摘要
目的:观察灯盏花素对人非小细胞肺癌A549细胞顺铂化疗敏感性的影响,并基于磷脂酰肌醇3-激酶/蛋白激酶B(phosphatidylinositol-3-kinase/protein kinase B,PI3K/AKT)通路探讨潜在作用机制。方法:以0、2、4、8、16、32μmol·L^(-1)灯盏花素和0、1、2、4、8、16μmol·L^(-1)顺铂干预A549细胞,MTT法检测细胞增殖活力,并计算半抑制浓度IC_(50灯盏花素)和IC_(50顺铂)。将A549细胞分为空白组、灯盏花素组、顺铂组、灯盏花素+顺铂组、灯盏花素+顺铂+LY294002组。培养48 h后,MTT法检测细胞活力,平板克隆实验检测细胞克隆形成能力,流式细胞术检测细胞凋亡,RT-PCR法检测细胞PI3K、AKT基因表达水平,Western blot法检测细胞PI3K、AKT、B细胞淋巴瘤-2(B-cell lymphoma-2,Bcl-2)、Bcl-2相关X蛋白(Bax)、半胱氨酸蛋白酶(Caspase)-9、Cleaved Caspase-9、Caspase-3、Cleaved Caspase-3蛋白表达水平。结果:灯盏花素和顺铂对A549细胞增殖活力的抑制作用均呈剂量依赖性,IC 50灯盏花素和IC_(50顺铂)分别为14.36μmol·L^(-1)和5.72μmol·L^(-1)。与空白组相比,其余组细胞增殖活力、克隆形成数目降低,凋亡率升高,PI3K mRNA、AKT mRNA水平降低(P<0.05)。与灯盏花素组或顺铂组相比,灯盏花素+顺铂组和灯盏花素+顺铂+LY294002组细胞增殖活力、克隆形成数目降低,凋亡率升高,PI3K mRNA、AKT mRNA水平降低(P<0.05)。与灯盏花素+顺铂组相比,灯盏花素+顺铂+LY294002组细胞增殖活力、克隆形成数目降低,凋亡率升高,PI3K mRNA、AKT mRNA水平降低(P<0.05)。与空白组相比,其余组细胞PI3K、AKT、Bcl-2蛋白表达水平降低,Bax、Cleaved Caspase-9、Cleaved Caspase-3蛋白表达水平及Bax/Bcl-2、Cleaved Caspase-9/Caspase-9、Cleaved Caspase-3/Caspase-3比值升高(P<0.05)。与灯盏花素组或顺铂组相比,灯盏花素+顺铂组和灯盏花素+顺铂+LY294002组细胞PI3K、AKT、Bcl-2蛋白表达水平降低,Bax、Cleaved Caspase-9、Cleaved Caspase-3蛋白表达水平及Bax/Bcl-2、Cleaved Caspase-9/Caspase-9、Cleaved Caspase-3/Caspase-3比值升高(P<0.05)。与灯盏花素+顺铂组相比,灯盏花素+顺铂+LY294002组细胞PI3K、AKT、Bcl-2蛋白表达水平降低,Bax、Cleaved Caspase-9、Cleaved Caspase-3蛋白表达水平及Bax/Bcl-2、Cleaved Caspase-9/Caspase-9、Cleaved Caspase-3/Caspase-3比值升高(P<0.05)。结论:灯盏花素可能通过抑制PI3K/AKT通路,抑制细胞增殖并促进其凋亡,从而增强A549细胞化疗敏感性。
Objective:To observe the effect of lanugoin on the sensitivity of human non-small cell lung cancer A549 cells to cisplatin chemotherapy,and to explore the potential mechanism of action based on phosphatidylinositol-3-kinase/protein kinase B(PI3K/AKT)pathway.Methods:A549 cells were intervened with 0,2,4,8,16,and 32μmol·L^(-1) luciferin and 0,1,2,4,8,and 16μmol·L^(-1) cisplatin,and the cell proliferation viability was detected by MTT assay,and the semi-inhibitory concentrations IC_(50 luciferin) and IC_(50 cisplatin) were calculated.The A549 cells were divided into blank group,lamparicin group,cisplatin group,lamparicin+cisplatin group,and lamparicin+cisplatin+LY294002 group.After 48 h of culture,cell viability was detected by MTT assay,cell clone formation ability was detected by plate cloning assay,cell apoptosis was detected by flow cytometry,cell expression levels of PI3K and AKT genes were detected by RT-PCR,and cell PI3K,AKT,B-cell lymphoma-2(Bcl-2),Bcl-2-related X protein(Bax),cysteine protease(Caspase)-9,Cleaved Caspase-9,Caspase-3,Cleaved Caspase-3 protein expression levels.Results:The inhibitory effects of both lamprophyllin and cisplatin on the proliferation viability of A549 cells were dose-dependent,with IC50 lamprophyllin and IC 50 cisplatin being 14.36μmol·L^(-1) and 5.72μmol·L^(-1),respectively.compared with the blank group,the proliferation viability and the number of clone formation of the cells in the rest of the group were reduced,the apoptosis rate was elevated,and the levels of PI3K mRNA and AKT mRNA were decreased(P<0.05).Compared with the lanugin group or the cisplatin group,the cell proliferation vigor,the number of clone formation decreased,the apoptosis rate increased,and the levels of PI3K mRNA and AKT mRNA decreased in the lanugin+cisplatin group and the lanugin+cisplatin+LY294002 group(P<0.05).Compared with the lanugin+cisplatin group,cell proliferation vigor,number of clone formation decreased,apoptosis rate increased,and PI3K mRNA,AKT mRNA levels decreased in the lanugin+cisplatin+LY294002 group(P<0.05).Compared with the blank group,the cellular PI3K,AKT,and Bcl-2 protein expression levels were reduced in the rest of the group,and the protein expression levels of Bax,Cleaved Caspase-9,Cleaved Caspase-3 and Bax/Bcl-2,Cleaved Caspase-9/Caspase-9,Cleaved Caspase-3/Caspase-3 ratio were elevated(P<0.05).Compared with the lanugin group or the cisplatin group,the cellular PI3K,AKT,and Bcl-2 protein expression levels were decreased in the lanugin+cisplatin group and the lanugin+cisplatin+LY294002 group,and the protein expression levels of Bax,Cleaved Caspase-9,and Cleaved Caspase-3 protein expression levels and the ratio of Bax/Bcl-2,Cleaved Caspase-9/Caspase-9,and Cleaved Caspase-3/Caspase-3 ratio were elevated(P<0.05).Compared with the lanugin+cisplatin group,the cellular PI3K,AKT,and Bcl-2 protein expression levels were decreased in the lanugin+cisplatin+LY294002 group,and the protein expression levels of Bax,Cleaved Caspase-9,Cleaved Caspase-3,and the ratios of Bax/Bcl-2,Cleaved Caspase-9/Caspase-3/Caspase-3 were increased(P<0.05).Conclusion:Lampetin may enhance the chemosensitivity of A549 cells by inhibiting cell proliferation and promoting apoptosis through inhibiting the PI3K/AKT pathway.
作者
韦辉
王天珩
高世奇
WEI Hui;WANG Tianheng;GAO Shiqi(The First Hospital of Handan,Handan Hebei China 056002)
出处
《中医学报》
2025年第5期1108-1115,共8页
Acta Chinese Medicine
基金
河北省医学科学研究课题项目(20220506)。
