黄芪皂苷Ⅱ对肾透明细胞癌细胞生长抑制作用及机制

doi:10.6040/j.issn.1671-7554.0.2022.0885

摘要/Abstract

摘要： 目的探讨黄芪皂苷Ⅱ(As Ⅱ)对肾透明细胞癌786-O细胞增殖的影响及潜在机制。方法噻唑兰(MTT)法和克隆实验检测As Ⅱ对786-O细胞的生长和集落形成的抑制作用。细胞流式术检测AS Ⅱ的凋亡诱导作用。随后,用免疫印迹法检测不同处理组细胞PI3K-AKT-mTOR通路蛋白及凋亡执行蛋白的变化,最后利用PI3K-AKT-mTOR通路激活剂IGF-1与As Ⅱ对细胞共处理后,流式细胞术检测细胞凋亡水平的变化,进一步验证PI3K-AKT-mTOR通路在凋亡诱导中的作用。结果 As Ⅱ对786-O细胞的增殖抑制效应具有浓度和时间依赖性,时间与浓度之间有交互作用(F时间=513.00, P<0.001;F浓度=1 678.00, P<0.001;F时间×浓度=18.23, P<0.001),并可抑制细胞集落形成。As Ⅱ可诱导细胞凋亡并诱导凋亡执行蛋白cleved caspase-3的表达(P_{0 μmol/L As Ⅱ组 vs 10 μmol/L As Ⅱ组}= 0.013 9,P_{0 μmol/L As Ⅱ组 vs 20 μmol/L As Ⅱ组}<0.001)。As Ⅱ可抑制PI3K、AKT、mTOR蛋白的磷酸化(p-PI3K: P_{0 μmol/L As Ⅱ组 vs 20 μmol/L As Ⅱ组}=0.032 4;p-mTOR: P_{0 μmol/L As Ⅱ组 vs 10 μmol/L As Ⅱ组}=0.004 1,P_{0 μmol/L As Ⅱ组 vs 20 μmol/L As Ⅱ组}<0.001;p-AKT:P_{0 μmol/L As Ⅱ组 vs 10 μmol/L As Ⅱ组}=0.003 2,P_{0 μmol/L As Ⅱ组 vs 20 μmol/L As Ⅱ组}=0.001 2)。IGF-1可以逆转As Ⅱ对AKT的磷酸化(P=0.006 8),并减弱AsⅡ对786-O细胞的凋亡诱导效应。结论 As Ⅱ可抑制肾癌780-O细胞的增殖和集落形成,并通过抑制PI3K-AKT-mTOR信号通路的磷酸化,上调cleved caspase-3的表达,进而诱导细胞凋亡。

关键词: 肾透明细胞癌, 黄芪皂苷Ⅱ, 肿瘤治疗, 凋亡

Abstract: Objective To investigate the effect of astragaloside Ⅱ(As Ⅱ)on the proliferation of renal clear cell carcinoma 786-O cells and its potential mechanism. Methods The inhibitory effect of As Ⅱ on the growth and colony formation of 786-O cells was detected by MTT assay and cloning assay. Cell flow cytometry was used to detect the apoptosis-inducing effect of AS Ⅱ. Subsequently, Western blotting was used to detect the changes of PI3K-AKT-mTOR pathway protein and apoptosis executioner protein in cells treated with different concentrations of As Ⅱ. After co-treatment of cells with PI3K-AKT-mTOR pathway activator IGF-1 and As Ⅱ, flow cytometry was used to detect the changes of apoptosis and further verify the role of PI3K-AKT-mTOR pathway in apoptosis induction. Results As Ⅱ inhibited the proliferation of 786-O cells in a concentration- and time-dependent manner, with an interaction between time and concentration(F_time=513.00, P<0.001; F_{concentration}=1 678.00, P<0.001; F_{time×concentration}=18.23, P<0.001), and inhibited cell colony formation. As Ⅱ induced apoptosis and increased the expression of cleved caspase-3, an apoptotic executioner protein(P_{0 μmol/L As Ⅱ group vs 10 μmol/L As Ⅱ group}=0.013 9,P_{0 μmol/L As Ⅱ group vs 20 μmol/L As Ⅱ group}<0.001). As Ⅱ inhibited the phosphorylation of PI3K, AKT and mTOR proteins(p-PI3K: P_{0 μmol/L As Ⅱ group vs 20 μmol/L As Ⅱ group}=0.032 4;p-mTOR: P_{0 μmol/L As Ⅱ group vs 10 μmol/L As Ⅱ group}=0.004 1,P_{0 μmol/L As Ⅱ group vs 20 μmol/L As Ⅱ group}<0.001;p-AKT:P_{0 μmol/L As Ⅱ group vs 10 μmol/L As Ⅱ group}=0.003 2,P_{0 μmol/L As Ⅱ group vs 20 μmol/L As Ⅱ group}=0.001 2). IGF-1 reversed the phosphorylation of AKT by As Ⅱ(P=0.006 8), and attenuated the apoptosis-inducing effect of As Ⅱ on 786-O cells. Conclusion As Ⅱ can inhibit the proliferation and colony formation of renal carcinoma 780-O cells, and up-regulate the expression of cleved caspase-3 by inhibiting the phosphorylation of PI3K-AKT-mTOR signaling pathway, which in turn induces apoptosis.

Key words: Clear cell renal carcinoma, Astragaloside Ⅱ, Cancer therapy, Apoptosis

中图分类号:

R737.11

赵凯,尹心宝,张宗亮,王振林,朱冠群,王科. 黄芪皂苷Ⅱ对肾透明细胞癌细胞生长抑制作用及机制[J]. 山东大学学报 (医学版), 2023, 61(1): 10-16.

ZHAO Kai, YIN Xinbao, ZHANG Zongliang, WANG Zhenlin, ZHU Guanqun, WANG Ke. Inhibitory effect and mechanism of astragaloside Ⅱ on renal clear cell carcinoma cells[J]. Journal of Shandong University (Health Sciences), 2023, 61(1): 10-16.

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