目的 探討2',4'-二羥基-3'-甲基-3-甲氧基查耳酮(C20)對人肝癌HepG2細胞的體外抗腫瘤作用及其潛在的作用機制。方法 通過CCK-8法、集落形成實驗、5-乙炔基-2'-脫氧尿苷(EdU)染色法檢測C20對人肝癌HepG2細胞增殖的影響;通過彗星實驗檢測C20(10 μmol·L^-1)對HepG2細胞DNA損傷的影響;通過流式細胞術檢測C20(5、10 μmol·L^-1)對HepG2細胞周期阻滯的影響;通過Hoechst染色和流式細胞術檢測C20(5、10 μmol·L^-1)對HepG2細胞凋亡的影響。借助Western blotting法檢測C20(5、10 μmol·L^-1)處理對HepG2細胞中與凋亡、DNA損傷、細胞周期阻滯相關蛋白表達水平的調控作用。結果 與對照組比較,C20顯著抑制HepG2細胞的活力(P<0.001),給藥48 h的半數(shù)抑制濃度(IC₅₀)為7.937 μmol·L^-1;5 μmol·L^-1 C20能夠顯著抑制HepG2細胞的集落形成能力(P<0.01);EdU染色結果顯示5、10 μmol·L^-1的C20能夠抑制人肝癌HepG2細胞的增殖能力;5、10 μmol·L^-1的C20顯著誘導HepG2細胞G₂/M期阻滯(P<0.001);5、10 μmol·L^-1的C20顯著促進HepG2細胞凋亡(P<0.001),并顯著上調Caspas-3、Caspase-9以及PARP的剪切水平(P<0.01);10 μmol·L^-1的C20能夠誘導HepG2細胞發(fā)生DNA損傷,并且5、10 μmol·L^-1的C20顯著上調γH2AX、p21的蛋白水平(P<0.01)。結論 C20能夠造成HepG2細胞發(fā)生DNA損傷,上調p21蛋白水平,導致細胞G₂/M期阻滯,并進一步誘發(fā)凋亡,發(fā)揮體外抗肝癌作用。

Objective To investigate the in vitro antitumor effects of 2',4'-dihydroxy-3'-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 (10 μmol·L^-1) on DNA damage of HepG2 cells were detected by comet assay. The effects of C20 (5, 10 μmol·L^-1) on cell cycle of HepG2 cells were detected by flow cytometry. The effects of C20 (5 and 10 μmol·L^-1) on apoptosis of HepG2 cells were detected by Hoechst staining and flow cytometry. The effects of C20 (5 and 10 μmol·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 7.937 μmol·L^-1 after 48 h of treatment; 5 μmol·L^-1 C20 significantly inhibited the colony-forming ability of HepG2 cells (P < 0.01); the EdU staining results showed that 5 and 10 μmol·L^-1 of C20 could inhibit the proliferation of human hepatocellular carcinoma HepG2 cells; 5 and 10 μmol·L^-1 of C20 significantly induced G₂/M phase arrest in HepG2 cells (P < 0.001); 5 and 10 μmol·L^-1 of C20 significantly promoted apoptosis in HepG2 cells (P < 0.001), and significantly upregulated the cleavage levels of Caspas-3, Caspase- 9, and PARP (P < 0.01); 10 μmol·L^-1 of C20 could induce DNA damage in HepG2 cells, and 5 and 10 μmol·L^-1 of C20 significantly upregulated the protein levels of γH2AX 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.

R965

國家自然科學基金項目(82074072);中央高?；究蒲袠I(yè)務費專項資金(2023-JYB-JBQN-051);北京中醫(yī)藥大學國家級人才精準培育計劃(JZPY202206);國家中醫(yī)藥管理局青年岐黃學者支持項目