Search

Objectives This study evaluated the endogenous matrix metalloproteinase (MMP) activity of demineralized dentin matrix following 1 or 5 min pretreatment by various collagen crosslinkers. Generic MMP activity assay, total protein analysis, in situ zymography, gelatin zymography and multiplex bead technology were used to evaluate matrix-bound MMP activity. Methods Six different crosslinkers; glutaraldehyde, riboflavin/UVA, riboflavin-5-monophospate/UVA, sumac berry extract, grape seed extract, and curcumin were used. Demineralized dentin beams were pretreated with respective crosslinkers for 1 or 5 min. Demineralized dentin beams with no crosslinker pretreatment served as control. The reduction in the total activity of dentin matrices were measured using generic MMP activity assay. Dentin slabs were used for in situ zymography and evaluated by using hydrolysis of self-quenched fluorescein-conjugated gelatin under confocal microscopy. Dentin beam extracts were used for total protein assay and multiplex analysis and powder extracts were used for gelatin zymography. Results MMP activity in crosslinker pretreated samples decreased significantly between 21% and 70%, whereas untreated control samples’ activity increased up to 84%. Zymograms confirmed a decrease in the gelatinolytic activity and in the amount of extractable total protein content. Multiplex analysis of extracts of crosslinker-treated dentin showed a reduction in the MMP-8, MMP-2 and MMP-9 release. Significance The result of this work suggests that the effect of the crosslinkers is source-dependent. The use of crosslinkers for as little as 1 min on demineralized dentin can inactivate the endogenous protease activity of dentin matrices. [ABSTRACT FROM AUTHOR]

Objectives This study evaluated the effect of dentin pretreatment with collagen crosslinkers on matrix metalloproteinase (MMP) and cathepsin K mediated collagen degradation. Methods Dentin beams (1 mm × 2 mm × 6 mm) were demineralized in 10% H 3 PO 4 for 24 h. After baseline measurements of dry mass, beams were divided into 11 groups ( n = 10/group) and, were pretreated for 5 min with 1% glutaraldehyde (GA); 5% GA; 1% grape-seed extract (GS); 5% GS; 10% sumac (S); 20 μM curcumin (CR); 200 μM CR; 0.l% riboflavin/UV (R); 0.5% R; 0.1% riboflavin-5-phosphate/UV (RP); and control (no pretreatment). After pretreatment, the beams were blot-dried and incubated in 1 mL calcium and zinc-containing medium (CM, pH 7.2) at 37 °C for 3, 7 or 14 days. After incubation, dry mass was reassessed and aliquots of the incubation media were analyzed for collagen C-telopeptides, ICTP and CTX using specific ELISA kits. Data were analyzed by repeated-measures ANOVA. Results The rate of dry mass loss was significantly different among test groups ( p < 0.05). The lowest 14 day mean dry mass loss was 6.98% ± 1.99 in the 200 μM curcumin group compared to control loss of dry mass at 32.59% ± 5.62, p < 0.05, at 14 days. The ICTP release over the incubation period (ng/mg dry dentin) ranged between 1.8 ± 0.51 and 31.8 ± 1.8. CTX release from demineralized beams pretreated with crosslinkers was significantly lower than CM (5.7 ± 0.2 ng/mg dry dentin). Significance The results of this study indicate that collagen crosslinkers tested in this study are good inhibitors of cathepsin K activity in dentin. However, their inhibitory effect on MMP activity was highly variable. [ABSTRACT FROM AUTHOR]

Objective This study tested whether treatment of demineralized dentin with polyacrylic acid (PAA) has any activatory or inhibitory activity on dentin matrix metalloproteinases (MMP)s or cathepsin K (CAT-K). Methods Dentin beams (1 mm × 2 mm × 6 mm; n = 10) were completely demineralized with EDTA. After initial dry mass assessment, the beams were dipped into 37% phosphoric acid (PA), PA + 2% benzalkonium chloride (BAC), PA + 2% chlorhexidine digluconate (CHX), 10% PAA, PAA + BAC or PAA + CHX for 20 s. Demineralized beams without treatment served as control. All beams were incubated in simulated body fluid (SBF) for 1 week and the dry mass loss was evaluated. Aliquots of SBF were used to analyze solubilized telopeptide fragments using ICTP as indicator of MMP-mediated collagen degradation and CTX for CAT-K-mediated degradation. Additional demineralized beams ( n = 10) were used to measure the influence of different chemical treatments on total MMP activity of EDTA-demineralized dentin using generic MMP assay. Data were analyzed by ANOVA ( α = 0.05). Results Dry mass loss ranged from 6% (PA) to 2% for (PA-BAC) or (PAA-BAC) ( p < 0.05). ICTP release of PAA-treated group was significantly higher ( p < 0.05) than the control, and not significantly different from the PA group ( p > 0.05). PA + CHX or PAA + CHX and PAA + BAC showed significantly lower ICTP than PA or PAA groups ( p < 0.05). CAT-K activity increased significantly after 10% PAA treatment compared to control ( p < 0.05) or to PA postreatment. Significance Demineralized dentin treated with 10% polyacrylic acid activated CAT-K more than 37% phosphoric acid; 2% chlorhexidine digluconate seems to be a better inhibitor of MMPs and CAT-K than 2% benzalkonium chloride. [ABSTRACT FROM AUTHOR]

This study analyzed the amounts of solubilized telopeptides cross-linked carboxyterminal telopeptide of type I collagen (ICTP) and C-terminal crosslinked telopeptide of type I collagen (CTX) derived from matrix-metalloproteinases (MMPs) and cysteine cathepsins (CTPs) subsequent to application of a filler-free (Res.A) or an ion-releasing resin (Res.B) to ethylenediaminetetraacetic acid (EDTA)-demineralized dentin with or without zoledronate-containing primer (Zol-primer) pre-treatment. The chemical modification induced following treatments and artificial saliva (AS) storage was also analyzed through attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR). Totally EDTA-demineralized specimens were infiltrated with Res.A or Res.B with or without Zol-primer pre-treatment, light-cured, and immersed in AS for up to 4 wk. ICTP release was reduced following infiltration with Res.B and further reduced when Res.B was used with Zol-primer; remarkable phosphate mineral uptake was attained after AS storage. CTX release was increased in Res.A- and Res.B-treated dentin. However, when Zol-primer was used with Res.A, the CTX release fell significantly compared to the other tested resin-infiltration methods. In conclusion, zoledronate offers an additional inhibitory effect to the ion-releasing resins in MMP-mediated collagen degradation. However, Zol-primer induces a modest reduction in CTX release only when used with resin-based systems containing no ion-releasing fillers. [ABSTRACT FROM PUBLISHER]

Matrix metalloproteinases (MMPs) cause collagen degradation in hybrid layers created by dentin adhesives. This in vitro study evaluated the feasibility of using a cross-linking agent, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), to inactivate soluble rhMMP-9, as an example of dentin MMPs, and matrix-bound dentin proteases. The inhibitory effects of 5 EDC concentrations (0.01-0.3 M) and 5 incubation times (1-30 min) on soluble rhMMP-9 were screened with an MMP assay kit. The same EDC concentrations were used to evaluate their inhibitory effects on endogenous proteinases from completely demineralized dentin beams that were incubated in simulated body fluid for 30 days. Decreases in modulus of elasticity (E) and dry mass of the beams, and increases in hydroxyproline content of hydrolysates derived from the incubation medium were used as indirect measures of matrix collagen hydrolysis. All EDC concentrations and pre-treatment times inactivated MMP-9 by 98% to 100% (p < 0.05) compared with non-cross-linked controls. Dentin beams incubated in 0.3 M EDC showed only a 9% decrease in E (45% decrease in control), a 3.6% to 5% loss of dry mass (18% loss in control), and significantly less solubilized hydroxyproline when compared with the control without EDC cross-linking (p < 0.05). It is concluded that EDC application for 1 min may be a clinically relevant and effective means for inactivating soluble rhMMP-9 and matrix-bound dentin proteinases if further studies demonstrate that EDC is not toxic to pulpal tissues. [ABSTRACT FROM PUBLISHER]