siRNA沉默LKB1基因激活Hedgehog信号通路及对人乳腺癌裸鼠移植瘤模型生长的实验研究

Silencing LKB1 by siRNA activated Hedgehog signaling pathway and the growth of xenografted breast carcinoma in nude mice

目的探讨应用小分子干扰RNA(small interfering RNA,siRNA)沉默抑癌基因LKB1对人乳腺癌细胞MDA-MB-231中Hedgehog信号通路相关因子的表达及人乳腺癌裸鼠移植瘤模型的肿瘤生长的影响。方法构建LKB1基因siRNA质粒LKB1-siRNA;建立LKB1表达抑制的MDA-MB-435细胞模型;裸鼠乳晕皮下接种,建立人乳腺癌裸鼠移植瘤动物模型;成瘤后,观察肿瘤体积变化、裸鼠生存时间;并用Western印迹法检测瘤组织中LKB1和Hedgehog信号通路中信号肽Shh、Sufu、膜受体Ptch、Smo、转录因子Gli1、Hip蛋白表达的变化。结果 LKB1-siRNA质粒组裸鼠的肿瘤体积明显增长(P<0.05);肿瘤内LKB1基因表达水平明显下降,而Hedgehog信号通路相关因子Shh、Gli1、Ptch、Smo的表达升高,Hedgehog信号通路抑制因子Sufu、Hip表达下降。结论 LKB1基因siRNA能够明显抑制人乳腺癌裸鼠移植模型的LKB1基因的表达,上调Hedgehog信号通路相关因子的表达,促进肿瘤生长。LKB1基因和Hedgehog信号通路在乳腺癌细胞中呈现负相关表达。

Objective To investigate the effect of silencing LKB1 by small interfering RNA（siRNA）on the expression of the correlation factor of Hedgehog signaling pathways in human breast cancer MDA-MB-231 cells and the growth of xenografted breast carcinoma in nude mice.Methods Plasmids of siRNA for LKB1 gene were constructed.RNA interference technique was used to silence LKB1 gene in breast carcinoma cells,xenografted tumor model was established in nude mice by subcutaneous inoculation of MDA-MB-231 cells.The tumor volume and survival time of nude mice were recorded.The expression of LKB1,Shh,Sufu,Gli 1,Ptch,Smo and Hip was measured by Western blotting.Results The tumor size was significantly increased in LKB1-siRNA treated group（P0.01）.Western blotting analysis showed that the expression of LKB1 in xenografted tumor was markedly decreased and the correlation factor of Hedgehog signaling pathways was significantly elevated compared with control groups.But the inhibiting factor of Hedgehog signaling pathways was markedly decreased.Conclusion LKB1-siRNA markedly silenced the expression of LKB1 gene,up-regulated the expression of the correlation factor of Hedgehog signaling pathways,advanced the growth of xenografted tumor.LKB1 and Hedgehog signaling pathways were negatively correlated in breast cancer cells.