Effects of Cathepsin K Inhibitors on Dentin Erosion: An in vitro Study.

Cathepsin K (catK) modulates the degradation of dentin collagen. This study aimed to evaluate the effects of catK inhibitors on dentin erosion. Dentin beams were eroded (4 times/d for 5 days) and immersed in deionized water (negative control), 0.1 M NaCl, 0.3 M NaCl, 0.5 M NaCl, or 1 µm odanacatib (each n = 16) for 30 min after each erosive challenge. Erosive dentin loss (EDL) and demineralized organic matrix (DOM) thickness were evaluated profilometrically. Additionally, dentin beams were demineralized, immersed in the respective solutions for 30 min each (n = 5), and then incubated in artificial saliva for 5 days. Dentin collage degradation was evaluated by quantifying the levels of the C-terminal peptide of type I collagen (CTX), C-terminal cross-linked telopeptide of type I collagen (ICTP), and hydroxyproline (HYP) in the incubation media. Significantly lower EDL and dentin collagen degradation (CTX, ICTP, and HYP) and thicker DOM layers were observed in the samples treated with 0.3 m NaCl and 1 µm odanacatib than in those treated with deionized water (all p < 0.05). The samples treated with 1 µm odanacatib showed significantly lower levels of CTX and HYP than those treated with 0.3 M NaCl (all p < 0.05). The present findings support the potential use of catK inhibitors in controlling dentin erosion.

The activity, distribution, and colocalization of cathepsin K and matrix metalloproteases in intact and eroded dentin.

OBJECTIVES: This study aimed to assess the activity, distribution, and colocalization of cathepsin K (catK) and matrix metalloproteases (MMPs) in both intact and eroded dentin in vitro. MATERIALS AND METHODS: Eroded dentin was obtained by consecutive treatment with 5% citric acid (pH = 2.3) for 7 days, while intact dentin remained untreated. Pulverized dentin powder (1.0 g) was extracted from both intact and eroded dentin using 5 mL of 50 mM Tris-HCl buffer (0.2 g/1 mL, pH = 7.4) for 60 h to measure the activity of catK and MMPs spectrofluorometrically. In addition, three 200-µm-thick dentin slices were prepared from intact and eroded dentin for double-labeling immunofluorescence to evaluate the distribution and colocalization of catK and MMPs (MMP-2 and MMP-9). The distribution and colocalization of enzymes were analyzed using inverted confocal laser scanning microscopy (CLSM), with colocalization rates quantified using Leica Application Suite Advanced Fluorescent (LAS AF) software. One-way analysis of variance (ANOVA) was used to analyze the fluorescence data related to enzyme activity (α = 0.05). RESULTS: The activity of catK and MMPs was significantly increased in eroded dentin compared with intact dentin. After erosive attacks, catK, MMP-2, and MMP-9 were prominently localized in the eroded regions. The colocalization rates of catK with MMP-2 and MMP-9 were 13- and 26-fold higher in eroded dentin, respectively, than in intact dentin. CONCLUSIONS: Erosive attacks amplified the activity of catK and MMPs in dentin while also altering their distribution patterns. Colocalization between catK and MMPs increased following erosive attacks. CLINICAL RELEVANCE: CatK, MMP-2, and MMP-9 likely play synergistic roles in the pathophysiology of dentin erosion.

Protecting primary teeth from dental erosion through bioactive glass.

OBJECTIVES: The present study aimed to evaluate the effectiveness of bioactive glass (BAG) in preventing dental erosion in primary teeth. METHODS: Enamel and dentin specimens (2 × 2 × 2 mm) were obtained from extracted primary teeth, which were randomly divided into the following groups based on the pretreatments (n = 12): DW (deionized water), NaF (2 % sodium fluoride), 2BAG (2 % BAG), 4BAG (4 % BAG), 6BAG (6 % BAG), and 8BAG (8 % BAG). The specimens were immersed in the respective solutions for 2 min and subjected to in vitro erosive challenges (4 × 5 min/d) for 5 d. The erosive enamel loss (EEL), erosive dentin loss (EDL), and the thickness of the demineralized organic matrix (DOM) were measured using a contact profilometer. The surface microhardness (SMH) was measured, and the percentage of SMH loss (%SMHL) was calculated. The surface morphology and mineral composition were evaluated by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS), respectively. RESULTS: After the erosive challenges, the EEL, EDL, and%SMHL of the 2BAG, 4BAG, 6BAG, and 8BAG groups significantly reduced, with the greatest reduction was observed in the 6BAG (EEL: 6.5 ± 0.2 µm;%SMHL in enamel: 12.8 ± 2.6; EDL: 7.9 ± 0.3 µm; %SMHL in dentin: 22.1 ± 2.7) and 8BAG groups (EEL: 6.4 ± 0.4 µm;%SMHL in enamel: 11.0 ± 1.9; EDL: 7.8 ± 0.5 µm; %SMHL in dentin: 22.0 ± 2.5) (P < 0.05). With increasing BAG concentrations, the number of surface deposits containing Ca, P, and Si increased. CONCLUSIONS: 6BAG was the most effective for preventing dental erosion in primary teeth and showed a particularly strong potential for dentin erosion prevention. CLINICAL SIGNIFICANCE: Bioactive glass, especially at a 6 % concentration, has proven effective in reducing erosive tooth wear and surface microhardness loss while also protecting demineralized organic matrix in primary dentin.

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

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.

Receptor-mediated cellular uptake of folate-conjugated fluorescent nanodiamonds: a combined ensemble and single-particle study.

Fluorescent nanodiamonds (FNDs) are nontoxic and photostable nanomaterials, ideal for long-term in vivo imaging applications. This paper reports that FNDs with a size of approximately 140 nm can be covalently conjugated with folic acid (FA) for receptor-mediated targeting of cancer cells at the single-particle level. The conjugation is made by using biocompatible polymers, such as polyethylene glycol, as crosslinked buffer layers. Ensemble-averaged measurements with flow cytometry indicate that more than 50% of the FA-conjugated FND particles can be internalized by the cells (such as HeLa cells) through receptor-mediated endocytosis, as confirmed by competitive inhibition assays. Confocal fluorescence microscopy reveals that these FND particles accumulate in the perinuclear region. The absolute number of FNDs internalized by HeLa cells after 3 h of incubation at a particle concentration of 10 microg mL(-1) is in the range of 100 particles per cell. The receptor-mediated uptake process is further elucidated by single-particle tracking of 35-nm FNDs in three dimensions and real time during the endocytosis.

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.