Assessment of enamel erosion and protective effect of salivary pellicle by surface roughness analysis and scanning electron microscopy.

PURPOSE: To assess dental erosion caused by 0.1% and 1.0% citric acid in vitro and to estimate the protective influence of experimentally formed salivary pellicle. MATERIALS AND METHODS: Bovine enamel slabs (n = 80) were polished and embedded in epoxy resin. For the formation of pellicle layer 40 specimens were immersed for 24 h in pooled human saliva. Erosion was caused by immersion in citric acid solution for 1, 5, 10 and 30 min. Erosive alterations on the pellicle-covered and non-covered enamel specimens were scored as a change (delta) of surface roughness parameters Ra, Rt and RzDIN using contact profilometer and observed in scanning electron microscope. RESULTS: Profilometric analysis of eroded enamel specimens emphasized the aggressiveness of even low concentrated citric acid with a short period of challenge. The change of roughness parameters after 1-min immersion in 0.1% citric acid were 16.4, 182.6 and 132.2 nm for deltaRa, deltaRt and deltaRzDIN, respectively, and 54.8, 516.6 and 258.2 nm after 1-min immersion in 1.0% citric acid. Changes of the surface roughness were dependent on the exposure time and concentration of acidic solution. Pellicle layer significantly reduced the extent of erosive destruction, which was additionally documented on SEM-micrographs. Residual pellicle-like structures were detected after 5 min of immersion in 0.1% citric acid. However, there were no significant differences in pellicle-covered and non-covered enamel slabs measured profilometrically for 1.0% citric acid with 10 min and 30 min exposure time. CONCLUSION: The findings confirm the property of pellicle layer to resist against erosive influence of organic acids, which is, however, limited by duration of acidic treatment and concentration of erosive agent.

Sites of dental erosion are saliva-dependent.

Acid demineralization of teeth causes occlusal erosion and attrition and associated non-carious cervical lesions at sites relatively unprotected by saliva. Associations of occlusal pathology and cervical lesions were looked for in 450 patients with toothwear, and 174 subjects with cervical lesions were identified. Associations of occlusal attrition, or erosion, or no wear, with cervical lesions at 72 buccal and lingual sites were recorded from epoxy resin replicas of the subjects' dentitions (3241 teeth). Criteria used to discriminate occlusal erosion from attrition; and shallow from grooved and wedge-shaped cervical lesions were delineated by scanning electron microscopy (SEM). In the absence of occlusal pathology, cervical lesions were very rare (<1%). In the presence of occlusal pathology, cervical lesions were present in 27.71% of buccal sites as opposed to 2.61% of lingual sites. The commonest site of cervical lesions was the facial of maxillary incisors (36% of sites). The least common site was the lingual aspect of mandibular molars (1.7% of sites). These differences may reflect the normal protective role of serous saliva and salivary pellicle in a site-specific manner, on the lingual surfaces of mandibular teeth particularly, and do not support abfraction as the prime aetiology of cervical lesions.

Syndromes with salivary dysfunction predispose to tooth wear: Case reports of congenital dysfunction of major salivary glands, Prader-Willi, congenital rubella, and Sjögren's syndromes.

Four cases-of congenital dysfunction of the major salivary glands as well as of Prader-Willi, congenital rubella, and Sjögren's syndromes-were identified in a series of 500 patients referred for excessive tooth wear. Although there was evidence of consumption of highly acidic drinks, some occlusal parafunction, and unacceptable toothbrushing habits, salivary dysfunction was the salient factor predisposing a patient to tooth wear in these syndromal cases. The 500 subjects have been characterized either as having medical conditions and medications that predispose them to xerostomia or lifestyles in which workplace- and sports-related dehydration lead to reduced salivary flow. Normal salivation, by buffering capacity, clearance by swallowing, pellicle formation, and capacity for remineralization of demineralized enamel, protects the teeth from extrinsic and intrinsic acids that initiate dental erosion. Thus, the syndromes, unrelated in many respects, underline the importance of normal salivation in the protection of teeth against tooth wear by erosion, attrition, and abrasion.

Protective properties of salivary pellicles from two different intraoral sites on enamel erosion.

The purpose of this study was to investigate the protective effect and ultrastructure of salivary pellicles formed in vivo near the orifices of the ducts of parotid and submandibular/sublingual salivary glands. Pellicles were formed by exposing bovine enamel slabs to the oral environment at the buccal aspect of the upper first molars and at the lingual aspect of the lower incisors in 3 subjects over periods of 24 h. Enamel specimens with and without 24-hour pellicles were immersed in citric acid (0.1 and 1%) for periods ranging from 30 s to 5 min, and processed for measurement of surface microhardness (SMH) and transmission electron microscopy (TEM). In comparison to uncovered enamel specimen significantly less decrease in SMH due to acid exposure was observed in pellicle-coated enamel specimens. Pellicles formed at the buccal aspect of the upper molars were less effective in protecting the enamel against acid-induced softening as compared to pellicles formed at the lingual aspect of the lower incisors only after 5 min exposure in 1% citric acid. TEM analysis showed that pellicle layers were dissolved continuously due to acid exposure. However, even after 5 min exposure to 1% citric acid, a residual pellicle layer could be detected on the enamel surface. In conclusion, site-dependent differences of buccally and lingually in vivo formed 24-hour pellicles have minor importance concerning the pellicle-induced protection of the enamel surface against erosive changes.

Pulp-dentin biology in restorative dentistry. Part 5: Clinical management and tissue changes associated with wear and trauma.

Tooth wear occurs during normal mastication. Pathologic wear, including abrasion and erosion, may also take place. Formation of reactionary and reparative dentin and obturation of dentinal tubules are biologic responses that compensate for the loss of tissue. Physical trauma to teeth will affect the blood supply to the pulp. Extensive physical trauma may result in the formation of mineralized, often bonelike tissue in the pulp chamber. Minor trauma, such as that associated with orthodontic tooth movement, may exert transient effects on the pulp or it may result in permanent structural changes. No experimental data involving restorative work on traumatized teeth have been published, but available evidence suggests that special care may be necessary in the restoration of such teeth because their reaction patterns may differ from those in nontraumatized teeth.

The effect of saliva on fluoride release by a glass-ionomer filling material.

The initial aim of this study was to investigate the effect of saliva and the formation of pellicle on the fluoride release in vitro of the glass-ionomer filling material, Chemfil Superior. For the first study glass-ionomer discs of 6 mm in diameter and 1.5 mm thick were made. Ten discs were immersed in whole stimulated saliva each day for 10 min and 10 control discs were immersed in deionized water. For the remaining 23 h and 50 min of each day, over the 20-day experimental period, both test and control discs were placed in deionized water. A considerable amount of fluoride was released on the first day (14.5 ppm F control and 13.3 ppm F test). The concentration of fluoride released on the second day fell sharply to 5.3 ppm F for controls and 4.9 ppm F for tests. This release had almost reached a plateau by day 10 and at day 20 the pellets continued to release low levels of fluoride. The concentration of fluoride released was only slightly higher for controls than for test discs when both were immersed in deionized water until day 20. However, during the 10-min period between 1.5 and 2 times as much fluoride was released into the deionized water as into saliva until day 20 when the ratio fell to 1.2:1. The second experiment assessed fluoride release when specimens were incubated for 1 h using an identical protocol. Again, less fluoride was released from the saliva-coated specimens compared with the controls (17%), which was not substantially different to the comparable 10-min samples (13%). This study indicates that saliva retards the release of fluoride from glass-ionomer and that this retarding effect is still present when discs are subsequently immersed in water compared with those that were placed in water alone. This suggests that salivary deposits have formed within minutes of immersion in saliva. This retarding effect was observed throughout the study period with the exception of the 20-day samples which had been incubated in saliva for 10 min.

The effect of saliva or serum on Streptococcus mutans and Candida albicans colonization of hydroxylapatite beads.

OBJECTIVE: Several recent reports imply the possibility of cariogenicity of denture plaque containing Candida albicans. Hence the purpose of this study was to investigate the effects of salivary and serum pellicles on C. albicans and Streptococcus mutans colonization on hydroxylapatite beads. METHODS: The colonization of three isolates of C. albicans and two isolates of S. mutans was examined by the use of a bioluminescent adenosine triphosphate (ATP) assay based on the firefly luciferase-luciferin system. RESULTS: In the preliminary study, a good correlation was observed between the cell number and ATP amount of each isolate tested, and the results yielded a level of significance (P < 0.001; Student's t-test), confirming the validity of this method. When the relative ATP content of the 48 h colonization of both isolates of S. mutans were compared, a saliva pellicle was significantly more effective in promoting bacterial colonization than either uncoated or serum pellicle (ANOVA; P < 0.01). In contrast, in the case of colonization of C. albicans isolates, a serum pellicle was significantly more effective in promoting the colonization of C. albicans GDH 18 and GDH 19, than both uncoated specimens and saliva pellicle (ANOVA; P < 0.01). Similar trends were observed with C. albicans GDH 16, though significant differences were not observed (ANOVA; P > 0.05). CONCLUSION: The results suggest that the mechanism involved in fungal colonization on hydroxylapatite (HAP) should be different from that of mutans streptococci.

A mathematical model of the influence of salivary urea on the pH of fasted dental plaque and on the changes occurring during a cariogenic challenge.

Urea diffusing from saliva into dental plaque is converted to ammonia and carbon dioxide by bacterial ureases. The influence of normal salivary urea levels on the pH of fasted plaque and on the depth and duration of a Stephan curve is uncertain. A numerical model which simulates a cariogenic challenge (a 10% sucrose rinse alone or one followed by use of chewing-gum with or without sugar) was modified to include salivary urea levels from 0 to 30 mmol/l. It incorporated: site-dependent exchange between bulk saliva and plaque surfaces via a salivary film; sugar and urea diffusion into plaque; pH-dependent rates of acid formation and urea breakdown; diffusion and dissociation of end-products and other buffers (acetate, lactate, phosphate, ammonia and carbonate); diffusion of protons and other ions; equilibration with fixed and mobile buffers; and charge-coupling between ionic flows. The Km (2.12 mmol/l) and Vmax (0.11 micromol urea/min/mg dry weight) values for urease activity and the pH dependence of Vmax were taken from the literature. From the results, it is predicted that urea concentrations normally present in saliva (3-5 mmol/l) will increase the pH at the base of a 0.5-mm-thick fasted plaque by up to 1 pH unit, and raise the pH minimum after a sucrose rinse or sugar-containing chewing-gum by at least half a pH unit. The results suggest that plaque cariogenicity may be inversely related to salivary urea concentrations, not only when the latter are elevated because of disease, but even when they are in the normal range.

Experimental salivary pellicles formed on titanium surfaces mediate adhesion of streptococci.

The goal of this study was to characterize salivary components of titanium pellicles and to determine how experimental pellicles affect adhesion of several strains of streptococci to titanium surfaces. Titanium experimental pellicles were formed by incubation of fresh human parotid or human submandibular-sublingual saliva on pure titanium beads. Pellicle was recovered from the beads using sodium dodecyl sulfate buffer and was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting to identify adsorbed salivary components. Streptococcus anginosus, S. oralis, and S. salivarius recovered from in vivo titanium plaque and five reference strains of streptococci were used in adhesion assays to titanium beads with and without experimental salivary pellicles. The experimental pellicle formed on titanium was found to be composed of selected proteins from human parotid and human submandibular-sublingual saliva. Salivary alpha-amylase and proline-rich proteins were found in all experimental pellicles, while sIgA, high-molecular weight mucin, and proline-rich glycoproteins were detected in one of the experimental pellicles examined. Adhesion of fresh isolates and reference stains of S. anginosus, S. oralis, and S. salivarius to saliva-coated titanium was reduced compared to that of titanium without saliva coating. However, adhesion of laboratory strains of S. gordonii and S. sanguis was found to be significantly greater to experimental pellicles of human submandibular-sublingual saliva than was the adhesion of the fresh isolates, suggesting that streptococci-colonizing implant surfaces may be inherently less adhesive than other bacterial strains. This study found that salivary pellicles are selectively formed on titanium and mediate in vitro adhesion of streptococci.

Human oral microbial ecology and dental caries and periodontal diseases.

In the human oral cavity, which is an open growth system, bacteria must first adhere to a surface in order to be able to colonize. Ability to colonize a non-shedding tooth surface is necessary prior to any odontopathic or periodontopathic process. Complex microbe-host relationships occur and must be studied before the commensal-to-pathogenic nature of the human indigenous oral flora can be understood. Medical pathogens, if present in the appropriate host, always produce specific disease. Caries and periodontal diseases are conditional diseases, requiring numbers of certain indigenous species at various sites, particularly the tooth surface. In the case of caries, the condition is related to sugar consumption. Periodontal disease/s may require certain host and environmental conditions, such as local environment or nutritional factors in gingival crevicular fluids. Nonetheless, critical numbers of certain indigenous species must be present in order for these diseases to occur. The aim of this review is to understand the acquisition of the indigenous oral flora and the development of human dental plaque. The role of the salivary pellicle and adherence of indigenous bacteria to it are critical first steps in plaque development. Bacterial interactions with saliva, nutritional factors, growth factors, and microbial physiologic processes are all involved in the overall process of microbial colonization.

Computer modeling of the effects of chewing sugar-free and sucrose-containing gums on the pH changes in dental plaque associated with a cariogenic challenge at different intra-oral sites.

Variation in salivary access to different intra-oral sites is an important factor in the site-dependence of dental caries. This study explored, theoretically, how access is modified by chewing sugar-free and sugar-containing gums. A finite difference computer model, described elsewhere, was used. This allowed for diffusion and/or reaction of substrate, acid product, salivary buffers, and fixed-acid groups. Site-dependent saliva/plaque exchange was modeled in terms of a 100-microns-thick salivary film covering the plaque (a) flowing directly from the salivary ducts, (b) flowing from the intra-oral salivary pool, or (c) exchanging with the pool. Computed flow-velocities or rates of exchange were based on previous intra-oral measurements. The model was also tested against an in vitro study conducted by two of the authors. In addition, the three proposed models of saliva/plaque interaction were compared, and the effect of salivary film thickness investigate. Results suggested that: (1) although sugar-free gum chewed during a cariogenic challenge causes a rapid rise in plaque pH, sucrose-containing gums cause the pH, after a temporary rise resulting from increased salivary flow, to stay low for an extended period; (2) the computer model reproduced in vitro tests reasonably well; (3) although the three models of the plaque/saliva interaction start from different assumptions, two lead to closely related predictions; and (4) increasing the assumed salivary film thickness by a large amount (e.g., from 50 to 200 microns) caused no change in modeled Stephan curves, as long as these changes were accompanied by appropriate reductions in film velocity, in accord, theoretically, with the practical clearance data.

Lysozyme and lactoperoxidase inhibit the adherence of Streptococcus mutans NCTC 10449 (serotype c) to saliva-treated hydroxyapatite in vitro.

Lysozyme, lactoperoxidase and salivary peroxidase inhibit the metabolism and growth of mutans streptococci, but any possible effects on the adherence of these bacteria are unknown. In this study the effects of lysozyme and lactoperoxidase on the adhesion of 3H-labelled Streptococcus mutans (NCTC 10449, serotype c strain) to saliva-coated hydroxyapatite were studied at pH 5.0 and 7.0. Human whole saliva was either lysozyme-depleted and centrifuged, or sterilized and dialysed to achieve no detectable lysozyme and peroxidase activities; this modified saliva was used to form experimental pellicles. The incorporation of lysozyme (50-200 micrograms/ml) to the pellicle caused a significant (p < 0.01) reduction in the adherence of S. mutans without any loss of bacterial viability. Pretreatment of either saliva-coated apatite or S. mutans cells with lysozyme did not change the results but lysozyme bound more readily to bacteria than to the experimental pellicles. Also, lactoperoxidase (10-200 micrograms/ml) reduced significantly (p < 0.001) the adherence of S. mutans but, in contrast to lysozyme, in a dose-dependent way. The strongest inhibition of adhesion was found when both saliva-coated apatite and bacteria were pretreated with lactoperoxidase. This enzyme bound to experimental pellicles in preference to streptococci. A non-specific protein control, albumin, did not block the inhibition by lysozyme or lactoperoxidase. The inhibition of adherence of a serotype c strain of S. mutans to saliva-coated hydroxyapatite is a novel antibacterial mechanism for both lysozyme and lactoperoxidase.

Calcium fluoride formation on sound enamel using fluoride solutions with and without lactate.

The formation of calcium (Ca) fluoride (CaF2) on bovine enamel blocks during clinically relevant treatment times using neutral fluoride (F) solutions (0.26 mol/l F) with and without 0.1 mol/l lactate was investigated. Uncoated and pellicle-coated blocks were evaluated for alkali-soluble (1 mol/l KOH, three consecutive 24-hour treatments) Ca, PO4, and F after treatment by the F solutions for 0, 5, 15, 30, and 60 min. There was an overall time-related increase in F recovery, while Ca tended to remain at baseline levels. Less F was recovered from the pellicle-coated blocks. The addition of lactate to the F treatment solution did not result in an overall increase in alkali-soluble F recovery, but did result in the formation of cuboidal shaped crystals which closely approached the morphology of pure CaF2. A 1:2 stoichiometric ratio Ca:2F (mol:mol) was not established based on chemical analyses. The ultrastructural and elemental composition of surface deposits on the samples, as determined using scanning electron microscopy, X-ray diffraction, and energy-dispersive spectroscopy, established the presence of CaF2 after 24-hour F treatments; however, it was not possible to directly demonstrate the formation of CaF2 after clinically relevant treatment times.

Interactions between denture lining material, protein pellicles and Candida albicans.

The interactions between pellicles derived from saliva, serum, mucin and lysozyme deposited on lining material (tissue conditioner) and Candida albicans were investigated by monitoring pH changes associated with protein-free and protein-coated lining material and by ultrastructural observations of yeast colonization. No significant differences in pH reduction between culture media in contact with the protein-free, control lining materials and those coated with saliva, serum or mucin were observed after 120 h of incubation. However, scanning electron microscopy revealed that much greater numbers of the yeasts colonized the saliva- or serum-coated lining material than the lysozyme-, mucin-coated or control material. Hyphal invasion was observed in saliva-coated lining material. These results suggested that denture pellicle derived from saliva and/or serum may potentiate candidal colonization of denture lining materials.

Effect of whole saliva components on enamel demineralization in vitro.

The aim of the present study was to use an in vitro enamel demineralization model (1) to confirm that whole saliva pretreatment conferred acid resistance to dental enamel and (2) to determine whether this phenomenon was attributable to specific salivary proteins, minerals, lipids, or some combination of these. Crowns of human teeth, each with one exposed window, were prepared in groups of ten. They were each pretreated by immersion individually in 4 ml of either (1) clarified whole saliva for 18, 72, or 168 h, (2) dialyzed saliva (3500 MWCO membrane), (3) the "flow-through" fraction from a DEAE separation of whole saliva (neutral and basic proteins), (4) the "eluted" fraction of a DEAE separation of whole saliva (anionic proteins), or (5) a combination of salivary lipids and the DEAE "flow-through" fraction of whole saliva (neutral and basic proteins). Control groups were group 6 with no pretreatment, group 7 pretreated for 168 h in a borate buffer (5 mmol/l), and group 8 pretreated in a mineral solution containing calcium (0.7 mmol/l) and phosphate (2.6 mmol/l). The crowns were then demineralized for 7 d in vitro (0.1 mol/l acetate, 1 mmol/l Ca and phosphate, pH 5.0) to produce artificial caries-like lesions. Lesions were assessed by cross-sectional microhardness profiles, and mineral loss (delta Z, micron x vol% mineral) calculated. Mineral loss (delta Z) values decreased linearly with the square root of time of pretreatment by whole saliva, confirming a time-dependent protective effect of salivary pellicle against demineralization of enamel.(ABSTRACT TRUNCATED AT 250 WORDS)

Adsorption of whole saliva onto hydrophilic and hydrophobic solid surfaces: influence of concentration, ionic strength and pH.

The influence of the concentration of salivary proteinaceous material from solutions of whole saliva on the kinetics of in vitro pellicle formation were studied together with the effects of ionic strength, pH and certain substrate characteristics. The pellicle formation was monitored by an automated Rudolph ellipsometer, equipped with a He-Ne laser (wavelength 632.8 nm). The substrates compared in the study were hydrophilic negatively charged silica surfaces and hydrophobic methylated silica surfaces. The results show that the adsorption of salivary proteins is a very rapid process on both types of surfaces. Part of the formed biofilm, however, desorbed upon rinsing, indicating that the proteinaceous material was adsorbed with varying binding strengths. Larger adsorbed amounts were recorded on hydrophobic than on hydrophilic surfaces. Increase of ionic strength caused larger amounts to be adsorbed on both types of surfaces but change of pH did not affect the adsorption on either of the studied surfaces. Ellipsometry was found to be a suitable technique to monitor the adsorption of salivary proteins at solid/liquid interfaces.

[A method for assessing the efficacy of means for interdental space hygiene]. / Sposob otsenki éffektivnosti sredstv gigieny mezhzubnykh promezhutkov.

A method for assessment of the efficacy of various means for maintenance of interdental space hygiene is suggested, based on measurement of the activity of acid producing oral microflora expressed through local pH-metry values. Three types of dental threads were under study.

The hard tissue lesion revisited.

Recent research indicates that the thorough removal of calculus is still a critical step in the treatment of periodontal disease. Surgical access facilitates this in deep pockets. Repair is mediated by a long junctional epithelium as microscopically total calculus removal is rarely achieved. In intrabony defects the apical portion of the root surface which has not attracted deposits of calculus has the potential for connective tissue reattachment and regeneration if care is taken to preserve collagen fibres still attached to the root.

The effect of 2-O-stearoyl-glycerol 1,3-bisphosphate on the demineralization of dental enamel in vitro.

Parts of the buccal and lingual enamel surfaces of sound human third molars were treated with a test compound, and the non-treated parts served as control. After the treatment both parts were demineralized, and the artificial caries lesions were analyzed by means of quantitative microradiography. Treatment during only 1 minute with an aqueous solution of 1.5 mmol/l of the surface active compound 2-O-stearoyl-glycerol 1,3-bisphosphate (Glydip) resulted in a decrease in the rate of enamel demineralization by about 60%. When Glydip was dissolved in a toothpaste/water slurry it inhibited the demineralization by about 30%. When the enamel was covered by a salivary pellicle prior to the application of the slurry, the inhibition of the demineralization disappeared. However, when Glydip was applied in an aqueous solution, the presence of a salivary pellicle did not interfere with the decreasing effect of Glydip on the demineralization. It is concluded that Glydip has promising properties for its use as an anti-caries agent, in combination with fluoride.