反义ST6GalⅠ基因对HeLa细胞黏附和侵袭力的影响

Effects of ST6Gal Ⅰ antisense oligonucleotide-mediated gene silencing on cell adhesion and invasiveness of HeLa cells

目的研究人α2，6-唾液酸转移酶（ST6GalⅠ）反义寡核苷酸（ASODN）转染对ST6GalⅠ高表达的官颈癌HeLa细胞的黏附和侵袭力的影响。方法设计并合成靶向ST6GalⅠ的ASODN及正义寡核苷酸（SODN），用脂质体Lipofectamine 2000包裹后转染HeLa细胞。实验设空白对照组、脂质体对照组、SODN组和ASODN组。应用RT-PCR测定细胞中ST6GalⅠ mRNA水平，流式细胞术检测细胞表面α2，6-唾液酸含量，分别用CytoMatrix^TM细胞黏附试剂盒和细胞侵袭分析试剂盒，检测细胞对细胞外基质（ECM）的黏附与侵袭力。结果HeLa细胞被转染48h后，与空白对照组、脂质体对照组和SODN组相比，ASODN组细胞中ST6GalⅠ mRNA表达明显下调（P〈0．01）；与3个对照组相比，ASODN组细胞表面以，6-唾液酸的含量明显降低，同时细胞对ECM的黏附和侵袭力均降低（均P〈0．05）。结论靶向ST6GalⅠ的ASODN能下调HeLa细胞ST6GalⅠ基因的表达，降低细胞表面α2，6-唾液酸含量，继而降低细胞对ECM的黏附和侵袭力。

Objective To study the effects of antisense oligonucleotide （ASODN）targeting ST6Gal Ⅰ on cell adhesion and invasiveness of human cervical carcinoma cell line HeLa which over-expressed ST6Gal Ⅰ. Methods ASODN and sense oligonucleotide （SODN） targeting ST6Gal Ⅰ were designed and constructed, and transfected into a cervical cancer cell line, HeLa, by lipofectmine 2000. HeLa cells were cultured and divided into 4 groups：blank control group, liposome group, SODN group and ASODN group. RT-PCR was used to examine the ST6Gal Ⅰ mRNA expression. Flow cytometry was used to examine the amount of α2, 6-sialylation on the HeLa cell surface. The HeLa cell adhesion and invasiveness to extracellular matrix （ECM） were analyzed by using CytoMatrix^TM kit and cell invasion assay kit, respectively. Results The expression of ST6Gal Ⅰ mRNA in HeLa cells at 48 hrs after transfection in the ASODN group was significantly decreased in comparison with that in the blank control group, liposome group, and SODN group（P 〈0.01 ）. The amount of α2,6-sialylation on cell surface in ASODN group was significantly lower than that of the other 3 groups （ P 〈 0.05 ）. The adhesion and invasiveness of the cells in the ASODN group decreased remarkably, both significantly lower than those of the other 3 groups （ all P 〈 0.05）. Conclusion Specific ASODN targeting ST6Gal Ⅰ effectively inhibits HeLa cell ST6Gal Ⅰ expression, decreases the amount of or2,6-sialylation on cell surface and leads to a decline of cell adhesion and invasiveness to ECM. This result also established a fine base for further studying on anti-tumor treatment with antisense oligonucleotide.