过表达MMP1对根尖牙乳头干细胞骨/牙向分化能力的影响

摘要

Objective: To research the effect of matrix metalloproteinase 1(MMP1)overexpression on the osteo/odontogenic differ-entiation of human stem cells from the apical papilla(SCAPs).Methods: SCAPs were isolated and purified from human third molars with developing roots.Retrovirus expression plasmid PMSCV/MMP1 was constructed.The osteo/odontogenic differentiation capacity was tested by alkaline phosphatase(ALP)activity,alizarin red staining,Western blot assay,and real-time RT-PCR.Results: The ALP activity assay and alizarin red staining showed that ALP activity and mineralization capacity of MMP1 overexpression were de-creased.The results of RT-PCR and Western bolt indicated that the mRNA levels of COL1,OCN,RUNX2,OSX and DSPP were down-regulated and the protein levels of DSP,RUNX2,OSX and OCN were decreased.Conclusions: MMP1 overexpression downregulates the osteo/odontogenic differentiation potential of stem cells from apical papilla.%目的:研究过表达基质金属蛋白酶1(MMP1)对根尖牙乳头干细胞(SCAPs)骨/牙向分化能力的影响.方法:从牙根未发育完全的第三磨牙上摘取根尖牙乳头组织,采用酶消法和组织块法结合的方法分离纯化得到SCAPs.设计构建MMP1过表达质粒,转染SCAPs.通过碱性磷酸酶(ALP)活性检测试剂盒、茜素红染色、Western blot和实时定量RT-PCR检测MMP1过表达组和空载质粒组的骨/牙向分化相关指标的变化.结果:成功构建SCAPs MMP1过表达模型;ALP 活性检测和茜素红染色结果显示MMP1过表达组的ALP活性降低和矿化能力减弱,Western bolt结果显示MMP1过表达组的Runt相关转录因子2(RUNX2)、成骨细胞特异性转录因子Osterix(OSX)、牙本质涎蛋白(DSP)、骨钙素(OCN)的表达降低,实时定量RT-PCR结果显示Ⅰ型胶原蛋白基因(COL1)、OCN、OSX、RUNX2、牙本质涎磷蛋白基因(DSPP)表达降低.结论:过表达 MMP1 抑制SCAPs的骨/牙向分化潜能.