摘要
目的研究干细胞表面标记CD24基因对根尖牙乳头干细胞成骨分化能力的影响。方法利用CD24 shRNA基因敲除CD24进行丧失性功能研究;实时定量RT-PCR检测CD24的基因敲除效果;通过碱性磷酸酶(ALP)活性检测、茜素红染色及钙离子定量分析明确根尖牙乳头干细胞体外成骨分化能力的变化;实时荧光定量RT-PCR检测成骨分化相关基因-Ⅰ型胶原蛋白1A、Ⅰ型胶原蛋白1B、骨涎蛋白、骨桥蛋白和成骨相关转录因子RUNX2的表达,分析研究根尖牙乳头干细胞基因表达的改变。结果实时定量RT-PCR结果显示CD24可以在根尖牙乳头干细胞有效的被敲除;碱性磷酸酶ALP活性结果显示敲除CD24抑制根尖牙乳头干细胞碱性磷酸酶ALP活性;茜素红染色及钙离子定量分析结果显示敲除CD24明显抑制根尖牙乳头干细胞体外矿化能力;实时定量RT-PCR结果显示敲除CD24明显抑制I型胶原蛋白1A、Ⅰ型胶原蛋白1B、骨涎蛋白、骨桥蛋白和RUNX2的表达。结论基因沉默CD24可以通过抑制转录因子RUNX2及成骨分化基因的表达,从而抑制根尖牙乳头干细胞体外成骨分化功能。
Objective To investigate the effect of CD24 molecule（ CD24） on the osteogenic differentiation potential of stem cells from apical papilla（ SCAPs）. Methods Lentiviral CD24 ShRNA was used to silence the expression of CD24. Real-time fluorescence quantitative RT-PCR was used to detect the knockout effect of CD24. Alkaline phosphatase（ ALP） activity assay was used to detect the ALP activity,which was the early marker of osteogenic differentiation. Alizarin-red staining and calcium quantitative analysis were used to study the potential of SCAPs mineralization in vitro. The expression of osteogenic related genes was detected by real-time fluorescence quantitative RT-PCR,including collagen type I alpha 1 chain（ COL1 A1）,collagen type I alpha 2 chain（ COL1 A2）,bone sialoprotein（ BSP）,osteonectin（ ON） and runt-related transcription factor 2（ RUNX2）. The gene expression changes of SCAPs were analyzed.Results Real time RT-PCR results showed that CD24 could be silenced in SCAPs. ALP activity assay showed that the depletion of CD24 inhibited ALP activity. Alizarin-red staining and calcium quantitative analysis revealed that the knockout of CD24 obviously suppressed the mineralization of SCAPs in vitro. Real-time fluorescence quantitative RT-PCR displayed that the depletion of CD24 obviously depressed the expressions of COL1 A1,COL1 A2,BSP and ON in SCAPs. The RUNX2 expression was repressed in CD24 silenced SCAPs compared with control group. Conclusion The gene silencing of CD24 represses the osteogenic differentiation potential of SCAPs by targeting the RUNX2.
作者
王海锋
刘亚男
刘思思
宁美芝
幸丹
李文峰
曹钰
WANG Haifeng;LIU Ya＇nan;LIU Sisi;NING Meizhi;XING Dan;LI Wenfeng;CAO Yu(Department of Stomatology,Belting Bo＇ ai Hospital,China Rehabilitation Research Center,Beijing 10068,China)
出处
《口腔医学》
CAS
2018年第7期582-586,共5页
Stomatology
基金
国家自然科学基金面上项目(81670948)
中国康复研究中心重点项目(2017ZX-03)
关键词
CD24
根尖牙乳头干细胞
成骨分化
CD24 molecule （CD24）
stem cells from apical papilla （SCAPs）
osteogenic differentiation
