Immunohistochemical and biochemical assay of MMP-3 in human dentine

Objective: The function of endogenous MMP-3 and its distribution within the human dentine is unclear. Thus, the aim of the present study was to assay the presence and distribution of MMP-3 within human sound dentine by means of biochemical and immunohistochemical assays. Methods: Powdered dentine from extracted human teeth was prepared and (1) partially demineralised with 1% H3PO4 for 10 min or (2) untreated (control). The presence of MMP-3 was measured using a colorimetric assay system (QuantiSir (TM), Epigentek, USA). Additional cryo-fractured dentine fragments were processed for immunohistochemical identification of MMP-3 under FEI-SEM. Casein-zymography was used to investigate MMP-3 activity. Results: MMP-3 detected level was 2.732 ng/mu L in partially demineralised dentine powder, whilst it increased to 3.280 ng/mu L in mineralised dentine. The FEI-SEM analysis revealed positive immunolabelling patterns for MMP-3, predominantly localized on the intertubular collagen fibrillar network showing MMP-3 directly or indirectly bound to the collagen fibrils. Casein-zymograms showed positive proteolytic activity for MMP-3 in demineralised dentine powder. Conclusion: The results of the study clearly revealed the presence and distribution of MMP3 in human sound dentine. Whilst the presence was verified, its role is still unclear. Future studies are needed to investigate the possible involvement of MMP-3 in physiological and pathological condition of the dentine-pulp complex. (C) 2011 Elsevier Ltd. All rights reserved.