Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

Investigation of the mechanisms of coagulation factor Ⅶa-induced colon cancer 8W620 cell proliferation and migration

Objective To investigate the mechanisms that coagulation factor Ⅶa promotes proliferation and migration of a colon cancer cell line (SW620 cells) in vitro. Methods The expression of interleukin 8 (IL-8), tissue factor (TF), caspase-7 and p-p38 MAPK in SW620 cells treated with factor Ⅶa or protease activated receptor 2 aganist (PAR2-AP) was measured by ELISA, Western-blotting and QT-PCR. Results Factor Ⅶa and PAR2-AP induced IL-8 expression at both mRNA and protein levels, up-regulated TF mRNA expression and TF activity, but down-regulated caspese-7 mRNA and p-p38 MAPK levels in SW620 cells. The effects of factor Ⅶa were not only blocked by anti-TF but also by anti-PAR2 antibodies. Conclusion Factor Ⅶa binds to TF on cell surface, forming a complex which activates PAR2, then provoking IL-8 and TF expression, and suppresses caepase-7 expression, thus promotes the tumor cell proliferation and migration, p38 MAPK may negatively regulate this process.     

凝血因子Ⅶa促进SW620细胞增殖与迁移的机制探讨

目的 体外探讨凝血因子Ⅶa促进结肠癌细胞株SW620增殖与迁移的作用机制.方法 采用蛋白酶激活受体2激动剂(PAR2-AP)、凝血因子Ⅶa等处理SW620细胞,以实时定量PCR检测SW620细胞中白细胞介素8(IL-8)、组织因子(TF)及半胱氨酸蛋白酶7(caspase-7)mRNA的表达水平;以酶联免疫吸附试验(ELISA)检测细胞上清IL-8蛋白的含量;以Xa生成法检测细胞TF活性;以Western blot法检测细胞磷酸化p38丝裂原活化蛋白激酶(p-p38 MAPK)水平.结果 PAR2-AP、凝血因子Ⅶa能够增加SW620细胞中IL-8基因和蛋白的表达,上调TF mRNA水平及活性,下调caspase-7基因表达和p-p38 MAPK水平,单克隆抗TF及抗PAR2抗体均可抑制凝血因子Ⅶa的作用.结论 凝血因子Ⅶa与细胞表面TF形成复合物,通过活化PAR2,上调结肠癌细胞株SW620中IL-8、TF的表达,下调细胞caspase-7表达,从而促进细胞增殖与迁移能力,p38 IVIAPK在此过程中起负性调节作用.     

核因子κB在凝血因子Ⅶa促进结肠癌SW620细胞增殖迁移中的作用

目的 探讨核因子κB(NF-κB)在促进结肠癌SW620细胞增殖迁移过程中的作用及其可能的机制。方法 以凝血因子Ⅶa、NF-κB抑制剂二硫代氨基甲酸吡咯烷(PDTC)等处理结肠癌细胞株SW620,采用Western blot法检测细胞核NF-κB(p65)、细胞浆NF-κB抑制蛋白（IκB-o）和凋亡蛋白半胱氨酸蛋白酶7（caspase-7）的蛋白表达变化;采用流式细胞术检测SW620细胞的细胞周期变化;采用Transwell法测定SW620细胞的迁移能力;采用实时定量聚合酶链反应(PCR)检测白细胞介素8( IL-8)和组织因子(TF) mRNA的表达水平。结果凝血因子Ⅶa能够显著下调细胞浆中IκB-o的表达水平,并使细胞核内NF-κB的表达水平升高,单克隆抗TF和抗蛋白酶激活受体2(PAR2)抗体能够抑制凝血因子Ⅶa的这一作用。PDTC能够明显干预凝血因子Ⅶa对SW620细胞增殖、迁移的促进作用。PDTC能够明显干预凝血因子Ⅶa对SW620细胞中TF、IL-8 mRNA表达的促进作用和对caspase-7蛋白表达的下调作用。结论 凝血因子Ⅶa与细胞表面TF结合形成TF/Ⅶa复合物,活化受体PAR2,经NF-κB通路上调IL-8、下调caspase-7的表达,促进SW620细胞的增殖与迁移。TF/Ⅶa/PAR2/NF-κB通路还可进一步上调TF的表达,从而形成TF/Ⅶa/PAR2/NF-κB/TF正反馈通路。     

EGCG对因子Ⅶa促进SW620细胞增殖与迁移的干预作用及机制

目的探讨表没食子儿茶素没食子酸酯(EGCG)对因子Ⅶa促进SW620细胞增殖与迁移的干预作用及机制。方法用EGCG、因子Ⅶa等处理SW620细胞,应用流式细胞术、Transwell法分别检测细胞增殖周期及迁移能力;Western blotting检测细胞NF-κB抑制蛋白(IκB-α)、核NF-κB(p65/RelA)的蛋白水平变化。结果 EGCG(100 mg/L)能干预因子Ⅶa对SW620细胞增殖与迁移的促进作用;EGCG能抑制因子Ⅶa对IκB-α表达的下调作用及对核NF-κB(p65/RelA)表达的促进作用。结论 EGCG可能通过干预信号分子IκB-α和NF-κB(p65/RelA)的表达和调节作用,抑制因子Ⅶa对SW620细胞增殖与迁移的促进作用。     

Involvement of PKCα activation in TF/VIIa/PAR2-induced proliferation, migration, and survival of colon cancer cell SW620

Our previous study has demonstrated that protease-activated receptor 2 (PAR2) activation mediated by tissue factor (TF)/VIIa complex triggers the ERK1/2/NF-κB signaling pathway, which further contributes to the proliferation and migration of colon cancer cell line SW620. However, the detailed mechanisms remain unclear. This study was to investigate whether protein kinase Cα (PKCα) is involved in these events and the possible mechanism. The results revealed that PAR2-activating peptide or VIIa could induce time-dependent upregulation of PKCα phosphorylation in SW620 cells and PKCα translocation from the cytoplasm to the perinuclear region and nucleus. The activation of PKCα was sufficient to induce ERK1/2 and NF-κB phosphorylation. The VIIa effect was obviously blocked by both anti-TF and anti-PAR2 antibodies. The PKCα inhibitor, safingol, inhibited ERK1/2 phosphorylation and NF-κB activation that is induced by VIIa and abrogated the enhanced proliferation, migration, and survival of SW620 cells by VIIa treatment. Both safingol and PDTC (NF-κB inhibitor) could apparently rescue the effects of VIIa on expression of MMP-9, caspase-3, TF, and Bcl-2/bax in SW620 cells. Collectively, the data in this study suggest that TF/VIIa/PAR2-induced SW620 cell proliferation, migration, and survival are ascribed to the activation of PKCα, and these effects are achieved through PKCα downstream signaling pathways, ERK1/2 and NF-κB.     

Involvement of ERK1/2/NF-κB signal transduction pathway in TF/FVIIa/PAR2-induced proliferation and migration of colon cancer cell SW620

Our previous study has demonstrated that TF/FVIIa and protease-activated receptor 2 (PAR2) are closely related to the proliferation and migration of colon cancer cell line SW620. However, the detailed signaling cascades and underlying molecular mechanisms remain unclear. This study has investigated whether extracellular signal-regulated kinase 1 and 2 (ERK1/2) and nuclear factor kappaB (NF-κB) signaling pathways are involved in the events. The results revealed that PAR2-activating peptide (PAR2-AP) or FVIIa elicited time-dependent upregulation of ERK1/2 phosphorylation in SW620 cells, and the effect of FVIIa was significantly attenuated by anti-TF antibody. PAR2-AP or FVIIa also increased NF-κB (p65/RelA) levels among cell nuclear proteins and simultaneously decreased IκB-α levels in the cytoplasmic proteins. Such effects of FVIIa can be inhibited with anti-PAR2 or anti-TF antibodies. While ERK1/2 inhibitor (U0126) intervened with the regulatory effects of PAR2-AP and FVIIa on IκB-α/NF-κB (p65/Rel) expression in the cells, NF-κB inhibitor (PDTC) partially blocked the enhancing effects of PAR2-AP and FVIIa on the proliferating and migratory ability of SW620 cells. Furthermore, the regulatory effects of PAR2-AP and FVIIa on expressions of certain proteins (IL-8, caspase-7, and TF) were also significantly abolished by PDTC. Collectively, the data in this study suggest that the interaction between FVIIa and TF induces PAR2 activation, thereby triggers the ERK1/2 and IκB-α/NF-κB signal transduction pathway to regulate the gene expression of IL-8, TF, and caspase-7, and ultimately promotes SW620 cell proliferation and migration.