Structural and biomechanical changes to dentin extracellular matrix following chemical removal of proteoglycans.

Proteoglycans are biomacromolecules with significant biomineralization and structural roles in the dentin extracellular matrix. This study comprehensively assessed the mechanical properties and morphology of the dentin extracellular matrix following chemical removal of proteoglycans to elucidate the structural roles of proteoglycans in dentin. Dentin extracellular matrix was prepared from extracted teeth after complete tissue demineralization. Chemical removal of proteoglycans was carried-out using guanidine hydrochloride for up to 10 days. The removal of proteoglycans was determined by dimethylmethylene blue colorimetric assay and histological staining analyses using transmission electron microscopy and optical microscopy. The modulus of elasticity of dentin matrix was determined by a 3-point bending test method. Partial removal of proteoglycans induced significant modifications to the dentin matrix, particularly to type I collagen. Removal of proteoglycans significantly decreased the modulus of elasticity of dentin extracellular matrix (p < 0.0001). In conclusion, the subtle disruption of proteoglycans induces pronounced changes to the collagen network packing and the bulk modulus of elasticity of dentin matrix.

Influence of intraoral temperature and relative humidity on the dentin bond strength: an in situ study.

STATEMENT OF THE PROBLEM: The effect of the intraoral environment during adhesive restorative procedures remains a concern, especially in the absence of rubber dam isolation. OBJECTIVE: To evaluate the temperature and relative humidity (RH) at anterior and posterior intraoral sites and their effects on the dentin bond strength of two-step etch-and-rinse adhesive systems. METHODS: Sixty human molars were assigned to six groups according to the adhesive systems (Adper Single Bond Plus and One Step Plus) and intraoral sites (incisor and molar sites). The room condition was used as a control group. Dentin fragments were individually placed in custom-made acetate trays and direct composite restorations were performed. The intraoral temperature and RH were recorded during adhesive procedures. Then, specimens were removed from the acetate trays and sectioned to obtain multiple beams for the microtensile bond strength test. In addition, the adhesive interface morphology was evaluated through scanning electron microscopy. Intraoral conditions were statistically analyzed by paired Students' t-tests and the bond strength data by two-way analysis of variance and Tukey test (α = 0.05). RESULTS: The posterior intraoral site showed a significant increase in the temperature and RH when compared with the anterior site. However, both intraoral sites revealed higher temperatures and RH than the room condition. In regards to the adhesive systems, the intraoral environment did not affect the bond strength, and the One Step Plus system showed the highest bond strength means. CONCLUSION: Despite the fact that remarkable changes in the intraoral conditions were observed for both anterior and posterior sites, the intraoral environment was not able to compromise the immediate dentin bond strength. CLINICAL SIGNIFICANCE: Some conditions of intraoral temperature and relative humidity may not impair the dentin bond strength of two-step etch-and-rinse adhesive systems. Thus, an adequate relative isolation seems to be a good alternative under the specific clinical conditions in which rubber dam isolation is either impossible or very difficult to perform.

The effect of photopolymerization on the degree of conversion, polymerization kinetic, biaxial flexure strength, and modulus of self-adhesive resin cements.

STATEMENT OF PROBLEM: Understanding the effect of the degree of conversion on the mechanical properties of auto- and dual-polymerizing self-adhesive resin cements leads to a better estimation of their performance in different clinical scenarios. PURPOSE: The purpose of this study was to evaluate the effect of photopolymerization on the degree of conversion (DC) and polymerization kinetic of 4 dual-polymerized resin cements, 20 minutes after mixing, and its effects on the mechanical properties (biaxial flexural strength [FS] and modulus [FM]) after short-term aging. MATERIAL AND METHODS: Conventional (RelyX ARC and Clearfil Esthetic Cement) and self-adhesive resin cements (RelyX Unicem and Clearfil SA Cement) were applied to a Fourier infrared spectrometer to assess the DC (n=5) under the following 3 polymerization conditions: direct light exposure (dual-polymerizing mode), exposure through the prepolymerized disk, or autopolymerizing. The polymerization kinetic was recorded for 20 minutes. Then, disk-shaped specimens (n=11) were prepared to evaluate the effect of polymerization on the FS and FM in both extreme polymerization conditions (dual-polymerizing or autopolymerizing). Data were statistically analyzed by 2-way repeated measure ANOVA (DC) and by 2-way ANOVA (FS and FM), followed by the Tukey-Kramer post hoc test (α=.05). RESULTS: Autopolymerizing groups exhibited reduced DC means, whereas intermediate values were observed when resin cements were polymerized through the disk. All groups exhibited higher DC at the end of 20 minutes. The polymerization kinetic revealed a rising curve, and materials, when directly photopolymerized, reached a plateau immediately after light exposure. Regarding the flexural biaxial testing, most of the resin cements were affected by polymerization mode and differences among groups were product dependent. CONCLUSIONS: The resin cements achieved immediate higher DC and mechanical properties when photopolymerized. The total absence of photoactivation may still impair their mechanical properties even after short-term aging.

Mimicking the hierarchical functions of dentin collagen cross-links with plant derived phenols and phenolic acids.

Proanthocyanidins (PACs) are secondary plant metabolites that mediate nonenzymatic collagen cross-linking and enhance the properties of collagen based tissue, such as dentin. The extent and nature of cross-linking is influenced by the composition and specific chemical structure of the bioactive compounds present in certain PAC-rich extracts. This study investigated the effect of the molecular weight and stereochemistry of polyphenol compounds on two important properties of dentin, biomechanics, and biostability. For that, purified phenols, a phenolic acid, and some of its derivatives were selected: PAC dimers (A1, A2, B1, and B2) and a trimer (C1), gallic acid (Ga), its esters methyl-gallate (MGa) and propyl-gallate (PGa), and a pentagalloyl ester of glucose (PGG). Synergism was assessed by combining the most active PAC and gallic acid derivative. Mechanical properties of dentin organic matrix were determined by the modulus of elasticity obtained in a flexural test. Biostability was evaluated by the resistance to collagenase degradation. PACs significantly enhanced dentin mechanical properties and decreased collagen digestion. Among the gallic acid derivatives, only PGG had a significant enhancing effect. The lack of observed C1:PGG synergy indicates that both compounds have similar mechanisms of interaction with the dentin matrix. These findings reveal that the molecular weight of polyphenols have a determinant effect on their interaction with type I collagen and modulates the mechanism of cross-linking at the molecular, intermolecular, and inter-microfibrillar levels.

Effect of storage times and mechanical load cycling on dentin bond strength of conventional and self-adhesive resin luting cements.

STATEMENT OF PROBLEM: The lack of long-term bond stability between resin cements and dentin may compromise the success of indirect restorations. PURPOSE: The purpose of this study was to evaluate the effects of long-term storage in artificial saliva and mechanical load cycling on the microtensile bond strength of conventional and self-adhesive resin cements to dentin. MATERIAL AND METHODS: The occlusal dentin surfaces of 128 human molars were exposed and flattened. The teeth were assigned to 16 groups (n=8) according to resin cement and in vitro aging strategy. Two self-adhesive resin cements (RelyX Unicem and Clearfil SA Cement) and 2 conventional cementing systems (RelyX ARC and Clearfil Esthetic Cement) were used. Resin cements were applied to prepolymerized indirect resin disks, which were bonded to the dentin surfaces and light polymerized. The control groups were represented by immediate microtensile bond strength (24 hours) and aging methods were performed with mechanical load cycling or storage in artificial saliva (1 year and 2 years). Bonded beams were tested in tension until failure. Data (MPa) were analyzed by Proc Mixed for repeated measures and the Tukey-Kramer test (α=.05). RESULTS: The self-adhesive resin cements exhibited higher microtensile bond strength than conventional cementing systems for all conditions studied. The microtensile bond strength of RelyX ARC and self-adhesive resin cements did not decrease after storage in artificial saliva and mechanical load cycling. The Clearfil Esthetic Cement showed the lowest microtensile bond strength and a significant reduction after 2 years of storage in artificial saliva. CONCLUSIONS: The storage times and mechanical load cycling did not affect the microtensile bond strength of self-adhesives and RelyX ARC resin cements. The highest microtensile bond strength was obtained for self-adhesive resin cements, with no significant difference between them.

Cigarette smoke: effects on water sorption and solubility of restorative dental composites.

Although scientific evidence has shown the effects of tobacco on changes in the color of composite resins, the association between tobacco exposure and the physical properties of composite resins has not been thoroughly investigated. The purpose of this study was to evaluate the effects of cigarette smoke products on water sorption and solubility of microfilled, microhybrid, and nanofilled composite resins (Durafill VS, Filtek Z250, and Filtek Z350 XT, respectively). Ten discs were prepared of each material and divided into 2 groups (n =5), according to cigarette smoke exposure. Specimens were first desiccated until a constant mass was obtained (M1). Then half of the samples were immersed in deionized water while the other half were exposed daily to tobacco smoke, then washed and stored in deionized water. After 21 days, the resin discs were measured (M2) and placed in desiccators until constant mass was achieved (M3). Water sorption and solubility were calculated and the data was statistically analyzed. Water sorption revealed significant differences among the composite resins. The Filtek Z350 XT exhibited the highest water sorption, followed by Durafill VS and Filtek Z250. Cigarette smoke significantly increased water sorption for all products, but only the solubility of Durafill VS showed a significant difference. Filtek Z250 demonstrated significantly lower solubility than Durafill VS, and Filtek Z350 XT had intermediate values. These results indicated that water sorption and solubility varied among the products, and tobacco smoke may alter the physical properties of resin-based materials.

24 hours and 3-months bond strength between dual-cured resin cements and simplified adhesive systems.

This study evaluated the bonding compatibility between dual-cured resin cements and simplified adhesive systems (one-step self-etch and two-step etch & rinse), measured after 24 hours and 3 months. The occlusal dentin surfaces of 24 human third molars were exposed and flattened. Teeth were randomly assigned to 3 groups and treated with different combinations of adhesive system and resin cement [G1-Single Bond/Rely X ARC (SB/RX); G2-Excite DSC/Variolink II (EX/VR); G3-Adper Prompt/Rely X ARC (AD/RX)]. Indirect composite restorations were cemented on flattened surfaces, and sectioned to obtain multiple bonded beams for the microtensile bond strength test. The beams from each tooth were tested under tension after 24 hours and 3 months (ANOVA/ Tukey's test, alpha=5%). Failure patterns were evaluated with scanning electron microscopy. After 24h, AD/RX presented the lowest bond strength mean values. AD/RX specimens did not withstand three months storage. SB/RX and EX/VR presented similar bond strengths in both periods tested. The association AD/RX resulted in low bond strength mean values, especially after storage. Cementing indirect restorations using one-step self-etch adhesive systems and dual-cured resin cements would be clinically unreliable.

Evidence to the role of interflavan linkages and galloylation of proanthocyanidins at sustaining long-term dentin biomodification.

OBJECTIVES: The interactivity of proanthocyanidins (PACs) with collagen modulates dentin matrix biomechanics and biostability. Herein, PAC extracts selected based on structural diversity were investigated to determine key PAC features driving sustained effects on dentin matrices over a period of 18months. METHODS: The chemical profiles of PAC-rich plant sources, Pinus massoniana (PM), Cinnamomum verum (CV) and Hamamelis virginiana (HV) barks, as well as Vitis vinifera (VV) seeds, were obtained by diol HPLC analysis after partitioning of the extracts between methyl acetate and water. Dentin matrices (n=15) were prepared from human molars to determine the apparent modulus of elasticity over 18months of aging. Susceptibility of the dentin matrix to degradation by endogenous and exogenous proteases was determined by presence of solubilized collagen in supernatant, and resistance to degradation by bacterial collagenase, respectively. Data were analyzed using ANOVA and Games-Howell post hoc tests (α=0.05). RESULTS: After 18months, dentin matrices modified by PM and CV extracts, containing only non-galloylated PACs, were highly stable mechanically (p<0.05). Dentin matrices treated with CV exhibited the lowest degradation by bacterial collagenase after 1h and 18months of aging (p<0.05), while dentin matrices treated with PM showed the least mass loss and collagen solubilization by endogenous enzymes over time (p<0.05). SIGNIFICANCE: Resistance against long-term degradation was observed for all experimental groups; however, the most potent and long-lasting dentin biomodification resulted from non-galloylated PACs.

Galloyl moieties enhance the dentin biomodification potential of plant-derived catechins.

Proanthocyanidin-rich plant-derived agents have been shown to enhance dentin biomechanical properties and resistance to collagenase degradation. This study systematically investigated the interaction of chemically well-defined monomeric catechins with dentin extracellular matrix components by evaluating dentin mechanical properties as well as activities of matrix metalloproteinases (MMPs) and cysteine-cathepsins (CTs). Demineralized dentin beams (n=15) were incubated for 1h with 0.65% (+)-catechin (C), (-)-catechin gallate (CG), (-)-gallocatechin gallate (GCG), (-)-epicatechin (EC), (-)-epicatechin gallate (ECG), (-)-epigallocatechin (EGC) and (-)-epigallocatechin-3-gallate (EGCG). The modulus of elasticity (E) and the fold increase in E were determined by comparing specimens at baseline and after treatment. Biodegradation rates were assessed by differences in percentage of dry mass before and after incubation with bacterial collagenase. The inhibition of MMP-9 and CT-B by 0.65, 0.065 and 0.0065% of each catechin was determined using fluorimetric proteolytic assay kits. All monomeric catechins led to a significant increase in E. EGCG showed the highest fold increase in E, followed by ECG, CG and GCG. EGCG, ECG, GCG and CG significantly lowered biodegradation rates and inhibited both MMP-9 and CT-B at a concentration of 0.65%. Overall, the 3-O-galloylated monomeric catechins are clearly more potent than their non-galloylated analogues in improving dentin mechanical properties, stabilizing collagen against proteolytic degradation, and inhibiting the activity of MMPs and CTs. The results indicate that galloylation is a key pharmacophore in the monomeric and likely also in the oligomeric proanthocyanidins that exhibit high cross-linking potential for dentin extracellular matrix.

Dentin biomodification: strategies, renewable resources and clinical applications.

OBJECTIVES: The biomodification of dentin is a biomimetic approach, mediated by bioactive agents, to enhance and reinforce the dentin by locally altering the biochemistry and biomechanical properties. This review provides an overview of key dentin matrix components, targeting effects of biomodification strategies, the chemistry of renewable natural sources, and current research on their potential clinical applications. METHODS: The PubMed database and collected literature were used as a resource for peer-reviewed articles to highlight the topics of dentin hierarchical structure, biomodification agents, and laboratorial investigations of their clinical applications. In addition, new data is presented on laboratorial methods for the standardization of proanthocyanidin-rich preparations as a renewable source of plant-derived biomodification agents. RESULTS: Biomodification agents can be categorized as physical methods and chemical agents. Synthetic and naturally occurring chemical strategies present distinctive mechanism of interaction with the tissue. Initially thought to be driven only by inter- or intra-molecular collagen induced non-enzymatic cross-linking, multiple interactions with other dentin components are fundamental for the long-term biomechanics and biostability of the tissue. Oligomeric proanthocyanidins show promising bioactivity, and their chemical complexity requires systematic evaluation of the active compounds to produce a fully standardized intervention material from renewable resource, prior to their detailed clinical evaluation. SIGNIFICANCE: Understanding the hierarchical structure of dentin and the targeting effect of the bioactive compounds will establish their use in both dentin-biomaterials interface and caries management.

24 hours and 3-months bond strength between dual-cured resin cements and simplified adhesive systems / Avaliação da resistência de união de cimentos resinosos de dupla polimerização associados a sistemas adesivos simplificados após 24 horas e 3 meses

This study evaluated the bonding compatibility between dualcuredresin cements and simplified adhesive systems (one-step self-etch and two-step etch & rinse), measured after 24 hours and 3 months. The occlusal dentin surfaces of 24 human third molars were exposed and flattened. Teeth were randomly assigned to 3 groups and treated with different combinations ofadhesive system and resin cement [G1 - Single Bond/Rely X ARC (SB/RX); G2 - Excite DSC/Variolink II (EX/VR); G3 -Adper Prompt/Rely X ARC (AD/RX)]. Indirect composite restorations were cemented on flattened surfaces, and sectioned to obtain multiple bonded beams for the microtensile bond strength test. The beams from each tooth were tested under tension after 24 hours and 3 months (ANOVA/ Tukey’s test, α=5%). Failure patterns were evaluated with scanning electron microscopy. After 24h, AD/RX presented the lowest bond strength mean values. AD/RX specimens did not withstandthree months storage. SB/RX and EX/VR presented similar bond strengths in both periods tested. The associationAD/RX resulted in low bond strength mean values, especially after storage. Cementing indirect restorations using one-step self-etch adhesive systems and dual-cured resin cements wouldbe clinically unreliable.

Este estudo avaliou a compatibilidade de união entre cimentos resinosos de dupla polimerização e sistemas adesivos simplificados (adesivo autocondicionante - 1 passo- e adesivo total-each - 2 passos), após 24h e 3 meses. A superfície oclusal de 24 terceiros molares humanos foi exposta e planificada. Os dentes foram aleatoriamente divididos em 3 grupos usados de acordo a seguinte combinação entre sistema adesivo/cimento resinoso [G1 - Single Bond/Rely X ARC (SB/RX); G2 - Excite DSC/Variolink II (EX/VR); G3-Adper Prompt/Rely X ARC (AD/RX)]. Restaurações indiretas em resina composta foram cimentadas na superfície dentinária, e seccionados em múltiplos palitos para a realização do teste de resistência de união através do ensaio de microtração. Os palitos obtidos por dente foram submetidos ao teste após 24 horas e 3 meses (ANOVA/ Tukey test, α=5%). O padrão de fraturafoi observado no microscópio eletrônico de varredura. Após 24 horas, AD/RX apresentou menor valor médio de resistência de união. Os corpos-de-prova do grupo AD/RX não resistiram aoarmazenamento de 3 meses. SB/RX e EX/VR demonstraram similarresistência de união em ambos os períodos avaliados. A associação AD/RX resultou em menores valores médios deresistência de união, especialmente após o armazenamento. A cimentação de restaurações indiretas utilizando sistemas autocondicionantes (1 passo) e cimentos resinosos de dupla polimerização não foi considerada clinicamente confiável.