PGC-1α-shRNA慢病毒载体对牙髓干细胞分化的影响  

Effects of PGC-1α gene silencing by PGC-1α-shRNA mediated with lentiviral vector on the differentiation of dental pulp stem cells

作　　者：王子露[1] 曹灵[1] 郑阳玉[1] 杨迷芳[1] 王舒舒[1] 于金华[1] 

机构地区：[1]南京医科大学口腔医学研究所,江苏南京210029

出　　处：《口腔生物医学》2011年第2期67-70,共4页Oral Biomedicine

摘　　要：目的:构建过氧化物酶体增殖物激活受体γ辅激活因子1α(peroxisome proliferators activated receptor γ coactivator 1α,PGC-1α)短发夹RNA(short hairp in RNA,shRNA)慢病毒表达载体,体外评价其对牙髓干细胞(dental pulp stem cells,DPSC)分化的影响。方法:根据shRNA设计原则,设计合成用于体内干扰PGC-1α编码序列的shRNA,重组入慢病毒载体PLKO.1。将构建正确的PLKO.1/PGC-1α-shRNA感染DSPC,实时荧光定量PCR检测PGC-1α的表达,细胞化学染色和实时荧光定量PCR检测其对DPSC分化的影响。结果:测序证实重组质粒构建成功;体外试验表明,DPSC转染了PGC-1α-shRNA后PGC-1α表达水平降低,干扰效率达81%,而且细胞钙化结节增多,牙本质涎磷蛋白和Ⅰ型胶原的表达明显上调,与对照组相比具有显著性差异(P<0.05)。结论:慢病毒介导的PGC-1α-shRNA成功在体外抑制了目的基因的表达,并影响牙髓干细胞的分化。Objective：To construct a lentiviral plasmid of RNA interference of peroxisome proliferators activated receptor γ coactivator 1α（PGC-1α）gene,and evaluate the interference effect of PGC-1α-shRNA on the differentiation of dental pulp stem cells（DPSC） in vitro.Methods：According to the target sequence and design principle of shRNA,the short hairpin RNA was designed and synthesized to interfere the PGC-1α expression,and the shRNA duplex was ligated into the PLKO.1 vector.DPSC was transfected with PLKO.1/PGC-1α-shRNA,and the expression level of PGC-1α was detected by real time RT-PCR;cytochemical stainings and real time RT-PCR were used to investigate the differentiation potential of DPSC.Results：The recombinant vector was successfully constructed and confirmed by sequencing.In vitro experiment demonstrated that the expression level of PGC-1α was significantly downregulated,and the interfering efficiency was 81%;The down-regulation of PGC-1α not only significantly increased the calcified nodules,but also significantly increased the expression level ofdentin sialosphoprotein and collagen type Ⅰ.Conclusions：RNA interference can successfully interfere with the expression of PGC-1α and affect the differentiation of DPSC.

关 键 词：短发夹RNA 慢病毒载体 过氧化物酶体增殖物激活受体γ辅激活因子1α 牙髓干细胞 

分 类 号：Q786[生物学—分子生物学]