Advances in Dental Research

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The Dentin Disc Surface: A Plausible Model for Dentin Physiology and Dentin Sensitivity Evaluation D.G. Gillam, N.J. Mordan and H.N. Newman ADR 1997 11: 487 DOI: 10.1177/08959374970110041701 The online version of this article can be found at: http://adr.sagepub.com/content/11/4/487

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THE DENTIN DISC SURFACE: A PLAUSIBLE MODEL FOR DENTIN PHYSIOLOGY AND DENTIN SENSITIVITY EVALUATION D.G. GlLLAM1 N J . MORDAN2 H.N. NEWMAN1

'Department of Periodontology 2 Electron Microscopy Unit Eastman Dental Institute for Oral Health Care Sciences University of London 256 Gray's Inn Road London WC1X 8LD, United Kingdom Adv Dent Res 11(4):487-501, November, 1997

Abstract—Dentin sensitivity (DS) is a painful clinical condition which may affect 8-35% of the population. Various treatment modalities have claimed success in relieving DS, although at present there does not appear to be a universally accepted desensitizing agent. Current opinion based on Brannstrom's Hydrodynamic Theory would suggest that following exposure of the dentin surface (through attrition, abrasion, or erosion), the presence of open dentinal tubules, patent to the pulp, may be a prerequisite for DS. The concept of tubule occlusion as a method of dentin desensitization, therefore, is a logical conclusion from the hydrodynamic theory. The fact that many of the agents used clinically to desensitize dentin are also effective in reducing dentin permeability tends to support the hydrodynamic theory. This paper reviews the in vitro evaluation of desensitizing agents, the techniques used to characterize their effects on the prepared dentin surface, and the ability of these agents to reduce permeability through tubule occlusion, and presents recent findings from ongoing research based on the Pashley Dentin Disc model. It can be concluded that the use of this model to determine surface characteristics, and reductions in dentin permeability through tubule narrowing or occlusion, provides a useful screening method for evaluating potential desensitizing agents. Interpreting changes observed in vitro is difficult, and extrapolation to the clinical situation must be tempered with caution. Key words: Dentin, surface characteristics, tubule occlusion, desensitizing agents, electron microscopy. Presented at "Advances in the Characterization of Surface and Subsurface Areas of Dental Hard Tissues'', a workshop sponsored by the Council of Europe and the Deutsche Forschungsgemeinschaft (German Research Agency), November 13-17, 1996, at the University of Mainz, Germany

D

entin sensitivity (DS), or hypersensitivity, may be defined as a transient pain arising from exposed dentin, typically in response to chemical, thermal, tactile, or osmotic stimuli, which cannot be explained by any other dental defect or pathology (Addy et aL, 1985). The reported prevalence of the condition is 835%, depending on the population studied (Gillam, 1992; Gillam et aL, 1994), and peak prevalence is in the third and fourth decades (Graf and Galasse, 1977; Flynn et aL, 1985; Addy, 1990, 1992; Fischer et aL, 1992). Epidemiological surveys indicate that the majority of sensitive surfaces are at the buccal-cervical margins (Graf and Galasse, 1977; Flynn et aL, 1985; Fischer et aL, 1992), although other surfaces may also be affected (Robb and Smith, 1992; Chabanski et aL, 1995). Any site of exposed dentin may exhibit sensitivity, but not all exposed dentin is sensitive. The condition may affect any tooth (Graf and Galasse, 1977; Flynn et aL, 1985; Fischer et aL, 1992; Chabanski et aL, 1995). Although DS is common, its etiology is poorly understood (Addy and West, 1994), though the exposure of dentin due to gingival recession is of primary importance. Current evidence (Pashley, 1990a,b; Vongsavan and Matthews, 1994) favors the 'Hydrodynamic Theory', originally postulated in the nineteenth century and later developed by Brannstrom (1963). This theory suggests that dentin tubules act as capillary tubes and that the fluid within them obeys the laws of fluid movement. The rapid movement of fluid in dentin tubules, in response to certain stimuli, may cause distortion of intradental nerves and generate a pain response. Scanning Electron Microscopy (SEM) studies have demonstrated patency of the tubules on the outer dentin surface in sensitive teeth (Brannstrom, 1965; Ishikawa, 1969; Absi et aL, 1987; Yoshiyama et aL, 1989, 1990). Current treatments tend to concentrate on two approaches, to occlude the tubules and to block neural transmission. Although the mechanisms of pain transmission across dentin are not fully understood, both dentin permeability and sensitivity are reduced when the dentin tubules are occluded (Pashley et aL, 1978a,b). Most treatments, which range from toothpastes (over-the-counter) to dentin bonding agents (in-office), are therefore based on the hydrodynamic mechanism of stimulus transmission and attempt to inhibit sensitivity either by sealing the tubules, by altering their contents, or by creating insoluble calcium complexes, thus forming mechanical or chemical plugs. Most of the experimental evidence for the mechanisms of stimulus transfer across dentin and the presumed mode of action of the various desensitizing agents have been derived from animal or laboratory studies. This usually involved cavity preparation in coronal and radicular dentin or coronal dentin sections with a view to studying effects on micro-


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leakage, conditioning, adhesion or bonding, tubule occlusion, hydraulic conductance, and fluid flow. Specimens were subsequently qualitatively or quantitatively evaluated by a variety of techniques, including: light microscopy; confocal, scanning, and transmission electron microscopy; atomic force microscopy; environmental microscopy; scanning electrochemical microscopy; and image analysis, as well as identification of the resultant chemical components on the altered dentin surface by x-ray microanalysis and Fourier transform infrared photoacoustic spectroscopic analysis. All these techniques have their benefits and limitations, and so results have to be treated with caution, especially with extrapolation to the clinical situation. This paper reviews the in vitro evaluation of desensitizing agents, the techniques used to characterize their effects on the prepared dentin surface, and their ability to reduce permeability through tubule occlusion, and presents recent relevant findings from ongoing research with the Pashley Dentin Disc model. DENTIN PERMEABILITY AND SENSITIVITY Formation and structure Dentin is a mineralized connective tissue, like all mineralized tissues laid down as an organic matrix into which minerals are subsequently deposited. Formation begins in the late 'Bell Stage' of tooth germ development, when the cells of the inner enamel epithelium induce the cells of the dental papilla to differentiate into odontoblasts. The cells of the papilla are of mesodermal origin, but it has been claimed that ectomesenchymal cells, that develop from the neural crest, may be present and that these may be precursors of the odontoblasts (Linde, 1984). The odontoblasts are tall columnar cells with an oval nucleus located at the proximal end of the cell. Initially, the odontoblast shows several cytoplasmic extensions at the surface adjacent to the amelodentinal junction, but as dentinogenesis progresses, these foldings develop into a single process ultimately responsible for the tubular nature of dentin. The odontoblasts secrete an organic matrix, predentin, rich in collagen, glycoprotein, and proteoglycan complexes. Hydroxyapatite is then deposited, and the foci of mineralization grow into spherical structures called calcospherites, which eventually fuse with adjacent spheres to give a solid mass. There is no overall consensus with regard to the length of the odontoblast process. Thomas and Payne (1983) indicated that it extended between a third and half the length of the tubule, while ten Cate et al. (1985) concluded that the odontoblast process extended to the amelo-dentinal junction and that conflicting evidence was due to different interpretations of scanning and transmission electron microscopy data or possibly due to the way that the specimens were prepared (Holland, 1994). The number of tubules per unit area varies depending on location, because of the decreasing area of the dentin surface in a pulpal direction. In humans there are some 65,000 tubules per mm2 near the pulp, but only 15,000 tubules per mm2 near the periphery of the tooth (Linde, 1984). The tubules follow a sinuous course from the amelo-dentinal junction and from the cementodentinal junction and are conical, being wider at the pulpal

ADV DENT RES NOVEMBER 1997

end than at the periphery. A recent study (Mjor and Nordahl, 1996) indicated that the mean number of tubules in the middle part of the root was significantly lower than in the middle part of the crown. This study also showed that the tubules were interconnected by an intricate and profuse system of canaliculi that branch off from the main tubules at a variety of angles. Changes occur in the dentin as a result of age, trauma, and post-extraction. Secondary dentin is deposited throughout life, and the formation of peritubular dentin can ultimately result in partial or complete obturation of the tubules, producing dead tracts and areas of sclerosed dentin. In response to trauma, an irregular layer of secondary dentin is deposited. This layer has fewer tubules and, since they are not continuous with those in primary dentin, provides an effective barrier to diffusion (Holland, 1994). Following extraction, the dentin becomes more brittle than that of vital teeth (Fitzgerald, 1976). Permeability of dentin The presence of the tubules in dentin makes the tissue permeable, especially when the outer protective layer of enamel or cementum is removed. Permeability may be demonstrated by dye penetration experiments and measured by hydraulic conductance (Pashley, 1990b). Superficial dentin has fewer tubules per mm2 than deeper dentin, while thick dentin offers more resistance to diffusion than thin dentin (Pashley, 1985a,b). Patients with dentin sensitivity have patent dentin tubules on the outer surface of the dentin (Brannstrom, 1965; Ishikawa, 1969; Absi et al., 1987; Yoshiyama et al., 1989, 1990). The buccal cervical area of the root is commonly quoted as being most sensitive (Graf and Galasse, 1977; Flynn et al, 1985; Addy, 1990, 1992; Fischer et ah, 1992; Chabanski et al., 1995). However, although the tubules may be patent externally, there is no evidence that they are patent through to the pulp chamber. Mechanisms of stimulus transmission across dentin The exact mechanisms responsible for dentin sensitivity are still unclear (Addy and West, 1994; Gillam, 1995), but Pashley and Parsons (1987) suggested that dentin sensitivity transmission could be explained according to three main hypotheses: (1) Nerve endings or nociceptors that respond directly when dentin is stimulated are located throughout the dentin. (2) Odontoblasts, being chemically or electrically related to nerves, function as receptors, generating nerve impulses. (3) Stimuli applied to dentin, producing a displacement of dentin tubule contents, could excite mechanosensitive nerve endings near the pulpal ends of the tubules (Hydrodynamic Mechanism). Current evidence (Pashley, 1990a; Vongsavan and Matthews, 1994) supports the Hydrodynamic Theory, which is generally credited to Brannstrom (1963). However, a review of the literature by Rosenthal (1990) indicated that in the midnineteenth century, Blandy (1850) reported the work of a Dr. John Neill, who had suggested that dentin consisted of hollow fluid-filled tubules and that compression of the fluid in these tubules affected the pulp. Later, Gysi (1900) proposed that


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fluid movement in either direction within dentin tubules produced pain and that the movement of fluid away from the pulp was induced by salt, sugar, or alcohol. He also suggested that coagulating the protein within the tubules would relieve dentin sensitivity. Kramer (1955) proposed the Hydrodynamic Theory, although neither he nor Neill was convinced that fluid movement was an acceptable explanation for pain transmission in dentin. It was the later work by Brannstrom which finally led to the acceptance of the Hydrodynamic Theory. The Hydrodynamic Theory proposed that dentin could be thought of as composed of hollow tubules containing fluid or semifluid material. This fluid would obey the same physical laws as fluid in fine capillary tubes (Poiseuille's Law). If the displacement of the fluid was rapid enough, then a shift of fluid in either direction could deform the odontoblast layer or stimulate nerve fibers and give rise to pain. Recent investigations in the cat (Vongsavan and Matthews, 1991, 1992) appear to provide further support for a hydrodynamic mechanism. However, the evidence is not conclusive, and Narhi (1985) suggested that until fluid flow in the tubules could be measured in vivo, the hydrodynamic mechanism remained unsubstantiated. A recent review (Gillam, 1995) suggested that despite general acceptance of the Hydrodynamic Theory, other theories could not be excluded and that further investigation was required. Physiological pulpal defense mechanisms The pulp has a few natural defenses to protect it from irritating stimuli: (a) Irregular, atubular dentin may be produced at the pulpal wall. This is by no means an impermeable barrier, but it does result in a relatively efficient seal of the pulpal endings of the tubules, which may block the transmission of pain-producing stimuli. It is unfortunate that in some patients, for unknown reasons, the formation of secondary dentin can be delayed or absent (Brannstrom et al., 1979). (b) In areas with exposed non-sensitive dentin, irregular dentin was not only produced, but many tubules were also sealed with mineral deposits (Brannstrom et al., 1979). Brannstrom and Garberoglio (1980) demonstrated that naturally attrited teeth had tubule lumina which were either partially or completely obliterated. In the dentinexposed teeth, the obliteration ranged from no to complete reduction of the tubule lumen. They concluded: "The tubules of sclerotic zones beneath regions of attrition were occluded by peritubular dentin-like material. The results also supported the view that the oral environment contributed to the development of dentinal sclerosis and obliteration of the tubule lumen under attrited dentin." Another mechanism which may occlude dentin tubules is the adsorption of salivary and/or plasma proteins to the dentinal tubules (Pashley et al., 1982). These macromolecules may gain access to tubules from the oral end via saliva or plaque and to the pulpal ends of the tubules via dentinal fluid, respectively. It is possible that protein adsorption to the walls of the tubules may reduce their functional radii to the point where less fluid flow would occur in response to stimuli, thereby reducing or eliminating

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dentin sensitivity (Michelich et al., 1978). This would suggest that small amounts of fibrinogen leaking across pulpal vessels and slowly permeating dentinal tubules from the pulpal side may, over days to weeks, reduce dentin permeability (Michelich et al., 1978; Pashley et al, 1982). Although the total protein concentration of saliva is much less than that of plasma, the total amount passing over dentin surfaces per day may be greater. Many salivary proteins are highly charged glycoproteins which tend to exist in a highly extended form, and dentinal tubules can adsorb glycoproteins (Pashley et al., 1982). Removal of enamel or cementum permits dentinal fluid and substances dissolved in it to seep slowly toward the dentin surface. Over time, this would allow continual deposition of plasma proteins contained in dentinal fluid on the walls of the tubules. Since saliva is saturated with respect to most forms of insoluble calcium phosphate at normal salivary flow rate and pH, numerous physicochemical mechanisms tend to occlude tubules with a wide variety of crystal types. This may lower the hydraulic conductance of the exposed dentin (Pashley, 1986). The formation of peritubular dentin by calcium crystals is the tooth's physiological way of minimizing dentinal sensitivity. It will decrease the fluid flow within the tubules and, according to the hydrodynamic theory, the pain from exogenously applied stimuli (Berman, 1985). (c) Karlsson and Penney (1975) postulated that an element of natural desensitization occurs in the in vivo situation, since not all patients develop sensitivity after periodontal surgery. These investigators stimulated exposed root surfaces in dogs and recorded pain response by a positive jaw-jerk reflex. Although this was a crude method, they found that the sensitivity decreased with time. They suggested that the acquired pellicle formed an effective thermal or chemical barrier which could resist dentinal conduction of common oral stimuli. Blockage of the tubules at the dentin surface may also occur by other means, e.g., toothpaste components, calculus, and oral debris (Hiatt and Johansen, 1972). Kerns et al. (1991) suggested that natural desensitization can occur from the deposition of salivary minerals around tubules, which would result in tubule diameter reduction and subsequent reduction in fluid flow. Sensitivity may occur periodically, or be of short duration because of a positive response by the tooth's defense mechanisms. When both ends of the tubules are sealed, the dentin has the potential to become nonsensitive again. The oral environment may, however, exacerbate an already established dentin exposure. The physiopathological interplay of causal factors is complicated and may be difficult to comprehend, but the role of environmental factors is important (Brannstrom, 1992). Brannstrom (1992) proposed some factors which may contribute not only to the initiation, but also to the persistence of sensitivity. There may be a poor host response due to impairment of the pulp and poor blood supply. Formation of a barrier to block the tubule may be prevented and the tubules remain open. Saliva may be incapable of producing a calcified pellicle which would plug the orally exposed tubules. Third, protective calcium salts and underlying hard


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tissue may be continuously removed from the surface by erosion due to acidic dietary or medicinal products. This would also prevent the development of a calcified pellicle. Fourth, frequent and overzealous brushing with abrasive dentifrices may mechanically remove periodontal tissue, exposing many tubules. Finally, eccentric occlusal loading due to bruxism (abfraction), incorrect design of dental restorations, and movement of teeth may impair the pulpal vascular and nervous systems. Any of these factors in combination may prevent the development of a smear layer, and the pulp would be unable to develop a complete dentin barrier, and sensitivity would then persist.

TECHNIQUES REPORTED IN THE LITERATURE USED TO EVALUATE DENTIN MORPHOLOGY AND THE SURFACE/ SUBSURFACE CHARACTERISTICS INDUCED BY DESENSITIZING AGENTS Permeability studies In 1974, Pashley and co-workers published the first experimental work utilizing a laboratory method to measure dentin permeability by hydraulic conductance, which can be determined by a number of variables, including the pressure moving fluid across the dentin, the length of the dentinal tubules, the viscosity of the fluid, and the radius of the tubule (Pashley, 1990a,b). In this work, a split-chamber device was described wherein thin slices (approximately 1 mm) of coronal dentin from extracted human third molars were placed between fixed surface area plexiglass reservoirs, one end of which could be connected to a source of hydrostatic pressure or treatment solution and the other end to a means of measuring flow rate or to collect diffusal fluid. Movement through a micropipette was found to be an accurate flow meter (Outwaite et ah, 1974; Sena, 1990). Using this device, investigators determined the fundamental properties of dentin permeability (Pashley et ah, 1978a; Reeder et ah, 1978). Because this technique provided the ability for variables such as thickness, surface area, and applied pressure to be standardized, its use significantly advanced the understanding of dentin permeability (Sena, 1990). Pashley et ah (1978a) determined the contribution of regional area to total dentinal fluid flow resistance. The most important finding was that 86% of the resistance to dentinal fluid flow was attributable to the surface characteristics of dentin, strongly suggesting that alteration of permeability by surface agents could be a useful treatment (Sena, 1990). More recently, Ciucchi et ah (1995) reported that it was possible to measure dentin fluid flow, the hydraulic conductance of dentin, and the estimated tissue pressure in vivo. Levinkind et al. (1992), however, suggested that, instead of measuring permeability from hydraulic conductance, which is essentially bulk movement of fluid, it is possible to measure permeability from the ion movement within an electrolyte conductance system (electrochemical impedance measurements). Other investigators have also attempted to measure fluid flow by confocal optical microscopy (Watson and Griffiths, 1993) and scanning electrochemical microscopy (SECM) (Gilbert et al., 1993;


Macpherson et ah, 1995). Scanning electron microscopy, x-ray microanalysis, and image analysis Most attention for the treatment of DS has been directed at blocking the hydrodynamic mechanism by occluding tubules (Addy et ah, 1985; Pashley, 1985a,b). However, how the numerous compounds actually achieve their effects has received limited attention (Greenhill and Pashley, 1981; Pashley et ah, 1984). SEM can be used to measure uptake onto the dentin surface (Addy and Mostafa, 1988, 1989; Absi et ah, 1989a,b, 1995; Cuenin et ah, 1991; Knight et ah, 1993; Dijkman et ah, 1994) or the characteristics of the smear layer (Eick et ah, 1970; Eick, 1992; Dautel-Morazin et ah, 1994) and hybrid layers following etching (Eick et ah, 1993; Titley et ah, 1995), and x-ray microanalysis can be used to characterize the nature of the deposits on the surface (Ling et ah, 1997). Other investigators have used SEM and/or TM to evaluate both dentin morphology in sensitive and nonsensitive dentin and/or the surface characteristics following application of desensitizing agents (Yoshiyama et ah, 1989, 1990, 1992, 1994; Oyama and Matsumoto, 1991; Ishikawa et ah, 1994), as well as the ultrastructure of the resin-dentin interdiffusion zone (Van Meerbeek et ah, 1993), or x-ray tomographic microscopy and atomic force microscopy to evaluate mineral distribution and dimensional changes in human dentin during mineralization (Kinney et ah, 1995). Ide et al. (1994) used confocal optical microscopy to determine the surface characteristics induced by selected desensitizing toothpastes. Confocal optical microscopy has also been used to assess resin composite systems (del Wilmot et ah, 1994), the influence of handling techniques on the hybrid layer with bonding agents (Griffiths and Watson, 1993), and the sealing ability of selected dentin bonding agents (Griffiths and Watson, 1995). Fourier transform infrared photoacoustic spectroscopy has been used to evaluate dentin morphology in abrasion/erosion cavities (Mixson et ah, 1995). SEM and image analysis have been utilized together to quantify dentin tubule morphology and/or following application of desensitizing agents and restorative materials, or the effect of smear layers on dentin permeability (Levinkind et ah, 1992; Marchetti et ah, 1992; Arends et ah, 1995; Me Andrew and Kourkouta, 1995; Rimondini et ah,

\995\\p et ah, 1997). Animal studies Kim (1986) used a (cat) neurophysiological model to evaluate DS. Briefly, animals were anesthetized and two deep dentin cavities prepared on the buccal surface of an anterior tooth. These served as electric recording cavities. A third deep-cut cavity prepared on the lingual surface was used for the delivery of solutions of therapeutic agents. Results indicated that the K+ ion was the most effective ingredient in sensory nerve activity reduction when compared with many other agents, e.g., sodium, lithium, and aluminum compounds. This finding lends credence to the hypothesis that sensory nerve reduction was achieved by increasing the extracellular K+ concentration. K+ ions presumably depolarize


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the nerve fiber membrane, eliciting an initial increase in action potential. After the initial depolarization, the nerve fibers cannot repolarize due to the maintained high levels of extracellular K+, and thus a sustained depolarized state occurs. During this state, few or no action potentials can be evoked. In physiological terms, this phenomenon is sometimes called axonal accommodation (Kim, 1986). Two criticisms, however, have been raised in relation to Kim's (1986) work: (1) The animal model needs human confirmation. (2) Potassium ions as delivered from a dentifrice could not traverse the length of dentin tubules in sufficient quantity and at a rate adequate to depolarize the pulpal nerve. The actual clinical situation is in contrast to Kim's experimental model in which deep-cut cavities were used and the therapeutic agent traveled through only a thin slice of dentin. In addition, the opposing pulpal pressure producing outward flow of dentinal fluids is another possible modifying factor which may prevent the ingress of substances (Sena, 1990). Investigations of the permeability of (cat) dentin have shown that outward fluid flow through exposed dentin in vivo can be sufficient to reduce substantially the inward diffusion of substances into the tubules (Vongsavan and Matthews, 1991, 1992). Final clarification of the ability of potassium to reach the pulp requires further work, but the work of Kim (1986) has raised the possibility of combination therapies, i.e., neural and physical, and as such has provided a sensitive model to test novel experimental agents (Sena, 1990). For example, the use of potassium oxalate as a desensitizing agent has been claimed to combine the tubule-occluding properties of calcium salt crystals and the inhibitory properties of potassium ions on intradental nerves. This provides an example of how drugs or chemicals can be selected for a therapeutic effect (Pashley, 1986). To date, however, no evidence has been provided to suggest that the efficacy of potassium-containing products in vivo is due to the inhibitory properties of potassium ions. Indeed, as mentioned above, the ingress of the potassium ions may be prevented by the presence of a smear layer on the root surface and/or the outward flow of dentinal fluid. Other components of the potassium-containing system may contribute to its effectiveness in relieving discomfort from DS in addition to tubule occlusion properties. SEM and dye penetration studies Absi et al. (1987) attempted to link dentin permeability and the in vivo evaluation of therapeutic agents. Their technique involved combining SEM and classic dye penetration techniques for a quantitative study of the patency of dentin tubules in sensitive and non-sensitive areas of human teeth. In this study, teeth classified as non-sensitive or sensitive after suitable stimulation were examined by SEM. Sensitive teeth showed highly significantly increased numbers of tubules per unit area (approximately 8x) compared with nonsensitive teeth. Tubule diameters were significantly wider (approximately 2x) in sensitive compared with non-sensitive teeth. Importantly, the link between open tubules and

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transmission to the pulp was provided by subsequent dye penetration experiments in which both the intensity and propensity for penetration were significantly greater in the sensitive specimens. These investigators also indicated that the permeable dentin was not uniform but seemed to be clustered into discrete regions. This was consistent with clinical observation of DS which appeared to be localized to specific areas of the tooth surface (Pashley, 1992). There appears, however, to be no available evidence to suggest that 'hypersensitive' dentin is more permeable than 'nonsensitive' dentin. Narhi et al. (1992) have also reported that, in some human teeth, dentin hypersensitivity is not abolished even when the tubules are blocked, which suggests that other factors apart from tubule occlusion are involved in the prevention of stimuli transmission across dentin. In vivo studies

Replication

techniques.—In

the treatment of DS,

therapeutic agents may either enter patent dentinal tubules to modify neural response at the pulp or, alternatively, partially or totally occlude tubules (Pashley, 1985). Although more attention has been given to compounds which might occlude tubules, there is a lack of knowledge as to how apparently effective agents achieve their action. A novel approach to this problem has recently been reported by Absi et al. (1989a), who developed a replica technique to study 'sensitive' and 'non-sensitive' cervical dentin. In an in vitro pilot study, silicone impressions were taken of extracted human teeth which had been root-planed and acid-etched to expose the tubules. Epoxy resin casts from these impressions were examined by SEM and compared with original tooth surfaces. Excellent correlation was detected between the original and replica SEMs in terms of tubule counts and resolution of surface details such as tubule diameters as low as 1 (xm, illustrating patent tubules. The second part of the study compared original and replica impression SEMs of teeth categorized clinically as 'sensitive' and 'non-sensitive', and demonstrated the ability of the technique to distinguish the types of dentin surface involved. The in vivo results demonstrated that naturally occurring surface details of dentin could be reproduced by the replication technique (Absi et al., 1989a). Other investigators have had less success with this method, since it relies on the investigator's ability to clean plaque from tooth surfaces without creating a smear layer (Pashley, 1992). Duke and Lindemuth (1991) with regard to tubule patency in sensitive dentin, observed that non-sensitive teeth were frequently coated with an amorphous and/or crystalline smear layer absent in sensitive dentin. The investigation by Absi et al. (1989a) was noteworthy for a number of reasons. First, it confirmed the presence of patent tubules and their role in the hydrodynamic mechanism of sensitivity. Second, it may also assist the clinician in the diagnosis of sensitivity. Most importantly, it provides an opportunity for the direct evaluation, in vivo, of the effects of therapeutic agents postulated to function by tubule occlusion (Sena, 1990).

Dentin

biopsy

technique.—Another


approach to

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TABLE 1 ACTIVE INGREDIENTS AND MANUFACTURER INFORMATION OF DIFFERENT TEST AGENTS Test Agent

Manufacturer

Active Ingredients

Butler Protect

John O. Butler Co., USA

Potassium oxalate

Sensodyne Sealant

Block Drug Corp., Jersey City, NJ, USA

Ferric oxalate

Boots Sensitive Teeth Mouthwash

Boots Company Pic Nottingham, England, UK

(a) Cetyl pyridinium chloride (b) Potassium citrate (c) Sodium fluoride, 0.05%

Colgate FluoriGard Gel

Colgate-Palmolive, Ltd., Guildford, Surrey, UK

(a) Stannous fluoride, 0.4% (b) Abrasive-free

Sensodyne F toothpaste

Stafford-Miller Ltd., Welwyn Garden City, UK

(a) Potassium chloride 3.75% (b) Sodium monofluorophosphate, 0.8%

Sensodyne Mint toothpaste

Stafford-Miller Ltd., Welwyn Garden City, UK

Strontium chloride hexahydrate, 10%

Elgydium Mint Gel toothpaste

Pierre Fabre, UK

(a) Chlorhexidine, 0.004% (b) Nicomethanol fluorhydrate, 0.85%

Macleans Sensitive toothpaste

SmithKline Beecham, Brentford, Middlesex, UK

(a) Sodium fluoride, 0.23% (b) Strontium acetate hemihydrate, 8%

Mentadent Sensitive toothpaste

Elida Gibbs, Portman Square, London, UK

(a) Tripotassium citrate, 5.35% (b) Sodium monofluorophosphate, 0.8%

Boots Formula F toothpaste

Boots Company Pic Nottingham, England, UK

(a) Potassium nitrate, 5.0% (b) Sodium monofluorophosphate, 0.8%

Macleans Freshmint toothpaste

SmithKline Beecham, Brentford, Middlesex, UK

(a) Sodium monofluorophosphate, 0.8% (b) Calcium glycerophosphate, 0.13% (c) Triclosan

Brushing only

nil

Brushing

identifying whether sensitive root surfaces have exposed, patent tubules, was reported by Yoshiyama et al. (1989, 1990). They identified regions of sensitivity clinically and then biopsied the 'sensitive dentin' using a hollow (1-mm inside diameter) core-producing diamond bur. Examination of the surfaces of cylindrical specimens by SEM revealed that 75% of the tubules were open, in contrast to only 24% in the 'non-sensitive' dentin biopsies. They also fractured the biopsies to examine the subsurface tubule contents. 'Sensitive' dentin exhibited relatively open lumina, while the tubules of 'non-sensitive', exposed dentin were partially occluded with mineral deposits. TEM was used to study the tubule lumen ultrastructure in superficial dentin. Only 15% of 'sensitive' dentin but 81% of 'non-sensitive' dentin tubules

were occluded with highly electron-dense material which appeared to be continuous with peritubular dentin. In situ studies Brannstrom and Garberoglio (1980) examined tubules below a superficial dentinal attritional surface. They included not only vital teeth but also young intact premolars implanted into dentures. In this in situ study, the enamel on the buccal cusps of the extracted teeth was removed and the dentin exposed to attrition for 3 yrs. The results supported the view that salivary components contributed to the development of dentin sclerosis and obliteration of the tubule lumen under attrited dentin. Kerns et al. (1991) also evaluated tubule occlusion longitudinally by various clinical procedures,


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including scaling and root planing and the application of potassium oxalate. The treated dentin discs were then incorporated into removable dentures worn by the donors, and the samples were evaluated at 1 wk by SEM. This study demonstrated that the above clinical procedures are relatively short-lived in their tubule-occluding effects. Significant loss of the smear layer and oxalate coating was seen in samples 1 wk after their production. Significantly, the non-treated acidetched slices were occluded by growth of deposits from salivary minerals, which appears to confirm earlier work by Brannstrom and Garberoglio (1980). Kuroiwa et al. (1994) examined the effect of brushing, with or without an abrasive toothpaste, on human cervical dentin blocks attached to resin plates which were exposed to the oral environment for 8 wks. SEM results indicated that brushing with a toothpaste containing calcium-based abrasive caused most tubules to open, while brushing without a toothpaste resulted in occlusion with an organic pellicle. It was suggested that a non-abrasive toothpaste might prevent DS.

QUALITATIVE AND QUANTITATIVE ANALYSIS OF DESENSITIZING AGENTS AND THEIR EFFECTS ON THE DENTIN SURFACE/ SUBSURFACE AND FLUID FLOW IN EXPERIMENTS BASED ON THE DENTIN DISC MODEL Several studies were undertaken to establish the applicability of the dentin disc model in the assessment of desensitizing agents based on the methodology of Greenhill and Pashley (1981) as modified by Mordan et al (1997). Caries-free, surgically extracted maxillary and mandibular third molars were fixed in 3% glutaraldehyde in 0.1 M sodium cacodylate buffer solution (pH 7.4) at 4°C for up to one week. After all organic matter was cleaned from the fixed tooth surface with a single-edged razor blade, the teeth were stored in 0.1 M sodium cacodylate buffer solution and subsequently sectioned mesio-distally into discs approximately 1 mm thick by means of a diamond sectioning saw. All dentin discs were stored in fresh sodium cacodylate buffer solution (4°C) until required. Prior to application of the selected in-office and over-the-counter (OTC) desensitizing agents (Table 1), dentin discs were ultrasonicated in distilled water for 30 sec, etched in 6% citric acid for 2 min, and rinsed in distilled water. Each disc was marked (on either side) to assist in identification and orientation of both test and control (sides) and then broken into experimental test and control sections by means of a dental chisel. Both tooth and discs were kept moist at all times. All desensitizing agents and techniques (laser) were applied to the experimental test sections in accordance with the manufacturers' instructions for clinical application, or were brushed on with an electric toothbrush and toothpaste for 2 min, following the protocol of Mordan et al. (1997). After application, the specimens were dried in a desiccator for 2 days, discs were attached to aluminum Cambridge stubs with carbon-conducting cement, coated with a thin layer of gold/palladium in a Polaron E5000 sputter-coater, and subsequently viewed in a Cambridge Stereoscan 90B SEM at

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a constant working distance of 10 mm. Variable accelerating voltages (10 K or 15 K) were selected to optimize image quality. Micrographs were taken from selected fields in the central portion of each disc at various magnifications (500x, lOOOx, or 2000x) for qualitative analysis. For each agent, a treated disc half was further fractured into two quarters by means of dental pliers without touching the treated disc surface, allowed to dry in a desiccator for 2 days, and examined under high resolution in a Hitachi 600 Field Emission SEM to determine evidence of depth of tubule penetration of the various applications. The smear layer, which totally obliterated the dentin tubules often observed on discs prepared with the diamond saw, was created as a result of the mechanical cutting procedure. Cracks were almost certainly artefacts produced by the extreme vacuum and drying procedure required for SEM examination (Pashley et al., 1981). Ultrasonication with distilled water failed to remove the superficial debris of the smear layer completely. Citric acid (6%) demonstrated a much greater efficiency in removing the smear layer, thus leaving patent dentin tubules and the intertubular surface exposed. It can be postulated that, in the in vivo situation, the smear layer may prevent or significantly reduce dentin permeability, as evidenced in the permeability studies of Pashley and coworkers (1978a,b, 1981, 1982, 1984, 1992), presumably by restricting the surface area available for diffusion of both small and large particles. Brushing is the usual mode of application of toothpastes. In one of our studies (Ling et al., 1997), we chose a twominute brushing or application time to apply a selection of desensitizing agents including toothpastes and sealants. Previous studies have used a twominute brushing or treatment time (Kerns et al, 1991; Knight et al, 1993; Ide et al., 1994), and this was considered reasonable for observing any changes produced by the test agents. Saliva supernatant, pre-treatment of the discs, and rinsing in a rotator attempted to simulate the in vivo situation. Rotation with saliva was carried out to investigate the degree of deposit retention over a period of time. SEM was utilized for qualitative analysis of the surface deposit, as well as x-ray microanalysis to characterize the elemental compositions of different specimens and of the deposits on treated discs. The finding of carbon in specimens mounted on carbon-coated aluminum stubs, and of gold/palladium, calcium, phosphorus, carbon, and oxygen on treated dentin discs was not surprising. This was almost certainly due to penetration of the electron beam into the dentin discs and was unavoidable because of the need to produce a conductive specimen source in the SEM preparation procedure. This raised a problem in differentiating between exogenous and endogenous elements, such as calcium, phosphorus, oxygen, and carbon. Interpretation of the spectra could be difficult if there are overlaps of peaks. A small peak might not show if there is a marked weight percentage difference between two elements. In this study (Ling et al., 1997), peaks of the elements detected in the compounds were significantly spread away from the natural elements of dentin structure and the gold/palladium used in specimen preparation. Under these circumstances, a


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reasonable correlation between the applied substance and the SEM preparation deposit on the dentin surface could be made. The crystal-like structures seen on the disc surface, which occluded almost all the tubule orifices following treatment with Sensodyne Sealant, probably reflected the tubuleoccluding potential of ferric oxalate. The different shapes of the crystal-like structures probably represent different stages of maturation. Sena (1990) proposed that the mode of action of ferric oxalate was possibly by interaction of the available ions, which include ferric, calcium, oxalate, and phosphate. This would result in the precipitation of calcium oxalate and ferric phosphate salts which occluded the dentin tubules. Xray microanalysis results from this study appeared to agree with the suggestion (Sena, 1990) that this precipitation is a possible mode of action for ferric oxalate, although the relative proportions of the two different salts were not established. This might also explain the finding of Yeh et al. (1990) that a one-minute application of Sensodyne Sealant reduced dentin permeability by 97% in the dentin disc model. The results of the present study, however, were in contrast to those of Knight et al. (1993), in which SEM evaluation of both the initial and washed specimens revealed minimal obliterating effect following a two-minute application. These investigators used a post-application one-minute water spray from an air-water syringe and 30 sec of air-blasting, which probably reduced the retention of the crystal-like structures. It should be stated that the manufacturer's recommendation for rinsing the tooth following ferric oxalate application is 10 sec. Differences between the methodology of the Knight et al. (1993) study and our study (which followed the manufacturer's recommendation) may be one of the reasons for the contrast in the results. Treatment of the dentin discs with Butler Protect did not consistently produce any significant degree of tubule occlusion. Potassium oxalate has been reported to reduce dentin sensitivity by a dual mechanism, namely, (1) by occluding tubule orifices with a fine, acid-resistant layer of calcium oxalate crystals, and (2) by an inhibitory effect of potassium ions on sensory nerve activity (SNA). The results of this study, however, failed to reproduce the results of Pashley and Galloway (1985), in which oxalate treatment of a smear layer of a dentin disc resulted in the formation of a layer of crystalline precipitates. The results of Kerns et al. (1991) also indicated the formation of numerous crystals of calcium oxalate completely obscuring the tubules. However, both studies involved the in vitro application of 30% dipotassium oxalate. In the clinical situation, it is doubtful that the Butler Protect solution would have such a high oxalate concentration. This difference in oxalate concentration probably accounted for the differences in results. Moreover, the long-term retention of the oxalate crystals was questioned by Kerns et al. (1991), who demonstrated that dentin surfaces treated with potassium oxalate showed few oxalate crystals after 7 days and that the effects of the oxalate solutions might be relatively short-lived. Absi et al. (1990) reported that brushing alone took several hours to form a smear layer. In the present study,


brushing with an electric toothbrush for 2 min had virtually no visible effect on the etched dentin surface. On this basis, where brushing was the only mode of application, any observable change was due to the test agent, and this provided a useful control. The in situ investigation by Kerns et al. (1991) indicated that etched dentin chips (no active treatment) incorporated into patients' dentures did not show any significant change after 1 wk, and that at 4 wks, the tubules were almost completely occluded. The apparent lack of evidence of any deposits after application of the Boots Sensitive Teeth Mouthwash and the 'abrasive free' Colgate FluoriGard Gel on the dentin discs would appear to suggest that they do not have tubule-occluding properties and may work by an entirely different mode of action, which could not be detected by this in vitro model. Two important aspects of this study were the surface changes produced by the toothpastes and the effects of rotation with saliva supernatant. The active ingredients of toothpaste, with the exception of potassium salts, have usually been considered to be effective tubule-occluding agents, but the results of the present study would appear to contradict this. The presence of silica and calcium by x-ray microanalysis, both on the disc surface and within the tubules, suggested that the abrasive component rather than the actice ingredient per se may be responsible for tubule occlusion. The effects produced by these abrasive components are probably dependent not only on the type, particle size, and shape of abrasive but also on the formulation of the toothpaste. Chemical and physical interactions between ingredients may lead to considerable differences in retention of abrasive components when rinsed with saliva. Almost all toothpastes, with the exception of Sensodyne F, formed an initial granular deposit on the dentin surface which partially or completely occluded the tubules. Sensodyne Mint toothpaste, which contained the silica-based diatomaceous earth, was largely removed by washing. The retention of surface deposits of silica-based Elgydium Mint gel, Macleans Sensitive, and Mentadent Sensitive toothpastes was markedly reduced by rinsing. Boots Formula F appeared less affected. Deposit of the benchmark fluoride toothpaste, Macleans Freshmint, was little influenced by rinsing, and xray microanalysis indicated that calcium was the main element. The tubule-occluding effect produced by this calcium-based toothpaste was comparable with, if not superior to, that of the other desensitizing toothpastes. The results of this study support the findings of Addy and Mostafa (1989) that the tubule occlusion effects of the test toothpastes were almost certainly produced by the abrasive and/or other components. The fact that most of the active ingredients of the test agents in the present study failed to demonstrate any tubule-occluding properties is also consistent with the results of Addy and Mostafa (1988), who reported that of the agents tested, only the tin ion produced marked changes on the dentin surface. Takahashi (1986) also failed to detect any obvious changes produced by the various salts included in a reported investigation with co-workers; aluminum lactate was the only compound with a distinct tubule occlusion effect. Interpreting changes observed in


PLAUSIBILITY OF THE DENTIN DISC MODEL

vitro, however, is difficult, and extrapolation to the clinical situation must be tempered with caution. Different mechanisms of action other than tubule occlusion, which would not be normally simulated in this model, may also be responsible for the reduction of DS in vivo. The importance of outward fluid flow in areas of exposed dentin (Vongsavan and Matthews, 1991, 1992) cannot be overlooked. If their findings were extrapolated to the human model, then it could be postulated that the outward fluid flow would prevent the ingress of chemical substances into the tubule. This raised the question as to whether the proposed mode of action of potassium-containing salts (i.e., the inward diffusion of potassium ions from the tooth surface to effect desensitization of the pulpal nerves) is valid as discussed above. Pearce et al. (1994) have suggested that a commercially formulated fluoride toothpaste would be a more appropriate control than a so-called 'active minus' control. However, the formulated toothpaste may also have some therapeutic potential. Our results in vitro clearly demonstrated the tubuleoccluding potential of a commercially formulated fluoride toothpaste. If this result were mimicked in the en vivo situation, this would explain, in part, the lack of statistical significance between the test and control agents shown in their study. Furthermore, because of the number of commercially available fluoride formulations which may have different therapeutic potentials, it would be difficult (at present) to select one as the 'so-called' gold standard/ benchmark formulation for a control in clinical trials designed to determine the efficacy of desensitizing agents. A further study (Gillam et al, 1996) investigated the potential tubule-occluding effects of 5 selected in-office desensitizing agents: Sensodyne Sealant (Block Drug Co, Inc., Jersey City, NJ, USA), Butler Protect (John O. Butler Company, Chicago, IL, USA), OxaGel (Art-Dent Ind e Com de Produtos Odontologicos, Brazil), AllBond 2 (Bisco Inc., Itasca, USA), and One-Step (Bisco Inc., Itasca, USA). The results of this study indicated that all of the applied desensitizing agents produced some occlusion of the tubules, although the levels of coverage and occlusion varied between the products. A qualitative assessment of the visible surfaces by SEM showed that ferric oxalate, the active ingredient of Sensodyne Sealant, produced initial crystal-like structures which occluded a high proportion of the tubules across the dentin disc surface. All-Bond 2 and One-Step (both lightcured primer systems) showed uneven coverage across the disc surface, with both occlusion and reduction in tubule diameter. The use of a surface conditioner did not appear to have a noticeable effect when used in conjunction with AllBond 2 and One-Step. These three products, however, appeared to be more effective than either the Butler Protect (potassium oxalate) or Oxa-Gel (potassium oxalate in a gel), with which there was a marked decrease in the levels of both coverage and tubule occlusion. However, evaluation of the fractured discs demonstrated that all desensitizing agents penetrated the tubules to various degrees. This study demonstrated that, following application of the 5 selected desensitizing agents to the dentin disc surface, there were coverage and deposition which occluded the tubule orifices,

495

although this effect varied between products. Evaluation of the surface characteristics alone, however, may lead to a misrepresentation of the potential effects of these products, and evidence of tubule penetration demonstrated by the results from the fractured discs should be provided together with results from suitable clinical studies.

THE SURFACE CHARACTERISTICS OF LASER APPLICATION ON DENTIN BY MEANS OF THE FOTONA TWINLIGHT DENTAL LASER (Nd:Yag, Er:Yag, HeNe) Apart from the application of toothpastes and practitionerapplied products such as sealants, varnishes, and primers to the dentin surface, there have been both in vitro and in vivo studies of the effects of laser on dentin (Wakabayashi and Matsumoto, 1988; Wilder-Smith, 1988; White et al., 1990, 1991; Manton et al, 1992; Renton-Harper and Midda, 1992; Pashley et al, 1992; Gelskey et al, 1993; Gerschman et al, 1994). It has been assumed that hard lasers reduce DS by a surface effect (smear layer creation) which occludes the tubules. However, soft lasers may act by interfering with the polarity of cell membranes. Olsen (1981) has proposed that low-level laser therapy (LLLT) exerts its effect by stimulating the sodium/potassium pump in cell membranes which maintains the potential difference across the membrane. Stimulation of the pump is thought to hyperpolarize the membrane, thus increasing the nerve threshold of firing, and also increasing the pain threshold. The surface effects of the pulsed Nd:YAG laser on enamel and dentin have been reported by Cox et al (1994), who confirmed similar findings by Scheinen and Kantola (1969) using the CO2 laser, demonstrating cratering of the enamel and dentin surfaces together with fusion of the surface layer. Our present laser study (McCarthy et al, 1997), using a Fotona Twinlight Dental Laser ([Fotona d.d. Stegene, Ljubljana, Slovenia] [Nd:Yag, Er.Yag, HeNe]) compared the effects on root surfaces and dentin discs at various power settings (therapeutic values for the treatment of DS) as recommended by the manufacturer. Er.Yag radiation at 60 mJ, 80 mJ, and 100 mJ, at a frequency of 2 Hz for 10 sec, produced craters with open tubules in dentin discs, but closed tubules in root surfaces. Nd:Yag radiation at 3.5 W, 3.75 W, or 4 W at an 80-Hz frequency for 25 sec produced irregular melting and resolidification in all specimens, with some tubule occlusion. The HeNe radiation produced no apparent effects. There was no evidence of a smooth, glazed layer of melted and resolidified dentin at the chosen power levels. The conclusion that can be drawn from these experiments is that laser radiation may reduce DS by sealing dentin tubules, but this could not be confirmed by this study.

QUANTITATIVE STUDY ON THE SURFACE CHARACTERISTICS OF SELECTED DESENSITIZING AGENTS BY MEANS OF SEM AND IMAGE ANALYSIS Previous studies where investigators assessed the tubule-


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TABLE 2 SUMMARIES OF SEM AND X-RAY MICRO ANALYSIS RESULTS OF DENTIN DISCS TREATED WITH DIFFERENT TEST AGENTS, AND THE EFFECT OF SALIVA ROTATION Test Agent O-hr Rotation

X-ray Microanalysis

SEM Observation 6-hr Rotation

Brushing only

patent dentinal tubules


no analysis

Boots Sensitive Teeth Mouthwash



no analysis

Colgate FluoriGard Gel



no analysis

Sensodyne F

granular deposit around the tubule orifices and on the surface, most tubules remained patent irregular deposit of diatomaceous earth, some tubules partially occluded

reduced deposit, both around the tubule orifices and on the surface

no analysis

particles largely reduced, some tubules still occluded

no analysis

Elgydium Mint Gel

irregular granular deposit, tubules partially or completely occluded, some deposit inside the tubules

considerable reduction in deposit, many tubule openings exposed, halolike appearance at the periphery

silica, oxygen, carbon

Macleans Sensitive

irregular granular deposit over the entire surface, tubules partially or completely occluded

less granular deposit, many tubules largely unblocked, halo-like appearance around the tubules

silica, oxygen

Mentadent Sensitive

large and very fine granular deposits, tubules partially occluded

many tubules unblocked, very fine deposit apparent around the tubule orifices

silica, oxygen

Boots Formula F

irregular large and small granular deposit, most tubules occluded

reduced granular deposit, larger granular deposit absent, most tubules partially unblocked

no analysis

Macleans Freshmint

small irregular and rodshaped particles, tubules occluded by membranelike sheath

little change in deposit, tubules partially occluded by membrane-like sheath

calcium, oxygen, carbon, silica

Sensodyne Mint

From Ling etal. (1997).

occluding effects of desensitizing agents by viewing dentin discs by SEM provided only descriptive terms—for example, partial or complete blockage of tubules. Other investigators described their results with symbols, such as +, ++, +++, etc.

(Absi et aL, 1992, 1995; Addy and Urquhart, 1992) or provided indices based on percentages of occluded tubules (McAndrew and Kourkouta, 1995; Rimondini et aL, 1995), although some results were measured by graticules (Absi et


PLAUSIBILITY OF THE DENTIN DISC MODEL

aL, 1987). In essence, although these investigations claimed that their methods were quantitative, in reality, they were mainly descriptive (qualitative) studies. The aim of the present quantitative study (Ip et aL, 1997) was to improve the methodology involved in the assessment of dentin morphology and the surface characteristics of surfaces treated with selected desensitizing agents by means of image analysis (Quantimet 520, Leica, UK). Three desensitizing agents—namely, Butler Protect, Macleans Sensitive toothpaste, and Colgate FluorideGard (Gel-Kam)—were used. Two different desensitizing agents (Butler Protect and Sensodyne Sealant) were used in a pilot study to determine reproducibility for the proposed model. Both studies were repeated as a test for reproducibility. Micrographs were taken from four areas on both test and control sections in the central portion of the disc, and 6 randomly selected fields were taken from each of the 4 SEM negatives (magnification, lOOOx; working distance, 10 mm). Measurements of number of tubules, width of tubule lumen, tubule patent area, and proportion of patent area against field area were obtained and compared. Comparison of control and test specimens indicated that differences in number of tubules, patent tubule areas, width of tubules, and percentage of patent areas could be assessed quantitatively. Furthermore, this method also demonstrated differences between the tubule-occluding properties of the selected desensitizing agents. Another of the interesting findings was the observation that desensitizing agents previously reported as having limited or little effect on the dentin surface (Ling et aL, 1997)—namely, potassium oxalate and Gel-Kam—provided significant reduction in tubule width and area when analyzed by image analysis. This would appear to support the observation of Gillam et aL (1996) that descriptive or qualitative examination of the dentin surface alone may be insufficient for the evaluation of the tubule-occluding properties of selected desensitizing agents.

DISCUSSION The concept of tubule occlusion as a method of dentin desensitization is a logical conclusion from the hydrodynamic theory. The fact that many of the agents used clinically to desensitize dentin are also effective in reducing dentin permeability tends to support the hydrodynamic theory. Theoretically, all agents which occlude dentin tubules should reduce dentin permeability and, therefore, decrease DS. However, the converse of that statement is not necessarily true. Not all agents that decrease dentin sensitivity do so by occluding dentin tubules. Currently, there are at least two recognized mechanisms of action of desensitizing agents. One involves blocking fluid movement by occluding tubules. The other involves blocking pulpal nerve activity by altering the excitability of the sensory nerves (Pashley, 1986). It has been shown that the morphological variability of dentin tubule size and density throughout the tooth necessitates careful choice of dentin disc and careful control procedures (Mordan et aL, 1997). The divergence of tubules which occurs between the pulp and the dentin-enamel

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junction makes the tubule density vary widely, depending on distance from the pulp. Another complicating factor is that tubule diameter is largest at the pulp side of the dentin and smallest at the enamel side. When the dentin discs were prepared, the difference in orientation of cut tubules resulted in different morphological appearances of the central and peripheral areas. For meaningful comparisons and accurate interpretation, only those tubules from the central portions on either side of the fractured disc were examined. In the various studies by Gillam and co-workers, any change on the test side was compared with the corresponding control side on the same disc. Therefore, any variation in tubule orifices was attributable to either the agents under study and/or the application procedures. Most studies on tubule occlusion have focused on coronal dentin, where important variables such as dentin surface area, thickness, and surface characteristics can be controlled. The validity of data collected in vitro, however, is open to criticism. The hydraulic conductance of radicular dentin has been found to be much lower than that of coronal dentin, and tubule density and diameter correlated well with measured hydraulic conductance (Fogel et aL, 1988). If the design of the experiment would allow for a curved treatment surface, it might be preferable to use cervical dentin blocks for the screening of potential desensitizing agents (Kerns et aL, 1991; Dragolich et aL, 1993). For the purpose of previous and ongoing studies, however, where brushing of a flat dentin surface with toothpastes and standardized comparisons of the different treatment procedures were required, a dentin disc model prepared from the coronal tooth structure appeared appropriate, given the variation in tubule density and diameter. Difficulties in the interpretation of the in vitro results, however, may also arise with the use of extracted teeth designed to reproduce the effects of chemicals on dentin in vivo. For example, the presence or absence of a smear layer, and the penetration of acid, enzymes, and exogenous materials into dentinal tubules in vitro are likely to be much greater than in vivo (Vongsavan and Matthews, 1991). Experiments may also give misleading results when such tests are carried out on dried, etched dentin in vitro, as opposed to dentin in which the ends of the tubules are filled with fluid, as will probably be the state in vivo, despite attempts to dry the exposed dentin surface (Tao and Pashley, 1989; Dragolich etaL, 1993). Despite these apparent limitations, the dentin disc model is nevertheless a recognized adequate screening model for potential tubule-occluding agents. The dentin disc has been used extensively as a model for assessing the surface deposition and tubule-occluding effects of desensitizing agents as well as the effects of these agents on fluid flow through dentin (hydraulic conductance) (Pashley et aL, 1978a,b, 1984, 1987; Greenhill and Pashley, 1981; Addy and Mostafa, 1988, 1989; Cuenin etaL, 1991; Knight etaL, 1993; Kuroiwa et aL, 1994; Ling et aL, 1997). It should be recognized, however, that the hydrodynamic theory is based on the concept of fluid dynamics in capillary tubes, which may not necessarily be true in the light of the Mjor and Nordahl (1996) observation of an intricate network of


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intercommunicating dentinal tubules. Detailed characterization of the experimental dentin surface/subsurface following etching and application of desensitizing agents ranging from toothpastes to dentin-bonding materials may, however, be complicated by various factors such as the age, condition and source of the dentin specimen, the groove size of the specimen following cutting, the presence or absence of a smear layer, the density, diameter, direction, and orientation (cross-sectional or longitudinal) of the tubules, the presence or absence of highly mineralized peritubular dentin, and the variation of the intricate branching tubule system as well as the types and locations of these branches (Marshall, 1993; Mjor and Nordahl, 1996; Mordan et aL, 1997). The resultant variations in the disc surface morphology following etching, therefore, make the choice and evaluation of suitable controls a significant factor in the experimental design (Mordan et aL, 1997). Results from the studies by Gillam and co-workers using this methodology demonstrated that, following application of the various OTC and in-office dentifrices to the surfaces of dentin discs, there was coverage, albeit to different levels, although not all the products occluded the tubules. Qualtitative evaluation of the surface effects alone may, however, lead to a misrepresentation of the potential effects of these products in vitro, as indicated by the results from the fractured disc half/halves, where tubule penetration was clearly observed, even in the specimens where the surface deposition was less evident. It may be concluded that the dentin disc provides a versatile and readily available model for DS studies. Its reliability depends upon the precise location of the disc within the tooth and to the position on the disc (Mordan et aL, 1997). Those agents which do not block tubule orifices in the dentin disc model may still, however, be effective in vivo, since they may operate via a mechanism other than occlusion of tubules, or they may block tubules in vivo by a mechanism which cannot be simulated in this laboratory model.

ACKNOWLEDGMENTS The authors would like to acknowledge the contributions from various colleagues for data reported in this paper: Mrs. P.M. Barber, Dr. D. McCarthy, Dr. GJ. Pearson, Dr. T.Y.Y. Ling, Dr. T. Ip, Mr. N. Khan, and Mr. J. Critchell.

REFERENCES Absi EG, Addy M, Adams D (1987). A study of the patency of dentinal tubules in sensitive and non sensitive cervical dentine. / Clin Periodontol 14:280-284. Absi EG, Addy M, Adams D (1989a). Dentine hypersensitivity: The development and evaluation of a replica technique to study sensitive and non-sensitive cervical dentine. J Clin Periodontol 16:190-195. Absi EG, Addy M, Adams D (1989b). Dentine hypersensitivity. Uptake of various sensitizing toothpastes onto dentine in vitro SEM investigation (abstract). J Dent Res 68(Spec Iss):573.


Absi EG, Addy M, Adams D (1990). Does toothbrushing remove or produce a smear layer on dentine? An S.E.M. investigation (abstract). J Dent Res 69(Spec Iss):965. Absi EG, Addy M, Adams D (1992). Dentine hypersensitivity—the effect of toothbrushing and dietary compounds on dentine in vitro: an SEM study. J Oral Rehabil \9\\Q\-\\Q. Absi EG, Addy M, Adams D (1995). Dentine hypersensitivity: uptake of toothpastes onto dentine and effects of brushing, washing and dietary acid—SEM in vitro study. J Oral Rehabil 22:175-182. Addy M (1990). Etiology and clinical implications of dentine hypersensitivity. Dent Clin North Am 34:503-514. Addy M (1992). Clinical aspects of dentine hypersensitivity. Proc Finn Dent Soc 88(Suppl l):23-30. Addy M, Mostafa P (1988). Dentine hypersensitivity: I. Effects produced by the uptake in vitro of metal ions, fluoride and formaldehyde onto dentine. J Oral Rehabil 15:575-585. Addy M, Mostafa P (1989). Dentine hypersensitivity. II. Effects produced by the uptake in vitro of toothpastes onto dentine. J Oral Rehabil 16:35-48. Addy M, Urquhart E (1992). Dentine hypersensitivity: its prevalence, aetiology and clinical management. Dent Update (Dec):407-412. Addy M, West N (1994). Etiology, mechanisms and management of dentine hypersensitivity. Curr Opin Periodontol 71-77. Addy M, Mostafa P, Absi EG, Adams D (1985). Cervical dentin hypersensitivity. Etiology and management with particular reference to dentifrices. In: Proceedings of Symposium on Hypersensitive Dentin. Origin and Management. University of Michigan. Rowe NH, editor. Edinburgh and London: E. and S. Livingstone Limited, pp. 147-167. Arends J, Stokroos I, Jongebloed WG, Ruben J (1995). The diameter of dentinal tubules in human coronal dentine after demineralization and air drying. Caries Res 29:118121. Berman LH (1985). Dentinal sensation and hypersensitivity. A review of mechanisms and treatment alternatives. / Periodontol 56:216-222. Blandy AA (1850). On the sensitivity of teeth. Am J Dent Sci 1:22-28. Brannstrom M (1963). A hydrodynamic mechanism in the transmission of pain producing stimuli through the dentine. In: Sensory mechanism in dentine. Anderson DJ, editor. Oxford: Pergammon Press, pp. 73-79. Brannstrom M (1965). The surface of sensitive dentine. OdontRev 16:293-299. Brannstrom M (1992). Etiology of dentin hypersensitivity. Proc Finn Dent Soc 88(Suppl 1):7-13. Brannstrom M, Garberoglio R (1980). Occlusion of dentinal tubules under superficial attrited dentin. Swed Dent J 4:8191. Brannstrom M, Johnson G, Nordenvall KJ (1979). Transmission and control of dentinal pain: resin impregnation for the desensitization of dentin. J Am Dent Assoc


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