Release of bio‐active dentine extracellular matrix components by histone deacetylase inhibitors (HDACi)

Aim To characterize dentine matrix component (DMC) release and smear layer removal by histone deacetylase inhibitors (HDACis). Methodology DMCs were extracted from powdered human dentine over 14 days using three HDACis, valproic acid (VPA), trichostatin A (TSA) and suberoylanilide hydroxamic acid (SAHA) and compared with a control extractant, 10% (w/v) EDTA. Protein compositions of the resultant extracts were analysed by 1D-polyacrylamide gel electrophoresis (1D-PAGE), TGF-β-1 and MMP-9 ELISAs and a high-throughput growth factor antibody array. Dentine discs with a standardized smear layer were prepared from human molars and treated with EDTA (17% w/v), polyacrylic acid (PA) (20% v/v) and the experimental HDACis prior to analysis by scanning electron microscopy. Parametric ELISA data were analysed using one-way anova and Tukey's post hoc test, whilst nonparametric smear layer data were analysed by Kruskal–Wallis test and Mann–Whitney U-test (P < 0.05). Results HDACis did not remove smear layer in the presence or absence of PA pre-treatment (P ≥ 0.478). 1D-PAGE analysis demonstrated different protein profiles for EDTA and HDACi extracts. All HDACi solutions released TGF-β-1 although less effectively than EDTA (P < 0.001), whilst MMP-9 was extracted in significantly higher concentration by EDTA and VPA compared with TSA (P < 0.012). Antibody array analysis demonstrated the ability of HDACis to extract a complex cocktail of established/novel growth factors from dentine, albeit significantly less efficiently than EDTA for certain cytokines (TGF-β-1, PDGF-AA, VEGF-A) and significantly more effectively for others (GDF-15, IGF-1, EGRF-1, NGFR, BDNF, SCF-R). Conclusions HDACi release a range of bioactive DMCs that could promote dentine repair processes in vivo; however, they are ineffective at removing smear layer.