Division of Restorative Dentistry and Periodontology, Dublin Dental University Hospital, Trinity College Dublin, Dublin, Ireland.
J Endod. 2012 Mar;38(3):339-45. doi: 10.1016/j.joen.2011.12.014. Epub 2012 Jan 9.
Histone deacetylase inhibitors (HDACis) alter the homeostatic balance between 2 groups of cellular enzymes, histone deacetylases (HDACs) and histone acetyltransferases (HATs), increasing transcription and influencing cell behavior. This study investigated the potential of 2 HDACis, valproic acid (VPA) and trichostatin A (TSA), to promote reparative processes in pulp cells as assayed by viability, cell cycle, and mineralization analyses.
VPA (0.125-5 mmol/L) and TSA (12.5-400 nmol/L) were applied to a pulp-derived cell population and compared with unsupplemented controls. Cell proliferation and viability were evaluated by trypan blue staining and cell counting, whereas cell cycle and apoptosis were analyzed by immunocytochemical staining with antibodies for p53, phosphorylated p53, Bcl-2 homologous antagonist/killer (BAK), caspase-3 and p21(WAF1/CIP), and DNA staining with Hoechst 33342. For mineralization analysis, cultures were stained with Alizarin red and quantified spectrophotometrically. Relative gene expression levels of mineralization associated markers were analyzed using reverse-transcriptase polymerase chain reaction. One-way analysis of variance and Tukey post hoc tests were applied to the data (P < .05).
VPA and TSA reduced cell proliferation dose dependently with no significant effect on cell viability except at 400 nmol/L TSA. The transcription factor p21(WAF1/CIP) was significantly increased at the highest concentration of TSA but not VPA. Significant increases (P < .05) in the apoptosis marker protein active caspase-3 and cell cycle alterations were only evident at the maximum concentrations of TSA/VPA, whereas HDACi-induced mineralization per cell was stimulated dose dependently with a significant increase in the expression of the dentinogenic-associated transcript, dentine matrix protein-1.
These results indicate that HDACis are capable of epigenetically modulating pulp cell behavior, signifying their therapeutic potential for augmenting biomaterials, and stimulating regenerative responses in the damaged pulp.
组蛋白去乙酰化酶抑制剂（HDACi）改变了组蛋白去乙酰化酶（HDACs）和组蛋白乙酰转移酶（HATs）这两组细胞酶之间的稳态平衡，增加了转录并影响了细胞行为。本研究通过活力、细胞周期和矿化分析，研究了 2 种 HDACi，丙戊酸（VPA）和曲古抑菌素 A（TSA）促进牙髓细胞修复过程的潜力。
将 VPA（0.125-5mmol/L）和 TSA（12.5-400nmol/L）应用于牙髓细胞群体，并与未补充对照组进行比较。通过台盼蓝染色和细胞计数评估细胞增殖和活力，通过针对 p53、磷酸化 p53、Bcl-2 同源拮抗剂/杀伤（BAK）、半胱天冬酶-3 和 p21（WAF1/CIP）的免疫细胞化学染色以及用 Hoechst 33342 进行 DNA 染色分析细胞周期和细胞凋亡。对于矿化分析，用茜素红染色并通过分光光度法进行定量。使用逆转录聚合酶链反应分析与矿化相关的标记物的相对基因表达水平。对数据进行单因素方差分析和 Tukey 事后检验（P <.05）。
VPA 和 TSA 剂量依赖性地降低细胞增殖，除 TSA 浓度为 400nmol/L 外，对细胞活力没有显著影响。最高浓度的 TSA 显著增加了转录因子 p21（WAF1/CIP），但 VPA 没有。只有 TSA/VPA 的最大浓度才会显著增加凋亡标志物蛋白活性半胱天冬酶-3 和细胞周期改变，而每细胞的 HDACi 诱导矿化则呈剂量依赖性增加，牙本质相关转录物、牙本质基质蛋白-1 的表达显著增加。
这些结果表明，HDACi 能够表观遗传地调节牙髓细胞行为，表明它们具有增强生物材料和刺激受损牙髓中再生反应的治疗潜力。
