2ʹ,4ʹ-二羟基-3ʹ-甲基-3-甲氧基查耳酮上调p21抑制人肝癌HepG2细胞的生长  

2′,4′-dihydroxy-3′-methyl-3-methoxychalcone inhibits growth of human hepatocellular carcinoma HepG2 cells through up-regulation of p21

作　　者：王龙燕 张云封[1,2] 王柱国 谭鹏 魏雪娇[1,2] 李军 胡仲冬 WANG Longyan;ZHANG Yunfeng;WANG Zhuguo;TAN Peng;WEI Xuejiao;LI Jun;HU Zhongdong(School of Chinese Materia Medica,Beijing University of Chinese Medicine,Beijing 100029,China;Modern Research Center for Traditional Chinese Medicine,Beijing Institute of Traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]北京中医药大学中药学院,北京100029 [2]北京中医药大学北京中医药研究院中药现代研究中心,北京100029

出　　处：《药物评价研究》2024年第5期994-1001,共8页Drug Evaluation Research

基　　金：国家自然科学基金项目(82074072);中央高校基本科研业务费专项资金(2023-JYB-JBQN-051);北京中医药大学国家级人才精准培育计划(JZPY202206);国家中医药管理局青年岐黄学者支持项目。

摘　　要：目的探讨2ʹ,4ʹ-二羟基-3ʹ-甲基-3-甲氧基查耳酮(C20)对人肝癌HepG2细胞的体外抗肿瘤作用及其潜在的作用机制。方法通过CCK-8法、集落形成实验、5-乙炔基-2′-脱氧尿苷(EdU)染色法检测C20对人肝癌HepG2细胞增殖的影响;通过彗星实验检测C20(10μmol·L^(−1))对HepG2细胞DNA损伤的影响;通过流式细胞术检测C20(5、10μmol·L^(−1))对HepG2细胞周期阻滞的影响;通过Hoechst染色和流式细胞术检测C20(5、10μmol·L^(−1))对HepG2细胞凋亡的影响。借助Western blotting法检测C20(5、10μmol·L^(−1))处理对HepG2细胞中与凋亡、DNA损伤、细胞周期阻滞相关蛋白表达水平的调控作用。结果与对照组比较,C20显著抑制HepG2细胞的活力(P<0.001),给药48 h的半数抑制浓度(IC50)为7.937μmol·L^(−1);5μmol·L^(−1) C20能够显著抑制HepG2细胞的集落形成能力(P<0.01);EdU染色结果显示5、10μmol·L^(−1)的C20能够抑制人肝癌HepG2细胞的增殖能力;5、10μmol·L^(−1)的C20显著诱导HepG2细胞G2/M期阻滞(P<0.001);5、10μmol·L^(−1)的C20显著促进HepG2细胞凋亡(P<0.001),并显著上调Caspas-3、Caspase-9以及PARP的剪切水平(P<0.01);10μmol·L^(−1)的C20能够诱导HepG2细胞发生DNA损伤,并且5、10μmol·L^(−1)的C20显著上调γH2AX、p21的蛋白水平(P<0.01)。结论C20能够造成HepG2细胞发生DNA损伤,上调p21蛋白水平,导致细胞G2/M期阻滞,并进一步诱发凋亡,发挥体外抗肝癌作用。Objective To investigate the in vitro antitumor effects of 2ʹ,4ʹ-dihydroxy-3ʹ-methyl-3-methoxychalcone(C20)on human hepatocellular carcinoma HepG2 cells and its potential mechanisms.Methods The effects of C20 on the proliferation of HepG2 cells were detected by CCK8 assay,colony formation assay,and 5-ethynyl-2′-deoxyuridine(EdU)staining assay.The effects of C20(10μmol·L^(−1))on DNA damage of HepG2 cells were detected by comet assay.The effects of C20(5,10μmol·L^(−1))on cell cycle of HepG2 cells were detected by flow cytometry.The effects of C20(5 and 10μmol·L^(−1))on apoptosis of HepG2 cells were detected by Hoechst staining and flow cytometry.The effects of C20(5 and 10μmol·L^(−1))on expression levels of proteins related to DNA damage,cell cycle,and apoptosis in HepG2 cells were detected by Western blotting.Results Compared with control group,C20 significantly inhibited the viability of HepG2 cells(P<0.001),with a half-maximal inhibitory concentration(IC50)of 7.937μmol·L^(−1) after 48 h of treatment;5μmol·L^(−1) C20 significantly inhibited the colony-forming ability of HepG2 cells(P<0.01);the EdU staining results showed that 5 and 10μmol·L^(−1) of C20 could inhibit the proliferation of human hepatocellular carcinoma HepG2 cells;5 and 10μmol·L^(−1) of C20 significantly induced G2/M phase arrest in HepG2 cells(P<0.001);5 and 10μmol·L^(−1) of C20 significantly promoted apoptosis in HepG2 cells(P<0.001),and significantly upregulated the cleavage levels of Caspas-3,Caspase9,and PARP(P<0.01);10μmol·L^(−1) of C20 could induce DNA damage in HepG2 cells,and 5 and 10μmol·L^(−1) of C20 significantly upregulated the protein levels ofγH2AX and p21(P<0.01).Conclusion We speculate that C20 treatment can induce DNA damage and upregulate p21 expression in HepG2 cells,leading to G2/M arrest and further inducing apoptosis,which is partially responsible for anti-hepatocellular carcinoma activity of C20.

关 键 词：2ʹ 4ʹ-二羟基-3ʹ-甲基-3-甲氧基查耳酮 肝癌 DNA损伤 G2/M周期阻滞 P21 

分 类 号：R965[医药卫生—药理学]