| Adhesion, as one of the most effective technique in recent stomotology development, is the rational foundation of current dental esthetic restoration and dental rehabilitation. By virtue of etching technique, the adhesion to enamel reveals favorable results. However, the effectiveness and durability of various kinds of dental bonding systems remain questionable because of its complicated structure.Collagen fibrils are not only the component of inorganic substance but also of bonding interface in dentin. With either removing the smear layer or maintaining it as the substrate for the bonding, adhesives interact with collagen fibril network exposed by dentin demineralization forming the hybrid layer. Due to the effects of water and enzymes on the elution of non-polymerized resin monomers and water-soluble molecules from the collagen fibrils within the hybrid layer of both etch-and-rinse and self-etching systems, adhesion failure is caused by the degeneration of the bonding interface where the exposed collagen fibrils are degraded. Thus, the effects of the degradation of collagen in hybrid layer on the bond strength and stability of dental adhesion are the crucial point whose management may further advance the adhesive mechanism.The present study evaluated the effects of acid etching on the type I collagen in dentin and analyzed the relationship between collagen degradation and bond strength on the basis of the trend of changing with time respectively. The bond strength was measured by microtensile bond strength test. Analysis of hybrid layer was carried out by the combination of colloidal gold technique and a field emission in-lens scanning electron microscope. Quantity of collagen degradation was determined by enzyme linked immunosorbent assay. The study provided direct theoretical and experimental basis to the durability investigation of adhesion to dentin.Major results and conclusions from the studies are as follows:1. Longer applications of 37% phosphoric acid may increase collagen degradation. 60 s application showed the most degradation of collagen, which was. 4.86(1.55) mg/g, followed by 30 s and 15 s, which were 2.76(0.87) mg/g and 1.93(0.88) mg/g. 10 s was 0.95(0.38) mg/g. The control group showed the least degradation of 0.06(0.03) mg/g. Significant differences were found among groups (P<0.005).2. Smear layer were removed but tubular orifices and collagen fibrils were covered by particles after dentin being etched with 37% phosphoric acid for 10 s, while tubular orifices clearly opened, major fibrils and globular particles were observed for 15 s group. Smoother surfaces of exposed collagen fibrils and fewer globular particles were found in the samples conditioned for 30 s than those conditioned for 15 s. In the 60 s group, it was found that the number of major fibrils decreased while minor branching fibrils increased indicating the intratubular structure collapsed and fibrils fractured.3. One-bottle self-etching adhesive got the weaker immediate bond strength, which was 24.07±7.47 MPa, than two-step self-etching and etch-and-rinse adhesive, which were 29.53±7.13 MPa and 29.81±7.34 MPa (P<0.05). Prolonged artificial saliva solutions immersion time can decrease the bond strength of all 3 adhesives. However, for the one-bottle self-etching adhesive, the decrease of bond strength was of no significant differences. The 1 m immersion bond strength of one-bottle self-etching adhesive was 22.11±7.16 MPa, which was significantly lower than that of the other two (P<0.05), while the 4 m immersion bond strength was 20.55±6.06 MPa, the mean of that was lower than that of etch-and-rinse adhesive, higher than that of two-step self-etching adhesive, but with no significant differences. For the other two adhesives, 1 m immersion bond strength were 28.88±10.52 MPa for two-step self-etching adhesive and 28.25±9.32 MPa for etch-and-rinse adhesive, despite of which were both lower than immediate bond strength, no significant difference could be found. 4 m immersion of two-step self-etching adhesive and etch-and-rinse adhesive were 18.96±5.31 MPa and 21.21±6.39 MPa showing the significantly weaker bond strength comparing with that of 1 m immersion and immediate (P < 0.05). There was no significant difference between the two-step self-etching and etch-and-rinse adhesive.4. The immediate collagen degradation of one-bottle self-etching system was 0.18±0.01 mg/g , significantly more than the other two, 0.16±0.01 mg/g for the two-step self-etching system and 0.15±0.01 mg/g for the total-etching system (P<0.05). The collagen degradation of 3 systems was increased by extending the application time. For the 1 m immersion, comparing with no statistical differences between the total-etching and one-bottle self-etching system, which was 0.18±0.01 mg/g and 0.19±0.02 mg/g, the collagen degradation of two-step self-etching system was 0.16±0.01 mg/g revealing significantly less than the former two (P<0.05). There were significant differences among all the systems immersed in artificial saliva solutions for 4 m, which were 0.26±0.01 mg/g for one-bottle self-etching system, 0.21±0.02 mg/g for two-step self-etching system, 0.19±0.02 mg/g for total-etching system (P<0.05). Total-etching system showed the large amount of collagen degradation in the 1 m immersion while self-etching systems in the 4 m. Besides, the collagen degradation of one-bottle self-etching system was significant higher than two-step self-etching system (P<0.05).5. There was negative correlation between the degradation of collagen fibrils and the bond strength of dentin adhesion (r=-0.65, P=0.003).6. The type I collagen not encapsulated can be immunohistochemically identified after staining with anti-type I collagen antibody and the secondary antibody conjugated with colloidal gold.7. The bonding interface of total-etching system can be observed with longer resin tags and variation in the thickness of hybrid layer. By mixing secondary and back-scattered electrons, an extensive labeling of collagen fibrils were seen in the hybrid layer. In addition, the labeling of the deepest area was more intense than that of the superficial. Comparing with the hybrid layer created by the two-step self-etching system, which was uniform and compact, the hybrid layer of porous defects and fractured resin tags was formed by the one-bottle self-etching system. Immunolabeling of the hybrid layer created by self-etching systems did not gradient. With longer application in immersing in artificial saliva sloution, the number of resin tags decreased, the labeling of all 3 systems became weaker. In contrast to total-etching system, cracks were evident in the bottom of hybrid layer created by self-etching system immersed for 4 m relative to 1 m.To concluded, the degradation of type I collagen plays an important role in dentin adhesion bond strength and stability. Etching dentin surface for 15 s was enough for demineralization; longer application may increase excessive collagen degradation resulting in weak adhesion. Immersed in artificial saliva solutions for longer time can worsen the integrity of bonding interface, increase the degradation of type I collagen, decrease the bond strength. Furthermore, negative correlation can be found between the latter two. |
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