Do matrix metalloproteinase and cathepsin K inhibitors work synergistically to reduce dentin erosion?

Abstract

Objectives: To evaluate the effects of matrix metalloproteinase (MMP) and cathepsin K (catK) inhibitors on resistance to dentin erosion.

Methodology: A total of 96 dentin specimens (3×3×2 mm) were prepared and randomly assigned into four groups (n=24): deionized water (DW); 1 µM odanacatib (ODN, catK inhibitor); 1 mM 1,10-phenanthroline (PHEN, MMP inhibitor); and 1 µM odanacatib + 1 mM 1,10-phenanthroline (COM). Each group was further divided into two subgroups for the application of treatment solutions before (PRE) and after erosive challenges (POST). All specimens were subjected to four daily erosive challenges for 5 d. For each erosive challenge, the specimens in subgroup PRE were immersed in the respective solutions before cola drinks, while the specimens in subgroup POST were immersed in the respective solutions after cola drinks (the immersion duration was 5 min in both cases). All specimens were stored in artificial saliva at 37°C between erosive challenges. The erosive dentin loss (EDL) was measured by profilometry. The residual demineralized organic matrix (DOM) of specimens was removed using type VII collagenase and evaluated by profilometry. Both the EDL and thickness of the residual DOM were statistically analyzed by two-way analysis of variance (ANOVA) and Bonferroni's test (α=0.05). The surface topography and transverse sections of the specimens were observed using SEM. MMPs and catK were immunolabeled in the eroded dentin and in situ zymography was performed to evaluate the enzyme activity.

Results: Significantly lower EDL was found in the groups ODN, PHEN, and COM than in the control group (all p<0.05), while no significant difference in EDL was found among the groups ODN, PHEN, and COM (all p>0.05). The application sequence showed no significant effect on the EDL of the tested groups (p=0.310). A significantly thicker DOM was observed in the group ODN than in the control group regardless of the application sequence (both p<0.05). The treatment with ODN, PHEN, and COM inhibited the gelatinolytic activity by approximately 46.32%, 58.6%, and 74.56%, respectively.

Conclusions: The inhibition of endogenous dentinal MMPs and catK increases the acid resistance of human dentin but without an apparent synergistic effect. The inhibition of MMPs and catK is equally effective either before or after the acid challenge.

Figure 1. Experimental flowchart

Figure 2. Representative SEM images (×5000) of the specimens (dentin surface) treated with the respective solutions in subgroups PRE (A, C, E, G) and POST (B, D, F, H)

Figure 3. Representative SEM images (×2000) of the specimens (transverse sections) treated with the respective solutions in subgroups PRE (A, C, E, G) and POST (B, D, F, H).

Figure 4. CLSM images of immunolabeled MMP-8 (A, B, C) and catK (D, E, F) in eroded dentin

Figure 5. CLSM images of the in situ zymography of the dentin slices without (A, B, C) or with (D, E, F, G, H, I, J, K, L) inhibitory treatment after incubation for 24 h