脐带间充质干细胞改性微环境对体外人牙髓细胞多能性的调控作用
The effects of human umbilical cord mesenchymal stem cells modified microenvironment on the pluripotency of human dental pulp cells in vitro
黄浩权 1刘路 1许喆桢 1韦曦1
作者信息
- 1. 中山大学光华口腔医学院附属口腔医院,广东省口腔医学重点实验室,广东广州510055
- 折叠
摘要
目的:探讨人脐带间充质干细胞(UCMSCs)改性微环境(UM)对人牙髓细胞(DPCs)多能性的调控作用.方法:建立UCMSCs和DPCs间接共培养体系并形成UM.采用流式细胞术、CCK8、SA-β-gal染色及RT-PCR检测细胞周期、增殖、衰老情况;RT-PCR及Western Blot检测细胞多能性相关因子Oct4、Sox2、c-Myc的表达水平;共培养7d后,对各组细胞进行成牙本质、成脂、成软骨向诱导,分别进行茜素红、油红0、阿尔辛蓝染色定量分析,并利用RT-PCR检测多向分化相关基因(DSPP、PPARγ、Sox9)的表达水平;采用ELISA检测UCMSCs的碱性成纤维细胞生长因子(bFGF)分泌量,并通过单独或联合使用bFGF以及FGF受体拮抗剂SU5402对DPCs进行预处理,检测UM和bFGF对DPCs多能性以及多向分化能力的影响.结果:UM能促进DPCs增殖及多能性相关因子基因和蛋白的表达水平,并提高DPCs的多向分化能力,同时抑制DPCs的衰老.SU5402抑制UM及bFGF对DPCs增殖、多能性因子、多向分化能力的促进作用(除UM对DPCs成软骨向分化的促进作用外),并促进DPCs衰老.结论:UM能促进DPCs增殖、提高多能性相关因子的表达、增强多向分化能力以及抑制衰老,可能与UCMSCs分泌bFGF有关.
Abstract
AIM:To investigate the effects of human umbilical cord mesenchymal stem cells modified microenvironnent (UM) on the pluripotency of human dental pulp cells (DPCs) in vitro.METHODS:DPCs were cultured in UM via an indirect co-culture system consist of UCMSCs and DPCs established by using transwell.DPCs proliferation,senescence,cell cycle distribution and pluripotent factors were detected by CCK-8,flow cytometry analysis,SA-β-gal staining,RT-PCR and western blot.Tri-lineage differentiation potential was compared by specifically staining quantitative analysis and RT-PCR respectively.To measure the content of bFGF secreted by UCMSCs,ELISA method was used.bFGF and SU5402 (FGF receptor inhibitor) were used respectively or in combination to prove the effects of UM and bFGF on DPCs.RESULTS:Proliferation,the level of mRNA and proteins related to pluripotentcy and the ability of multi-differentiation of DPCs were enhanced and the senescence of DPCs was depressed by UM and bFGF respectively or in combination.Except for the chondrogenic differentiation,SU5402 restrained the enhancement of UM on DPCs,but boosted cell senescence.CONCLUSION:UM promotes the pluripotency of DPCs mainly through bFGF secreted by UCMSCs.
关键词
牙髓细胞/脐带间充质干细胞/微环境/碱性成纤维细胞生长因子/多能性Key words
dental pulp cells/umbilical cord mesenchymal stem cells/microenvironment/basic fibroblast growth factor/pluripotency引用本文复制引用
基金项目
国家自然科学基金(81400499)
国家自然科学基金(81670983)
广东省科技计划(2015A020212004)
出版年
2017
NETL
NSTL
维普
万方数据