This study aimed to evaluate the activity of matrix metalloproteinases (MMPs) on primary and permanent dentin and monitor the degradation of dentin collagen over time. Twenty-four dentin beams with dimensions of 1.0 mm wide by 1.0 mm thick and 4.0 mm long will be obtained from sound human primary and third molars (n=12). The beams will be completely demineralized in 10% phosphoric acid for 18h at 3-4oC. The total initial MMP activity will be determined for each group (primary and permanent dentin) using a microcolorimetric assay (SensoLyte®). Then, the initial elastic modulus will be measured by flexural test with the demineralized beams immersed in deionized water and using a metal device with a 2.5 mm span between the support points. Subsequently, the initial dry mass of each beam will be determined after dehydration for 48 hours in a desiccator containing calcium sulfate. Then, the beams will be rehydrated in deionized water for 1h and storage in 300uL of artificial saliva for 7 days, 1 month or 2 months at 37oC. After each storage period the elastic modulus and the dry mass will be reassessed. The artificial saliva for each evaluation period will be frozen for measurement of the release of ICTP and CTX fragments produced by collagen hydrolysis mediated by MMPs and cysteine cathepsins, respectively.
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