Tsai M-C, Chen K-D, Wang C-C, Huang K-T, Wu C-H, Kuo I-Y, Chen L-Y, Hu T-H, Goto S, Nakano T, Dorling A, McVey J H, Chen C-L, Lin C-C
Division of Hepato-Gastroenterology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan; Graduate Institute of Clinical Medical Sciences, Chang Gung University College of Medicine, Kaohsiung, Taiwan.
Center for Translational Research in Biomedical Sciences, Liver Transplantation Program and Department of Surgery, Kaohsiung Chang Gung Memorial Hospital , Kaohsiung, Taiwan.
Cell Death Discov. 2015 Nov 30;1:15051. doi: 10.1038/cddiscovery.2015.51. eCollection 2015.
We previously demonstrated PAR2 starts upstreamed with tissue factor (TF) and factor VII (FVII), inhibited autophagy via mTOR signaling in HCC. However, the mechanism underlying for merging functions of PAR2 with the coagulation system in HCC progression remained unclear. The present study aimed to investigate the role of TF, FVII and PAR2 in tumor progression of HCC. The expressions of TF, FVII and PAR2 from HCC specimens were evaluated by immunohistochemical stains and western blotting. We found that the expression of FVII, but not TF and PAR2, directly related to the vascular invasion and the clinical staging. Importantly, a lower level of FVII expression was significantly associated with the longer disease-free survival. The addition of FVII but not TF induced the expression of PAR2 and phosphorylation of ERK1/2, whereas knockdown of FVII decreased PAR2 expression and ERK1/2 phosphorylation in HCC cell lines. Furthermore, levels of phosphor-TSC2 (Ser664) were increased after treatment with FVII and PAR2 agonist whereas these were significantly abolished in the presence of a potent and specific MEK/ERK inhibitor U0126. Moreover, mTOR knockdown highly reduced Hep3B migration, which could be reverted by FVII but not TF and PAR2. These results indicated that FVII/PAR2 signaling through MEK/ERK and TSC2 axis for mTOR activation has potent effects on the migration of HCC cells. In addition, FVII/PAR2 signaling elicits an mTOR-independent signaling, which promotes hepatoma cell migration in consistent with the clinical observations. Our study indicates that levels of FVII, but not TF, are associated with tumor migration and invasiveness in HCC, and provides clues that evaluation of FVII expression in HCC may be useful as a prognostic indicator in patients with HCC and may form an alternative target for further therapy.
我们之前证明，蛋白酶激活受体2（PAR2）在组织因子（TF）和凝血因子VII（FVII）的上游启动，通过哺乳动物雷帕霉素靶蛋白（mTOR）信号通路抑制肝癌中的自噬。然而，PAR2与凝血系统在肝癌进展中的合并功能的潜在机制仍不清楚。本研究旨在探讨TF、FVII和PAR2在肝癌肿瘤进展中的作用。通过免疫组织化学染色和蛋白质印迹法评估肝癌标本中TF、FVII和PAR2的表达。我们发现，FVII的表达而非TF和PAR2的表达与血管侵犯和临床分期直接相关。重要的是，较低水平的FVII表达与更长的无病生存期显著相关。添加FVII而非TF可诱导PAR2的表达和细胞外信号调节激酶1/2（ERK1/2）的磷酸化，而敲低FVII可降低肝癌细胞系中PAR2的表达和ERK1/2的磷酸化。此外，用FVII和PAR2激动剂处理后，磷酸化结节性硬化复合物2（TSC2，Ser664）的水平升高，而在存在强效特异性丝裂原活化蛋白激酶/细胞外信号调节激酶（MEK/ERK）抑制剂U0126的情况下，这些水平显著消除。此外，敲低mTOR可高度降低Hep3B细胞的迁移，FVII可使其恢复，但TF和PAR2则不能。这些结果表明，FVII/PAR2信号通过MEK/ERK和TSC2轴激活mTOR对肝癌细胞的迁移具有强大作用。此外，FVII/PAR2信号引发一种不依赖mTOR的信号，促进肝癌细胞迁移，这与临床观察结果一致。我们的研究表明，FVII的水平而非TF的水平与肝癌中的肿瘤迁移和侵袭相关，并提供线索表明评估肝癌中FVII的表达可能作为肝癌患者的预后指标有用，并且可能形成进一步治疗的替代靶点。
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