Objectives: We hypothesised that the application of two HDACis, Valproic acid (VPA) and Trichostatin A (TSA), would induce repair-associated responses in primary dental pulp (DPC) cultures at concentration levels which do not stimulate significant anti-proliferative, apoptotic or cytotoxic effects.
Methods: VPA (0.03mM-5mM) and TSA (3nM-400nM) were applied to primary rat DPCs (24hrs) and their stimulation of reparative processes was investigated using proliferation, viability, apoptosis, cell cycle and mineralization assays.
Results: Flow cytometry demonstrated that TSA (100nM, 400nM) significantly increased cell viability. Neither HDACi was cytotoxic, although cell-growth analysis revealed significant anti-proliferative effects at higher concentrations for VPA (>0.5mM) and TSA (>50nM). While high-content-analysis demonstrated that HDACi did not significantly induce caspase-3 or p21 activity, p53-expression was increased by VPA (3mM, 5mM) at 48hrs. HDACi-exposure induced mineralization per cell dose-dependently to a plateau level (VPA-0.125mM and TSA-25nM) with accompanying increases in mineralization/dentinogenic-associated gene expression at 5 (DMP-1, BMP-2/-4, Nestin) and 10 days (DSPP, BMP-2/-4). Both HDACis, at a range of concentrations, significantly stimulated osteopontin and BMP-2 protein expression at 10 and 14 days further supporting the ability of HDACi to promote differentiation
Conclusion: HDACi exert different effects on primary compared with transformed DPCs and promote epigenetically-induced mineralization and differentiation events without cytotoxic effects. These results highlight the potential benefits of low concentration HDACi for topical application in vital pulp therapy.