Erdoğan Mehmet Kadir, Ağca Can Ali, Aşkın Hakan
Department of Biology, Faculty of Arts and Sciences, Bingol University, Bingol, Turkey.
Department of Molecular Biology and Genetics, Faculty of Arts and Sciences, Bingol University, Bingol, Turkey.
Nutr Cancer. 2022;74(2):660-676. doi: 10.1080/01635581.2021.1900301. Epub 2021 Jul 26.
The aim of this study was to investigate the antitumor effects of quercetin and luteolin combined with 5-Fluorouracil (5-FU) in HT-29 human colorectal cancer cells. Cell viability induced by quercetin, luteolin and combination of these compounds with 5-FU were determined by MTT assay, also Cell death detection Elisa assay and fluorescence microscopy were performed to investigate apoptotic effects. Hu-VEGF Elisa assay was employed to determine the effects of treatments on angiogenesis. Western blot and qRT-PCR analysis were performed to investigate effects on p53, Bax, Bcl-2, p38 MAPK, mTOR, PTEN, and Akt proteins and genes. The results indicated that quercetin, luteolin and combinations of these compounds with 5-FU inhibited the growth of HT 29 cells. Compared to the control, apoptosis were triggered 8.1 and 10.1 fold in HT-29 cells, that treated with quercetin + 5-FU and luteolin + 5-FU, respectively. VEGF amount significantly decreased by combined treatments. qRT-PCR and western blot results demonstrated that quercetin, luteolin and the combinations of these flavonoids with 5-FU, modulate the apoptotic pathways in HT-29 cells. The increase in p53, Bax, p38 MAPK, and PTEN gene expression levels compared to the control group was 1.71, 1.42, 3.26, and 3.29-fold with 5-FU + L treatment, respectively, while this increase was 8.43, 1.65, 3.55, and 3.54-fold with 5-FU + Q treatment, respectively. In addition, when the anti-apoptotic Bcl-2, mTOR, and Akt gene expression levels were normalized as 1 in the control group, they were 0.28, 0.41, and 0.22 with 5-FU + L treatment, and 0.32, 0.46, and 0.39, respectively, with 5-FU + Q treatment. These findings suggested that quercetin and luteolin synergistically enhanced the anticancer effect of 5-FU in HT 29 cells and may therefore minimize the toxic effects of 5-FU in the clinical treatment of colorectal cancer.
本研究旨在探讨槲皮素和木犀草素与5-氟尿嘧啶（5-FU）联合应用对HT-29人结肠癌细胞的抗肿瘤作用。采用MTT法测定槲皮素、木犀草素以及这些化合物与5-FU联合作用诱导的细胞活力，同时进行细胞死亡检测ELISA分析和荧光显微镜观察以研究凋亡效应。采用人血管内皮生长因子（Hu-VEGF）ELISA分析来确定各处理对血管生成的影响。进行蛋白质印迹法（Western blot）和定量逆转录聚合酶链反应（qRT-PCR）分析以研究对p53、Bax、Bcl-2、p38丝裂原活化蛋白激酶（p38 MAPK）、雷帕霉素靶蛋白（mTOR）、磷酸酶和张力蛋白同源物（PTEN）以及蛋白激酶B（Akt）蛋白和基因的影响。结果表明，槲皮素、木犀草素以及这些化合物与5-FU的组合均抑制HT-29细胞的生长。与对照组相比，用槲皮素+5-FU和木犀草素+5-FU处理的HT-29细胞凋亡分别触发了8.1倍和10.1倍。联合处理使血管内皮生长因子（VEGF）量显著降低。qRT-PCR和蛋白质印迹法结果表明，槲皮素、木犀草素以及这些黄酮类化合物与5-FU的组合可调节HT-29细胞中的凋亡途径。与对照组相比，5-FU+木犀草素（5-FU+L）处理使p53、Bax、p38 MAPK和PTEN基因表达水平分别增加1.71倍、1.42倍、3.26倍和3.29倍，而5-FU+槲皮素（5-FU+Q）处理时分别增加8.43倍、1.65倍、3.55倍和3.54倍。此外，当对照组中抗凋亡的Bcl-2、mTOR和Akt基因表达水平归一化为1时，5-FU+L处理时分别为0.28、0.41和0.22，5-FU+Q处理时分别为0.32、0.46和0.39。这些发现表明，槲皮素和木犀草素可协同增强5-FU对HT-29细胞的抗癌作用，因此在结直肠癌临床治疗中可能会将5-FU的毒性作用降至最低。
