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作 者:万宗淼[1] 张斌[1] 肖俊[1] 刘远[1] 戴闽[1]
出 处:《南昌大学学报(医学版)》2017年第1期13-18,22,共7页Journal of Nanchang University:Medical Sciences
基 金:江西省自然科学基金青年项目(20151BAB215001)
摘 要:目的探究转录因子Brachyury上调基质金属蛋白酶(MMPs)与促进肺癌细胞转移的机制。方法建立Brachyury过表达和低表达的肺癌稳转细胞株,分析Brachyury转录因子对NSCLC细胞侵袭能力的影响;通过RT-PCR和real-time PCR技术比较23个MMPs的表达差异;原发性NSCLC组织标本免疫组织化学验证Brachyury与MMPs表达的关系。CHIP-PCR实验检测Brachyury对MMPs成员启动子是否存在调控位点。结果Brachyury转录因子能显著促进肺癌细胞的侵袭能力;Brachyury高表达的稳转细胞内MMP12、MMP24表达显著增加323%(P=0.02)、183%(P=0.40)。Brachyury低表达的稳转细胞内MMP7、MMP9、MMP10、MMP12和MMP19显著降低34%(P=0.09)、43%(P=0.08)、56%(P=0.07)、77%(P=0.002)和23%(P=0.04);且免疫组织化学进一步验证了Brachyury与MMPs的表达具有显著一致性。CHIP-PCR结果显示MMP12启动子区域(转录起始位点上游5′-266/-1443bp 3′)存在PCR显著扩增。结论多个基质金属蛋白酶的表达与转录因子Brachyury表达相关,MMP12可能与Brachyury的作用更加明显,这可能是Brachyury能促进NSCLC细胞转移的机制之一。Objective To investigate the mechanisms of upregulation of matrix metalloproteinases(MMPs) and promotion of lung cancer cell metastasis by transcription factor brachyury . Methods Lung cancer cell lines with over-expression and down-expression of brachyury were constructed to observe the effect of brachyury on invasion ability of non-small cell lung cancer (NSCLC) cells. The expression of 23 MMPs was detected by RT-PCR and real-time PCR . The re-ationship between brachyury and MMPs was analyzed by immunohistochemistry in primary NSCLC specimens. The potential brachyury-binding sites in promoters of MMPs were determinedby CHIP-PCR. Results Brachyury promoted the invasion of lung cancer cells. The expression ofMMP12 and MMP24 was increased by 323% ( P = 0. 02) and 183% P = 0. 40) in cells with brachyury over-expression, respectively. However, the expression of MMP7, MMP9, MMP10, MMP12 and MMP19 was decreased by 34% (P= 0. 09),43% (P= 0. 08),56% (P= 0. 07),77% (P= 0. 002) and 23% (P= 0. 04) in cells with brachyury down-expression, respectively. Immuno- histochemistry contirmed a remarkable consistency in the expression of brachyury and MMPs. CHIP-PCR showed positive amplitication of MMP12 promoter sequence(-266/-1443 bp). Conclu-sion Multiple MMPs, especially MMP12,are correlated with the expression of transcription fac-tor brachyury . The correlation may be one of mechanisms of brachyury promoting metastasis me-tastasis.
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