杨益民, 李黔宁, 应大君, 龚自力, 成戎川, 吕敏, 刘勇, 周竹娟, 郑健. 形成三链DNA的寡核苷酸抑制切应力诱导的大鼠内皮细胞组织因子表达J. 药学学报, 2006, 41(9): 808-813.
引用本文: 杨益民, 李黔宁, 应大君, 龚自力, 成戎川, 吕敏, 刘勇, 周竹娟, 郑健. 形成三链DNA的寡核苷酸抑制切应力诱导的大鼠内皮细胞组织因子表达J. 药学学报, 2006, 41(9): 808-813.
YANG Yi-min, LI Qian-ning, YING Da-jun, GONG Zi-li, CHENG Rong-chuan, Lü Min, LIU Yong, ZHOU Zhu-juan, ZHENG Jian. Triplex-forming oligonucleotide inhibits the expression of tissue factor gene in endothelial cells induced by the blood flow shear stress in ratsJ. Acta Pharmaceutica Sinica, 2006, 41(9): 808-813.
Citation: YANG Yi-min, LI Qian-ning, YING Da-jun, GONG Zi-li, CHENG Rong-chuan, Lü Min, LIU Yong, ZHOU Zhu-juan, ZHENG Jian. Triplex-forming oligonucleotide inhibits the expression of tissue factor gene in endothelial cells induced by the blood flow shear stress in ratsJ. Acta Pharmaceutica Sinica, 2006, 41(9): 808-813.

形成三链DNA的寡核苷酸抑制切应力诱导的大鼠内皮细胞组织因子表达

Triplex-forming oligonucleotide inhibits the expression of tissue factor gene in endothelial cells induced by the blood flow shear stress in rats

  • 摘要: 目的检测针对组织因子(TF)基因启动子区切应力反应元件(shear stress response element,SSRE)设计的反向硫代形成三链DNA的寡核苷酸(antiparallel phosphorothioate triplex-forming oligonucleotide,apsTFO)对大鼠颈总动脉狭窄处内皮细胞TF表达的影响。方法采用硅胶管套扎法建立SD大鼠颈总动脉中段重度狭窄模型。应用原位杂交和免疫组织化学方法结合图像分析系统测定狭窄段内膜TF,Egr-1和Sp1的mRNA表达及蛋白合成。术前0.5 h在尾静脉注射0.5 mg·kg-1针对TF的SSRE结合位点设计合成的GT20-apsTFO,GT20-psTFO,GT21-apsTFO及部分绿色荧光素(FITC)标记的apsTFO,术后4 h检测血管内皮细胞内的TFO,术后6 h检测TFO对TF的抑制效果及对Egr-1和Sp1的影响。结果给予TFO后,在血管内皮细胞核中可见荧光分布。GT20-apsTFO和GT21-apsTFO对TF的mRNA表达和蛋白合成有显著性抑制(P<0.05),且GT20-apsTFO的抑制作用较强,与GT21-apsTFO组相比差异有显著性(P<0.05),GT20-psTFO对TF的mRNA表达和蛋白合成无明显抑制(P>0.05)。GT20-apsTFO,GT20-psTFO和GT21-apsTFO对Egr-1及Sp1的mRNA表达和蛋白合成无抑制作用。结论apsTFO能部分进入血管内皮细胞并在一定程度上抑制TF基因的表达,而不影响Egr-1和Sp1基因的表达。

     

    Abstract: AimTo study the effect of antiparallel phosphorothioate triplex-forming oligonucleotide (apsTFO) matching with the shear stress response element (SSRE) of tissue factor (TF) gene promoter region on the expression of TF in endothelial cells (ECs) of rat common carotid artery stenosis. Methods The model of common carotid artery middle segment stenosis was established by silica gel pipe loop ligation in SD rats. The mRNA expression and protein synthesis of TF, early growth response-1 (Egr-1) and specificity protein 1 (Sp1) were measured by in situ hybridization (ISH) and immunohistochemistry (IHC) technique. GT21-apsTFO, GT20-apsTFO, GT20-psTFO and FITC-labeled apsTFO, matching with the SSRE of TF gene promoter region, were designed, and intravenously injected into rats at 0.5 h before operation. TFO was detected 4 h after the operation, and the mRNA expression and protein synthesis of TF, Egr-1 and Sp1 were detected 6 h after the operation. ResultsThere were much fluorescence in vascular tissue, especially in the nuclear of ECs 4.5 h after the injection of apsTFO. The mRNA expression and protein synthesis of TF reduced by 22%-23% with injection of GT20-apsTFO 6.5 h after stenosis (P<0.01) and by 10%-11% with GT21-apsTFO at the same time (P<0.05). The inhibition by GT20-apsTFO was stronger than that of the GT21-apsTFO (P<0.05). The expression of TF was not inhibited by the GT20-psTFO (P>0.05). The mRNA expression and protein synthesis of Egr-1and Sp1 did not change in the rat treated with GT20-apsTFO, GT20-psTFO and GT21-apsTFO (P>0.05). ConclusionapsTFO could mero-inhibit the expression of TF gene but could not change the expression of Egr-1 and Sp1 protein.

     

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