u-PAR反义核酸载体的构建与高侵袭人前列腺癌细胞PC-3M亚系的转染

目的:为了特异封闭PC-3M高侵袭亚系尿激酶型纤溶酶原激活物受体(u-PAR)的表达,观察其对侵袭能力的抑制效应。方法: 应用RT-PCR获得u-PAR的cDNA片段,反向插入pcDNA3质粒载体中,构建u-PAR反义核酸载体并导入高侵袭亚系中;借助RT-PCR和免疫组化检测转染细胞u-PAR的表达。 结果:u-PAR反义核酸载体转染株的u-PAR mRNA和蛋白质水平明显低于对照组,抑制率分别为53%和73%,同时其体外侵袭能力亦明显降低,抑制率为79%。 结论: u-PAR反义核酸在PC-3M高侵袭亚系中发挥了特异封闭作用,为进一步观察u-PAR反义核酸抑制高侵袭前列腺癌细胞亚系的侵袭效应提供了细胞模型。

Construction of antisense RNA expression plasmid for u-PAR and its transfection to highly invasive PC-3M cell subclones

AIM: To inhibit specifically the u-PAR expres sion in highly invasive cell subclones and block its function in those cells in vasion. METHODS: A cDNA fragment of u-PAR obtained by RT-PCR was i nserted i nto a plasmid vector named pcDNA3 in the reverse direction. Then an antisense RN A expression plasmid for u-PAR was constructed and transfected into highly invas ive cell subclones. The u-PAR expression in resistant cells was examined by RT-P CR and immunohistochemical assay. RESULTS: Compared to the control cells, the content of mRNA and protein of u-PAR in transfected cells decreased sharply, and the rate of inhibit ion was 53% and 73%, respectively. CONCLUSION: The results indicated that an antisense vector for u -PAR might played a specific inhibitory role in the cells. This model is useful for observing the inhibitory effects of the antisense vector for u-PAR on invasi on by highly invasive cell subclones of human prostate carcinoma.