Effects of MMP inhibitors incorporated within dental adhesives

Abstract

Matrix metalloproteinase (MMP) inhibition has been shown to reduce adhesive bond degradation when applied as a pre-conditioner, adding to clinical steps in the placement of adhesives, but their incorporation within dental adhesives has not been fully explored. This study examined the effect of including 2 MMP inhibitors (BB94 and GM6001) within the primers of 3 commercially available adhesives. Fluorometric assay and zymography showed that adhesives with MMP inhibitors had high affinity toward both synthetic fluorogenic FRET peptides (95%) and dentin powder substrates, respectively. The immediate microtensile bond strength was enhanced for 2 types of adhesives following the addition of both inhibitors. However, no changes were detected between the control and the inhibitor groups following 3-month storage. The modified two-step etch-and-rinse and single-step systems showed less Rhodamine B penetration to the "hybrid layer" and to the "adhesive", respectively. The incorporation of BB94 and GM6001 within the primers resulted in the inhibition of dentin MMPs with improved initial bond strength and enhanced sealing ability.

Figure 1.

The efficacy of inhibitor-doped commercial adhesives on MMP activity. (a) The effects on recombinant human (rh) MMP-1 and MMP-2 activity. The modified PB and GB showed significant inhibitory effects on MMP-1 and MMP-2. (b) Gelatin zymogram for OB. The highlighted areas represent the protease activity consistent with MMP-2 activity with gelatin. No reaction was observed when both MMP inhibitors adhesives were mixed with dentin powder. Molecular masses, expressed in kDa, are reported in the std lane.

Figure 2.

The evaluation of micropermeability of simulated pulpal fluids into the adhesive interface. (a) The micropermeability index: (1) completely intact hybrid layer with no dye reaching it, (2) the dye reaches the base of the hybrid layer, (3) the dye is completely infiltrated within the hybrid layer, and (4) the dye is reaching the adhesive layer. (b) Micropermeability scores for OB groups. (c) Micropermeability scores for PB groups. (d) Micropermeability scores for GB groups. *Significantly different from the control.

Figure 3.

TSM images of the interface morphology and examples of the 9 experimental groups’ micropermeability. Each row represents the most common features observed in the resin-dentin interfaces created with the 3 adhesive systems used in this study. The TSM images in each column represent the interface morphology (1), control adhesives without MMP inhibitors (2), adhesives with GM6001 (3), and adhesives with BB94 (4). (A1) TSM image captured in fluorescence mode (Rhodamine B excitation/emission), showing the interfacial characterization of the OB-bonded dentin specimens. It is possible to observe long resin tags (rt) and a 5- to 8-µm hybrid layer (HL) localized beneath a thick adhesive layer (a). The OB-bonded dentin showed dye penetration (micropermeability) within the entire thickness of the HL (A2), while the micropermeability of the resin-bonded interface created with the OB doped with GM6001 (A3) or BB94 (A4) was detected only inside the dentinal tubules. (B1) The interfacial characterization of the PB-bonded dentin specimens, also showing, in this case, a clear hybrid layer (HL) localized beneath a thick adhesive layer (a). This type of interface showed micropermeability within the entire thickness of the HL for the control group (B2). PB doped with GM6001 (B3) and BB94 (B4) had less dye reaching the hybrid layer. (C1) A TSM image showing the interfacial characterization of the GB-bonded dentin specimens. It is possible to observe short resin tags (rt) and a 1- to 2-µm hybrid inter-diffusion layer (HDL) localized beneath a thick adhesive layer (a) characterized by the clear presence of phase separation. This type of resin-bonded interface was affected by severe micropermeability within both the hybrid inter-diffusion and part of the adhesive layer, created with GB only (C2). The incorporation of GM6001 (C3) and BB94 (C4) resulted in less dye reaching the adhesive layer.