圈套寡核苷酸抑制NIH3T3细胞α2Ⅰ型胶原基因表达的研究

目的 研究激活剂蛋白1(activator protein-1,AP—1)圈套寡核苷酸(decoy-oligodeoxynu-cleotides,Decoy-ODNs)对成纤维细胞α2I型胶原表达的影响，探讨病理性腋痕的基因治疗。方法 设计合成针对AP—1的Decoy-ODNs，用阳离子脂质体转染NIH3T3细胞，观察Decoy-ODNs在细胞中的分布；用凝胶迁移变动分析(electrophoretic mobility shift assay,EMSA)研究Decoy-ODNs对AP—1的抑制作用，采用RT—PCR观察其对细胞胶原合成的影响。结果 AP—1的Decoy-ODNs可在体外竞争抑制核转录因子AP—1的活性；阳离子脂质体可以将Decoy-ODNs转染进入细胞浆及细胞核从而发挥作用；Decoy-ODNs作用24h后，NIH3T3细胞α2I型胶原mRNA的表达明显降低。结论 De—coy-ODNs可以通过拮抗核转录因子AP—1的活性而抑制α2I型胶的表达。

An experimental study of inhibiting effect of decoy oligodeoxynucleotides on the gene expression of collagen in NIH3T3 cells

OBJECTIVE: To investigate the effect of activator protein-1 (AP-1) decoy-oligodeoxynucleotides (Decoy-ODNs) on the expression of fibroblast alpha2 type I collagen, so as to explore the gene therapy of pathologic scar. METHODS: Decoy-ODNs targeting AP-1 were designed and synthesized. NIH3T3 cells were transfected by cationic liposomes. The distribution of Decoy-ODNs in the cells was investigated. The inhibiting effects of Decoy-ODNs on AP-1 were determined by electrophoretic mobility shift assay (EMSA). And the effects of Decoy-ODNs on the collagen synthesis in the cells were analyzed by RT-PCR. RESULTS: AP-1 Decoy-ODNs could competitively inhibit the AP-1 in vitro activity. Cationic liposomes could play roles by effectively transfecting Decoy-ODNs into the plasma and nucleus. The mRNA expression of fibroblast alpha2 type I collagen decreased evidently after 24 hours of Decoy-ODNs action. CONCLUSION: Decoy-ODNs could inhibit the mRNA expression of fibroblast alpha2 type I collagen by antagonizing AP-1.