Department of Endodontics, College of Stomatology, Tianjin Medical University, Num12, Road Qixiangtai, Heping district, Tianjin, 300070, China.
Department of Stomatology, Baodi Clinical College of Tianjin Medical University, Tianjin Baodi Hospital, Tianjin, China.
J Mol Histol. 2017 Dec;48(5-6):367-377. doi: 10.1007/s10735-017-9735-2. Epub 2017 Sep 13.
Odontoblasts have been suggested to contribute to nociceptive sensation in the tooth via expression of the transient receptor potential (TRP) channels. The TRP channels as a family of nonselective cation permeable channels play an important role in sensory transduction of human. In this study, we examined the expression of transient receptor potential vanilloid-1 (TRPV1), transient receptor potential vanilloid-2 (TRPV2) and transient receptor potential vanilloid-3 (TRPV3) channels in native human odontoblasts (HODs) and long-term cultured human dental pulp cells with odontoblast phenotyoe (LHOPs) obtained from healthy wisdom teeth with the use of immunohistochemistry (IHC), immunofluorescence (IF), quantitative real-time polymerase chain reaction (qRT-PCR),western blotting (WB) and immunoelectron microscopy (IEM) assay. LHOPs samples were made into ultrathin sections, mounted on nickel grids, floated of three TRPV antibodies conjugated with 10 nm colloidal gold particles and observed under IEM at 60,000 magnifications. The relative intracellular distributions of these three channels were analyzed quantitatively on IEM images using a robust sampling, stereological estimation and statistical evaluation method. The results of IHC and IF convinced that TRPV1, TRPV2 and TRPV3 channels were expressed in native HODs and (LHOPs). The result of qRT-PCR and WB confirmed that the gene and protein expression of TRPV1, TRPV2, and TRPV3 channels and TRPV1 mRNA are more abundantly expressed than TRPV2 and TRPV3 in HODs (P < 0.05). Quantitative analysis of IEM images showed that the relative intracellular distributions of these three channels are similar, and TRPV1, TRPV2 and TRPV3 proteins were preferential labeled in human odontoblast processes, mitochondria, and endoplasmic reticulum. Thus, HODs could play an important role in mediating pulp thermo-sensation due to the expression of these three TRPV channels. The difference of relative intracellular distributions of three channels suggests that special structures such as processes may have an important role to sensing of the outer stimuli first.
成牙本质细胞被认为通过表达瞬时受体电位 (TRP) 通道参与牙齿的伤害感受。TRP 通道作为非选择性阳离子可渗透通道家族，在人类感觉转导中发挥重要作用。在这项研究中，我们使用免疫组织化学（IHC）、免疫荧光（IF）、实时定量聚合酶链反应（qRT-PCR）、Western blot（WB）和免疫电镜（IEM）检测法，检测了天然人成牙本质细胞（HODs）和长期培养的具有成牙本质细胞表型的人牙髓细胞（LHOPs）中瞬时受体电位香草醛 1（TRPV1）、瞬时受体电位香草醛 2（TRPV2）和瞬时受体电位香草醛 3（TRPV3）通道的表达。LHOPs 样本制成超薄切片，安装在镍网格上，用 10nm 胶体金标记的三种 TRPV 抗体漂浮，在 60,000 倍放大倍率下在 IEM 下观察。使用稳健采样、立体学估计和统计评估方法，在 IEM 图像上对这三种通道的相对细胞内分布进行定量分析。IHC 和 IF 的结果证实 TRPV1、TRPV2 和 TRPV3 通道在天然 HODs 和 LHOPs 中表达。qRT-PCR 和 WB 的结果证实 TRPV1、TRPV2 和 TRPV3 通道的基因和蛋白表达以及 TRPV1 mRNA 在 HODs 中比 TRPV2 和 TRPV3 更丰富（P<0.05）。IEM 图像的定量分析表明，这三种通道的相对细胞内分布相似，TRPV1、TRPV2 和 TRPV3 蛋白优先标记在人成牙本质细胞突起、线粒体和内质网中。因此，由于这三种 TRPV 通道的表达，HODs 可能在介导牙髓热敏感觉中发挥重要作用。三种通道的相对细胞内分布差异表明，突起等特殊结构可能在对外界刺激的感知中发挥重要作用。
