## 4 Discussion  
Dentin is a tissue characterized by a high organic content, which includes collagen fibers and odontoblasts in dentin tubules; these latter are filled with dentinal fluid [7,21]. It is well-known that the hydrolysis of collagen fibrils during storage in water could decrease the bond strength of the resin-dentin interface [7,18,21]. In this study, the bond strength for all the tested adhesive systems, which were applied following their manufacturers' instructions, was higher after 24 h of storage in water than after long-term storage. Immediately (after 24 h) the SBS test showed no significant differences (\(p>0.05\)) between the tested adhesives. The adhesives SU and IPM preserved the bond strength values after a six-month storage period, whilst the SBS of IPU, IB, and FB decreased after long-term storage (\(p<0.05\)). Therefore, the first hypothesis that there would be no difference between the tested adhesives in bond strength after long-term storage must be rejected. A possible explanation might be correlated to the different compositions of the tested materials, which influence the quality of the hybrid layer they create [2,22]. Indeed, the stability and durability of bond strength of IPM and SU could be in part attributed to the quality of 10-MDP used within their formulations. The formation of stable and insoluble nano-layered 10-MDP-calcium salts can significantly improve the bond strength and its durability over time [23]. Initial bond strength and its durability was shown to be important in the functional monomers that have more intense monomer-calcium formation than those creating lower monomer-calcium formation [23,24]. The results of our study are in accordance with those of [PERSON] et al. [25] and [PERSON] et al. [2]; they reported that SU when it is applied in self-etch mode could preserve the bond strength after 12 months of an aging storage period. FB had lower bond strength after long storage, although it contains MDP; this may be due to the quality of the MDP used in the adhesive [26]. In this study, SEM was used to measure the thickness of the resin layer of the five tested adhesives applied onto dentin; the mean thickness ranged between 10 and 17 \(\upmu\)m. SU and IB were significantly thicker than the other adhesives (\(p<0.05\)). These findings might be explained and related to the adhesive fillers, which could play a crucial role in the flowability of such adhesives. However, no correlation was found between the thicknesses and SBS values of the tested adhesives. It has been reported that the bond strength of resin-dentin interface is independent from the thickness of hybrid and/or adhesive layer [27]. Conversely, the quality of hybrid layer is an important factor that can influence the dentin bond strength, whilst a thick adhesive layer does not offer any increase in bond strength [28]. Figure 4: Representative photos of scanning electron microscopy showed the voids into the IBond Universal adhesive layer (white arrows) after a 6-months water-storage period at 37 \({}^{\circ}\)C. (**a**): x500 magnification; (**b**): x3000 magnification. All the adhesive systems used in this study created resin tags, which infiltrated several microns into dental tubules. Therefore, we hypothesized that all such adhesives can sufficiently dissolve the smear layer and allow the resin adhesive to enter into the tubules. However, the results of tag lengths were not significantly different (\(p>0.05\)). These findings could be explained by the fact that all the tested adhesives have a mild pH. Thus, the second hypothesis that the adhesive thickness and its resin infiltration into dental tubules would play no important role on the bond strength durability must also be rejected. All the measures for the evaluation of the tag penetration were performed in correspondence of the center of the composite-cylinder in order to prevent the effect of tubules position (density and diameter). Tubules diameters and density increase from the dentin-enamel junction to central dentin area [29].