Hydroxamate-Based Histone Deacetylase Inhibitors as Potential Mediators to Induce Dentine Regeneration by Human Dental Pulp Cell

Abstract

Human dental pulp cells (hDPCs) have shown their plasticity when treated with the hydroxamate-based histone deacetylase (HDAC) inhibitor members, Trichostatin A (TSA), and suberoylanilide hydroxamic acid (SAHA). However, a comparison of their potency to stimulate odontoblast-like differentiation and mineralization has not been reported. The aim of our study was to confirm and compare these TSA and SAHA effects. Primary hDPCs cultured with/without various TSA or SAHA concentrations were evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), ALP activity, alizarin red staining, and scratch wound healing assays. The inhibitory effect of TSA and SAHA on inhibiting the activity of HDAC was evaluated by HDAC activity assay. Odontoblast-related gene expression was determined using RT-qPCR. The MTT assay indicated that TSA or SAHA did not affect hDPC viability. TSA or SAHA treatment-induced odontoblast-like differentiation as evidenced by a significant increase in alkaline phosphatase activity and mineral deposition after 400 nM TSA or 1 μM SAHA treatment. A significant increase in nuclear factor I C, kruppel like factor 4, dentin matrix acidic phosphoprotein 1, dentin sialophosphoprotein, collagen type I alpha 1 chain, alkaline phosphatase (ALPL), integrin-binding sialoprotein, bone gamma-carboxyglutamate protein, vascular endothelial growth factor A, and cyclin-dependent kinase inhibitor 1A gene expression analyzed by RT-qPCR, at 24, 72 h, 7, and 10 days of treatment. The activity of HDAC in hDPCs culture was significantly inhibited after 72 h TSA and SAHA treatment. The scratch wound healing assay displayed enhanced cell migration at 72 h after TSA or SAHA treatment. Our findings demonstrated that TSA and SAHA have similar stimulatory effects in inducing HDPC odontogenic differentiation and mineralization and propose another potential use of TSA and SAHA to promote dentin regeneration.

Tissue engineering approaches for dental pulp regeneration: The development of novel bioactive materials using pharmacological epigenetic inhibitors

“…TSA is a potent pan-HDAC isoform inhibitor, non-selectively targeting various HDACs, which has garnered significant attention within dental pulp regenerative contexts [ 346 ]. TSA demonstrates significant anti-proliferative effects in primary DPCs when applied at 100 nM and 400 nM without affecting cell viability [ 347 , 348 ], reflecting previous reports using TSA at much lower concentrations [ 349 , 350 ]. Notably, TSA-induced caspase-3 and -9 cleavage were observed in DPSCs, a process associated with apoptosis [ 351 ].…”

“…Discrepancies exist between studies regarding the effects of VPA on specific gene expression patterns due to variations in experimental methods and models. For instance, BMP-2 and DMP-1 upregulation was observed in VPA-treated primary DPCs under osteogenic induction [ 347 , 348 ], an effect which was not mirrored in non-induced MDPC-23 [ 353 ], underscoring the importance of representative experimental frameworks moving forward.…”

“…SAHA, also known as vorinostat, is another pan-HDACi like TSA, with FDA approval for treating T-cell lymphoma. Previous research has shown that benzamides, such as MS-275, and hydroxamic acids, like TSA and SAHA, exhibit cytotoxic effects under constant, high-concentration exposure in cell culture [ 346 , 348 ] and in primary DPCs [ 363 ]. Moreover, a negative correlation exists between HDACi treatment (VPA and SAHA) and bone density, an effect that has been attributed to derive from the susceptibility of immature osteoblasts to HDACi-mediated cytotoxicity, a property not held by terminally differentiated osteoblasts [ [364] , [365] , [366] ].…”

Inhibition of histone deacetylases class I improves adipogenic differentiation of human periodontal ligament cells

“…For example, Jin et al [35] reported that 50 nM increased growth, while 100 nM had a cytotoxic effect on DPSCs. On the contrary, Sulistyowati et al [36] reported that 200 nM TSA is an appropriate dose to treat DPCs in vitro without affecting their cell viability. Regarding VPA, a wide range of concentrations (0.01-100 mM) have been reported in human dental pulp cells [33,34,37] pointing out that higher VPA concentrations have an antiproliferative effect.…”

Silica-based silver nanocomposite 80S/Ag as Aggregatibacter actinomycetemcomitans inhibitor and its in vitro bioactivity

“…Moreover, the increased pH values after immersion in simulated body fluid would help neutralize the acidic oral environment, where the bacteria damage teeth and the surrounding regions. 27 Various genes were involved in odontoblast-like cell differentiation and mineralization, namely the dentine remineralization and regeneration, 28 such as dentin matrix acidic phosphoprotein 1 (DSPP), dentin sialophosphoprotein (DSP), dentine matrix protein 1 (DMP-1), collagen type I alpha 1 chain (COL1A1), alkaline phosphatase (ALPL), integrin-binding sialoprotein (IBSP), bone gamma-carboxyglutamate protein (BGLAP), vascular endothelial growth factor A (VEGFA), cyclin dependent kinase inhibitor 1A (CDKN1A), etc . Additionally, GPC-1(glypican-1) and TGF-β1(transforming growth factor beta-1) have been reported to be related to the early stages of odontoblast-like cell differentiation, 29 with the downregulation of the GPC-1 gene observed for the formation of reparative dentine.…”