黄芩素通过PI3K/AKT通路增强三氧化二砷对肝癌细胞的促凋亡作用  

Baicalein enhances As_2O_3 induced pro-apoptotic effects in primary liver cancer via PI3K/AKT pathway

作　　者：周珍[1] 刘少平[1] 周朗 梁青 刘尚勤[1] 周芙玲[1] 马梓[1] 何莉[1] Zhou Zhen;Liu Shaoping;Zhou Lang;Liang Qing;Liu Shangqin;Zhou Fuling;Ma Zi;He Li(Zhongnan Hospital of Wuhan University,Hubei Wuhan 430071,China;Hubei Institute of Science and Technology,Hubei Xianning 437000,China;Wuhan No.4 Hospital,Hubei Wuhan 430071,China)

机构地区：[1]武汉大学中南医院,湖北武汉430071 [2]湖北科技学院,湖北咸宁437000 [3]武汉市第四医院,湖北武汉420071

出　　处：《现代肿瘤医学》2019年第10期1661-1668,共8页Journal of Modern Oncology

基　　金：国家自然科学基金资助项目(编号:81800206)

摘　　要：目的:探讨黄芩素增强As_2O_3对肝癌细胞的促凋亡作用及其可能的作用机制。方法:采用MTT方法检测不同作用浓度及作用时间的黄芩素、As_2O_3单用和联用对肝癌高转移细胞HCCLM9、肝癌低转移细胞MHCC97L的增殖抑制率,并根据联合用药q值判断两药联用的性质;流式细胞术检测肝癌细胞的周期分布;Tunel技术检测肝癌细胞的凋亡情况;Real-Time PCR和Western blot检测肝癌细胞凋亡相关蛋白caspase-3、caspase-6、caspase-9的mRNA和蛋白质水平以及PI3K、AKT和P-AKT蛋白的表达情况。结果:黄芩素、As_2O_3单药及联合后均能明显抑制HCCLM9和MHCC97L细胞增殖能力,但两药联合作用更强,且呈明显时间和剂量依赖性效应。两药单用及联合均能够明显导致肝癌细胞G_0/G_1期停滞,Tunel检测到细胞凋亡率升高,且两药联合作用强于单药。两种药物同时处理后肝癌细胞的caspase-3、caspase-6及caspase-9的mRNA和蛋白水平表达显著升高而PI3K、p-AKT蛋白表达下降。结论:黄芩素增强As_2O_3对肝癌细胞的促凋亡的作用,同时引起PI3K、p-AKT表达的下调及caspase-3、caspase-6及caspase-9表达的上调。推测黄芩素可能是通过激活Caspase凋亡信号,同时抑制PI3K/AKT通路增强As_2O_3对肝癌细胞促凋亡的作用。Objective:To investigate baicalein's enhancing effects on As 2O 3 induced apoptosis in primary liver cancer and the potential underlying mechanism.Methods:The inhibition effects of baicalein and As 2O 3 single agent and combination were investigated using the MTT approach in two primary liver cancer cell lines,the HCCLM9 and MHCC97L showing distinctly different metastatic potential(high vs low).The proliferation inhibition rates calculated and the q value was used to determine the effect by combined treatment.The flow cytometry was performed to analyze the cell cycle status of primary liver cancer cells.The Tunnel assay was performed to analyze the apoptosis of primary liver cancer cells.The real-time PCR and Western blot analysis were performed to measure the mRNA and protein expression of apoptosis-related proteins(caspase-3,caspase-6 and caspase-9)and proteins involved in the PI3K/AKT pathway(PI3K,AKT and p-AKT)were detected.Results:Baicalein or As 2O 3 alone can both significantly inhibit the proliferation of HCCLM9 and MHCC97L cell lines,and the combined treatment showed enhanced inhibition effects in a time-and dose-dependent manner.Baicalein or As 2O 3 alone can both significantly induce G 0/G 1 blockade and apoptosis,and the combined treatment showed enhanced effects.After the co-treatment of the two agents,the mRNA and protein expression of caspase-3,caspase-6 and caspase-9 was significantly increased,and that of PI3K and p-AKT significantly reduced.Conclusion:Baicalein enhances the pro-apoptotic effects of As 2O 3,causing the down-regulation of PI3K and p-AKT and up-regulation of caspase-3,caspase-6 and caspase-9.It is speculated that baicalein enhances As 2O 3's pro-apoptotic effects via Caspase apoptotic signals and PI3K/AKT pathway.

关 键 词：黄芩素 三氧化二砷 肝癌 凋亡 

分 类 号：R735.7[医药卫生—肿瘤]