摘要
目的:探讨黄精黄芪复方抑制肺癌进展的作用机制。方法:采用CCK-8法检测不同浓度黄精黄芪复方处理下人肺癌A549和H1299细胞的增殖抑制率并计算黄精黄芪复方的半抑制浓度(half maximal inhibitory concentration,IC_(50));用携带Cre酶的腺病毒对C57BL/6小鼠(基因型:KRAS^(G12D/+);TP53^(flox/flox))滴鼻1次,构建原发性肺癌小鼠模型,予以黄精黄芪饲料喂养以直接检测黄精黄芪复方在体内对肺癌组织的作用,采用免疫组织化学染色法检测小鼠肺组织的病理进展;生物信息学分析提示黄精黄芪复方通过爱帕琳肽(apelin)-过氧化物酶体增殖物激活受体γ共激活剂1-alpha(peroxisome proliferator-activated receptor gamma coactivator 1-alpha,PGC1α)-解偶联蛋白1(mitochondrial brown fat uncoupling protein 1,UCP1)信号通路影响肺癌进展;采用实时荧光定量PCR和蛋白质印迹法检测黄精黄芪复方对A549和H1299细胞中apelin-PGC1α-UCP1信号通路相关基因mRNA和蛋白的表达水平的影响;分别采用ATP检测试剂盒和流式细胞术检测黄精黄芪复方对A549和H1299细胞ATP总产量及线粒体活性氧(reactive oxygen species,ROS)生成的影响;采用siUCP1沉默UCP1表达,用2160诱导UCP1过表达,并检测A549和H1299细胞ATP总产量及线粒体ROS的变化,以进一步验证黄精黄芪复方是否通过apelin-PGC1α-UCP1信号通路影响肺癌进展。结果:黄精黄芪复方可明显抑制A549和H1299细胞的增殖,IC_(50)分别为10.66 mg/mL和9.66 mg/mL;在小鼠肺原位癌模型中,黄精黄芪复方可明显抑制肿瘤的生长,并下调apelin-PGC1α-UCP1信号通路,抑制促肺癌基因UCP1的表达;在A549和H1299细胞中黄精黄芪复方能显著抑制apelin、PGC1α和UCP1的表达(P<0.05)、促进ATP合成(P<0.0001)和ROS产生并恢复线粒体氧化磷酸化,抑制有氧糖酵解(P<0.01);沉默UCP1能增加A549和H1299细胞的ATP合成(P<0.01)和线粒体ROS产生,并降低有氧糖酵解关键酶己糖激酶2(hexokinase 2,HK2
Objective:To investigate the mechanism of polygonatum and astragalus compound(PA)in inhibiting the progression of lung adenocarcinoma.Methods:CCK-8 assay was used to assess the inhibitory rate of proliferation in A549 and H1299 cells treated with PA at different concentrations and to calculate the half maximal inhibitory concentration(IC_(50)).C57BL/6 mice(KRAS^(G12D/+);TP53^(flox/flox))were treated with adenovirus carrying Cre enzyme via nasal inhalation to establish a mouse model of primary lung adenocarcinoma.The model mice were fed with PA-containing diet to directly observe the effect of PA on the lung adenocarcinoma tissue.Immunohistochemical staining was used to examine the pathological status of the lung tissue.Bioinformatics analysis indicated that PA affects the progression of lung adenocarcinoma through the apelin-peroxisome proliferator-activated receptor gamma coactivator 1-alpha(PGC1α)-mitochondrial brown fat uncoupling protein 1(UCP1).Real-time quantitative PCR and Western blotting analysis were used to study the effect of PA on the mRNA and protein expression levels of apelin-PGC1α-UCP1 signaling pathway related genes.An ATP detection kit and flow cytometry were used to evaluate the effect of PA on the ATP and mitochondrial ROS production,respectively,in A549 and H1299 cells.siUCP1 was used to silent the expression of UCP1 while Z160 was used to induce UCP1 overexpression in A549 and H1299 cells,and the changes in ATP and mitochondrial ROS production were examined to further investigate whether PA acts on apelin-PGC1α-UCP1 signaling pathway to affect the progression of lung adenocarcinoma.Results:PA could obviously inhibit the proliferation of A549 and H1299 cells with the IC_(50)values of 10.66 mg/mL for A549 cells and 9.66 mg/mL for H1299 cells.In the mouse primary lung adenocarcinoma model,PA could effectively inhibit the growth of tumor,downregulate apelin-PGC1α-UCP1 signaling pathway and inhibit the expression of lung adenocarcinoma-promoting gene UCP1.In A549 and H1299 cells,PA could sig
作者
王宗灿
郑天盛
韦梦铃
庄文彬
李明
王菲
岳利多
范理宏
WANG Zongcan;ZHENG Tiansheng;WEI Mengling;ZHUANG Wenbin;LI Ming;WANG Fei;YUE Liduo;FAN Lihong(Shanghai Tenth People's Hospital Clinical College,Nanjing Medical University,Nanjing 211100,Jiangsu Province,China;Shanghai Tenth People's Hospital of Tongji University,Tongji University School of Medicine,Shanghai 200072,China;School of Integrative Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200072,China;Nantong University Medical School,Nantong 226000,Jiangsu Province,China;Department of Integrated Traditional Chinese and Western Medicine,Shanghai Tenth People's Hospital of Tongji University,Shanghai 200072,China;Institute of Energy Metabolism and Health,School of Medicine,Tongji University,Shanghai 200072,China)
出处
《肿瘤》
CAS
2024年第2期180-194,共15页
Tumor
基金
国家自然科学基金面上项目(31770131)
一带一路国际合作项目(20400750600)
国家中医药管理局,新型冠状病毒感染中医药应急专项(2023ZYLCYJ02-6)
上海市中西医结合创新旗舰医院建设基金(No.ZY(2021-2023)-0205-05)
