Arzu Tezvergil-Mutluay

State of the art etch-and-rinse adhesives

OBJECTIVES The aim of this study was to explore the therapeutic opportunities of each step of 3-step etch-and-rinse adhesives. METHODS Etch-and-rinse adhesive systems are the oldest of the multi-generation evolution of resin bonding systems. In the 3-step version, they involve acid-etching, priming and application of a separate adhesive. Each step can accomplish multiple goals. Acid-etching, using 32-37% phosphoric acid (pH 0.1-0.4) not only simultaneously etches enamel and dentin, but the low pH kills many residual bacteria. RESULTS Some etchants include anti-microbial compounds such as benzalkonium chloride that also inhibits matrix metalloproteinases (MMPs) in dentin. Primers are usually water and HEMA-rich solutions that ensure complete expansion of the collagen fibril meshwork and wet the collagen with hydrophilic monomers. However, water alone can re-expand dried dentin and can also serve as a vehicle for protease inhibitors or protein cross-linking agents that may increase the durability of resin-dentin bonds. In the future, ethanol or other water-free solvents may serve as dehydrating primers that may also contain antibacterial quaternary ammonium methacrylates to inhibit dentin MMPs and increase the durability of resin-dentin bonds. The complete evaporation of solvents is nearly impossible. SIGNIFICANCE Manufacturers may need to optimize solvent concentrations. Solvent-free adhesives can seal resin-dentin interfaces with hydrophobic resins that may also contain fluoride and antimicrobial compounds. Etch-and-rinse adhesives produce higher resin-dentin bonds that are more durable than most 1 and 2-step adhesives. Incorporation of protease inhibitors in etchants and/or cross-linking agents in primers may increase the durability of resin-dentin bonds. The therapeutic potential of etch-and-rinse adhesives has yet to be fully exploited.

Optimizing dentin bond durability: control of collagen degradation by matrix metalloproteinases and cysteine cathepsins

OBJECTIVES Contemporary adhesives lose their bond strength to dentin regardless of the bonding system used. This loss relates to the hydrolysis of collagen matrix of the hybrid layers. The preservation of the collagen matrix integrity is a key issue in the attempts to improve the dentin bonding durability. METHODS Dentin contains collagenolytic enzymes, matrix metalloproteinases (MMPs) and cysteine cathepsins, which are responsible for the hydrolytic degradation of collagen matrix in the bonded interface. RESULTS The identities, roles and function of collagenolytic enzymes in mineralized dentin has been gathered only within last 15 years, but they have already been demonstrated to have an important role in dental hard tissue pathologies, including the degradation of the hybrid layer. Identifying responsible enzymes facilitates the development of new, more efficient methods to improve the stability of dentin-adhesive bond and durability of bond strength. SIGNIFICANCE Understanding the nature and role of proteolytic degradation of dentin-adhesive interfaces has improved immensely and has practically grown to a scientific field of its own within only 10 years, holding excellent promise that stable resin-dentin bonds will be routinely available in a daily clinical setting already in a near future.

Strategies to prevent hydrolytic degradation of the hybrid layer-A review.

OBJECTIVE Endogenous dentin collagenolytic enzymes, matrix metalloproteinases (MMPs) and cysteine cathepsins, are responsible for the time-dependent hydrolysis of collagen matrix of hybrid layers. As collagen matrix integrity is essential for the preservation of long-term dentin bond strength, inhibition of endogenous dentin proteases is necessary for durable resin-bonded restorations. METHODS Several tentative approaches to prevent enzyme function have been proposed. Some of them have already demonstrated clinical efficacy, while others need to be researched further before clinical protocols can be proposed. This review will examine both the principles and outcomes of techniques to prevent collagen hydrolysis in dentin-resin interfaces. RESULTS Chlorhexidine, a general inhibitor of MMPs and cysteine cathepsins, is the most tested method. In general, these experiments have shown that enzyme inhibition is a promising approach to improve hybrid layer preservation and bond strength durability. Other enzyme inhibitors, e.g. enzyme-inhibiting monomers, may be considered promising alternatives that would allow more simple clinical application than chlorhexidine. Cross-linking collagen and/or dentin matrix-bound enzymes could render hybrid layer organic matrices resistant to degradation. Alternatively, complete removal of water from the hybrid layer with ethanol wet bonding or biomimetic remineralization should eliminate hydrolysis of both collagen and resin components. SIGNIFICANCE Understanding the function of the enzymes responsible for the hydrolysis of hybrid layer collagen has prompted several innovative approaches to retain hybrid layer integrity and strong dentin bonding. The ultimate goal, prevention of collagen matrix degradation with clinically applicable techniques and commercially available materials may be achievable in several ways.

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.

The anti-MMP activity of benzalkonium chloride

OBJECTIVE This study evaluated the ability of benzalkonium chloride (BAC) to bind to dentine and to inhibit soluble recombinant MMPs and bound dentine matrix metalloproteinases (MMPs). METHODS Dentine powder was prepared from extracted human molars. Half was left mineralized; the other half was completely demineralized. The binding of BAC to dentine powder was followed by measuring changes in the supernatant concentration using UV spectrometry. The inhibitory effects of BAC on rhMMP-2, -8 and -9 were followed using a commercially available in vitro proteolytic assay. Matrix-bound endogenous MMP-activity was evaluated in completely demineralized beams. Each beam was either dipped into BAC and then dropped into 1 mL of a complete medium (CM) or they were placed in 1 mL of CM containing BAC for 30 days. After 30 days, changes in the dry mass of the beams or in the hydroxyproline (HYP) content of hydrolysates of the media were quantitated as indirect measures of matrix collagen hydrolysis by MMPs. RESULTS Demineralized dentine powder took up 10-times more BAC than did mineralized powder. Water rinsing removed about 50% of the bound BAC, whilst rinsing with 0.5M NaCl removed more than 90% of the bound BAC. BAC concentrations 0.5wt% produced 100% inhibition of soluble recombinant MMP-2, -8 or -9, and inhibited matrix-bound MMPs between 55 and 66% when measured as mass loss or 76-81% when measured as solubilization of collagen peptide fragments. CONCLUSIONS BAC is effective at inhibiting both soluble recombinant MMPs and matrix-bound dentine MMPs in the absence of resins.

Incremental layers bonding of silorane composite: The initial bonding properties

OBJECTIVES Lack of oxygen inhibition layer of silorane composite with cationic polymerization raises the question of the bonding of incremental layers of the composite. This study aimed to evaluate the bond strength of the silorane composite layers. METHODS Fresh, 20 s, 5 min aged silorane composite (Silorane, 3M-ESPE) was used as substrate to adhere new silorane composite. For a comparison, dimethacrylate-based composite resin (Z250, 3M-ESPE) was adhered to the silorane composite with or without intermediate adhesive resin. As a control, dimethacrylate composite with oxygen inhibition layer was attached to fresh dimethacrylate composite. The bonded specimens (n=12/group) were water stored for 24 h. The shear bond strengths (SBS) were measured with a crosshead speed of 1.0 mm/min. Failure modes were assessed. Data were analysed by ANOVA, Tukeys post hoc tests and Chi-square tests (p=0.05). RESULTS Dimethacrylate-dimethacrylate composite resin combination showed the highest mean SBS (33.0 MPa) values with no adhesive failures. Fresh silorane-silorane SBS was slightly lower (26.7 MPa) and was further decreased by aging the substrate for 20s (25.4 MPa) and 5 min (22.4 MPa). The percent of adhesive failures increased from 25% to 75%, respectively. The failure modes were significantly different (Chi-square, p<0.001). Silorane-dimethacrylate composite showed the lowest (4.0 MPa) SBS among the groups, which was increased significantly by use of phosphate-methacrylate-based intermediate resin (p<0.05). CONCLUSION In order to bond dimethacrylate composite to silorane composite, a phosphate-methacrylate-based intermediate resin is required. The silorane composite showed slightly lower incremental bonding properties than conventional dimethacrylate composites.

Effect of Phosphoric Acid on the Degradation of Human Dentin Matrix

This study determined if dentin proteases are denatured by phosphoric acid (PA) used in etch-and-rinse dentin adhesives. Dentin beams were completely demineralized with EDTA for 30 days. We “acid-etched” experimental groups by exposing the demineralized dentin beams to 1, 10, or 37 mass% PA for 15 sec or 15 min. Control beams were not exposed to PA but were incubated in simulated body fluid for 3 days to assay their total endogenous telopeptidase activity, by their ability to solubilize C-terminal crosslinked telopeptides ICTP and CTX from insoluble dentin collagen. Control beams released 6.1 ± 0.8 ng ICTP and 0.6 ± 0.1 ng CTX/mg dry-wt/3 days. Positive control beams pre-incubated in p-aminophenylmercuric acetate, a compound known to activate proMMPs, released about the same amount of ICTP peptides, but released significantly less CTX. Beams immersed in 1, 10, or 37 mass% PA for 15 sec or 15 min released amounts of ICTP and CTX similar to that released by the controls (p > 0.05). Beams incubated in galardin, an MMP inhibitor, or E-64, a cathepsin inhibitor, blocked most of the release of ICTP and CTX, respectively. It is concluded that PA does not denature endogenous MMP and cathepsin activities of dentin matrices.

Degree of conversion of dual-cure luting resins light-polymerized through various materials.

Objective. The aim of this study was to investigate the degree of monomer conversion of four dual-cure luting resins irradiated through various restorative materials or dentin. Material and methods. RelyX ARC (3M-ESPE), RelyX Unicem (3M-ESPE), Variolink 2 (Ivoclar,Vivadent), and Panavia F 2.0 (Kuraray) were mixed in accordance with the manufacturers instructions. They were placed under the disks (thickness 1.5 mm) representing a metal restoration, a composite restoration (Sinfony D A3), a fiber-reinforced composite (EverStick 0.5 mm + 1.0 mm Sinfony D A3) restoration, and dentin. Five specimens (thickness 0.6 mm) in each group were irradiated through the disks for 40 s (Optilux-501, 800 mW/cm2). Light polymerization of the dual-cure luting resin without the covering disk was used as control. The degree of monomer conversion (DC%) was determined by Fourier transform infrared spectroscopy (FT-IR)/ATR spectrometry from the bottom of the resin. The infrared spectra were recorded at every 5.2 s for 15 min beginning from the mixing of the resin. Results. ANOVA revealed significant differences in DC% between the luting resins tested (p<0.001) and the different restorations (p<0.001). RelyX ARC showed the highest degree of conversion 15 min after the start of polymerization, whereas Panavia F 2.0 and RelyX Unicem showed the lowest. Conclusions. The degree of conversion of dual-cured luting resins differed significantly. Furthermore, the restorative material significantly influenced the DC% of the dual-cure luting resin underneath.

Biomimetic remineralization as a progressive dehydration mechanism of collagen matrices--implications in the aging of resin-dentin bonds.

Biomineralization is a dehydration process in which water from the intrafibrillar compartments of collagen fibrils are progressively replaced by apatites. As water is an important element that induces a lack of durability of resin-dentin bonds, this study has examined the use of a biomimetic remineralization strategy as a progressive dehydration mechanism to preserve joint integrity and maintain adhesive strength after ageing. Human dentin surfaces were bonded with dentin adhesives, restored with resin composites and sectioned into sticks containing the adhesive joint. Experimental specimens were aged in a biomimetic analog-containing remineralizing medium and control specimens in simulated body fluid for up to 12 months. Specimens retrieved after the designated periods were examined by transmission electron microscopy for the presence of water-rich regions using a silver tracer and for collagen degradation within the adhesive joints. Tensile testing was performed to determine the potential loss of bond integrity after ageing. Control specimens exhibited severe collagen degradation within the adhesive joint after ageing. Remineralized specimens exhibited progressive dehydration, as manifested by silver tracer reduction and partial remineralization of water-filled microchannels within the adhesive joint, as well as intrafibrillar remineralization of collagen fibrils that were demineralized initially as part of the bonding procedure. Biomimetic remineralization as a progressive dehydration mechanism of water-rich, resin-sparse collagen matrices enables these adhesive joints to resist degradation over a 12-month ageing period, as verified by the conservation of their tensile bond strength. The ability of the proof of concept biomimetic remineralization strategy to prevent bond degradation warrants further development of clinically relevant delivery systems.

The importance of size-exclusion characteristics of type I collagen in bonding to dentin matrices.

The mineral phase of dentin is located primarily within collagen fibrils. During development, bone or dentin collagen fibrils are formed first and then water within the fibril is replaced with apatite crystallites. Mineralized collagen contains very little water. During dentin bonding, acid-etching of mineralized dentin solubilizes the mineral crystallites and replaces them with water. During the infiltration phase of dentin bonding, adhesive comonomers are supposed to replace all of the collagen water with adhesive monomers that are then polymerized into copolymers. The authors of a recently published review suggested that dental monomers were too large to enter and displace water from collagen fibrils. If that were true, the endogenous proteases bound to dentin collagen could be responsible for unimpeded collagen degradation that is responsible for the poor durability of resin-dentin bonds. The current work studied the size-exclusion characteristics of dentin collagen, using a gel-filtration-like column chromatography technique, using dentin powder instead of Sephadex. The elution volumes of test molecules, including adhesive monomers, revealed that adhesive monomers smaller than ∼1000 Da can freely diffuse into collagen water, while molecules of 10,000 Da begin to be excluded, and bovine serum albumin (66,000 Da) was fully excluded. These results validate the concept that dental monomers can permeate between collagen molecules during infiltration by etch-and-rinse adhesives in water-saturated matrices.