摘要
目的:探讨Paris saponinⅦ(PSⅦ)抑制非小细胞肺癌(non-small cell lung cancer, NSCLC)的作用机制。方法:通过CCK-8法、台盼蓝拒染法和平板集落形成实验测定PSⅦ在体外对NSCLC细胞的毒性作用;流式细胞术、DAPI染色检测凋亡;免疫荧光、Western blot及自噬抑制剂共处理检测自噬;Western blot及AMPK抑制剂共处理明确PSⅦ的调控通路;分子对接分析PSⅦ与AMPK的结合位点;微尺度热泳动实验和基于靶点稳定性的药物亲和反应实验检验亲和力。结果:PSⅦ在低微摩尔浓度下能够抑制NSCLC细胞生长和增殖;PSⅦ能够诱导NSCLC细胞发生凋亡和自噬;PSⅦ能够激活NSCLC细胞中AMPK/mTORC1信号通路;PSⅦ与AMPK具有直接亲和力。结论:PSⅦ能够直接结合并激活AMPK,进而抑制mTORC1及其下游的信号分子,诱导NSCLC自噬及凋亡,从而抑制NSCLC增殖。
Objective To explore the mechanism of Paris Saponin Ⅶ(PSⅦ) in inhibiting non-small cell lung cancer(NSCLC). Methods CCK-8 assay, trypan blue assay and clone formation assay were used to determine the inhibitory effect of PSⅦ on the proliferation of NSCLC cells in vitro. Flow cytometry and DAPI staining were used to detect the effect of PSⅦ on apoptosis of NSCLC. Western blot, immunofluorescence and autophagy inhibitor co-treatment were used to detect autophagy. Western blot and AMPK inhibitor co-treatment were used to determine the regulatory pathway of PSⅦ. Molecular docking was used to analyze the binding site of PSⅦ to AMPK. Microscale thermophoresis and drug affinity responsive target stability assays were used to test affinity. Results PSⅦ could inhibit the growth and proliferation of NSCLC cells at low concentration. PSⅦ can induce apoptosis and autophagy in NSCLC cells. PSⅦ can activate the AMPK/mTORC1 signaling pathway in NSCLC cells. PSⅦ has direct affinity with AMPK. Conclusion PSⅦ can directly bind and activate AMPK, thereby inhibiting mTORC1 and its downstream signaling molecules, inducing autophagy and apoptosis of NSCLC, and thereby inhibiting the proliferation of NSCLC.
作者
金鑫
谭苗
任宇亮
向雨晨
万芳
刘莹
JIN Xin;TAN Miao;REN Yu-liang;XIANG Yu-chen;WAN Fang;LIU Ying(School of Basic Medical Sciences,Hubei University of Medicine,Shiyan,Hubei 442000,China;Hubei Key Laboratory of Wudang Local Chinese Medicine Research,Hubei University of Medicine,Shiyan,Hubei 442000,China)
出处
《湖北医药学院学报》
CAS
2022年第4期350-356,381,F0003,F0002,共10页
Journal of Hubei University of Medicine
基金
国家自然科学基金(82072928)
湖北医药学院研究生科技创新项目(YC2022031)
大学生创新创业训练计划项目(202110929001,202210929007)。
