Xiu Zhiru, Li Yiquan, Fang Jinbo, Han Jicheng, Li Shanzhi, Li Yaru, Yang Xia, Song Gaojie, Li Yue, Jin Ningyi, Zhu Yilong, Zhu Guangze, Sun Lili, Li Xiao
Academician Workstation of Jilin Province, Changchun University of Chinese Medicine, Changchun, People's Republic of China.
Medical College, Yanbian University, Yanji, People's Republic of China.
J Hepatocell Carcinoma. 2023 Apr 11;10:611-629. doi: 10.2147/JHC.S395617. eCollection 2023.
To explore the effects of Esculetin on liver cancer and explore potential mechanisms of Esculetin-inducing cells death.
Esculetin's effects on the proliferation, migration and apoptosis of HUH7 and HCCLM3 cells were detected by using CCK8, crystal violet staining, wound healing, Transwell and Annexin V-FITC/PI. Flow cytometry, fluorescence staining, Western blot, T-AOC, DPPH radical scavenging assay, hydroxyl radical's inhibitory capability and GSH test were used to examine the esculetin's effects on the ROS level, the oxidation-related substances and proteins' expression in hepatoma cells. In vivo experiment was performed by xenograft model. Ferrostatin-1 was used to determine the death way of hepatoma cells induced by esculetin. Live cell probe, Western blot, Fe content, MDA, HE staining, Prussian blue staining and immunohistochemistry were used to examine the ferritinophagy-related phenomenon induced by esculetin in hepatoma cells. The relationship between esculetin and NCOA4-mediated ferritinophagy was confirmed through gene silence and overexpression, immunofluorescence staining and Western blot.
Esculetin suppressed the proliferation, migration and apoptosis of HUH7 and HCCLM3 cells significantly, influenced the oxidative stress level, altered the autophagy and iron metabolism levels in cells, and produced a ferritinophagy-related phenomena. Esculetin increased the levels of cellular lipid peroxidation and reactive oxygen species. In vivo, esculetin could decrease tumour volume, promote LC3 and NCOA4 expressions, suppresse hydroxyl radical's inhibiting capacity and GSH, increase Fe and MDA levels, decrease antioxidant proteins expression in tumour tissue. In addition, Esculetin could also increase the iron deposition of tumour tissues, promote ferritinophagy, and induce tumours' ferroptosis.
Esculetin has an inhibitory effect on liver cancer in vivo and in vitro through triggering NCOA4 pathway-mediation ferritinophagy.
探讨七叶亭对肝癌的影响，并探究七叶亭诱导细胞死亡的潜在机制。
采用CCK8法、结晶紫染色法、伤口愈合实验、Transwell实验及Annexin V-FITC/PI法检测七叶亭对HUH7和HCCLM3细胞增殖、迁移及凋亡的影响。运用流式细胞术、荧光染色、蛋白质免疫印迹法、总抗氧化能力检测、DPPH自由基清除实验、羟自由基抑制能力检测及谷胱甘肽检测，研究七叶亭对肝癌细胞中活性氧水平、氧化相关物质及蛋白质表达的影响。通过异种移植模型进行体内实验。使用铁死亡抑制剂Ferrostatin-1确定七叶亭诱导肝癌细胞的死亡方式。采用活细胞探针、蛋白质免疫印迹法、铁含量检测、丙二醛检测、苏木精-伊红染色、普鲁士蓝染色及免疫组织化学法，检测七叶亭在肝癌细胞中诱导的铁自噬相关现象。通过基因沉默和过表达、免疫荧光染色及蛋白质免疫印迹法，证实七叶亭与NCOA4介导的铁自噬之间的关系。
七叶亭显著抑制HUH7和HCCLM3细胞的增殖、迁移及凋亡，影响氧化应激水平，改变细胞自噬和铁代谢水平，并产生铁自噬相关现象。七叶亭增加细胞脂质过氧化和活性氧水平。在体内，七叶亭可减小肿瘤体积，促进LC3和NCOA4表达，抑制羟自由基抑制能力及谷胱甘肽水平，增加铁和丙二醛水平，降低肿瘤组织中抗氧化蛋白表达。此外，七叶亭还可增加肿瘤组织中铁沉积，促进铁自噬，并诱导肿瘤发生铁死亡。
七叶亭通过触发NCOA4途径介导的铁自噬，对体内外肝癌均有抑制作用。
