摘要
目的探讨八角莲醇提物对肾透明细胞癌OS-RC-2细胞增殖、凋亡的影响及作用机制。方法八角莲给药,CCK-8及克隆形成实验检测给药后OS-RC-2细胞增殖情况;划痕实验及Transwell实验检测细胞迁移情况;活性氧(Reactive oxygen species,ROS)检测试剂盒检测细胞ROS水平。网络药理学进行八角莲及ROS共同靶点的获取,并进行KEGG分析。流式细胞仪检测细胞凋亡及周期水平,Western blot对MAPK信号通路关键蛋白及凋亡相关蛋白进行检测。结果八角莲可明显抑制OS-RC-2细胞增殖、迁移能力,并升高细胞ROS水平。网络药理学分析显示,共得到165个八角莲-ROS的共同靶点,KEGG分析发现,MAPK信号通路变化明显。Western blot结果显示,八角莲给药后,明显降低JNK蛋白水平及p-ERK/ERK比值;降低caspase-9及Bcl-2蛋白水平,而升高cleaved caspase-9及Bax蛋白表达。流式结果显示,八角莲给药可促进细胞凋亡,降低G 1期而升高G 2/M期细胞量。回复实验结果显示,ROS抑制剂NAC与八角莲联合给药可明显逆转八角莲单独给药对OS-RC-2的细胞活力及凋亡作用,以及对细胞凋亡关键蛋白、MAPK信号通路蛋白的表达改变。结论八角莲可能通过影响ROS水平发挥MAPK信号通路抑制作用,并进一步对OS-RC-2细胞产生促凋亡及抑制增殖作用。
Aim To explore the effects of ethanol extraction of Dysosma versipellis on the proliferation and apoptosis of renal clear cell carcinoma OS-RC-2 cells and the underlying mechanism.Methods After treated with Dysosma versipellis,the proliferation of OS-RC-2 cells was detected by the CCK-8 assay and clone formation assay.The migration rate of cells was detected by thewound healing assay and Transwell assay.The level of ROS was detected by the reactive oxygen detection kit.The common targets between Dysosma versipellis and ROS were obtained by the network pharmacology.The above common targets were analyzed by KEGG.The apoptosis rate and cell cycle were detected by the flow cytometry,and the key proteins of MAPK signaling pathway and the levels of apoptosis related proteins were measured by Western blot.Results The results showed that Dysosma versipellis significantly inhibited the cell viability and migration ability of OS-RC-2 cells,and up-regulated the level of ROS.Network pharmacology analysis showed a total of 165 common targets between Dysosma versipellis and ROS.KEGG analysis of the common targets revealed that there were significant changes in the MAPK signaling pathway.The results of Western blot showed that after treated with Dysosma versipellis,the protein level of JNK and the ratio of p-ERK/ERK were down-regulated.Besides,the protein level of caspase-9 and Bcl-2 declined,while the levels of cleaved caspase-9 and Bax were promoted.The flow cytometry results showed that Dysosma versipellis could significantly promote the apoptosis rate,down-regulate the cells in G1-phase,while up-regulate the cells in G 2/M-phase.The results of the rescue experiment showed that co-administration of NAC and Dysosma versipellis could significantly reverse the cell viability and apoptosis rate,the level of apoptotic related proteins,as well as the protein levels of MAPK pathway,when compared to treated with Dysosma versipellis alone in OS-RC-2.Conclusion In summary,Dysosma versipellis may inhibit the MAPK signaling pathway via the changes in ROS,further promoting apoptosis rate and decline cell proliferation in OS-RC-2 cell line.
作者
游赣花
何恋
李凯
杨萌
文周
刘旺培
朱建国
YOU Gan-hua;HE Lian;LI Kai;YANG Meng;WEN Zhou;LIU Wang-pei;ZHU Jian-guo(Dept of Scientific Research/Dept of Pharmacy,the Second People′s Hospital of Guizhou Province,Guiyang 550002,China;College of Biological and Environmental Engineering,Guiyang University,Guiyang 550005,China;Graduate School,Zunyi Medical University,Zunyi Guizhou 563000,China;Chemistry Room,Shaoguan Institute for Food and Drug Control,Shaoguan Guangdong 521028,China;Dept of Urology Surgery,Guizhou Provincial People′s Hospital,Guiyang 550000,China)
出处
《中国药理学通报》
CAS
CSCD
北大核心
2023年第12期2305-2313,共9页
Chinese Pharmacological Bulletin
基金
国家自然科学基金资助项目(No 82160551)
贵州省高层次创新型人才(黔科合平台人才No[2018]5639)
贵州省中医药管理局项目(No QZYY-2021-168)。
