归芪益元膏联合顺铂调节AMPK/ULK1信号通路对Lewis肺癌荷瘤小鼠肿瘤组织的影响  

Effects of Guiqi Yiyuan Ointment Combined with Cisplatin on Regulating the AMPK/ULK1 Signaling Pathway in Tumor Tissue of Lewis Lung Cancer Bearing Mice

作　　者：李尚 梁建庆 靳银霞 杨楠 薛春霞 李娟 李金田[2,3] LI Shang;LIANG Jianqing;JIN Yinxia;YANG Nan;XUE Chunxia;LI Juan;LI Jintian(School of Basic Medicine,Gansu University of Chinese Medicine,Lanzhou 730000,China;Key Laboratory of Dunhuang Medicine and Transformation,Ministry of Education,Lanzhou 730000,China;Clinical College of Traditional Chinese Medicine,Gansu University of Chinese Medicine,Lanzhou 730000,China)

机构地区：[1]甘肃中医药大学基础医学院,甘肃兰州730000 [2]敦煌医学与转化教育部重点实验室,甘肃兰州730000 [3]甘肃中医药大学中医临床学院,甘肃兰州730000

出　　处：《中国中医药信息杂志》2024年第9期110-116,共7页Chinese Journal of Information on Traditional Chinese Medicine

基　　金：国家自然科学基金(82160872);甘肃省教育厅“双一流”科研重点项目(GSSYLXM-05);甘肃省中医药研究中心开放课题(zyzx-2020-zx17);2022年度中医学一级学科“岐黄英才”导师专项基金博导项目(ZYXKBD-202202);2022年度中医学一级学科“岐黄英才”导师专项基金硕导项目(ZYXKSD-202211);敦煌医学与转化教育部重点实验室开放课题(DHYX22-06)。

摘　　要：目的 基于AMPK/ULK1信号通路观察归芪益元膏联合顺铂对Lewis肺癌荷瘤小鼠的抑瘤作用及作用机制。方法 小鼠右侧腋下接种Lewis肺癌细胞建立皮下肿瘤模型。将小鼠随机分为空白组、模型组、顺铂组和中药高、中、低剂量+顺铂组,每组10只,顺铂组腹腔注射顺铂5 mg/kg(每7 d一次),中药高、中、低剂量+顺铂组在顺铂组基础上分别予归芪益元膏7.0、3.5、1.75 g/kg灌胃,每日1次,连续14 d。HE染色观察小鼠肿瘤组织形态,免疫组化测定肿瘤组织p-AMPK、p-ULK1蛋白表达,透射电镜观测肿瘤组织自噬小体数量,RT-PCR测定小鼠肿瘤组织p62、Beclin-l、Atg5、LC3 mRNA表达,Western blot检测肿瘤组织Beclin-1、p62、Atg5、LC3和AMPK/ULK1信号通路蛋白表达。结果 与模型组比较,各给药组小鼠瘤质量明显降低(P<0.05);肿瘤组织出现片状坏死区域且自噬小体数量增加,Beclin-l、Atg5、LC3 mRNA表达升高,Beclin-1、Atg5、LC3Ⅱ/Ⅰ、AMPK、p-AMPK、ULK1、p-ULK1蛋白表达升高,p62 mRNA和蛋白表达降低(P<0.05)。与顺铂组比较,中药高、中剂量+顺铂组小鼠瘤质量降低(P<0.05),抑瘤率增加;中药高、中、低剂量+顺铂组小鼠肿瘤组织ULK1蛋白和Atg5 mRNA表达明显升高(P<0.05);中药高、中剂量+顺铂组Beclin-1、LC3 mRNA表达明显升高(P<0.05),Atg5、LC3Ⅱ/Ⅰ、AMPK、p-AMPK、p-ULK1蛋白表达明显升高(P<0.05),p62蛋白表达明显降低(P<0.05);中药高剂量+顺铂组p62 mRNA表达明显降低(P<0.05),Beclin-1蛋白表达明显升高(P<0.05)。结论 归芪益元膏联合顺铂可能通过激活AMPK/ULK1信号通路促进肿瘤细胞自噬,进而抑制Lewis肺癌荷瘤小鼠皮下肿瘤生长。Objective To observe the anti-tumor effect and mechanism of Guiqi Yiyuan Ointment combined with cisplatin in Lewis lung cancer bearing mice based on AMPK/ULK1 signaling pathway.Methods The mice were inoculated with Lewis lung cancer cells to establish a subcutaneous tumor model in the right axillary subcutaneous area,and were randomly divided into blank group,model group,cisplatin group and Guiqi Yiyuan Ointment high-,medium-and low-dosage+cisplatin groups,with 10 mice in each group.The cisplatin group received intraperitoneal injection of 5 mg/kg(once every 7 days)cisplatin,while the Guiqi Yiyuan Ointment high-,medium-and low-dosage+cisplatin groups received gavage of 7.0,3.5 and 1.75 g/kg Guiqi Yiyuan Ointment on the basis of cisplatin group,once a day,for 14 consecutive days.HE staining was used to observe the morphology of tumor tissue,immunohistochemical method was used to detect the protein expression of p-AMPK and p-ULK1 in tumor tissue,transmission electron microscopy was used to observe the number of autophagosomes,RT-PCR was used to detect the expression of p62,Beclin-1,Atg5 and LC3 mRNA in tumor tissue,Western blot was used to detect the expression of Beclin-1,p62,Atg5,LC3 and AMPK/ULK1 signaling pathway proteins in tumor tissue.Results Compared with the model group,the tumor mass of mice in all administration groups decreased significantly(P<0.05);the tumor tissue showed patchy necrotic areas and an increase in the number of autophagosomes,the mRNA expressions of Beclin-1,Atg5,and LC3 increased,the protein expressions of Beclin-1,Atg5,LC3Ⅱ/Ⅰ,AMPK,p-AMPK,ULK1 and p-ULK1 increased,and the expression of p62 mRNA and protein decreased(P<0.05).Compared with the cisplatin group,the tumor mass of mice in Guiqi Yiyuan Ointment high-,medium-dosage+cisplatin groups decreased(P<0.05),the tumor inhibition rate increased;the expressions of ULK1 protein and Atg5 mRNA in tumor tissue of Guiqi Yiyuan Ointment high-,medium-and low-dosage+cisplatin groups significantly increased(P<0.05),the expressions of Beclin-1

关 键 词：肺癌 归芪益元膏 顺铂 细胞自噬 AMPK/ULK1信号通路 

分 类 号：R285.5[医药卫生—中药学]