摘要
目的 探讨乙酰肝素酶 (HPSE)反义寡脱氧核苷酸对人乳腺癌细胞株MDA435侵袭力的抑制作用。方法 设计合成互补于HPSEmRNA起始密码区的硫代反义寡脱氧核苷酸AS ODN及相应的无义对照寡脱氧核苷酸NS ODN ,以阳离子脂质体Lipofectin包埋后转染MDA435细胞 ,以逆转录 聚合酶链反应 (RT PCR)检测转染后HPSEmRNA表达的变化 ,以Western印迹检测HPSE蛋白表达的变化 ,以定量Matrigel侵袭实验检测MDA435细胞侵袭力的变化。结果 与正常对照组、脂质体对照组和NS ODN相应浓度组相比 ,AS ODN各浓度组对HPSEmRNA和蛋白的表达及细胞侵袭力均有明显抑制作用 (P <0 0 1) ,其抑制效应在不同浓度组间差别显著 (P <0 0 1)。AS ODN在终浓度为 0 1μmol/L、0 2 μmol/L和 0 4μmol/L时对MDA435细胞的侵袭力抑制率分别为 34 0 %、5 7 8%和 79 7%。结论 HPSE反义寡脱氧核苷酸通过下调HPSEmRNA和蛋白的表达可显著抑制人乳腺癌细胞株MDA435的侵袭力 ,并呈剂量依赖性。
Objective To evaluate the inhibitory effect of heparanase antisense oligodeoxynucleotide (AS-ODN) on the invasiveness of human mammary carcinoma cell line MDA435. Methods The AS-ODN complementary to the start codon region of heparanase mRNA and its control, scrambled nonsense oligodeoxynucleotide (NS-ODN) were designed and synthesized and phosphorothioated. The ODNs were embedded in cationic liposome Lipofectin and transfected into MDA435 cells. The total RNAs and proteins were extracted from the cells 48 hours after transfection and then semi-quantitative RT-PCR and Western bloting were performed to evaluate the heparanase gene and protein expression levels respectively. The invasiveness of transfected MDA435 cells were measured quantitatively by Matrigel invasion assays. Results The heparanase gene and protein expression and invasiveness of MDA435 cells treated with AS-ODN of different final concentrations were significantly decreased compared with that of the controls (P<0.01). Besides, the inhibitory effects were significantly different between the cells treated with AS-ODN of different concentrations (P<0.01). The invasiveness inhibition rates were 34.0%, 57.8% and 79.7% at the cells treated with AS-DON of the final concentrations of 0.1 μmol/L, 0.2 μmol/L, and 0.4 μmol/L, respectively. Conclusion Heparanase AS-DON complementary to the start codon region of heparanase mRNA has a significant inhibitory effect on the invasiveness of human mammary carcinoma cell line in a dose-dependent manner.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2003年第3期204-207,共4页
National Medical Journal of China
