Preliminary Studies On The Improvement Of The Durability Of Etch-and-rinse Dentin Bonds By Transient Proanthocyanidins Preconditioning

Over the past decades, the immediate bonding efficacy has been greatlyimproved with the the introduction of the wet bonding technique and continuingadvances in bonding systems. However, the long-term bonding durability of theadhesive restorations is not satisfactory, due to the particularity of dentinstructures and the complexity of oral environment.Recently, many experimental strategies have been proposed to prolong thelifespan of tooth-colored resin composites, including the use of hydrophobicadhesive, antibacterial adhesive, ethanol-wet bonding technique, inhibitors ofcollagenolytic enzymes and biomimetic remineralization. However, eachstrategy has its merits and limitations, and no specific treatment approach mayovercome the critical barriers currently encountered in dentin bonding.As the major component of the dentin organic matrix, type I collagen is aheterotrimeric molecule composed of two1chains and one2chain.Intermolecular cross-links are the basis for the stability, tensile strength, andviscoelasticity of the collagen fibrils. It has been demonstrated that the application of exogenous cross-linking agents to several connective tissues ishelpful to modify the structures of collagen fibrils, and improve theirdegradation resistance as well as stabilization. Proanthocyanidins （PA）, as a typeof natural cross-linking agents, is superior to synthetic crosslinkers because ofits good biocompatibility and moderate reaction rate, and has received anincreasing attention. Therefore, attempts have been made to biomodifydemineralized dentin collagen by PA in the present study. PA-based agents wereprepared in different polar solvents, and preconditioning time was reduced toevaluate the effect of PA preconditioning on demineralized dentin matrix,durability of resin-dentin bonds, using a more clinical relevant procedure. Also,the interaction mechanism between PA and dentin was explored in the presentstudy, in expectation of providing a new strategy for the improvement of dentinbonding durability.PART I. Morphologic and physiochemical properties of demineralizeddentin matrix treated with PAPA-based preconditioners with different concentrations （5%,10%,15%）were prepared by adding powdered grape seed extract enrich ofproanthocyanidins to various solvents [distilled water （DW）, anhydrous ethanol（ET） or acetone （AC）]. Demineralized dentin specimens were preconditioned for20s,30s,60s or120s, followed by the evaluation of their cross-linkingdegrees, mechanical properties and micromorphologies. The cross-linkingdegree and ultimate tensile strength of the demineralized dentin collagenexhibited concentration-and time-dependent increase after preconditioning.Specimens pretreated by10%or15%PA preconditioner for more than30spresented significantly higher cross-linking degree and ultimate tensile strengths than those of the blank controls （preconditioned solely by solvents, not withPA）, irrespective of the solvent （P <0.05）. The field emission scanning electronmicroscopy （FESEM） confirmed that the demineralized dentin collagen was in ahomogeneous and regular arrangement after preconditioning and maintainedexpanding, regardless of the surface moisture conditions. There are somedifferences among groups pretreated by PA preconditioners in differentsolvents, which suggested that the solvents may have influence on theperformance of PA.PART II. The effect of transient PA preconditioning on the immediateproperties of resin-dentin bondsTwo commercial dental adhesive systems, Adper^TMSingle Bond2（SB,water/ethanol-based） and Prime&Bond NT （NT, water/acetone-based） wereused in the present study. PA-based preconditioners were applied ondemineralized dentin for120s before the application of the bonding agent.Microtensile bond strength （μTBS）, failure modes and degree of conversionwere evaluated via microtensile testing, FESEM and Fourier transform infraredspectroscopy, respectively. It was shown that the μTBS of SB was improvedafter cross-linking treatment and presented concentration-dependent increase.10%and15%PA groups （in either solvent） displayed significantly higher μTBSthan non-treated controls （P <0.05）. When using NT, only15%PA-DW and10%PA-AC groups presented statistically higher μTBS （P <0.05）. Therefore,SB adhesive system was chosen in microtensile testing of specimens pretreatedby PA in reduced time （60s or30s）, to evaluate the clinical feasibility of PApreconditioners. It has been proved from the results that15%PApreconditioning improved μTBS even in reduced, clinically applicable treatmentduration （60s）（P <0.05）, whilst no statistically significant differences were found in μTBS between the shortest pretreated group and non-treated controls（P>0.05）. The degrees of conversion were not significantly affected by PApretreatment, which indicated that PA pretreatment had no adverse effects on thecuring characteristics of the test adhesive systems.PART III. The effect of transient PA preconditioning on the durabilityof resin-dentin bondsBased on the previous resμlts, the preferred parameters （concentration:10%,15%; solvent: ethanol） of PA-based preconditioner were screened out, withwhich the immediate μTBS was significantly higher than that of the negativecontrol. The μTBS values, the amount of collagen degradation and nanoleakagewere evaluated to investigate the potential effect of PA on the stability ofresin-dentin bonds against thermal cycling and MMPs degradation. It wasshown that the μTBS values of all groups were decreased afterdegradation,while PA-pretreated groups still exhibited significantly higher μTBSthan non-treated controls, especially specimens pretreated by15%PApreconditioner for120s （P <0.05）. The biodegradation rates and the amount ofnanoleakage of PA-pretreated groups were also significantly lower thannon-treated controls （P <0.05）. Specimens with PA pretreatment presentedcompact hybrid layer even after degradation, while some narrow gaps werefound in hybrid layer of non-treated specimens. It was suggested that PApretreatment may have some protective effect on the resin-dentin bondinginterface against thermal cycling and enzymatic degradation.PART IV. Exploration of mechanism of PA-dentin interactionRaman spectroscopy was used to investigate the mechanism of PA-dentininteraction. In addition, it was examine whether transient PA pretreatment was beneficial for the enhancement of dentin resistance against collagenolysis byboth MMPs and sodium hypochlorite. It was found that PA pretreatment todemineralized dentin induced more crosslinks of collagen mirofibrils, inhibitedgelatinolytic/collagenolytic activities and reducing the susceptibility of dentincollagen to sodium hypochlorite degradation. The results suggested that PA canact in several ways altogether to improve the stability of rensin-dentin bondinginterface.To sum up, PA pretreament may be a potential strategy to increase thedurability of etch-and-rinse dentin bonds.