抗菌肽Dermaseptin-PP协同化疗药物抗肿瘤并逆转A549/DDP细胞顺铂耐药性  

Dermaseptin-PP Combined with Chemotherapeutic Agents Treats Tumors and Reverses Cisplatin Resistance of A549/DDP Cells

作　　者：闫丽文 胡祖成 彭凤栖 胡文均 胡海燕 陈基杰 陆洋[1] YAN Li-wen;HU Zu-cheng;PENG Feng-qi;HU Wen-jun;HU Hai-yan;JAKKREE Tangthianchaichana;LU Yang(School of Chinese Materia Medica,Beijing University of Chinese Medicine,Beijing 102488,China;Chulabhorn International College of Medicine,Thammasat University,Pathum Thani 12120,Thailand)

机构地区：[1]北京中医药大学中药学院,北京102488 [2]泰国国立法政大学朱拉蓬国际医学院,巴吞他尼府12120

出　　处：《中国现代中药》2024年第10期1727-1738,共12页Modern Chinese Medicine

基　　金：国家中医药管理局高水平中医药重点学科建设项目(zyyzdxk-2023272)。

摘　　要：目的:探究抗菌肽Dermaseptin-PP对多种化疗药物体外抗肿瘤活性的协同增效作用,以及其对人非小细胞肺癌顺铂(DDP)耐药细胞株A549/DDP耐药性的逆转作用,并进一步探究其作用机制。方法:采用噻唑蓝(MTT)染色法检测不同质量浓度的Dermaseptin-PP对人非小细胞肺癌A549、H157细胞或小鼠乳腺癌4T1细胞的增殖抑制作用,考察低毒质量浓度Dermaseptin-PP与多烯紫杉醇(DTX)、阿霉素(DOX)或DDP联用对A549、H157或4T1细胞的协同抗肿瘤作用;采用细胞计数试剂盒-8(CCK-8)法检测低毒质量浓度Dermaseptin-PP对A549/DDP细胞DDP耐药性的逆转作用。采用流式细胞术检测Dermaseptin-PP对DDP诱导A549细胞凋亡的影响,采用乳酸脱氢酶(LDH)释放实验及碘化丙啶(PI)摄取实验考察Dermaseptin-PP对A549和A549/DDP细胞膜完整性的影响,最后通过扫描电镜观察Dermaseptin-PP处理后A549和A549/DDP细胞膜形态的变化。结果:Dermaseptin-PP可在体外抑制A549、H157和4T1细胞增殖且呈剂量依赖性;与低毒质量浓度Dermaseptin-PP(0.002、0.020、0.200μg·mL^(-1))联用后,DTX、DOX及DDP对A549细胞的增殖抑制率均有不同程度的提高,半数抑制浓度(IC50)均有所降低,药物联合指数(CI)基本均小于0.9,其中Dermaseptin-PP与DDP的协同作用最强(CI<0.4)。此外,Dermaseptin-PP与DDP联用对H157和4T1细胞的CI也均小于0.9,表现为协同作用。低毒质量浓度Dermaseptin-PP(0.2、2.0、4.0μg·mL^(-1))均能显著逆转A549/DDP细胞的DDP耐药性,逆转倍数(RF)分别为2.83、5.09、9.73。相比于单用DDP,与0.2μg·mL^(-1) Dermaseptin-PP联用可显著提高A549细胞的凋亡率(P<0.05);不同质量浓度的Dermaseptin-PP均可显著增加A549、A549/DDP细胞的LDH释放率和PI摄取率,且呈浓度依赖性;扫描电镜观察发现,经2×IC50的Dermaseptin-PP处理后,A549细胞的细胞膜明显受损、质膜解体、细胞形态发生改变,A549/DDP细胞的细胞膜也出现明显孔洞。结论:Dermaseptin-PP对�Objective:To investigate the effects of the antimicrobial peptide Dermaseptin-PP combined with chemotherapeutic agents on the tumor cells in vitro and the cisplatin(DDP)resistance of A549/DDP cells,and further explore the underlying mechanisms.Methods:The methyl thiazolyl tetrazolium(MTT)assay was employed to determine the inhibitory effects of different concentrations of Dermaseptin-PP on the proliferation of A549,H157 or 4T1 cells and the inhibitory effects of low concentrations of Dermaseptin-PP in combination with docetaxel(DTX),doxorubicin(DOX)or DDP on A549,H157 or 4T1 cells.The cell-counting kit 8(CCK-8)assay was used to determine the reversal effects of low concentrations of Dermaseptin-PP on the DDP resistance of A549/DDP cells.Flow cytometry was employed to measure the effect of Dermaseptin-PP on DDP-induced apoptosis of A549 cells.The lactate dehydrogenase(LDH)release assay and propidium iodide(PI)uptake assay were employed to investigate the effect of Dermaseptin-PP on the membrane integrity of A549 and A549/DDP cells.Finally,the morphological changes of A549 and A549/DDP cell membranes after Dermaseptin-PP treatment were observed by scanning electron microscopy.Results:Dermaseptin-PP inhibited the proliferation of A549,H157,and 4T1 cells in vitro in a dose-dependent manner.In combination with low concentrations(0.002,0.020,and 0.200μg·mL^(-1))of Dermaseptin-PP,DTX,DOX,and DDP demonstrated increased inhibition rates on the proliferation of A549 cells,with reduced half maximal inhibitory concentrations(IC50)and the combination indexes(CIs)lower than 0.9.The combination of Dermaseptin-PP with DDP demonstrated the strongest synergistic effect(CI<0.4).In addition,Dermaseptin-PP combined with DDP showed a synergistic effect,with the CIs lower than 0.9 for both H157 and 4T1 cells.Low concentrations(0.2,2.0,and 4.0μg·mL^(-1))of Dermaseptin-PP all significantly reversed the DDP resistance of A549/DDP cells,with the reversal folds(RFs)of 2.83,5.09,and 9.73,respectively.Compared with DDP alone,the combinati

关 键 词：抗菌肽 Dermaseptin-PP 化疗 抗肿瘤 耐药 顺铂 

分 类 号：R285[医药卫生—中药学]