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薯蓣皂苷元对肿瘤坏死因子诱导的单核细胞组织因子表达的抑制作用研究

Diosgenin inhibits tumor necrosis factor-induced tissue factor activity and expression in THP-1 cells via down-regulation of the NF-κB,Akt,and MAPK signaling pathways

  • 摘要: 目的:研究薯蓣皂苷元对肿瘤坏死因子(TNF-)诱导的THP-1细胞中组织因子(TF)促凝活性及表达的作用,并探讨其可能的作用机制。方法:采用TNF-(10 ngmL-1)诱导THP-1细胞活化,采用改良的发色底物法测定TF的促凝活性;应用逆转录聚合酶链式反应(RT-PCR)及荧光定量聚合酶链式反应(qPCR)测定TF的mRNA表达;Western blotting分析TF蛋白及相关信号通路激酶表达水平。结果:薯蓣皂苷元(0.01,0.1和1 molL-1)预处理,可浓度依赖性地抑制TNF-诱导的TF促凝活性,经计算其IC50为0.25 molL-1;薯蓣皂苷元明显抑制TNF-诱导的TF mRNA和蛋白表达。同时,薯蓣皂苷元1 molL-1对于TNF-诱导的5-30 min NF-B/p65、IKK、Akt、ERK和JNK的激活,也有一定的抑制作用。结论:薯蓣皂苷元可明显抑制TNF-诱导的THP-1细胞TF的活性及表达,其机制可能与下调NF-B/p65、IKK、Akt、ERK和JNK的磷酸化有关。

     

    Abstract: AIM:To investigate whether diosgenin could modulate tissue factor(TF) procoagulation activity,expression,and related signal transduction pathways.METHODS:Human THP-1 monocytic cells were exposed to tumor necrosis factor-(TNF-,10 ngmL-1) with or without diosgenin(0.01,0.1,and 1 molL-1) for 2 h or 5 h to induce TF procoagulant activity and expression,which were determined by the simplified chromogenic assay,reverse transcription-polymerase chain reaction(RT-PCR),real-time quantitative PCR,and Western blotting assays.In addition,the activation of the NF-B,Akt,and MAPK signaling pathways were also measured by Western blotting.RESULTS:Diosgenin significantly inhibited TNF--induced TF procoagulant activity at concentrations of 0.01 to 1 molL-1 with IC50 of 0.25 molL-1.It also reduced protein expression and mRNA accumulation of TF dose-dependently in activated THP-1 cells.TNF- stimulated significantly phosphorylation on Ser536 of NF-B/p65,Ser473 of Akt at 5-15 min,and activations of IKK- and ERK at 15-30 min.Diosgenin(1 molL-1) could inhibit the phosphorylation of NF-B/p65,IKK-,Akt,ERK,and JNK,but had no remarkable effects on IB and p38 phosphorylation in THP-1 cells.CONCLUSION:Diosgenin inhibits TNF--induced TF activity and expression in monocytes,partly due to its down-regulation of the phosphorylation of NF-B/p65,IKK-,Akt,ERK,and JNK.

     

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