Qi S C, Cui C, Yan Y H, Sun G H, Zhu S R
Center of Stomatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei; Department of Stomatology, The Tenth People's Hospital of Tongji University, Shanghai.
Int Endod J. 2013 Dec;46(12):1153-63. doi: 10.1111/iej.12112. Epub 2013 Apr 19.
To investigate the expression of high-mobility group box 1 (HMGB1) in human dental pulp tissues and the effects of HMGB1 on proliferation and odontoblastic differentiation of human dental pulp cells (hDPCs).
Immunohistochemical assay, immunofluorescence staining and flow cytometric analysis were used to detect the expression of HMGB1 in the human dental pulp and hDPCs, respectively. The proliferation of hDPCs was examined by CCK-8 after culturing human primary hDPCs in the presence of HMGB1 with different doses. Odontoblastic differentiation of hDPCs was determined using alkaline phosphatase (ALPase) activity assay and mineralized nodule formation. Important mineralization-related genes such as ALP, dental sialophosphoprotein (DSPP) and dental matrix protein-1 (DMP-1) were determined by real-time polymerase chain reaction. Western blot analysis was performed to determine the difference in expressions of DMP-1 and DSP with or without the presence of exogenous HMGB1. Simultaneously, messenger RNA and protein levels of HMGB1 and RAGE were also detected. The protein level of HMGB1 in the supernatants was quantified using ELISA analysis.
HMGB1 was found in human dental pulp tissue and in the nuclei of hDPCs. During hDPC odontoblastic differentiation, HMGB1 translocated from the nuclei to the cytoplasm and then secreted out from hDPCs. Exogenous HMGB1 promoted hDPC proliferation and mineralized nodule formation. It up-regulated the activity of ALPase and the mRNA and protein levels of dentine matrix protein-1 (DMP-1), alkaline phosphatase (ALP), dentine sialophosphoprotein (DSPP) and receptor for advance glycation end (RAGE) of hDPCs.
HMGB1 promoted the proliferation and odontoblastic differentiation of hDPCs.
研究高迁移率族蛋白B1（HMGB1）在人牙髓组织中的表达及其对人牙髓细胞（hDPCs）增殖和向成牙本质细胞分化的影响。
分别采用免疫组织化学分析、免疫荧光染色和流式细胞术检测HMGB1在人牙髓和hDPCs中的表达。在不同剂量的HMGB1存在下培养人原代hDPCs后，用CCK-8检测hDPCs的增殖情况。通过碱性磷酸酶（ALPase）活性测定和矿化结节形成来确定hDPCs的成牙本质细胞分化。通过实时聚合酶链反应测定重要的矿化相关基因，如碱性磷酸酶（ALP）、牙涎磷蛋白（DSPP）和牙基质蛋白-1（DMP-1）。进行蛋白质免疫印迹分析以确定有无外源性HMGB1时DMP-1和DSP表达的差异。同时，还检测了HMGB1和晚期糖基化终末产物受体（RAGE）的信使核糖核酸和蛋白质水平。用酶联免疫吸附测定（ELISA）分析定量上清液中HMGB1的蛋白质水平。
在人牙髓组织和hDPCs的细胞核中发现了HMGB1。在hDPCs向成牙本质细胞分化过程中，HMGB1从细胞核转移到细胞质，然后从hDPCs中分泌出来。外源性HMGB1促进了hDPCs的增殖和矿化结节的形成。它上调了hDPCs的ALPase活性以及牙本质基质蛋白-1（DMP-1）、碱性磷酸酶（ALP）、牙涎磷蛋白（DSPP）和晚期糖基化终末产物受体（RAGE）的信使核糖核酸和蛋白质水平。
HMGB1促进了hDPCs的增殖和成牙本质细胞分化。
