Toshikazu Uchiyama

The Inhibitory Effects of Quaternary Ammonium Methacrylates on Soluble and Matrix-bound MMPs

Matrix metalloproteinases (MMPs) bound to dentin contribute to the progressive degradation of collagen fibrils in hybrid layers created by dentin adhesives. This study evaluated the MMP-inhibiting potential of quaternary ammonium methacrylates (QAMs), with soluble rhMMP-9 and a matrix-bound endogenous MMP model. Six different QAMs were initially screened by a rhMMP-9 colorimetric assay. For the matrix-bound endogenous MMPs, we aged demineralized dentin beams for 30 days in calcium- and zinc-containing media (CM; control), chlorhexidine, or QAMs in CM to determine the changes in dry mass loss and solubilization of collagen peptides against baseline levels. The inhibitory effects of QAMs on soluble rhMMP-9 varied between 34 and 100%. Beams incubated in CM showed a 29% decrease in dry mass (p < 0.05), whereas beams incubated with QAMs showed only 0.2%-6% loss of dry mass. Significantly more solubilized collagen was detected from beams incubated in CM (p < 0.05). It is concluded that QAMs exhibited dentin MMP inhibition comparable with that of chlorhexidine, but required higher concentrations.

Chlorhexidine binding to mineralized versus demineralized dentin powder

OBJECTIVES The purposes of this work were to quantitate the affinity and binding capacity of chlorhexidine (CHX) digluconate to mineralized versus demineralized dentin powder and to determine how much debinding would result from rinsing with water, ethanol, hydroxyethylmethacrylate (HEMA) or 0.5M NaCl in water. METHODS Dentin powder was made from coronal dentin of extracted human third molars. Standard amounts of dentin powder were tumbled with increasing concentrations of CHX (0-30 mM) for 30 min at 37 degrees C. After centrifuging the tubes, the supernatant was removed and the decrease in CHX concentration quantitated by UV-spectroscopy. CHX-treated dentin powder was resuspended in one of the four debinding solutions for 3 min. The amount of debound CHX in the solvents was also quantitated by UV-spectroscopy. RESULTS As the CHX concentration in the medium increased, the CHX binding to mineralized dentin powder also increased up to 6.8 micromol/g of dry dentin powder. Demineralized dentin powder took up significantly (p<0.01) more CHX, reaching 30.1 micromol CHX/g of dry dentin powder. Debinding of CHX was in the order: HEMA<ethanol<0.05 M NaCl<water. The highest CHX binding to demineralized dentin occurred at 30 mM (1.5 wt.%). SIGNIFICANCE As CHX is not debound by HEMA, it may remain bound to demineralized dentin during resin-dentin bonding. This may be responsible for the long-term efficacy of CHX as an MMP inhibitor in resin-dentin bonds.

Inhibition of MMPs by alcohols.

OBJECTIVES While screening the activity of potential inhibitors of matrix metalloproteinases (MMPs), due to the limited water solubility of some of the compounds, they had to be solubilized in ethanol. When ethanol solvent controls were run, they were found to partially inhibit MMPs. Thus, the purpose of this study was to compare the MMP-inhibitory activity of a series of alcohols. METHODS The possible inhibitory activity of a series of alcohols was measured against soluble rhMMP-9 and insoluble matrix-bound endogenous MMPs of dentin in completely demineralized dentin. Increasing concentrations (0.17, 0.86, 1.71 and 4.28 mol/L) of a homologous series of alcohols (i.e. methanol, ethanol, propanols, butanols, pentanols, hexanols, the ethanol ester of methacrylic acid, heptanols and octanol) were compared to ethanediol, and propanediol by regression analysis to calculate the molar concentration required to inhibit MMPs by 50% (i.e. the IC(50)). RESULTS Using two different MMP models, alcohols were shown to inhibit rhMMP-9 and the endogenous proteases of dentin matrix in a dose-dependent manner. The degree of MMP inhibition by alcohols increased with chain length up to 4 methylene groups. Based on the molar concentration required to inhibit rhMMP-9 fifty percent, 2-hydroxyethylmethacrylate (HEMA), 3-hexanol, 3-heptanol and 1-octanol gave the strongest inhibition. SIGNIFICANCE The results indicate that alcohols with 4 methylene groups inhibit MMPs more effectively than methanol or ethanol. MMP inhibition was inversely related to the Hoys solubility parameter for hydrogen bonding forces of the alcohols (i.e. to their hydrophilicity).