title: MMP activity in the hybrid layer detected with in situ zymography.
abstract: Dentinal proteases are believed to play an important role in the degradation of hybrid layers (HL). This study investigated the HL gelatinolytic activity by in situ zymography and functional enzyme activity assay. The hypotheses were that HLs created by an etch-and-rinse adhesive exhibit active gelatinolytic activity, and MMP-2 and -9 activities in dentin increase during adhesive procedures. Etched-dentin specimens were bonded with Adper Scotchbond 1XT and restored with composite. Adhesive/dentin interface slices were placed on microscope slides, covered with fluorescein-conjugated gelatin, and observed with a multi-photon confocal microscope after 24 hrs. Human dentin powder aliquots were prepared and assigned to the following treatments: A, untreated; B, etched with 10% phosphoric acid; or C, etched with 10% phosphoric acid and mixed with Scotchbond 1XT. The MMP-2 and -9 activities of extracts of dentin powder were measured with functional enzyme assays. Intense and continuous enzyme activity was detected at the bottom of the HL, while that activity was more irregular in the upper HL. Both acid-etching and subsequent adhesive application significantly increased MMP-2 and -9 activities (p < 0.05). The results demonstrate, for the first time, intrinsic MMP activity in the HL, and intense activation of matrix-bound MMP activity with both etching and adhesive application.
authors.affiliation: Department of Medical Sciences, Unit of Dental Sciences and Biomaterials, University of Trieste, Italy. annalisa.mazzoni@unibo.it
authors.forename: A
authors.initials: A
authors.lastname: Mazzoni
authors.forename: F D
authors.initials: FD
authors.lastname: Nascimento
authors.forename: M
authors.initials: M
authors.lastname: Carrilho
authors.forename: I
authors.initials: I
authors.lastname: Tersariol
authors.forename: V
authors.initials: V
authors.lastname: Papa
authors.forename: L
authors.initials: L
authors.lastname: Tjäderhane
authors.forename: R
authors.initials: R
authors.lastname: Di Lenarda
authors.forename: F R
authors.initials: FR
authors.lastname: Tay
authors.forename: D H
authors.initials: DH
authors.lastname: Pashley
authors.forename: L
authors.initials: L
authors.lastname: Breschi
mesh_terms: D000134:Acid Etching, Dental; D003094:Collagen / Q000378:metabolism; D001840:Dental Bonding; D003804:Dentin / Q000201:enzymology*; D003806:Dentin Permeability; D017220:Dentin-Bonding Agents / Q000737:chemistry / Q000493:pharmacokinetics*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006868:Hydrolysis; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D018613:Microscopy, Confocal; D036641:Microscopy, Fluorescence, Multiphoton; D019279:Resin Cements / Q000737:chemistry / Q000493:pharmacokinetics
chemical_list: D017220:Dentin-Bonding Agents; D019279:Resin Cements; D003094:Collagen; C041330:Scotchbond; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9