5-羟色胺受体3拮抗剂通过IκBα/NF-κB信号通路抑制脂多糖诱导RAW264.7细胞炎症反应  

5-HT_(3)R Antagonists Prevent LPS-Induced Inflammation in RAW264.7 Cells by Inhibiting the IκBα/NF-κB Signaling Pathway

作　　者：邓邺云 任晓曦[1] 刘康瑞 陈茜 张建亮[1] DENG Ye-Yun;REN Xiao-Xi;LIU Kang-Rui;CHEN Xi;ZHANG Jian-Liang(Department of Neurobiology,School of Basic Medical Sciences,Capital Medical University,Beijing 100069,China)

机构地区：[1]首都医科大学基础医学院神经生物学系,北京100069

出　　处：《中国生物化学与分子生物学报》2023年第5期750-758,共9页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金(No.32071164);北京市自然科学基金(No.7192018)资助。

摘　　要：在败血症等炎症相关疾病中,巨噬细胞过度产生的炎症因子在疾病的发病机制中发挥关键作用。5-羟色胺受体3(5-hydroxytryptamine receptor 3,5-HT_(3)R)拮抗剂,近来被发现在免疫系统中具有抑制炎症的作用,但其具体机制尚不清楚。本文使用脂多糖(lipopolysaccharides,LPS)刺激小鼠巨噬细胞系RAW 264.7细胞,引发巨噬细胞炎症相关蛋白质的表达和炎症因子的释放。结果显示,在RAW 264.7细胞中,与无处理组相比,LPS以剂量和时间依赖的方式促进炎症相关蛋白质的表达和炎症因子的释放。进一步使用5-HT_(3)R拮抗剂格拉司琼(granisetron,GA)进行预处理,检测其抗炎作用。蛋白质免疫印迹和酶联免疫吸附测定的结果表明,与LPS处理组相比,随着GA浓度的升高,其抑制LPS诱导的RAW 264.7细胞炎症的效果越好(P<0.05)。同时,细胞活力和细胞毒性的结果也表明,所使用的GA对细胞不造成损伤。另外,通过免疫荧光实验和双荧光素酶检测表明,GA可以抑制LPS诱导的NF-κB的核移位和转录活性;进一步的蛋白质免疫印迹结果表明,GA可以通过抑制IκBα的磷酸化而抑制其降解,从而抑制NF-κB的功能。总之,本文的结果提示,GA可以通过抑制IκBα/NF-κB信号通路,抑制RAW 264.7巨噬细胞中炎症相关蛋白质的表达和炎症因子的释放。本研究结果为进一步探究5-HT_(3)R拮抗剂抗炎的分子机制,以及研发治疗败血症等炎症相关疾病的药物提供科学依据。The inflammatory factors overproduced by macrophages play a key role in the pathogenesis of inflammation-related diseases such as sepsis.Recently,5-hydroxytryptamine receptor 3(5-HT_(3)R)antagonists have been found to inhibit inflammation in the immune system,but the specific mechanism remains unclear.In this study,lipopolysaccharide(LPS)was used to stimulate RAW 264.7 cells,a mouse macrophage cell line,to induce the expression of inflammatory-related proteins and the release of inflammatory factors.The results showed that compared with the untreated group,LPS promoted the expression of inflammatory-related proteins and the release of inflammatory factors in a dose and time-dependent manner in RAW 264.7 cells.Further,RAW 264.7 cells were pretreated with the 5-HT_(3)R antagonist Granisetron(GA)to detect its anti-inflammatory effect.Compared with LPS-stimulating group,Western blotting and enzyme-linked immunosorbent assays showed that,with the increase of the concentration of GA,the inhibition effect of GA on LPS-induced inflammation of RAW 264.7 cells were better(P<0.05).Cell viability and cytotoxicity test results also showed that GA did not cause damage to cells.In addition,the immunofluorescence and dual luciferase results showed that GA inhibited the nuclear translocation and transcriptional activity of NF-κB induced by LPS.Further Western blotting results showed that GA prevented the degradation of IκBαby inhibiting its phosphorylation,thereby inhibiting the function of NF-κB.In conclusion,our results indicated that GA inhibits the expression of inflammation-related proteins and the release of inflammatory factors in RAW 264.7 macrophages by inhibiting the IκBα/NF-κB signal pathway,which provide a scientific basis for further studies on the regulatory mechanism of 5-HT_(3)R antagonists in anti-inflammation and developing drugs to treat sepsis and other inflammation-related diseases.

关 键 词：败血症 抗炎 5-羟色胺受体3 格拉司琼 IκBα/NF-κB信号通路 

分 类 号：R34[医药卫生—基础医学] Q789[生物学—分子生物学]