黄芪-莪术药对通过调控缺氧诱导因子及肿瘤干细胞化抑制结肠癌进展并增强5-FU疗效的机制研究

Astragali Radix-Curcumae Rhizoma inhibits colon cancer progression and enhances 5-FU efficacy by regulating hypoxia-inducible factors and tumor stem cells

该研究基于动物和细胞模型研究黄芪-莪术药对通过调控缺氧诱导因子及肿瘤干细胞化抑制结肠癌进展并增强5-氟尿嘧啶(5-FU)疗效的机制。建立裸鼠HCT116皮下移植瘤模型,将造模成功的24只裸鼠随机分为模型组、5-FU组、黄芪-莪术组和联合给药组,治疗后每2 d测量1次肿瘤体积,采用Western blot方法检测裸鼠HCT116皮下移植瘤组织缺氧核心区关键靶点表皮生长因子受体(epidermal growth factor receptor,EGFR)、二氢嘧啶脱氢酶(dihydropyrimidine dehydrogenase,DPYD)、胸苷酸合酶(thymidylate synthase,TYMS)以及缺氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)、缺氧诱导因子-2α(hypoxia-inducible factor-2α,HIF-2α)和肿瘤干细胞标志物CD133、SRY盒转录因子2(SRY-box transcription factor 2,SOX2)的蛋白表达情况。结果表明黄芪-莪术药对能减缓肿瘤生长速度,并显著提高5-FU的抑瘤率。黄芪-莪术药对明显降低肿瘤中EGFR和DPYD的蛋白表达,联合给药组中EGFR和TYMS的蛋白表达量显著降低。与模型组相比,黄芪-莪术药对可以显著抑制移植瘤组织缺氧核心区中HIF-1α、HIF-2α、SOX2、CD133的蛋白表达,与5-FU组相比,联合给药进一步抑制了HIF-1α、HIF-2α、SOX2的蛋白表达。在体外实验中,缺氧后HCT116细胞内HIF-1α、HIF-2α的蛋白表达量显著增加。与单独给予5-FU(1.38μmol·L^(-1))相比,单独使用黄芪-莪术(40 mg·mL^(-1))或5-FU联合黄芪-莪术均能更显著地抑制HIF-1α、HIF-2α、TYMS的蛋白表达。结果显示黄芪-莪术药对降低了结肠癌细胞中缺氧应答分子的表达,并减少了结肠癌的干细胞性质,从而起到了协同增强5-FU对结肠癌的治疗作用。

The animal and cell models were used in this study to investigate the mechanism of Astragali Radix-Curcumae Rhizoma(HQEZ) in inhibiting colon cancer progression and enhancing the efficacy of 5-fluorouracil(5-FU) by regulating hypoxia-inducible factors and tumor stem cells.The animal model was established by subcutaneous transplantation of colon cancer HCT116 cells in nude mice,and 24 successfully modeled mice were randomized into model,5-FU,HQEZ,and 5-FU+HQEZ groups.The tumor volume was measured every two days.Western blot was employed to measure the protein levels of epidermal growth factor receptor(EGFR),dihydropyrimidine dehydrogenase(DPYD),and thymidylate synthase(TYMS),the key targets of the hypoxic core region,as well as the hypoxia-inducible factors HIF-1α and HIF-2α and the cancer stem cell surface marker CD133 and SRY-box transcription factor 2(SOX2).The results of animal experiments showed that HQEZ slowed down the tumor growth and significantly increased the tumor inhibition rate of 5-FU.Compared with the model group,HQEZ significantly down-regulated the protein levels of EGFR and DPYD,and 5-FU+HQEZ significantly down-regulated the protein levels of EGFR and TYMS in tumors.Compared with the model group,HQEZ significantly down-regulated the protein levels of HIF-1α,HIF-2α,SOX2,and CD133 in the hypoxic core region.Compared with the 5-FU group,5-FU+HQEZ lowered the protein levels of HIF-1α,HIF-2α,and SOX2.The cell experiments showed that the protein le-vels of HIF-1α and HIF-2α in HCT116 cells elevated significantly after low oxygen treatment.Compared with 5-FU(1.38 μmol·L~(-1)) alone,HQEZ(40 mg·mL~(-1)) and 5-FU+HQEZ significantly down-regulated the protein levels of HIF-1α,HIF-2α,and TYMS.In conclusion,HQEZ can inhibit the expression of hypoxia-responsive molecules in colon cancer cells and reduce the properties of cancer stem cells,thereby enhancing the therapeutic effect of 5-FU on colon cancer.