乙酰肝素酶反义寡核苷酸对人乳腺癌细胞株侵袭力的抑制作用

The inhibitory effect of heparanase antisense oligodeoxynucleotides on the invasiveness of human mammary carcinoma cell line

目的 探讨乙酰肝素酶 (HPSE)反义寡脱氧核苷酸 (AS ODN )对人乳腺癌细胞株侵袭力的抑制活性。方法 设计合成分别互补于HPSEmRNA 5 '未翻译区和起始密码区的AS ODN1、AS ODN2及相应对照NS ODN1、NS ODN 2 ,在脂质体介导下以 2 5 0nmol/L终浓度转染人乳腺癌MDA43 5细胞 ,以RT PCR和Westernblot分别检测HPSEmRNA和蛋白表达 ,以基质凝胶侵袭实验检测细胞侵袭力。结果 AS ODN 2组的HPSEmRNA相对表达量、蛋白表达量和侵袭细胞数(0 .62± 0 .0 4、12 .70± 1.70、61.0 0± 9.0 0 )与脂质体对照组 (1.60± 0 .0 4、3 6.2 0± 2 .10、178.0 0±17.0 0 )和NS ODN2组 (1.48± 0 .0 3、3 5 .5 0± 2 .3 0、174.0 0± 2 0 .0 0 )相比较均显著降低 (P <0 .0 1) ;而AS ODN 1组与脂质体对照组和NS ODN1组相比较差异均无显著性 (P >0 .0 5 )。结论 互补于起始密码区的HPSEAS ODN可通过下调HPSE表达显著抑制人乳腺癌细胞株MDA43 5的侵袭力。

Objective To evaluate the inhibitory effect of heparanase antisense oligodeoxynucleotide (AS-ODN) on the invasiveness of human mammary carcinoma cell line.Methods AS-ODN1 and AS-ODN2,complementary to the 5′untranslation region and to the start codon region of heparanase mRNA respectively,and their controls,NS-ODN1 and NS-ODN2,were designed and synthesized.The ODNs with the final concentration of 250 nmol/L were delivered into MDA435 cells by cationic liposome.After transfection the heparanase gene and protein expression level were evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot,respectively and the cell invasiveness was measured by Matrigel invasion assay.Results The heparanase gene expression level,protein expression level and invasion cells in AS-ODN2 group (0.62±0.04,12.70±1.70 and 61.00±9.00,respectively) decreased significantly compared with those in liposome control group (1.60±0.04,36.20± 2.10 and 178.00±17.00) and in NS-ODN2 control group (1.48±0.03,35.5±2.3 and 174.00± 20.00) (P<0.01).In contrast,those in AS-ODN1 group were not significantly different from those in liposome control group and in NS-ODN1 control group (P>0.05).Conclusion Heparanase AS-ODN complementary to the start codon region could significantly inhibit the invasiveness of human mammary carcinoma cell line MDA435 by downregulating heparanase expression.