长链非编码RNATP53TG1促进根尖牙乳头干细胞成牙本质及成骨分化  

Long noncoding RNA TP53TG1 promotes odontogenic and osteogenic differentiation of stem cells from the apical papilla

作　　者：黎庭悦 郭倩 何文喜 吴家媛 Li Tingyue;Guo Qian;He Wenxi;Wu Jiayuan(Stomatological Medical School of Zunyi Medical University,Zunyi 563000,Guizhou Province,China;State Key Laboratory of Military Stomatology,Key Laboratory of Shaanxi Province Stomatology,Department of Dental Pulp Diseases,The Third Affiliated Hospital of Air Force Medical University,Xi’an 710032,Shaanxi Province,China;Department of Stomatology,Special Medical Center,Air Force Medical University,Beijing 100142,China)

机构地区：[1]遵义医科大学口腔医学院,贵州省遵义市563000 [2]军事口腔医学国家重点实验室,陕西省口腔医学重点实验室,空军军医大学第三附属医院牙体牙髓病科,陕西省西安市710032 [3]空军军医大学特色医学中心口腔科,北京市100142

出　　处：《中国组织工程研究》2025年第36期7776-7782,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(82460190),项目负责人:吴家媛;贵州省科技计划项目(黔科合基础-ZK[2022]一般638),项目负责人:吴家媛;北京市自然科学基金项目(7242140),项目负责人:何文喜;陕西自然科学基础研究计划-重点项目(2022JZ-42),项目负责人:何文喜。

摘　　要：背景:长链非编码RNATP53TG1(lncRNATP53TG1)参与调控多种癌症细胞的增殖、迁移、侵袭、凋亡,但在其他细胞中的作用报道甚少。目的:探讨lncRNATP53TG1对人根尖牙乳头干细胞增殖和分化的影响及途径。方法:分离、培养人根尖牙乳头干细胞,转染lncRNA TP53TG1过表达慢病毒,RT-qPCR检测lncRNA TP53TG1的过表达效率,Western blot检测PI3K、AKT、ERK、P38、Smad3及其磷酸化蛋白的相对表达量。将人根尖牙乳头干细胞分为慢病毒空载体组和过表达lncRNATP53TG1组,采用CCK-8法检测细胞增殖情况;成骨诱导第5天采用碱性磷酸酶染色检测碱性磷酸酶活性,成骨诱导第21天采用茜素红染色检测矿化结节形成情况;成骨诱导第3,7,14天采用RT-qPCR检测牙本质涎磷蛋白、Runt相关转录因子2、牙本质基质蛋白1、骨涎蛋白的mRNA表达水平;成骨诱导第3,7,14天采用Westernblot检测牙本质涎磷蛋白和Runt相关转录因子2的蛋白表达水平。结果与结论:①RT-qPCR检测结果显示慢病毒成功整合到根尖牙乳头干细胞基因组中,Western blot检测结果显示,过表达lncRNA TP53TG1上调了p-PI3K、p-AKT的蛋白水平而不影响其他通路磷酸化蛋白的表达;②从细胞培养第3天开始,过表达lncRNA TP53TG1显著促进根尖牙乳头干细胞的增殖;③在诱导根尖牙乳头干细胞牙源性分化过程中,过表达lncRNA TP53TG1促进成牙本质及成骨分化相关基因和蛋白的表达,碱性磷酸酶活性和矿化结节形成显著增加;④结果表明:lncRNA TP53TG1可能通过激活PI3K/AKT信号通路促进根尖牙乳头干细胞成牙本质及成骨分化。BACKGROUND:Long noncoding RNA TP53TG1(lncRNA TP53TG1)is involved in regulating the proliferation,migration,invasion,and apoptosis of various cancer cells,but there are few reports on its role in other cells.OBJECTIVE:To investigate the effects and pathways of lncRNA TP53TG1 on the proliferation and differentiation of human stem cells from the apical papilla.METHODS:Human stem cells from the apical papilla were isolated and cultured,and then transfected with lncRNA TP53TG1 overexpression lentivirus.RT-qPCR was used to detect the overexpression efficiency of lncRNA TP53TG1.Western blot assay was used to detect the relative expression levels of PI3K,AKT,ERK,P38,Smad3,and their phosphorylated proteins.Human stem cells from the apical papilla were divided into the empty lentiviral vector transfection group and the lncRNA TP53TG1 overexpression group.CCK-8 assay was used to measure the cell proliferation.Alkaline phosphatase activity was detected by alkaline phosphatase staining on day 5 of osteogenic induction.Formation of mineralized nodules was detected by alizarin red staining on day 21 of osteogenic induction.RT-qPCR was used to detect the mRNA expression levels of dentin sialophosphoprotein,Runt-related transcription factor 2,dentin matrix protein 1,and bone sialoprotein on days 3,7,and 14 of osteogenic induction.Western blot assay was used to detect the protein expression levels of dentin sialophosphoprotein and Runt-related transcription factor 2 on days 3,7,and 14 of osteogenic induction.RESULTS AND CONCLUSION:(1)RT-qPCR results showed that the lentivirus was successfully integrated into the genome of stem cells from the apical papilla.Western blot assay results showed that overexpression of lncRNA TP53TG1 up-regulated the protein levels of p-PI3K and p-AKT without affecting the expression of phosphorylated proteins in other pathways.(2)Starting from day 3 of cell culture,overexpression of lncRNA TP53TG1 significantly promoted the proliferation of stem cells from the apical papilla.(3)In the process of inducin

关 键 词：根尖牙乳头干细胞 长链非编码RNA lncRNA TP53TG1 病毒转染 成牙本质向分化 PI3K/AKT信号通路 工程化干细胞 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学] R781