目的探讨孤儿核因子受体(Nur77)在胃癌组织中表达及其与患者预后状况之间联系,研究Nur77在胃癌细胞侵袭与迁移进程中的作用机制。方法利用Oncomine数据库在线分析Nur77在胃癌及胃黏膜组织中mRNA表达情况;基于Human Protein Atlas网站数...目的探讨孤儿核因子受体(Nur77)在胃癌组织中表达及其与患者预后状况之间联系,研究Nur77在胃癌细胞侵袭与迁移进程中的作用机制。方法利用Oncomine数据库在线分析Nur77在胃癌及胃黏膜组织中mRNA表达情况;基于Human Protein Atlas网站数据对比胃癌及正常胃组织中Nur77蛋白表达分布特征;GEPIA2在线分析工具评估Nur77与患者总生存期之间联系;蛋白免疫印迹法(Westernblot)比较正常胃黏膜上皮细胞GES-1与胃癌细胞AGS、MKN-45中Nur77蛋白表达差异;分别通过siRNA靶向干扰以及质粒转染上调Nur77表达和靶向下调IL-6验证Nur-77与IL-6之间表达调控关系;运用划痕实验检测干扰Nur77表达前后胃癌细胞的迁移能力改变;将胃癌细胞分空载体组和过表达Nur77组以及空载体转染后干扰IL-6处理组,过表达Nur77结合干扰IL-6处理组(质粒转染24 h后siRNA继续处理24 h)经Transwell小室实验检测Nur-77及IL-6在胃癌细胞迁移和侵袭过程中作用;利用Westernblot检测Nur77表达对NF-κB/IL-6信号通路活化的影响。结果Oncomine数据库在线分析显示,与正常胃黏膜组织相比胃癌组织中Nur77的mRNA表达水平显著升高(P<0.05);免疫组化结果显示,胃癌组织中Nur77多表达于胃癌细胞细胞核内;Nur77的高表达与胃癌患者不良预后相关(P<0.05);胃癌细胞中高表达的Nur77参与IL-6的表达调控;划痕实验和Transwell小室结果显示,Nur77可能通过IL-6参与胃癌细胞的迁移和侵袭进程(P<0.05);Westernblot结果显示,Nur77通过调控p-p65、p65、p-Stat3、Stat3的表达参与NF-κB/IL-6信号通路活化。结论胃癌中高表达Nur77与患者不良预后密切相关;Nur77可能通过调控NF-κB/IL-6信号途径活化参与胃癌细胞侵袭与迁移进程。展开更多
OBJECTIVE Evidience appears that parthenolide(PN) induces anti-tumor effects by NF-κB signal pathway.MCL3 the derivative of PN,is sesquiterpene lactone synthesized by the group of Professor Pan Xiandao.The study was ...OBJECTIVE Evidience appears that parthenolide(PN) induces anti-tumor effects by NF-κB signal pathway.MCL3 the derivative of PN,is sesquiterpene lactone synthesized by the group of Professor Pan Xiandao.The study was to explore the anti-tumor activity and mechanism of MCL3 in glioma.METHODS The effect of MCL3 on the proliferation of glioma cell lines was examined by MTT assay.Apoptotic activity was investigated by flow cytometry.The Transwell cell invasion assay was used to determine the effect of MCL3 on the G422 cell invasive ability.The effect of MCL3 on the angio.genesis was analyzed by a capillary-like tube formation assay.The subcutaneously transplanted and orthotopic G422 cell xenograft models were used to detect the effect of MCL3 on tumor growth in vivo.The pathological changes were analyzed by H&E staining.Protein level related to the NF-κB signal pathway was dertimined by Western blotting.The effect of MCL3 on the NF-κB transcriptional activity was examined by a dual-luciferase reporter assay.RESULTS The anti-proliferative activity was observed following treatment with MCL3 for 96 h in G422,U-87 MG,U251 and Hs683 cell lines,and the IC50 was 8.94 μmol·L^(-1),6.44 μmol·L^(-1),14.8 μmol·L^(-1),18.9 μmol·L^(-1),respectively.The percentage of apop.totic cells increased in MCL3-treated G422 cells,and the apoptosis rate was 26.4%(the apoptosis rate was 5.68% in control group).MCL3 could inhibit the invasion in G422 cells,and the invasive inhibition rate was 43.63%(P<0.01) at 10.0 μmol·L^(-1).MCL3 inhibited tube formation of EA.hy926 cells,and the inhibitory rate was 81.67%(P<0.01) at 10.0 μmol·kg^(-1).At 40.00 mg·kg^(-1),MCL3 supressed tumor growth by79.03%(P<0.01) in tumor weight in subcutaneously transplanted G422 xenograft models,and by 69.97%(P<0.01) in volume in orthopotic G422 xenograft models.H&E staining demonstrated that MCL3 could decrease tumor angiogenesis and invasion,increased necrosis of tumor cells.The dualluciferase reporter assay showed that MCL3 inhibited NF-κB transcriptional actvity,and the inhibition rate was 50.07%(P<0.05) at 10.0 μmol·L^(-1) compared with control.Moreover,MCL3 inhibited the phos.phorylation of NF-κB in nuclear mediated by supression of phosphorylated IKKα/β and IκB,and decreased the expression of IL-6 regulated by NF-κB.Eventually,the phosphorylation of State3 decreased following the administration of MCL3,resulting in the downregulation of State3 taget genes,including HIF,VEGF,FAK,MMP-2,MMP-9,Bcl-2 and Bcl-xL.CONCLUSION The anti-tumor effect of MCL3 was partly due to the inhibition of NF-κB/IL-6/State3 pathway in glioma.展开更多
文摘目的探讨孤儿核因子受体(Nur77)在胃癌组织中表达及其与患者预后状况之间联系,研究Nur77在胃癌细胞侵袭与迁移进程中的作用机制。方法利用Oncomine数据库在线分析Nur77在胃癌及胃黏膜组织中mRNA表达情况;基于Human Protein Atlas网站数据对比胃癌及正常胃组织中Nur77蛋白表达分布特征;GEPIA2在线分析工具评估Nur77与患者总生存期之间联系;蛋白免疫印迹法(Westernblot)比较正常胃黏膜上皮细胞GES-1与胃癌细胞AGS、MKN-45中Nur77蛋白表达差异;分别通过siRNA靶向干扰以及质粒转染上调Nur77表达和靶向下调IL-6验证Nur-77与IL-6之间表达调控关系;运用划痕实验检测干扰Nur77表达前后胃癌细胞的迁移能力改变;将胃癌细胞分空载体组和过表达Nur77组以及空载体转染后干扰IL-6处理组,过表达Nur77结合干扰IL-6处理组(质粒转染24 h后siRNA继续处理24 h)经Transwell小室实验检测Nur-77及IL-6在胃癌细胞迁移和侵袭过程中作用;利用Westernblot检测Nur77表达对NF-κB/IL-6信号通路活化的影响。结果Oncomine数据库在线分析显示,与正常胃黏膜组织相比胃癌组织中Nur77的mRNA表达水平显著升高(P<0.05);免疫组化结果显示,胃癌组织中Nur77多表达于胃癌细胞细胞核内;Nur77的高表达与胃癌患者不良预后相关(P<0.05);胃癌细胞中高表达的Nur77参与IL-6的表达调控;划痕实验和Transwell小室结果显示,Nur77可能通过IL-6参与胃癌细胞的迁移和侵袭进程(P<0.05);Westernblot结果显示,Nur77通过调控p-p65、p65、p-Stat3、Stat3的表达参与NF-κB/IL-6信号通路活化。结论胃癌中高表达Nur77与患者不良预后密切相关;Nur77可能通过调控NF-κB/IL-6信号途径活化参与胃癌细胞侵袭与迁移进程。
文摘OBJECTIVE Evidience appears that parthenolide(PN) induces anti-tumor effects by NF-κB signal pathway.MCL3 the derivative of PN,is sesquiterpene lactone synthesized by the group of Professor Pan Xiandao.The study was to explore the anti-tumor activity and mechanism of MCL3 in glioma.METHODS The effect of MCL3 on the proliferation of glioma cell lines was examined by MTT assay.Apoptotic activity was investigated by flow cytometry.The Transwell cell invasion assay was used to determine the effect of MCL3 on the G422 cell invasive ability.The effect of MCL3 on the angio.genesis was analyzed by a capillary-like tube formation assay.The subcutaneously transplanted and orthotopic G422 cell xenograft models were used to detect the effect of MCL3 on tumor growth in vivo.The pathological changes were analyzed by H&E staining.Protein level related to the NF-κB signal pathway was dertimined by Western blotting.The effect of MCL3 on the NF-κB transcriptional activity was examined by a dual-luciferase reporter assay.RESULTS The anti-proliferative activity was observed following treatment with MCL3 for 96 h in G422,U-87 MG,U251 and Hs683 cell lines,and the IC50 was 8.94 μmol·L^(-1),6.44 μmol·L^(-1),14.8 μmol·L^(-1),18.9 μmol·L^(-1),respectively.The percentage of apop.totic cells increased in MCL3-treated G422 cells,and the apoptosis rate was 26.4%(the apoptosis rate was 5.68% in control group).MCL3 could inhibit the invasion in G422 cells,and the invasive inhibition rate was 43.63%(P<0.01) at 10.0 μmol·L^(-1).MCL3 inhibited tube formation of EA.hy926 cells,and the inhibitory rate was 81.67%(P<0.01) at 10.0 μmol·kg^(-1).At 40.00 mg·kg^(-1),MCL3 supressed tumor growth by79.03%(P<0.01) in tumor weight in subcutaneously transplanted G422 xenograft models,and by 69.97%(P<0.01) in volume in orthopotic G422 xenograft models.H&E staining demonstrated that MCL3 could decrease tumor angiogenesis and invasion,increased necrosis of tumor cells.The dualluciferase reporter assay showed that MCL3 inhibited NF-κB transcriptional actvity,and the inhibition rate was 50.07%(P<0.05) at 10.0 μmol·L^(-1) compared with control.Moreover,MCL3 inhibited the phos.phorylation of NF-κB in nuclear mediated by supression of phosphorylated IKKα/β and IκB,and decreased the expression of IL-6 regulated by NF-κB.Eventually,the phosphorylation of State3 decreased following the administration of MCL3,resulting in the downregulation of State3 taget genes,including HIF,VEGF,FAK,MMP-2,MMP-9,Bcl-2 and Bcl-xL.CONCLUSION The anti-tumor effect of MCL3 was partly due to the inhibition of NF-κB/IL-6/State3 pathway in glioma.
