Study On The Protective Effect Of Epigallocatechin Gallate On The Demineralized Dentin Matrix

Integrate dentin structure plays an important role in maintaining both a healthy toothand oral enviroment. Different from the enamel, dentin is more prone to an quick andprogressive damage under the attacks from oral bacteria, enzymes and various mechanicaland chemical stimulations. Dentin collagens is the net framework, which maintaines thedentin structure and prevents the progress of various tooth tissue diseases. Under thecondition of various pathological situations, such as dental carious, dentin erosion etc, thedentin could be demineralized and the collagen matrix be exposed. These naked andunprotected dentin collagens can be attacked by the matrix metalloproteinases (MMPs)that originated from dentin or saliva, being degraded and inducing the progressive damageof dentin. It has been considered that the collagen degradation that induced by theproteinases as an important endogenous factor. Therefore, the usage of MMPs inhibitor on the dentin surfaces should be an effective solution to protect the dentin matrix.Epigallocatechin gallate is a nature origin MMPs inhibitor which has showed an inhibitoryeffect on the activity of MMP-2, MMP-9from tumor cells. The inhibit effect of EGCG should bedue to its combination with the collagen by hydrogen or hydrophobic bond, which induce themodification of the secondary structure of collagenases. However, it is still unknown that whetherEGCG can effectively inhibit the activities of MMPs in dentin and protect the dentin matrix bypreventing collagen degradation. This is a quite meaningful research project and needed to befurther studied.Therefore, we designed three sections of experiments in the present study. In the first section,we detected the effect of EGCG on the activity of exogenous MMP-8and dentinal MMPs usingMicroplate reader to ensure whether EGCG can effectively inhibit the activities of the proteasesrelated to dentin collagen degradation. In the second section, a PH cycling with the term of6daysand twice for each day was applied to imitate the etching stimulation during the dentinal cariesprocess. In order to study the effect of EGCG treatment on the anti-demineralization capability ofdentin, the SEM and CLSM were applied to detect the untra-structure of dentin surfaces and thedemineralization pattern respectively, as well as the surface microhardness and rate ofdecalcification of dentin to analyze the demineralization degree. In the third section, bothlong-term artificial salvia storage and thermocycling treatment were applied to imitate the agingchallenge in oral environment. Using microtensile bonding strength test and the failure modesdetection, we try to find whether EGCG pretreatment can stabilize the dentinal bonding interfacesregardless of the adhesive type (both total-etching system and self-etching system). The followingresults could be obtained:1. The use of EGCG with concentration up to200μg/ml can significantly inhibit the activity ofexogenous MMP-8within the observing term of48hrs, which showed no significance withthe results of the two positive controls:0.2%CHX and0.4mM1-10phenanthroline. Theinhibiting effect increased with the increase of EGCG concentration.2. Within the observing term of48hrs, the use of EGCG can significantly inhibit the activities of dentinal MMPs and the inhibitory effect was concentration dependent. With the concentrationup to400μg/ml the inhibitory effect of EGCG was better than that of0.2%CHX-the positivecontrol.3. All the tested dentin showed demineralizations to different degree after PH cycling aging. Thegroup treated with400μg/ml EGCG showed a similar pattern with those of the two positivecontrols (treatments with0.2%CHX and1mgNaF), which is inferior than that of the negativecontrol treated with deionized water. In the three experimental groups, the dentin1surfacespresented a low degree of demineralization, with undetectable demineralization for both theintertubular dentin and the peritubular dentin, as well as the deposition of mineral particlesinside the tubules.4. The calcium depletion rate and the reducing surface microhardness of group with EGCGtreatment presented an inferior dentin demineralization in both the dentin decalcificationquantities and the decrease number of surface microhardness than that of the negative controlgroup (P<0.05). The dentin demineralization in EGCG group had no significant differencewith those of the two positive groups (treatments with0.2%CHX and1mgNaF, P>0.05).5. Concerning the total-etching adhesive system Adper Single Bond2, the pretreatment with400μg/ml EGCG after the acid etching resulted in a significantly higher bond strength than ofthe negative control group after the artificial saliva storage for6months(P<0.05), which wasnot significantly different with positive control group that pretreated with0.2%CHX (P>0.05)6. Concerning the self-etching adhesive system Clearfil SE Bond, the treatment with400μg/mlEGCG between the SE Primer and SE Bond resulted in a significantly higher bond strengththan of the negative control group after the thermocyclng challenge for5000times (P<0.05),which was not significantly different with positive control group that pretreated with0.2%CHX (P>0.05).