Latorre Karen L, Baldion Paula A
Grupo de Investigaciones Básicas y Aplicadas en Odontología (IBAPO), Universidad Nacional de Colombia, Bogotá, Colombia.
Int J Dent. 2020 Dec 29;2020:8813979. doi: 10.1155/2020/8813979. eCollection 2020.
Dentinal hypersensitivity is a frequent reason for dental consultation, and its pathophysiology has not been fully clarified. Previous findings have made it possible to establish a relationship between the cellular sensory capacity and the activation of the polymodal transient receptor potential vanilloid 1 (TRPV1), which is responsible for the nociceptive response and whose desensitization could cause analgesia. Thus, the objective of this study was to determine the expression, localization, and functional activity of TRPV1 in human odontoblasts-like-cells (hOLCs) and the effect of eugenol (EUG) on its activation and desensitization. Human dental pulp stem cells (hDPSCs) were obtained from third molars and were characterized using flow cytometry, and their differentiation potential toward the osteoblastic, chondrogenic, and adipogenic lineages was investigated. Subsequently, the hDPSCs underwent odontogenic differentiation for 7, 14, and 21 days, and their phenotype (odontogenic markers dentin matrix protein-1 (DMP-1) and dentin sialoprotein (DSP)) was evaluated using immunofluorescence. The TRPV1 gene expression in hOLCs was estimated using RT-qPCR, and its localization was analyzed using immunofluorescence. Half-maximal effective concentration (EC50) from both eugenol (EUG) and capsaicin (CAP) was determined; in addition, receptor activation was evaluated against chemical, thermal, and pH stimuli. For the statistical analysis, a one-way ANOVA with a Tukey post hoc test ( < 0.05) was used. After establishing the model of hOLCs and the membrane location of TRPV1, its chemical activation with EUG and CAP was demonstrated, as well as its thermal activation at ≥ 43°C and with an acidic (<6) or basic pH (between 9 and 12). Receptor desensitization was achieved after 20 min of exposure to two concentrations of EUG (603.5 and 1000 M). These findings represent a stepping-stone for the construction of a pulp pain study model oriented toward a therapeutic alternative for the treatment of dentinal hypersensitivity.
牙本质过敏是牙科就诊的常见原因，其病理生理学尚未完全阐明。以往的研究结果使得建立细胞感觉能力与多模式瞬时受体电位香草酸受体1（TRPV1）激活之间的关系成为可能，TRPV1负责伤害性反应，其脱敏可导致镇痛。因此，本研究的目的是确定TRPV1在人成牙本质细胞样细胞（hOLCs）中的表达、定位和功能活性，以及丁香酚（EUG）对其激活和脱敏的影响。从第三磨牙获取人牙髓干细胞（hDPSCs），并使用流式细胞术对其进行表征，研究其向成骨、软骨和成脂谱系的分化潜能。随后，hDPSCs进行7、14和21天的牙源性分化，并使用免疫荧光评估其表型（牙源性标记物牙本质基质蛋白-1（DMP-1）和牙本质涎蛋白（DSP））。使用RT-qPCR估计hOLCs中TRPV1基因的表达，并使用免疫荧光分析其定位。确定了丁香酚（EUG）和辣椒素（CAP）的半数最大有效浓度（EC50）；此外，针对化学、热和pH刺激评估受体激活情况。对于统计分析，使用单因素方差分析和Tukey事后检验（<0.05）。在建立hOLCs模型和TRPV1的膜定位后，证明了其与EUG和CAP的化学激活，以及在≥43°C和酸性（<6）或碱性pH（9至12）下的热激活。在暴露于两种浓度的EUG（603.5和1000μM）20分钟后实现了受体脱敏。这些发现为构建针对牙本质过敏治疗替代方案的牙髓疼痛研究模型奠定了基础。
