非小细胞肺癌患者血清MMP-9表达及siRNA靶向抑制A549细胞侵袭与转移的研究

目的:探讨非小细胞肺癌患者血清中MMP-9表达与肿瘤转移的关系;MMP-9siRNA对肺腺癌细胞系A549中MMP-9表达及细胞侵袭与转移抑制作用的影响。方法:ELISA法检测32例非小细胞肺癌(NSCLC)患者及20名健康者空腹血清MMP-9浓度,同时设计合成针对MMP-9的特异性siRNA,采用oligofectamine转染A549细胞,RT-PCR法检测转染细胞后MMP-9mRNA的表达,Transwell实验检测细胞侵袭情况。结果:NSCLC患者血清中MMP-9浓度〔(63.44±7.84)ng/mL〕比正常对照组〔(21.57±3.29)ng/mL〕明显升高(P<0.01),且浓度随病理分期增加而增强〔Ⅰ/Ⅱ:(45.40±5.30)ng/mL,Ⅲ/Ⅳ:(89.80±5.02)ng/mL,P<0.01〕,发生淋巴结转移的患者血清MMP-9浓度〔(82.26±5.09)ng/mL〕明显高于未发生淋巴结转移〔(35.92±4.68)ng/mL〕的患者P<0.01;RT-PCR法显示,转染MMP-9siRNA后,细胞中MMP-9mRNA表达较阴性对照组和空白组明显降低,P<0.05。RNA干扰MMP-9基因后A549细胞的侵袭能力也有明显下降,P<0.01。结论:MMP-9可作为NSCLC患者有无发生浸润转移的良好预测指标,靶向MMP-9的siRNA不仅能特异性的降低MMP-9mRNA及蛋白的表达,且能抑制人A549细胞的侵袭和迁移。

Expression of serum MMP-9 level in non-small cell lung cancer and inhibition of siRNA targeting MMP-9 on ivasion and migration of A549 cells

OBJECTIVE:To investigate correlation of serum total MMP-9 levels with human non-small cell lung cancer(NSCLC) metastasis,the effect of MMP-9 siRNA on MMP-9 expression of A549 cells and the inhibition effect on A549 cell invasion and migration.METHODS: Serum total MMP-9 levels were detected in 32 patients with human non-small cell lung cancer and 20 healthy volunteers by ELISA with monoclonal antibody against human total MMP-9.To design and synthesize siRNA targeting MMP-9,then transfect it into A549 cells with oligofectamine.RT-PCR was used to observe the expression of MMP-9 mRNA.The transwell experiment was used to detect the invasion ability of cells.RESULTS: The level of MMP-9 was significantly elevated in cancer samples((63.44±7.84) ng/mL) compared with normal serum((21.57±3.29) ng/mL,P<0.01) and became stronger with the TNM-stage increasing(Ⅰ/Ⅱ:(45.40±5.30) ng/mL,Ⅲ/Ⅳ:(89.80±5.02) ng/mL,P<0.01).A significant difference was found between levels in the presence((82.26±5.09) ng/mL) versus the absence of metastasis((35.92±4.68) ng/mL,P<0.01).RT-PCR indicated that the expressions of MMP-9 mRNA in A549 cells interfered by MMP-9 siRNA were significantly lower than those of other two groups(P<0.05).The invasion and migration ability of A549 cells were also decreased after siRNA interference of MMP-9 gene(P<0.01).CONCLUSIONS:It suggests that MMP-9 can be used as a marker for the detection of metastasis.The targeting RNA interference not only decreases the expression of MMP-9 mRNA,but also inhibits the invasion and migration ability of A549 cells.