Collagen cross-linkers on dentin bonding: Stability of the adhesive interfaces, degree of conversion of the adhesive, cytotoxicity and in situ MMP inhibition

Viviane Hass, Issis Virginia Luque-Martinez, Mario Felipe Gutierrez, Camila Guimarães Moreira, Valéria Bisinoto Gotti, Victor Pinheiro Feitosa, Garrit Koller, Michel Fleith Otuki, Alessandro D. Loguercio*, Alessandra Reis

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

116 Scopus citations


Objective To investigate the effect of collagen cross-links on the stability of adhesive properties, the degree of conversion within the hybrid layer, cytotoxicity and the inhibition potential of the MMPs' activity. Methods The dentin surfaces of human molars were acid-etched and treated with primers containing: 6.5 wt% proanthocyanidin, UVA-activated 0.1 wt% riboflavin, 5 wt% glutaraldehyde and distilled water for 60 s. Following, dentin was bonded with Adper Single Bond Plus and Tetric N-Bond; and restored with resin composite. The samples were sectioned into resin-dentin "sticks" and tested for microtensile bond strength (μTBS) after immediate (IM) and 18-month (18 M) periods. Bonded sticks at each period were used to evaluate nanoleakage and the degree of conversion (DC) under micro-Raman spectroscopy. The enzimatic activity (P1L10 cross-linkers, P1L22 MMPs' activities) in the hybrid layer was evaluated under confocal microscopy. The culture cell (NIH 3T3 fibroblast cell line) and MTT assay were performed to transdentinal cytotoxicity evaluation. Data from all tests were submitted to appropriate statistical analysis (α = 0.05). Results All cross-linking primers reduced the degradation of μTBS compared with the control group after 18 M (p > 0.05). The DC was not affected (p > 0.213). The NL increased after 18 M for all experimental groups, except for proanthocyanidin with Single Bond Plus (p > 0.05). All of the cross-link agents reduced the MMPs' activity, although this inhibition was more pronounced by PA. The cytotoxicity assay revealed reduced cell viability only for glutaraldehyde (p < 0.001). Significance Cross-linking primers used in clinically relevant minimized the time degradation of the μTBS without jeopardizing the adhesive polymerization, as well as reduced the collagenolytic activity of MMPs. Glutaraldeyde reduced cell viability significantly and should be avoided for clinical use.

Original languageEnglish
Pages (from-to)732-741
Number of pages10
JournalDental Materials
Issue number6
StatePublished - 1 Jun 2016
Externally publishedYes

Bibliographical note

Publisher Copyright:
© 2016 Academy of Dental Materials.


  • Dental bonding
  • Dentin collagen
  • Glutaraldehyde
  • Proanthocyanidin
  • Riboflavin


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