知母皂苷AⅢ通过JAK-2/STAT3/PD-L1信号通路增加A549/DDP对顺铂的敏感性  

Timosaponin AⅢ promotes cisplatin sensitivity in A549/DDP cells via JAK-2/STAT3/PD-L1 signaling pathway

作　　者：李嘉旗 卲轶群 许玲[1] 姚嘉麟 LI Jia-qi;SHAO Yi-qun;XU Ling;YAO Jia-lin(Dept of Oncology,Yueyang Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China;Dept of Urology,Yueyang Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China)

机构地区：[1]上海中医药大学附属岳阳中西医结合医院肿瘤一科,上海200437 [2]上海中医药大学附属岳阳中西医结合医院泌尿外科,上海200437

出　　处：《中国药理学通报》2023年第4期658-664,共7页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 82104720);上海中医药大学预算内项目(No 2020LK054)。

摘　　要：目的 探索知母皂苷AⅢ(timosaponin AⅢ,TA3)增加非小细胞肺癌顺铂耐药株A549/DDP细胞顺铂敏感性的作用机制。方法 选取A549/DDP细胞作为主要研究对象,利用CCK-8法检测TA3、顺铂以及两药联用对A549/DDP细胞增殖的影响;组别分为空白组、TA3组、顺铂组、TA3+顺铂组;应用细胞克隆形成检测联合用药对细胞增殖的影响;利用流式细胞检测术检测细胞凋亡率和周期分布;qRT-PCR和Western blot检测C-myc, cleaved-caspase-3,caspase-3基因mRNA和蛋白的相对表达量。Western blot检测用药后对细胞中JAK-2/STAT3通路蛋白表达的影响。流式细胞术检测细胞PD-L1的表达。结果 CCK8、克隆形成结果表明TA3组明显增加顺铂对A549/DDP细胞的敏感性;与顺铂组单独使用组相比,TA3和顺铂联用明显增加A549/DDP细胞凋亡率(P <0.05),诱导细胞周期阻滞在G0/G1期,同时抑制抗凋亡蛋白C-myc的mRNA和蛋白表达水平,促进凋亡蛋白caspase-3的mRNA和蛋白表达水平(P <0.05)。Western blot实验结果表明两药联用可以明显下调p-JAK2和pSTAT3蛋白表达水平(P <0.05)。流式细胞术结果显示TA3与顺铂联用能够抑制顺铂诱导的PD-L1的表达。结论TA3能增加A549/DDP对顺铂的敏感性,其机制可能与抑制JAK-2/STAT3信号通路和下调PD-L1的表达相关。Aim To explore the mechanism of timosaponin AⅢincreasing cisplatin sensitization of non-small cell lung cancer cisplatin-resistant A549/DDP cells.Methods The cell proliferation was detected by CCK8,the cell apoptosis was detected by Annexin V/PI,and the cell cycle distribution was measured by PI.The effect of the combined drug on cell proliferation was detected by cell clone formation.The mRNA expression levels of C-myc,cysteinyl aspartate-specific protease-3(caspase-3)were detected by reverse transcription polymerase chain reaction(RT-PCR).Western blot and Flow Cytometry were performed for the mechanism study.Results The results of CCK8 and clone formation showed that the timosaponin AⅢgroup significantly increased the sensitivity of cisplatin to A549/DDP cells.Compared with the cisplatin group alone,the combination of timosaponin AⅢand cisplatin significantly elevated the apoptosis rate of A549/DDP cells(P<0.05),induced cell cycle arrest in G 0/G 1 phase,reduced the mRNA and protein expression levels of the anti-apoptotic protein C-myc,and promoted the mRNA and protein expression levels of the apoptotic protein caspase-3(P<0.05).The results of Western blot showed that timosaponin AⅢ+cisplatin could significantly down-regulate the protein expression levels of p-JAK2 and p-STAT3(P<0.05).The results of flow cytometry showed that timosaponin AⅢcombined with cisplatin could inhibit the expression of PD-L1 induced by cisplatin.Conclusions Timosaponin AⅢcan increase the sensitivity of A549/DDP to cisplatin,and its mechanism may be related to the inhibition of JAK-2/STAT3 signaling pathway and the down-regulation of PD-L1 expression.

关 键 词：TA3 肺癌 顺铂 JAK2/STAT3/PD-L1 耐药 凋亡 

分 类 号：R284.1[医药卫生—中药学] R329.25[医药卫生—中医学] R345.57R734.2R979.1