靶向TrKA基因的SiRNA干扰对乳腺癌细胞生长的影响

    Effect of SiRNA interference to TrKA gene on the growth of breast cancer cells

    • 摘要: 目的:观察TrKA对乳腺癌细胞生长的影响,并初步探讨其分子机制,为乳腺癌治疗提供新的药物靶点。方法:荧光定量PCR和Western-blot检测干扰前后乳腺癌细胞株中TrKA的表达水平;MTT法检测干扰后对神经生长因子的促细胞生长作用的影响;Western-blot检测干扰后caspase-3的蛋白活性变化。结果:干扰后TrKAmRNA和蛋白表达分别下降73.8%和80.5%(P<0.05);NGF能促进乳腺癌细胞株MCF-7增殖(P<0.01);MCF-7+NGF组在OD490nm值,各时间点高于MCF-7组(P<0.01);干扰后抑制了NGF的促细胞增殖作用,TrKA-SiRNA和TrKA-SiRNA+NGF组在490nm处,各时间点均低于MCF-7(P<0.01);Western-blot结果显示,与MCF-7组相比TrKA-SiRNA和TrKA-SiRNA+NGF组caspase-3蛋白明显活化,cleavedcaspase-3增加(P<0.05)。结论:TrKA的SiRNA干扰有效抑制乳腺癌细胞的生长,并有助于细胞趋向凋亡,提示NGF-TrKA生物学轴对乳腺癌的生长起重要作用,阻断此生物学轴可能是乳腺癌治疗新的靶点。

       

      Abstract: Objective: To observe the effect of TrKA to the growth of breast cancer cells,investigate the molecular mechanism and provide new target of medicine for the treatment of breast cancer.Methods: The expression of TrKA mRNA and protein was detected by Real Time-PCR and Western-blot before and after interference.The proliferated effect of nerve growth factorl(NGF) was tested by MTT and the protein expression of caspase-3 was detected by Western-blot after RNAi.Results: The levels of TrKA mRNA and protein were decreased by 74.7% and 80.5% respectively after RNAi(P< 0.01);NGF could stimulate the proliferation of MCF-7 cells.The OD490 of the group of MCF-7+NGF was higher than that of the group of MCF-7 at each time point(P<0.01).The effect of NGF to the proliferation of cells was inhibited after RNAi.The absorption of the group TrKA-SiRNA and TrKA-SiRNA+NGF were lower than the group MCF-7 at each time(P<0.01);the protein of caspase-3 was activated in the group of TrKA-SiRNA and TrKA-SiRNA+NGF compared with MCF-7.The protein expression of cleaved caspase-3 was increased(P<0.01).Conclusions: Transfection of TrKA gene SiRNA can inhibit cellular proliferation and promote apoptosis.This suggests that the biologic axis of NGF-TrKA plays a siginificant role in the growth of breast cancer and the axis of interference may be a new target for the breast cancer treatment.

       

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