Biological considerations of dental materials as orifice barriers for restoring root-filled teeth.

There is ample published literature regarding the technical aspects of restoring root-filled teeth, but little concerning the biological impacts, consequences, and criteria for the selection of direct restorative materials following endodontic treatment. The provision of an effective coronal seal in addition to a sound root filling is known to be important in the prevention of root canal infection. This review seeks to explore the evidence concerning the selection of dental materials in the restoration of root-filled teeth, specifically with a close examination of the properties of commonly used materials as orifice barriers. © 2023 Australian Dental Association.

Oral health research funding in relation to disease burden in Australia.

BACKGROUND: This study aims to investigate and compare the major Australian government research funding schemes for oral health science with other disciplines from the burden of disease perspective. METHODS: Major government research funding scheme outcomes were identified. An innovative index of Fair Research Funding (FRF) was developed to examine the extent to which National Health and Medical Research Council funding is aligned with the disease burden. In addition to comparing different diseases, overall governmental research funding for different areas of oral health sciences was explored. RESULTS: Oral disorders with $15 million NHMRC grant funds (2017-2021) and FRF of 10.7 has the lowest and most inequitable amount of Australian government support in relation to disease burden. The share of oral health science in the Australian Research Council and Medical Research Future Fund was very minimal, with $3.43 and $1.88 million respectively. CONCLUSION: Governmental research funding for oral health sciences is inequitable according to the disease burden. More dedicated oral health sciences research funding schemes are essential. Funding for prevention-focused public oral health programmes is a vital requirement towards reducing the inequalities in population oral health.

Analysing the bioactive makeup of demineralised dentine matrix on bone marrow mesenchymal stem cells for enhanced bone repair.

Dentine matrix has proposed roles for directing mineralised tissue repair in dentine and bone; however, the range of bioactive components in dentine and specific biological effects on bone-derived mesenchymal stem cells (MSCs) in humans are less well understood. The aims of this study were to further elucidate the biological response of MSCs to demineralised dentine matrix (DDM) in enhancing wound repair responses and ascertain key contributing components. Dentine was obtained from human teeth and DDM proteins solubilised with ethylenediaminetetraacetic acid (EDTA). Bone marrow derived MSCs were commercially obtained. Cells with a more immature phenotype were then selected by preferential fibronectin adhesion (FN-BMMSCs) for use in subsequent in vitro assays. DDM at 10 µg/mL reduced cell expansion, attenuated apoptosis and was the minimal concentration capable of inducing osteoblastic differentiation. Enzyme-linked immunosorbent assay (ELISA) quantification of growth factors indicated physiological levels produced the above responses; transforming growth factor ß (TGF-ß1) was predominant (15.6 ng/mg DDM), with relatively lower concentrations of BMP-2, FGF, VEGF and PDGF (6.2-4.7 ng/mg DDM). Fractionation of growth factors from other DDM components by heparin affinity chromatography diminished osteogenic responses. Depletion of biglycan from DDM also attenuated osteogenic potency, which was partially rescued by the isolated biglycan. Decorin depletion from DDM had no influence on osteogenic potency. Collectively, these results demonstrate the potential of DDM for the delivery of physiological levels of growth factors for bone repair processes, and substantiate a role for biglycan as an additional adjuvant for driving osteogenic pathways.

Growth Factor Liberation and DPSC Response Following Dentine Conditioning.

Liberation of the sequestrated bioactive molecules from dentine by the action of applied dental materials has been proposed as an important mechanism in inducing a dentinogenic response in teeth with viable pulps. Although adhesive restorations and dentine-bonding procedures are routinely practiced, clinical protocols to improve pulp protection and dentine regeneration are not currently driven by biological knowledge. This study investigated the effect of dentine (powder and slice) conditioning by etchants/conditioners relevant to adhesive restorative systems on growth factor solubilization and odontoblast-like cell differentiation of human dental pulp progenitor cells (DPSCs). The agents included ethylenediaminetetraacetic acid (EDTA; 10%, pH 7.2), phosphoric acid (37%, pH <1), citric acid (10%, pH 1.5), and polyacrylic acid (25%, pH 3.9). Growth factors were detected in dentine matrix extracts drawn by EDTA, phosphoric acid, and citric acid from powdered dentine. The dentine matrix extracts were shown to be bioactive, capable of stimulating odontogenic/osteogenic differentiation as observed by gene expression and phenotypic changes in DPSCs cultured in monolayer on plastic. Polyacrylic acid failed to solubilize proteins from powdered dentine and was therefore considered ineffective in triggering a growth factor-mediated response in cells. The study went on to investigate the effect of conditioning dentine slices on growth factor liberation and DPSC behavior. Conditioning by EDTA, phosphoric acid, and citric acid exposed growth factors on dentine and triggered an upregulation in genes associated with mineralized differentiation, osteopontin, and alkaline phosphatase in DPSCs cultured on dentine. The cells demonstrated odontoblast-like appearances with elongated bodies and long extracellular processes extending on dentine surface. However, phosphoric acid-treated dentine appeared strikingly less populated with cells, suggesting a detrimental impact on cell attachment and growth when conditioning by this agent. These findings take crucial steps in informing clinical practice on dentine-conditioning protocols as far as treatment of operatively exposed dentine in teeth with vital pulps is concerned.

A novel ex vivo culture model for inflammatory bone destruction.

Pathological alterations in the balance of bone metabolism are central to the progression of inflammatory bone diseases such as periodontal disease. We have developed and characterized a novel ex vivo murine mandible model of inflammatory bone destruction. Slices of mandible were cultured for 14 days in the presence or absence of P. gingivalis lipopolysaccharide (LPS) or pro-inflammatory cytokines. Following culture, cell viability and tissue histomorphometry were assessed with quantification of matrix proteins, resident osteoclasts, ligament cells, monocytes, macrophages, and neutrophils. In the absence of inflammatory factors, culture viability, osteoclasts, and matrix components were maintained. LPS or TNFα stimulation demonstrated an increase in cellular proliferation, monocyte cells, osteoclast differentiation, and matrix degradation. Pathophysiological bone metabolism can be induced via exposure to LPS and direct influence of TNFα within the model despite the absence of systemic circulation, providing a model for inflammatory bone destruction and investigation of the effects of novel therapeutics.

Ex vivo antimicrobial efficacy of strong acid electrolytic water against Enterococcus faecalis biofilm.

AIM: To observe the antimicrobial effect of strong acid electrolytic water (SAEW) against an Enterococcus faecalis (E. faecalis) biofilm when used as a root canal irrigant. METHODOLOGY: The effect of SAEW, sodium hypochlorite (5.25%; NaOCl) and sodium chloride (0.9%; normal saline) on E. faecalis biofilm vitality on coverslips was observed by confocal laser scanning microscopy (CLSM). Thirty-five root canals from extracted human teeth were sterilized prior to contamination with E. faecalis for four weeks. Bacterial samples were collected with sterile paper points and plated onto BHI agar plates for 48 h. Root canal walls were observed by scanning electron microscopy before and after instrumentation, together with root canal irrigation with SAEW, NaOCl or normal saline, with or without ultrasonic vibration. Antimicrobial effectiveness was established by counting colony-forming units and analysed by two-way anova. RESULTS: Confocal laser scanning microscopy revealed that SAEW decreased E. faecalis biofilm vitality, and the proportion of dead bacteria increased in accordance with increasing treatment time. Most bacteria in the biofilms were killed after 10-min treatment. No significant difference was observed between SAEW and NaOCl groups at the same treatment time (P > 0.05) or in the susceptibility of E. faecalis to SAEW and NaOCl (P > 0.05) in extracted human teeth with or without ultrasonic activation. SAEW and NaOCl were more effective against E. faecalis biofilm than normal saline, and antimicrobial efficacy was significantly enhanced by ultrasonic vibration (P < 0.05). CONCLUSIONS: Strong acid electrolytic water effectively killed E. faecalis in a biofilm both on coverslips and in the root canals of extracted human teeth.

An ex vivo culture model for orthodontically induced root resorption.

OBJECTIVES: Root resorption is a ubiquitous although undesirable sequela to orthodontic treatment. Current methods to investigate the pathophysiology have certain limitations. In pursuit to understand and develop treatment modalities for orthodontically induced root resorption, the ability to manipulate cells within their natural extracellular matrix in a three dimensional organotypic model is invaluable. The study aimed to develop a laboratory-based organotypic model to investigate the effect of orthodontic forces on the periodontium. METHODS: Mandibular slices of male Wistar rats were maintained in Trowel-typed cultures at 37°C in 5% carbon dioxide in air for 7 days with test specimens subjected to compressive forces at 50 g and 100g by stainless steel springs. Tissue architecture and cell viability were maintained under culture conditions. RESULTS: Osteoclast numbers increased significantly in both test groups whilst odontoclasts increased in the 50 g group. Immunohistochemistry demonstrated increased dentine sialoprotein expression in both test groups, suggesting changes in mineralization-related activity due to mechanical strain. CONCLUSION: The study showed initial cellular and molecular changes of key markers that relate to root resorption in response to mechanical loading. CLINICAL SIGNIFICANCE: Severe root resorption may occur when forces applied are heavy or transmitted over an extended period and could lead to mobility and tooth loss. This ex vivo model can be used to investigate cellular and molecular processes during orthodontic tooth movement which may advance the clinical management of root resorption.

The effect of low intensity pulsed ultrasound in a 3D ex vivo orthodontic model.

OBJECTIVES: This study investigated the effects of low intensity pulsed ultrasound (LIPUS) on dentoalveolar structures during orthodontic force application using a novel organ culture system. METHODS: Mandibles were dissected from 28-day-old male Sprague Dawley rats, sliced into 1.5mm and cultured at 37°C and 5% CO(2), prior to application of a 50g force to each mandible slice. Slices were randomly divided into three groups of control, 5 and 10min LIPUS application and cultured for five days before histological and histomorphometrical analysis. RESULTS: Cementum and predentine thickness and subodontoblast and periodontal ligament cell counts were increased in the ultrasound groups, with increases statistically significant in the 10min treated groups. Odontoblasts remained viable during LIPUS exposure and osteoclast activity was increased by LIPUS. CONCLUSIONS: LIPUS may influence remodelling of the dentine-pulp complex and associated tissues during orthodontic force application ex vivo.

TGF-beta1 exposure from bone surfaces by chemical treatment modalities.

Growth factors are known to be sequestered to the mineralised matrix of bone. The aim of this study was to investigate the ability of citric acid, EDTA, calcium hydroxide and sodium hydroxide to release active growth factors from bone surfaces, able to promote osteoblast differentiation. All chemical treatments increased surface levels of TGF-beta1 (used as a biomarker of growth factor release), compared to control bone surfaces treated with PBS. Differences were observed in the kinetics of TGF-beta1 exposure at the surface and its subsequent release into the aqueous environment for the different chemical treatments. Surface levels of growth factor following chemical treatment were low, but of sufficient concentration to stimulate cell expansion and osteoblast differentiation of bone marrow stromal cells grown on EDTA and calcium hydroxide treated surfaces compared to PBS treated surfaces. The increased osteogenic potential on these surfaces may relate to an increase in growth factor availability and changes to the surface chemistry and topography.

Hunter-Schreger Band patterns and their implications for clinical dentistry.

Hunter-Schreger Bands (HSBs) are an optical phenomenon visualised when a cut or fractured enamel surface is viewed under reflected light. These bands demonstrate the synchronous decussation of individual or groups of enamel prisms. While the role of HSB patterns has been investigated in comparative anatomical studies, until recently there has been little consideration of HSB patterns in human teeth. The aim of this paper is to consider the significance of HSB patterns in the human dentition and in relation to clinical dentistry. It is concluded that within the human dentition, HSB patterns have evolved to optimise resistance to attrition, abrasion and tooth fracture. It appears that certain aspects of HSB packing densities and distributions have beneficial roles in enamel bonding. Hunter-Schreger Band patterns seem to passively facilitate conditions such as abfraction and cracked tooth syndrome.

Gabriel-Philippe de la Hire and the discovery of Hunter-Schreger bands.

Hunter-Schreger bands are an optical phenomenon observed in mammalian tooth enamel. Familiar to all current and former students of dental histology, this optical phenomenon appears as alternating patterns of dark and light bands when cut enamel is viewed under reflected light. The discovery of this important feature of mammalian enamel has been historically credited to two eighteenth-century investigators, Hunter and Schreger. A re-evaluation of the evidence would suggest that the bands were observed almost seventy years earlier by a French scientist, Gabriel-Philippe de la Hire, and subsequently confirmed by the famous French dentist Pierre Fauchard. This article reviews the contribution of de la Hire, as well as that of Fauchard, Hunter and Schreger, to the early recognition among the scientific community of what would now be referred to as 'enamel microstructure'.

Treatment planning for replacing missing teeth in UK general dental practice: current trends.

The aim of this study was to examine the confidence, barriers and attitudes towards the replacement of missing teeth by general dental practitioners (GDPs). The perceived impact of the recently introduced National Health Service (NHS) contract on the provision of prosthodontic treatments was also considered. Pre-piloted postal questionnaires were mailed to 500 GDPs in Wales. Open- and closed-ended questions were utilised to establish confidence, adequacy of training and attitudes towards treatments for replacing missing teeth. Two hundred and seventeen completed questionnaires were received (response rate = 43.4%). Many respondents described themselves as 'confident' or 'very confident' in the provision of removable partial dentures (RPDs) (acrylic = 100%, metal based = 99.5%), cantilever resin-bonded bridges (94.4%) and conventional bridgework (98.6%). GDPs were 'not confident' providing fixed-fixed resin-bonded bridges (21.1%) or implants (81.4%). Financial barriers were identified to the provision of prosthodontic treatments, including comments such as "the new [National Health Service] contract does limit the treatments available". Privately funded patients were more likely to be offered a fixed bridge or implant replacement of a missing upper first molar, whereas non-privately funded patients were more likely to be offered no treatment (P < 0.01). Most respondents reported confidence at providing more routine forms of prosthodontic care such as RPDs and bridges. It appears that funding arrangements may have an impact on treatments offered to replace missing teeth, particularly under the current NHS contract.

Quality of communication and master impressions for the fabrication of cobalt chromium removable partial dentures in general dental practice in England, Ireland and Wales in 2009.

The aim of this study was to investigate the quality of communication and master impressions for the fabrication of cobalt chromium removable partial dentures (RPDs) in general dental practice in England, Ireland and Wales in 2009. Two hundred and ten questionnaires were distributed to 21 laboratories throughout England, Ireland and Wales. Information was collected regarding the quality of written communication and selection of master impression techniques for cobalt chromium partial dentures in general dental practice. One hundred and forty-four questionnaires were returned (response rate = 68%). Alginate was the most popular impression material being used in 58% of cases (n = 84), while plastic stock trays were the most popular impression tray, being used in 31% of cases (n = 44). Twenty-four per cent (n = 35) of impressions were not adequately disinfected. Opposing casts were provided in 81% of cases (n = 116). Written instructions were described as being 'clear' in 31% of cases (n = 44). In 54% of cases (n = 76), the technician was asked to design the RPD. Based on the findings of this study, written communication for cobalt chromium RPDs by general dental practitioners is inadequate. This finding is in breach of relevant contemporary legal and ethical guidance. There are also concerns in relation to the fabrication process for this form of prosthesis, particularly, in relation to consideration of occlusal schemes.

TGF-beta/extracellular matrix interactions in dentin matrix: a role in regulating sequestration and protection of bioactivity.

TGF-beta isoforms sequestrated in dentin matrix potentially provide a reservoir of bioactive molecules that may influence cell behavior in the dentin-pulp complex following tissue injury. The association of these growth factors with dentin matrix and the influence of such associations on the bioactivity of growth factors are still unclear. We used surface plasmon resonance technology in the BIAcore 3000 system to investigate the binding of TGF-beta isoforms 1 and 3 to purified decorin, biglycan, and EDTA soluble dentin matrix components. TGF-beta isoforms 1 and 3 were immobilized on sensorchips CM4 through amine coupling. For kinetic studies of protein binding, purified decorin and biglycan, isolated EDTA soluble dentin matrix, and dentin matrix immunodepleted of decorin and/or biglycan were injected over TGF-beta isoforms and allowed to interact. Programmed kinetic analysis software provided sensorgrams for each concentration of proteoglycan or dentin matrix extract injected. Purified decorin and biglycan and dentin matrix extract bound to the TGF-beta isoforms. However, the association with TGF-beta3 was much weaker than that with TGF-beta1. After immunoaffinity depletion of the dentin matrix extract, the level of interaction between the dentin matrix extract and TGF-beta was significantly reduced. These results suggest isoform-specific interactions between decorin/biglycan and TGF-beta isoforms 1 and 3, which may explain why TGF-beta3 is not detected in the dentin matrix despite being expressed at higher levels than TGF-beta1 in odontoblasts. These proteoglycans appear to play a significant role in TGF-beta/extracellular matrix interactions and may be important in the sequestration of these growth factors in the dentin matrix.

Quality of prescription and fabrication of single-unit crowns by general dental practitioners in Wales.

The aim of this investigation was to describe the quality of prescription and fabrication of single-unit crowns by general dental practitioners in Wales. One hundred pre-piloted questionnaires were distributed to commercial laboratories in Wales with large catchment areas, and 20 pre-piloted questionnaires were distributed to the production laboratory at the Cardiff Dental Hospital. Information was collected relating to the quality of prescription and master impressions for single-unit crowns. One hundred and seven completed questionnaires were returned (response rate = 89%). Sixty per cent (n = 64) of questionnaires related to single-unit crowns being made in general practice under private funding arrangements, 30% (n = 32) were being made in general dental practice under National Health Service (public) funding arrangements and 10% (n = 11) were collected from the Dental Hospital. Polyvinylsiloxane impression material was used to record the master impression in all cases (n = 107). Plastic stock trays were used to make the master impression in 79% of cases (n = 85), metal stock trays were used in 19% of cases (n = 20) and special trays were used in 2% of cases (n = 2). Eighty-five per cent (n = 91) of master casts were considered to be adequate for crown fabrication. Less than 50% of written instructions (n = 52) were considered 'clear' and of sufficient detail to adequately specify the planned crown. In 21% of cases (n = 22), the technician had to contact the dentist for clarification of the design prior to making the crown. While the quality of impression making for single-unit crowns was of a reasonable standard, the quality of the accompanying written communication was poor and more than one-half of written instructions examined failed to meet the requirements of the European Union Medical Devices Directive.

Stem cells and the dental pulp: potential roles in dentine regeneration and repair.

The dentine-pulp complex displays exquisite regenerative potential in response to injury. The postnatal dental pulp contains a variety of potential progenitor/stem cells, which may participate in dental regeneration. A population of multipotent mesenchymal progenitor cells known as dental pulp stem cells with high proliferative potential for self-renewal has been described and may be important to the regenerative capacity of the tissue. The nature of the progenitor/stem cell populations in the pulp is of importance in understanding their potentialities and development of isolation or recruitment strategies, and allowing exploitation of their use in regeneration and tissue engineering. Various strategies will be required to ensure not only effective isolation of these cells, but also controlled signalling of their differentiation and regulation of secretory behaviour. Characterization of these cells and determination of their potentialities in terms of specificity of regenerative response will form the foundation for development of new clinical treatment modalities, whether involving directed recruitment of the cells and seeding of stem cells at sites of injury for regeneration or use of the stem cells with appropriate scaffolds for tissue engineering solutions. Such approaches will provide an innovative and novel biologically based new generation of clinical treatments for dental disease.

British Orthodontic Society, Chapman Prize Winner 2003. A novel in vitro culture model to investigate the reaction of the dentine-pulp complex to orthodontic force.

OBJECTIVE: To develop a novel mandible slice organ culture model to investigate the effects of externally applied force on the dentine-pulp complex. DESIGN: In vitro organ culture. SETTING: School of Dentistry, Birmingham, UK. MATERIALS AND METHODS: Transverse 2 mm thick sections were cut from the mandibles of five 28-day-old male Wistar rats. Serial sections were used for control and test pairs. Springs made from 0.016-inch and 0.019 x 0.025-inch stainless steel wires were used to apply a 50 g tensile or compressive force, respectively, to test specimens. Control and test specimens were cultured for 5 days in a humidified incubator with 5% CO(2) at 37 degrees C and processed for routine histological investigation. Nine more rats were used to provide control and compression test pairs where the pulps were extirpated after 3 days culture and total RNA isolated for gene expression analysis by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Histology showed the dental and supporting tissues maintained a healthy appearance in the control cultures after culture. Histomorphometric analysis revealed a 20-27% increase in pulp fibroblast density in test specimens compared with controls. Gene expression analyses revealed up-regulation in the test groups of PCNA, c-Myc, Collagen 1alpha, TGF-beta1 and alkaline phosphatase, whilst expression of osteocalcin was reduced. CONCLUSIONS: The results demonstrated that the present organ culture technique provides a valuable in vitro experimental model for studying the effects of externally applied forces. These forces stimulated a cellular response in the pulp chamber characterized by altered gene expression and proliferation of fibroblasts; the latter being unaffected by the nature of the force in terms of compression or tension.

Fluoride-induced changes to proteoglycan structure synthesised within the dentine-pulp complex in vitro.

Fluoride is known to influence mineralisation patterns within dentine, where alterations in the post-translational modification of proteoglycans (PG) have been proposed as an implicating factor. In light of recent studies elucidating changing PG profiles in the transition of predentine to mineralised dentine, this study investigates the influence of fluoride on the major PG populations (decorin, biglycan and versican) within the pulp, predentine and dentine. Tooth sections from rat incisors were cultured for 14 days in the presence 0, 1 and 6 mM sodium fluoride and the PG extracted from the pulp, predentine and dentine matrices. PG species and corresponding metabolites were identified by their immuno-reactivity to antibodies against decorin, biglycan and versican. Component glycosaminoglycan chains were characterised with respect to their nature, chain length and disaccharide composition. Levels of PG extracted from pulp and predentine were reduced, particularly for biglycan. Fluoride did not influence levels of decorin or versican within predentine or dentine, although the processing of these macromolecules within pulp and predentine was affected, particularly at higher fluoride concentrations. Levels of dermatan sulfate were reduced within pulp and predentine, although the effect was less pronounced for predentine. Fluoride reduced sulfation of glycosaminoglycan chains within pulp and predentine tissues, with a notable reduction in Deltadi6S evident. In all three tissues, glycosaminoglycan chain length was reduced. Considering the various roles for PG in the dentine-pulp complex, either directly or indirectly in the mineralisation process, changes in the synthesis, structure and processing of the different PG species within the pulp, predentine and dentine matrices provides a further molecular explanation for the altered mineralisation patterns witnessed during fluorosis.

The influence of fluoride exposure on dentin mineralization using an in vitro organ culture model.

This study aimed to characterize fluoride-induced alterations in dentin mineralization within a dentin-pulp organ culture system. Tooth sections derived from male Wistar rat incisors were cultured in Trowel-type culture for 14 days, in the presence of 0 mM, 1 mM, 3 mM and 6 mM sodium fluoride. Tooth sections were processed and analyzed for uptake of fluoride, its subsequent effect on dentin mineralization by tetracycline hydrochloride incorporation and mineral composition, expressed as calcium/phosphorous (Ca/P) ratios. Tetracycline hydrochloride incorporation was demonstrated to decrease with increased fluoride exposure, accompanied by significant increases in both Ca/P ratios and fluoride incorporation. These findings provide further evidence that the established alterations in dentin formation during fluorosis are a consequence of disruption to the mineralization process, and provide a model system with which to investigate further the potential role the extracellular matrix plays in inducing the apparent changes in mineral composition.

The influence of fluoride on the cellular morphology and synthetic activity of the rat dentine-pulp complex in vitro.

Exposure to high fluoride concentrations in the immediate environment of the tissue is recognized to result in the post-translational modification of non-collagenous dentine extracellular matrix (ECM) components, potentially altering dentine mineralization. However, less is known about the effects of fluoride exposure on the morphology or metabolism of the cells associated with the dentine-pulp complex. This study examined the effects of fluoride exposure at defined concentrations on the cellular morphology and ECM synthetic activities of odontoblasts and pulpal fibroblasts by the culture of tooth sections from male Wistar rat incisors in Trowel-type cultures for up to 14 days, in the presence and absence of 6mM sodium fluoride. Histomorphometric analysis of the dentine-pulp complex of sodium fluoride-exposed tooth sections demonstrated no obvious gross morphological differences with respect to the odontoblasts and pulpal fibroblasts throughout the 14-day culture period, in comparison with unexposed tooth sections. No significant differences in odontoblast and pulpal fibroblast cell numbers were determined in the absence and presence of fluoride. Image analysis examination of odontoblast cytoplasmic:nuclear (C/N) ratios also showed no significant differences in fluoride-exposed and unexposed tooth sections, although reductions in the C/N ratios of pulpal fibroblasts were evident in fluoride-exposed sections at days 10 and 14. No significant differences in predentine width were observed in fluoride-exposed and unexposed tooth sections over the 14-day culture period. Autoradiography following [3H]proline incorporation into the dentine-pulp complex demonstrated inhibition of collagen synthesis, particularly by the odontoblasts in tooth sections exposed to 6mM sodium fluoride. These findings, in association with those from previous studies, imply that dentine ECM alterations may contribute to the altered mineralization of dentine during fluorosis, rather than secretory-related changes in odontoblast morphology.