosomes were easily recognized along the cell membrane facing the tooth surface.

junctional epithelium consists of two basal laminae. Internal lamina towards the tooth and external lamina towards the connective tissue.

 

 Attachment coronal to the CEJ :

Well developed dental cuticles were seen between the afibrillar cementum overlying the enamel and the junctional epithelium. Some areas had no evident dental cuticle. Where the JE apposed the afibrillar cementum without an intervening dental cuticle, there was between the basal lamina and the afibrillar cementum a thin dense line, the linear border.

Junctional epithelium:

Internal Basal lamina: Lamina lucida, lamina densa, sublamina lucida

The lamina lucida occupied the narrow space between the peripheral density and the lamina densa. The lamina densa appeared suspended between the lamina lucida and the clear sublamina lucida. The sublamina lucida lay between the lamina densa and the linear border or the dental cuticle. The border between the dental cuticle and the subjacent afibrillar cementum was highly irregular and unclear.  Linear border was not defined in areas where the dental cuticle was well developed. However, the linear border was sharply evident between the enamel and sublamina lucida, between the enamel and the dental cuticle and between the afibrillar cementum and the sub-lamina lucida.

 

 Attachment apical to the CEJ

Basal lamina retained a uniform width.  In some cases, the surfaces of the root cementum were covered with a dental cuticle of great thickness variation. The hemidesmosomes were larger than those apposing the basal lamina of the oral epithelium and were sharply outlined along the surfaces of the junctional epithelium. In all specimens the attachment of the lamina densa to the root cementum or the dental cuticle was mediated by a thin clear space, the sub-lamina lucida. The linear border was especially evident in areas devoid of the dental cuticle and where the root cementum lacked collagen fibrils. It was never observed between fibrillar cementum and the basal lamina or dental cuticle. Where the dental cuticle came in contact with afibrillar cementum, this line could not be detected.

Special Considerations of the Denial Cuticle and Hemidesmosomes

Under high-power electron microscopy the dental cuticle may be described as a relatively homogeneous, somewhat granular and electron-dense layer.

Throughout these observations, the hemi-desmosomes were well-preserved regardless of the fixation procedure used. In high- power views of selected areas, details of the hemidesmosomes contained dense pyramidal particles along the inner surface of the peripheral density. Fine filaments extended from the peripheral density into the lamina densa.

CONCLUSION: The investigation has once again confirmed the concept that an attachment apparatus consisting of multiple hemidesmosomes and a basal lamina promotes adhesion of the JE to the teeth. This attachment apparatus behaves as a unit which is maintained a short distance from the teeth or dental cuticle by an intervening sub-lamina lucida.

Stern 1981                    Article

  Review article discussing all the history and current concepts of the dentogingival junction (DGJ; the epithelial and CT attachments). 

  Overall concept is that the DGJ is dynamic rather than static, with a high rate of turnover. Primary junctional epithelium is derived from ameloblasts. During ameloblast histodifferentiation the cells pass through two phases: forming enamel in the first, and primary junctional epithelium in the second. In the ameloblast life cycle, after the enamel matrix secreting and mineralizing stages of amelogenesis, the ameloblasts become smaller, terminate the enamel-forming function, and begin to form the epithelial attachment.

Secondary junctional epithelium is derived from gingival epithelium. 

Basal lamina is composed of lamina densa (composed of epithelium and CT interfaces) and lamina lucida (between outer leaflet of epithelial cell membrane and tooth it has important role in epithelial attachment).

Intracellular edema decreases desmosomes and disruption of CT facing basal lamina.

Pocket formation is associated with a loss of cellular continuity in coronal portion of JE.

Healthy JE – no Rete pegs, Diseased – rete pegs,  Return to health – Rete pegs remain. 

The DGJ shows a capacity to repair/regenerate following plaque elimination and resolution of inflammatory infiltrate.

Ten Cate 1975                    No Article

P:  To focus attention on the CT component of the DGJ and its possible significance in the etiology of periodontal disease.

D: CT is important in determining the development, structure, and function of epithelium. In embryology, the mesenchyme (embryonic CT) has a key role in determining the epithelial response (Billingham and Silvers;1968 and Slavkin; 1972). All epithelium responds in the same manner to a change in its surrounding CT. Before the tooth erupts, its enamel surface is covered by the reduced dental epithelium, which consists of an inner layer of reduced ameloblasts and an outer layer of polygonal epithelial cells. As tooth erupts and breaks through the oral epithelium there are 2 theories about the origin of the DGJ:

1. Basal epithelial cells, from the pool of epithelial cells over the erupting tooth, migrate apically over the reduced dental epithelium.

2. Reduced dental epithelium becomes JE as a result of epithelial transformation.                                                             

 This JE is sig different from the gingival epithelium. It is non-keratinized, has a decreased nuclear-cytoplasmic ratio, a higher amount of rough endoplasmic reticulum, and has large extracellular spaces between the cells (18% of the epithelium total volume). JE has a higher rate of cell turnover compared to AG epithelium (Skougaard 1962). The CT supporting JE is different from the CT supporting gingival epithelium. The main difference is in the amount of collagen fibers, which are few in the CT supporting JE and presence of vesiculated fibroblasts. Many investigators have reported that inflammation is always present in CT supporting JE. At the time of eruption, the CT is removed and an acute inflammatory reaction occurs in this CT, therefore, it’s possible that the resultant JE exhibits most of the characteristics of epithelium supported by a disturbed CT. The wide intercellular spaces allow continued ingress of antigen, maintaining a low-grade inflammatory lesion. Attempting to keratinize the sulcular epithelium by repeated stimulation of the epithelial cells has proven to be ineffective. If the ideas proposed in this essay are correct, attention should be paid to modifying the CT, not the epithelium if a keratinized sulcular epithelium is desired.

BL: The CT of the JE dictates the epithelial changes.

Pollanen 2003                    Article

Purpose: To review the factors associated with periodontal tissue protection and destruction with special reference to the junctional epithelial cells.

Discussion: Junctional epithelium: Several features that contribute to preventing pathogenic bacterial flora form colonizing the sub-g tooth surface. It is firmly attached to the tooth but allows access of GCF, inflammatory cells and components of the host defense to the gingival margin. It has rapid turnover rate.

Epithelial attachment apparatus: Hemidesmosomes at the plasma membrane of the cells directly attached to the tooth (DAT cells) and the internal basal lamina on the tooth surface. Internal basement lamina proteins include laminin and Type VIII collagen. Hemidesmosomes may act as specific sites of signal transduction and participate in regulation of gene expression, cell proliferation and cell differentiation. Turnover of the JE cells: JE has two layers. The basal facing the CT and the suprabasal extending to the tooth surface. The turnover rate in nonhuman primates is about 5 days and approximately twice the rate of the oral gingival epithelium. Data show that DAT cells have a more important role in tissue dynamics and reparative capacity of the JE than previously reported. Their phenotype may be affected by the internal basal lamina matrix on the tooth surface.  The internal basement lamina appears to be relatively morphologically resistant to external challenges. It”s molecular structures may still be altered leading to changes in DAT cells function.  JE in the antimicrobial defense: Although JE cells layers provide a barrier against bacteria, many bacterial substances pass easily though the external basal lamina into the CT. The area covered by the dividing cells in JE, is at least 50 times larger than the area through which the epithelial cells desquamate into the gingival sulcus which create a strong funneling effect. JE cells have been found to contain enzyme-rich lysosomes. The role of these enzymes is not completely studied yet. PMNs comprise probably the most important defense mechanism at the gingival margin. The cell surface carbohydrates expressed by the JE cells are thought to respond to extracellular changes and allow cells to communicate with their environment. JE cells may also secrete antibodies supplementary to system-derived antibodies and antibodies produced locally. Role of GCF: GCF is an exudate and contains components of serum, inflammatory cells, CT epithelium and microbial flora. In the healthy sulcus the amount of GCF is very small, but its constituents participate in the normal maintenance function of the JE. During inflammation, GCF increases and its composition starts to resemble that of an inflammatory exudate.  It contributes to host defense by flushing bacterial colonies and their metabolites away from the sulcus. The main route for diffusion is through the external basement membrane and then through the JE in the sulcus. Bacteria and host-derived products found in GCF have been associated with the initiation and progression of periodontal disease. Role of PMNs: When they reach bacteria, they release contents of their granules and may adhere to individual bacteria to phagocytose them. They do not have the ability to remove dental plaque but rather form a protective wall against it. They can also cause tissue damage as a result of the variety of enzymes, oxygen metabolites and other components that are released from their granules. They have two main types of granules: the azurophilic (primary and the specific (secondary) containing different enzymes. Activated PMNs also generate H2O2 and highly reactive oxygen radicals with the potential to destroy bacteria and gingival cells. PMNs are more effective in aerobic conditions close to the gingival margin. Lactoferrin is an important antimicrobial protein present in the secondary granules of PMNs.  Role of host proteinases and inflammatory mediators:  Different cell types of periodontal tissues produce MMPs, plasminogen activator, cathepsins and elastase in response to bacteria and inflammatory mediators. At the same time, they also contribute to tissue destruction and to apical and lateral proliferation of JE in the CT. Regulation of proteinase activities is a complex process involving activation of latent precursor molecules as well as inhibition of the active enzymes. Cytokines IL-1, IL-6,TNF-and PG-E2 have been strongly associated with periodontal disease. Role of bacterial products: Bacterial substances have a multitude effect on several cell types, ranging from activation of cell functions to cell death. Lipoteichoic acids found mainly in Gram+ bacteria are thought to mediate bacterial adhesion to human cells and teeth. LPS and porin proteins of the walls of Gram- bacteria also cause several host responses. They stimulate leukocyte function, increase cytokine and inflammatory mediator production and activate the complement system. They also stimulate bone resorption, increase epithelial permeability and penetrate healthy gingival sulcular epithelium. Growth and mitotic activity of epithelial cells can be reduced because of lipoteicohoic acids, interfering with the renewal of JE leading to degeneration and detachment.

Hujoel 2001                    Article

BGRecent studies implicating p-itis as cause of syst dzs have reported that the surface area of perio pckts exposed to bact biofilm ranges from 50-200 cm². Since the root surf area of human dentition excluding 3^rd M is 75 cm², this estimate appear to be too large. The dentogingival surface area (DGES) comprise the JE & SE in health, & any pocket epi in dz.

P:To relate linear perio probing measurements to the DGES

M&M:Formulas to estimate the DGES from clinical and radiographic measurements were applied to a representative sample of a US adult pop 1985-86, a sample of 1021 indiv at their initial visit to a periodontist, and the participants of the VA Dental Longitudinal Study.

R:Individuals w/out periodontitis had a typical DGES of 5 cm². In p-itis, the mean DGES in the 3 samples ranged from 8 cm²(from1-29 cm²) to 20 cm² (from 2-44cm²).

D/BL:Depending on the pop studied, p-itis leads to a mean incr of 3-15 cm² in DGES. The mean DGES among indiv w/p-itis ranges from 8-20 cm², considerable smaller than the range of 50-200 cm²currently assumed. It needs to be shown whether absolute size of DGES or its incr in p-itis is of sufficient magnitude to represent a clin imp risk factor for syst health (if it’s causally related).

Messer 2006                 No Article

P: To measure the rate and strength of attachment of human epithelial cells and periodontal ligaments fibroblasts to tooth dentin.

M&M: Rate and strength of attachment of epithelial cells and PDL fibroblasts were measured. They were cultured individually and co-cultured to dentin surfaces to determine which cell type has a faster attachment rate and greater adhesive strength to human dentin. Longitudinal dentin slices were seeded with either epithelial cells or PDL fibroblasts for 2-24 hrs. The specimens were placed into a parallel plate flow chamber. Effluent fluid was collected and detached cells were counted. Co-cultures of PDL fibroblasts and epithelial cells at 3 seeding ratios (10:1, 1:1, 1:10) were also tested.

R: PDL fibroblasts showed a stronger attachment to dentin at 24 hrs, while epithelial cells attached to dentin equally well at 2 and 24 hrs. Epithelial cells were strongly attached after 2 hrs when compared to PDL fibroblasts, but less strongly attached after 24 hrs. When epithelial cells and PDL fibroblasts were seeded together, at ratios

1) 1:1, PDL fibroblasts appeared to be more strongly attached at 2 but not 24 hrs

2) 10 (PDL) :1 (Epi), PDL fibroblasts appeared to be more strongly attached at 2 and 24 hrs

3) 1 (PDL) : 10 (Epi), Epithelial cells appeared to be more strongly attached at 2 hrs, but PDL fibroblasts showed a trend of stronger attachment at 24 hrs

Cocultures (PDL fibroblasts : Epithelial cells)

	P > E	P > E	E > P
24 hrs	P > E		P > E

 Summary of strength of attachment

Cultures

2 hrs PDLF
24 hrs PDLF		24 hrs

BL: Epithelial cells attach more quickly to dentin surfaces than PDL fibroblasts, but do not demonstrate increased attachment strength over time. Epithelial cells and PDL fibroblasts do not act independently, because epithelial cells enhanced the attachment rate of PDL fibroblasts.

Can you quantify inflamed periodontal tissues?

Nesse 2008                     Article

To develop a classification of periodontitis (Periodontal Inflamed Surface Area –PISA-) and to evaluate its applicability in quantifying the amount of inflamed periodontal tissue. 

 A literature search was performed and revealed there were no classification systems that quantified the area of inflamed periodontal tissue. An Excel spreadsheet was developed that calculates PISA based on CAL, recession, and BOP. Population based mean values for both root surface area and root length are used in the Excel formula. Results reflect the surface area of bleeding pocket epithelium in square millimeters. Calculating PISA using Hujoel et al’s spreadsheet for ALSA;

• 1-ALSA (attachment loss surface area) was calculated.

• 2-RSA (recession SA) was calculated.

• 3-ALSA-RSA=PESA (periodontal SA)

• 4-BOP tested on six sites per tooth

• 5-PESA multiplied by the BOP sites per each tooth = PISA (periodontal inflamed SA) e.g. if 3 surfaces are BOP+ then PISA for each tooth= PESA multiplied by 3/6)

• 6- PISA for the whole mouth is the sum of PISAs of individual teeth.

• Then the system was applied to 3 pts; with healthy periodontium, local periodontitis and sever generalized periodontitis.

  PISA measures the surface area of bleeding pocket epithelium in square millimeters. But it is not a precise tool because:

-PISA is subject to operator errors associated with measuring CAL and BOP.

– Actual patients are likely to differ from the population means used in the formula.

-Only quantifies in two dimensions and is less accurate when gingival overgrowth or pseudo pockets are present.

-Does not take into account type of inflammation or flora and therefore cannot be used to predict the diseases that inflammation might cause

PISA is a classification system with some short comings. However, it is more accurate than any other classification systems currently used for quantifying inflammation. May serve as a method of classification for associating periodontitis as a risk factor for other diseases.

Describe the dimensions of the DGJ and the significance of the “biologic width” in dentistry.

Garguilo 1961                     No Article

To evaluate the measurements of the dentogingival junction during four phases of passive eruption.

: 30 jaws of human autopsies. A total of 287 teeth were measured. Measured surfaces: M, D, vestibular and oral. 6 measurements were made for each: 1) Depth of sulcus, 2) Length of attached epi, 3) Most apical point of epi attachment from the CEJ,

4) Distance from base of sulcus to CEJ, 5) Distance of CEJ from alveolar bone,

6) Distance from most apical point of epithelial attachment to alveolar bone (CT)

All surfaces were placed in one average value for the given measurement.  They evaluated these measurements at four different phases of passive eruption.  The following values were found:

Phase	Avg. Age	Length of dentogingival junction (mm)
Phase I	24.5yrs	3.23mm
Phase II	31.4yrs	3.06mm
Phase III	32.3yrs	2.41mm
Phase IV	39.7yrs	2.53mm

The dentogingival junction is a functional unit with 2 components: 1) CT fibrous attachment and 2) epithelial attachment. As we age the total measurement of the DGJ decreases.  During passive eruption the epithelial attachment diminishes. In correlating the epithelial attachment with age it is seen that there was less epithelial attachment with an increase in age; however, the CT component appeared to stay constant through all stages of passive eruption.  The following table is the total average magnitude of 3 of the measurements taken:

Sulcus Depth	0.69 mm
Attached epithelium	0.97 mm
Connective Tissue	1.07 mm

Vacek 1994                    No Article

PURPOSE: To provide information on the dimensions of the dentingingival junction (biologic width) and related structures.

METHODS:  10 preserved jaws from cadavers (age 54-78) were used to prepare 7 block segments of 2-3 teeth each.  Block segments were sectioned first in a M-D direction along the long axis and contacts of the teeth. The remaining F-L portions were then sectioned B-L along the long axis of teeth.  Sections were stained with Masson’s trichrome and eosin stain and histomorphologically measured with Zeiss interactive digital analysis system.  Measurements recorded were sulcus depth (SUL), epithelial attachment (EA), connective tissue attachment (CTA), and loss of attachment (LOA).

RESULTS: There were no mean differences between measurements for the tooth surfaces (BLMD) for SUL, LOA, EA, or CTA. LOA did not affect the CTA, or biologic width.  Molars showed a significantly greater biologic width than anterior teeth.  Tooth surfaces with subG restorations had sig longer EA, but no other sig differences.  Mean measurements: SD- 1.34mm, EA-1.14mm, CTA-0.77 mm, LOA- 2.92mm.

DISCUSSION:  The CTA varied in width, but with a more narrow range and variance than EA, SUL, or LOA. SubG restored teeth showed a longer EA. No correlation could be found between LOA and width of biologic width (CTA).  LOA should not be used as a guide to determine requirements for reestablishment of EA and CTA..  Biologic width was slightly wider in molars than anterior teeth, therefore these teeth may require a greater length of biologic width when restoring these teeth.

CONCLUSION/BL: No correlation was found between LOA and length of biologic width.  Of the dimensions measured, CTA (biologic width) had the least amount of variance.

Perez 2008                    Article

P: To provide and compare the clinical Supra Osseous Gingivae (SOG) dimensions around molar, premolar, canine, incisor teeth in the maxillary and mandibular arches as measured by trans-sulcular probing (TSP) in patients without a history of periodontitis.

M&M: 23 patients (mean age of 35 years), 8 males and 15 females were included in the study.  SOG dimensions were evaluated around incisor and canine (zone 1), premolars (zone 2), molars (zone 3). Inclusion criteria: free of gingival hyperplasia and overt signs of inflammation, adequate plaque control, absence of altered passive eruption, no attachment loss or history of periodontitis.  Clinical parameters (PI, GI, BOP, PD) were measured to establish gingival health. TSP was done using standardized periodontal probes, with marking-width differences <0.2mm. Calibration exercises to achieve >90% intraexaminer reproducibility of measurements were conducted before the study started. ANOVA was used for statistical analysis.

R: All clinical parameters were indicative of the absence of gingival inflammation. The overall mean of SOG was 3.75mm (vs. Gargiulo 1961, 2.73mm).

For the maxillary teeth, a statistically significant difference was found among SOG measurements by tooth type for mean overall and lingual dimensions. For mean facial SOG measures, the difference between SOG measures by tooth type approached statistical significance. When comparing site level measures of SOG  by tooth type, statistical significance was found for mid-facial, disto-facial, mesio-lingual, mid- lingual, and disto-lingual measures.

 For mandibular teeth, statistical significance was found among mean overall and mean lingual SOG measurements but not among mean SOG facial measurements when comparing by tooth type. When comparing site- level measures of SOG by tooth type, statistical significance was found for the disto-facial and the mesial-, mid-, and distal-lingual SOG measurements but not for the mesial- and mid-facial measurements.

For both arches there was a SSD for mean overall and mean lingual dimensions. SOG increased from anterior to posterior.

BL: The average 3mm figure used to estimate the amount of sound structure needed after crown lengthening to accommodate the restorative and SOG dimension should be considered to be pre-empted by the SOG dimensions of the particular tooth.

Novak 2008                    Article

To determine whether previously observed norms in the biological width (BW) apply in a previously untreated population with severe generalized chronic periodontitis.  The importance of understanding the variations in BW that may occur with periodontal pathology may impact our approach to surgical intervention in conserving the existing periodontal attachment is a clinical priority.

28 patients (29-45 years old) with severe generalized chronic periodontitis. Clinical and radiographic measurements (not standardized) were taken by calibrated examiners.  BW was determined from most coronal level of clinical attachment to the crest of the alveolar bone for interproximal surfaces only and compared to the histological BW previously reported.

  The clinical BW in subjects with severe generalized periodontitis was significantly greater than previously reported.  The mean clinical BW was 3.95mm vs. mean histological width of 2.04mm.  The greatest clinical BW was seen with pockets < 2mm with 5.02 mm BW. As PD increased, the associated mean BW tended to de-crease

<2mm 2-4mm 5-7mmm >7mm All sites Clinical Mean BW 5.02 – 2.48 4.16 – 1.32 3.33 – 1.17 3.21 – 1.29 3.95 – 1.04

  Mean clinical BW in subjects with severe chronic periodontitis seemed to be significantly greater than the histologic BW previously reported for subjects not demonstrating significant periodontal pathology.

Describe the pocket epithelium

Muller-Glauser & Schroeder 1982                    No Article

P: To examine the pocket epithelium: a Light microscope and Electron microscope study

M&M: 8 Beagle dogs with experimental periodontitis for periods of 4 to 21 days or up to 5 months were studied. Block biopsies of the 4^th premolar buccal gingiva were excised and processed for LM and EM exam.

R: Pocket epithelium was displaced from the teeth by deep subgingival plaque, which was close to the apical termination of the pocket. The pocket epithelium had very irregular shape with rete pegs (RP) penetrating more than half of the infiltrated CT. Occasionally, rete pegs branched to form irregular network, (some portions of 2 cells only). Subepithilial CT was collagen poor, highly vascularized and infiltrated by leukocytes (mainly plasma cells).

Variable thickness of epithelium: at coronal and mid-pocket regions—thin and occasionally ulcerated. It was thicker apically. 

Epithelial cells: pocket epithelium was non-keratinized. Atypical stratification.

Basal cells: cuboidal to cylindrical with characteristic flat processes along the basal lamina. Hemidesmosomes at the basal side, but mainly microvilli and gap junction laterally.

Suprabasal cells: Polygonal or elongated, more basophilic.

Superficial cells: Variably basophilic & electron dense. Numerous filament bundles. Low-density cells were seen only at the very surface, (often lacking cytoplasm. membrane). Microorganisms occasionally adhered to those cells.

Intercellular spaces: Mostly dilated spaces with infiltrated leukocytes, PMNs, lysosomes and cell fragments.  Microorganisms in the superficial intercellular spaces with openings to the pocket space.

Basal complex: Consisted of Lamina Lucida, Lamina densa, and anchoring fibrils. Cytoplasmic projections from epithelial cells in areas of basal lamina discontinuity.

Infiltrations: Mainly by Lymphocytes T and B, and plasma cells (basal & suprabasa