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

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

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。

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 nonsmall 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 ceils 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.