摘要
目的:利用RNA干扰技术在裸鼠体内抑制COX-2基因表达,探讨RNA干扰对胃癌治疗的特异性和相关机制。方法:设计靶向COX-2基因的siRNA,构建重组表达质粒pTZU6+1-siRNA-COX-2并导入裸鼠皮下移植瘤,在体外诱导RNA干扰,采用RT-PCR法、免疫组化法同时检测处理组和对照组COX-2基因表达,Western blotting检测了与细胞增殖相关信号分子p53、PCNA及Ki67蛋白表达。结果:pTZU6+1-siRNA-COX-2导入裸鼠皮下移植瘤后14天,肿瘤体积明显变小。COX-2的mRNA表达由98.36%下调到43.44%,COX-2蛋白表达由86.24%下调至47.59%。PCNA及Ki67蛋白表达均明显下调。对照组各指标均无明显变化。结论:siRNA明显抑制了COX-2表达,并抑制肿瘤生长,这可能与上调p53蛋白和下调PCNA及Ki67蛋白表达有关。
Objective: To investigate the specificity and inhibitory mechanism of RNA interference technology for gastric carcinoma gene therapy by inhibiting COX-2 in nude mice. Methods: Small interference RNAs targeting COX-2 gene were designed, pTZU6+1-siRNA-COX-2 vector was constructed and transfected into hypodermical gastric tumor in nude mice to induce RNAi. The changes of COX-2 gene expression were detected with RT-PCR and immunochemistry, and the expression of PCNA and Ki67 were tested by Western blot. Results: The expression of COX-2 was remarkably inhibited by RNAi in vivo. Tumor size decreased significantly after pTZU6+1-siRNA-COX-2 vector was injected to tumor; the mRNA level of COX-2 decreased from 98.36% to 43.44%, and its protein expression also reduced from 86.24% to 47.59%. PCNA and Ki67 proteins were down-regulated. In contrast, there were almost no changes in control group. Conclusion: RNAi inhibits the COX-2 gene expression and tumor growth with good specificity, which may be the result of up-regulating of p53 protein and down-regulating PCNA and Ki67 proteins.
出处
《重庆医科大学学报》
CAS
CSCD
2006年第4期552-555,565,共5页
Journal of Chongqing Medical University