Isolation of dentin matrix proteins by EDTA and calcium hydroxide

Objectives: Under the influence of EDTA (ethylenediaminetetraaceticacid) it is possible to dissolve regenerative factors from dentin matrix, which can be used for in vivo and in vitro transformation of pulp fibroblasts to odontoblast-like cells as well as for production of reparative dentin. The aim of this study was to compare the dentin matrix dissolving capacity of EDTA to that of calcium hydroxide, because EDTA is, unlike calcium hydroxide, not appropriate for clinical applications to maintain pulp viability.

Methods: Powdered dentin of human third molars, incubated with solutions of 10% EDTA and 23mM calcium hydroxide for one and 14 days respectively, followed by dialysis, lyophilization, DEAE (diethylaminoethyl cellulose) ion exchange chromatography and SDS (sodium dodecyl sulphate) paging were used. Protein content of chromatography fractions was estimated by using the Lowry method, detection of proteins after electrophoresis by Gonski silver staining.

Results: EDTA dissolving capacity was 1.5 times higher for 14 days of incubation and 3.5 times higher for one day incubation compared to calcium hydroxide. 14 day incubation with calcium hydroxide opposite one day resulted in 158% higher protein content, while incubation with EDTA for 14 days vs. one day resulted in 3.7% higher protein content only. However proteins with molecular weight of about 21.5 and 14.5 kDa, e.g. as growth factors, could be dissolved by the use of EDTA exclusively.

Conclusions: The method used is sufficient to show a time dependent dentin matrix dissolving capacity of EDTA and calcium hydroxide but does not allow to draw conclusions about the influence of calcium hydroxide on the growth factor mediated regeneration of the pulp-dentin complex.