岩藻多糖调控G3BP1/NF-κB信号通路影响胰腺癌细胞增殖

Fucoidan regulates the G3BP1/NF-κB signaling pathway and affects the proliferation of pancreatic cancer cells

目的:探讨岩藻多糖对胰腺癌的影响,并分析其机制。方法:MTT法分析细胞增殖抑制率,GEPIA数据库分析G3BP1在胰腺癌组织中的表达水平及与生存率的关系。qRT-PCR分析GTP酶激活蛋白结合蛋白1(Ras-GTPase-activating protein binding proteins,G3BP1)水平,Western blot法分析p-NF-κBp65、NF-κBp65和IκB-α水平。免疫共沉淀检测G3BP1与p-NF-κBp65之间相互作用。敲低或G3BP1过表达,观察其对岩藻多糖调控细胞增殖以及NF-κB信号通路的影响。裸鼠成瘤实验验证岩藻多糖对裸鼠体内瘤体的瘤重、瘤体积及G3BP1、p-NF-κBp65、NF-κBp65和IκBα水平的影响。结果:1~32μg/mL岩藻多糖抑制capan-1细胞增殖,岩藻多糖48 h IC_(50)为7.729μg/mL。G3BP1在胰腺癌肿瘤组织中的表达明显高于正常组织,G3BP1高表达患者的生存率低于G3BP1低表达患者。2、4、8μg/mL岩藻多糖能下调G3BP1、p-NF-κBp65,上调IκBα水平。Co-IP实验发现G3BP1与p-NF-κBp65相互结合,并且岩藻多糖作用后结合能力降低。敲低G3BP1能促进岩藻多糖抑制capan-1细胞增殖,下调G3BP1、p-NF-κBp65,上调IκBα水平(P<0.05);G3BP过表达能下调岩藻多糖抑制capan-1细胞增殖效果,上调G3BP1、p-NF-κBp65,下调IκBα水平(P<0.05)。体内实验显示,敲低G3BP1能促进岩藻多糖减少瘤体体积、瘤体质量,下调瘤体G3BP1、p-NF-κBp65,上调IκBα水平(P<0.05)。结论:岩藻多糖抑制capan-1细胞增殖,对体内移植瘤抑瘤效果显著,其机制与调控G3BP1/NF-κB信号通路有关。

Objective:To investigate the effect of fucoidan on pancreatic cancer and analyze its mechanism.Methods:Cell proliferation inhibition rate was analyzed by MTT method.The expression level of G3BP1 in pancreatic cancer and its relationship with survival rate were analyzed by GEPIA database.The levels of Ras-GTPase-activating protein binding proteins(G3BP1)were analyzed by qRT-PCR.The levels of p-NF-κBp65,NF-κBp65 and IκB-αwere analyzed by Western blot.The interaction between G3BP1 and p-NF-κBp65 was detected by co-immunoprecipitation.The effects of knockdown or overexpression of G3BP1 on fucoidan-regulated cell proliferation and NF-κB signaling pathway were observed.The effects of fucoidan on tumor weight,tumor volume and levels of G3BP1,p-NF-κBp65,NF-κBp65 and IκBαin nude mice were investigated.Results:1~32μg/mL fucoidan inhibited capan-1 cell proliferation,and the IC_(50)of fucoidan at 48 h was 7.729μg/mL.The expression of G3BP1 in pancreatic cancer tissues was significantly higher than that in normal tissues.The survival rate of patients with high expression of G3BP1 was lower than that of patients with low expression of G3BP1.2,4,8μg/mL fucoidan could down-regulate G3BP1,p-NF-κBp65 and up-regulate IκBαlevels.CO-IP experiments showed that G3BP1 bound to p-NF-κBp65,and the binding ability of fucoidan decreased.Knockdown of G3BP1 promoted fucoidan inhibition of capan-1 cell proliferation,down-regulated G3BP1,p-NF-κBp65,and up-regulated IκBαlevels(P<0.05).Overexpression of G3BP could down-regulate the inhibitory effect of fucoidan on capan-1 cell proliferation,up-regulate G3BP1,p-NF-κBp65,and down-regulate the level of IκBα(P<0.05).In vivo experiments showed that knockdown of G3BP1 could promote fucoidan to reduce tumor volume and mass,down-regulate tumor G3BP1 and p-NF-κBp65,and up-regulate IκBαlevel(P<0.05).Conclusion:Fucoidan can inhibit the proliferation of capan-1 cells and inhibit tumor in vivo transplantation,and its mechanism is related to the regulation of G3BP1/NF-κB signaling pathway.