摘要
2',4'-二甲氧基查尔酮(DMC)是卡瓦胡椒素B的结构修饰物,本课题以胃癌细胞MGC-803和HGC-27为研究对象,探讨DMC对胃癌细胞的体内外抗肿瘤作用及机制。采用CCK-8法、EdU染色法和Annexin V-FITC/PI双染流式细胞术检测胃癌细胞活力、增殖率和凋亡率,发现DMC抑制胃癌细胞活力和增殖,促进胃癌细胞凋亡;构建裸鼠胃癌细胞皮下移植瘤模型观察DMC对胃癌移植瘤生长的影响,发现DMC抑制裸鼠胃癌皮下移植瘤的生长,动物实验获得上海中医药大学动物伦理委员会的批准,伦理编号为PZSHUTCM2310110002。转录组学研究DMC对MGC-803细胞RNA表达的影响,发现生物功能富集于糖酵解;采用2-NBDG探针标记及流式细胞术和乳酸测试盒检测胃癌细胞葡萄糖摄取能力和乳酸生成与外排能力,发现DMC抑制胃癌细胞的葡萄糖摄取能力和乳酸生成与外排;通过蛋白印迹实验检测胃癌细胞增殖、凋亡及糖酵解相关蛋白的表达,发现DMC能上调胃癌细胞促凋亡蛋白cleaved caspase-9、cleaved caspase-3、cleaved PARP的表达,下调胃癌细胞增殖标志物Ki-67的蛋白表达,并抑制胃癌细胞糖酵解相关蛋白c-Myc、LDHA、GLUT3、PDHK1和MCT1的表达;通过Seahorse能量代谢分析仪检测实时糖酵解速率,发现DMC能下调胃癌细胞基础糖酵解速率和代偿糖酵解;通过c-Myc过表达稳转株MGC-803细胞的翻转实验明确DMC通过抑制c-Myc介导的葡萄糖摄取和糖酵解发挥胃癌抑制作用。综上所述,DMC可能通过调控胃癌细胞c-Myc及其靶基因糖酵解相关蛋白的表达,抑制胃癌细胞c-Myc介导的葡萄糖摄取功能和糖酵解,从而抑制胃癌细胞增殖,促进胃癌细胞凋亡,最终在体内外抑制胃癌的生长。
2',4'-Dimethoxychalcone(DMC)is a structural modifier of carvacrol B.In this study,gastric cancer cells MGC-803 and HGC-27 were used as the subjects to investigate the anti-tumor effect and mechanism of DMC on gastric cancer(GC)cells both in vitro and in vivo.DMC inhibited cell viability and cell proliferation and promoted cell apoptosis in GC cells,detected by CCK-8 assay,EdU staining and Annexin V-FITC/PI doublestaining flow cytometry.A nude mouse model of GC cell xenograft was constructed by subcutaneous injection with MGC-803 cells,for measuring the effect of DMC on the growth of GC in vivo,and DMC inhibited the growth of subcutaneous transplantation tumor in nude mice.The animal experiments were approved by the Animal Ethics Committee of Shanghai University of Traditional Chinese Medicine under the ethical number PZSHUTCM2310110002.The effect of DMC on the RNA expression of MGC-803 cells was detected by RNA-seq assay,and it was found that the biological function of DMC was enriched in glycolysis.DMC inhibited the glucose uptake capacity and lactate production and efflux of gastric cancer cells,detected by using 2-NBDG probe with flow cytometry and lactate(LD)test kit.Western blot assay was performed to detect the protein expression of proliferation,apoptosis,and glycolysis-related proteins in gastric cancer cells,and the results demonstrated that DMC up-regulated the protein expression of cleaved caspase-9,cleaved caspase-3,cleaved PARP,down-regulated Ki-67 protein expression,and inhibited the protein expression of c-Myc,LDHA,GLUT3,PDHK1 and MCT1 in gastric cancer cells.The Seahorse energy metabolism analyser was used to measure the rate of glycolysis,and it was found that DMC could down-regulate the basal glycolysis rate and compensatory glycolysis in gastric cancer cells.The c-Myc overexpressing cell line MGC-803 was used in the reversal experiment to further confirm that DMC suppressed gastric cancer growth through inhibiting c-Myc mediated glucose uptake and glycolysis.In conclusion,DMC may inhibit the protein expression of c-Myc and its target glycolysis-related genes,suppressed c-Myc-mediated glucose uptake and glycolysis in gastric cancer cells,thereby inhibited the cell proliferation and promoted cell apoptosis of gastric cancer cells,and thus finally inhibited the growth of gastric cancer in vivo and in vitro.
作者
李昱彤
李文标
张玉杰
潘巧岭
杜婷婷
吴辉
黄菲
费晓燕
吴晓俊
石海莲
LI Yu-tong;LI Wen-biao;ZHANG Yu-jie;PAN Qiao-ling;DU Ting-ting;WU Hui;HUANG Fei;FEI Xiao-yan;WU Xiao-jun;SHI Hai-lian(Shanghai Key Laboratory of Compound Chinese Medicines,Key Laboratory of Chinese Medicine Standardization,Ministry of Education,Key Research Laboratory of New Resources and Quality Evaluation of Chinese Medicines,State Administration of Traditional Chinese Medicine,Shanghai Research Centre for Standardization of Chinese Medicines,Institute of Traditional Chinese Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;Department I of Gastroenterology,Longhua Hospital,Shanghai University of Traditional Chinese Medicine,Shanghai 200032,China)
出处
《药学学报》
北大核心
2025年第5期1421-1431,共11页
Acta Pharmaceutica Sinica
基金
国家自然科学基金项目(82474132)
上海市自然科学基金(23ZR1463100,21ZR1462800)
上海市复方中药重点实验室开放课题(21DZ2270500)
研究生创新培养专项(Y2021088)。
