Department of General Thoracic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
J Exp Clin Cancer Res. 2011 May 28;30(1):63. doi: 10.1186/1756-9966-30-63.
The human coagulation trigger tissue factor (TF) is overexpressed in several types of cancer and involved in tumor growth, vascularization, and metastasis. To explore the role of TF in biological processes of lung adenocarcinoma, we used RNA interference (RNAi) technology to silence TF in a lung adenocarcinoma cell line A549 with high-level expression of TF and evaluate its antitumor effects in vitro and in vivo.
The specific small interfering RNA (siRNA) designed for targeting human TF was transfected into A549 cells. The expression of TF was detected by reverse transcription-PCR and Western blot. Cell proliferation was measured by MTT and clonogenic assays. Cell apoptosis was assessed by flow cytometry. The metastatic potential of A549 cells was determined by wound healing, the mobility and Matrigel invasion assays. Expressions of PI3K/Akt, Erk1/2, VEGF and MMP-2/-9 in transfected cells were detected by Western blot. In vivo, the effect of TF-siRNA on the growth of A549 lung adenocarcinoma xenografts in nude mice was investigated.
TF -siRNA significantly reduced the expression of TF in the mRNA and protein levels. The down-regulation of TF in A549 cells resulted in the suppression of cell proliferation, invasion and metastasis and induced cell apoptosis in dose-dependent manner. Erk MAPK, PI3K/Akt pathways as well as VEGF and MMP-2/-9 expressions were inhibited in TF-siRNA transfected cells. Moreover, intratumoral injection of siRNA targeting TF suppressed the tumor growth of A549 cells in vivo model of lung adenocarcinoma.
Down-regulation of TF using siRNA could provide a potential approach for gene therapy against lung adenocarcinoma, and the antitumor effects may be associated with inhibition of Erk MAPK, PI3K/Akt pathways.
人类凝血酶原酶组织因子（TF）在多种类型的癌症中过度表达，参与肿瘤生长、血管生成和转移。为了探讨 TF 在肺腺癌生物学过程中的作用，我们使用 RNA 干扰（RNAi）技术沉默 TF 在 TF 高表达的肺腺癌细胞系 A549 中，并在体外和体内评估其抗肿瘤作用。
设计针对人 TF 的特异性小干扰 RNA（siRNA）转染 A549 细胞。采用逆转录-聚合酶链反应（RT-PCR）和 Western blot 检测 TF 的表达。MTT 和集落形成实验检测细胞增殖。流式细胞术检测细胞凋亡。划痕愈合、迁移和基质胶侵袭实验检测 A549 细胞的转移潜能。Western blot 检测转染细胞中 PI3K/Akt、Erk1/2、VEGF 和 MMP-2/-9 的表达。体内实验观察 TF-siRNA 对裸鼠 A549 肺腺癌移植瘤生长的影响。
TF-siRNA 显著降低 TF 在 mRNA 和蛋白水平的表达。TF 在 A549 细胞中的下调呈剂量依赖性抑制细胞增殖、侵袭和转移，并诱导细胞凋亡。Erk MAPK、PI3K/Akt 通路以及 VEGF 和 MMP-2/-9 的表达均受到抑制。此外，TF 靶向 siRNA 的瘤内注射抑制了 A549 细胞在肺腺癌体内模型中的肿瘤生长。
使用 siRNA 下调 TF 可为肺腺癌的基因治疗提供一种潜在的方法，其抗肿瘤作用可能与抑制 Erk MAPK、PI3K/Akt 通路有关。
