Phosphoric acid concentration affects dentinal MMPs activity.
To evaluate whether the concentration of phosphoric acid (PA) has an effect on the proteolytic activity of sound human demineralized dentin. It is hypothesized that the activity of matrix-bound and extracted enzymes depends on the PA concentration used to demineralize dentin.



One-gram aliquots of mid-coronal human dentin powder were demineralized with 1wt%, 10wt% and 37wt% PA. Concentrations of released calcium were measured for each set of demineralization. Extracted MMP-2 was immunologically identified by western blot and its activity was determined by conventional gelatin zymography. Analysis of released hydroxyproline (HYP) and in situ zymography were performed to evaluate the activity of insoluble, bound-matrix enzymes.



The amount of released calcium from dentin powder treated with 37wt% PA was significantly higher (p≤0.05) than that obtained by dentin demineralization with 10wt% and 1wt% PA. Expression and activity of endogenous enzymes, extracted from or bound to dentin matrix, were detected for all samples regardless of the PA concentration. However, the expression and activity of extracted MMP-2 were significantly higher when dentin was treated with 10wt% PA (p<0.05), followed by 1wt% and 37wt% PA. Similarly, the highest concentration of released HYP (i.e. meaning higher percentage of collagen degradation) and the highest activity in in situ zymography were observed when dentin samples were treated with 10wt% PA (p<0.05).



It was confirmed that PA does not denature endogenous enzymes of dentin matrices, but it may somehow modulate the expression and activity of these enzymes in a concentration-dependent manner.



Endogenous proteases have been identified and suggested to be responsible for the digestion of dentin matrix when activated by the acidic components of dental adhesives. Proteolytic activity of dentinal MMPs showed to be dependent on phosphoric acid concentration. The clinically-used concentration (37%) does not inhibit MMPs activity, but slows it.
Department of Biomaterials and Oral Biology, School of Dentistry, University of São Paulo (USP), São Paulo, Brazil; Discipline of Dental Materials, School of Dentistry, University Nove de Julho (UNINOVE), São Paulo, Brazil.
A G
AG
DeVito-Moraes
Department of Biomaterials and Oral Biology, School of Dentistry, University of São Paulo (USP), São Paulo, Brazil.
C
C
Francci
Discipline of Restorative Dentistry, School of Dentistry, University of Passo Fundo (UPF), Passo Fundo, Brazil.
C M P
CM
Vidal
Department of Oral Biology, Bauru School of Dentistry, University of São Paulo (USP), Bauru, Brazil.
P M C
PM
Scaffa
Department of Biomaterials and Oral Biology, School of Dentistry, University of São Paulo (USP), São Paulo, Brazil.
D
D
Nesadal
Department of Biomaterials and Oral Biology, School of Dentistry, University of São Paulo (USP), São Paulo, Brazil; Discipline of Dental Materials, School of Dentistry, University Nove de Julho (UNINOVE), São Paulo, Brazil.
L C
LC
Yamasaki
Department of Biomaterials and Oral Biology, School of Dentistry, University of São Paulo (USP), São Paulo, Brazil.
J
J
Nicolau
Biomaterials and Biotechnology in Health Programs, Anhanguera University of São Paulo (UNIAN), São Paulo, Brazil.
F D
FD
Nascimento
Department of Oral Biology, College of Dental Medicine, Georgia Health Sciences University, Augusta, USA.
D H
DH
Pashley
Biomaterials and Biotechnology in Health Programs, Anhanguera University of São Paulo (UNIAN), São Paulo, Brazil. Electronic address: marcelacarrilho@gmail.com.
M R
MR
Carrilho
D003804:Dentin; D006801:Humans; D020782:Matrix Metalloproteinases; D010756:Phosphoric Acids; D017001:Tooth Demineralization
D010756:Phosphoric Acids; D020782:Matrix Metalloproteinases
Dentin; MMPs; Phosphoric acid; Proteases; Proteolytic activity