自噬在5-氟尿嘧啶联合奥沙利铂诱导结肠癌细胞铁死亡中的作用

doi:10.3969/j.issn.1004-616x.2026.01.002

摘要/Abstract

摘要： 目的：探讨自噬在5-氟尿嘧啶(5-FU)联合奥沙利铂(L-OHP)诱导人结肠癌细胞铁死亡中的作用。方法：首先通过CCK-8法检测不同浓度5-FU和L-OHP分别单独作用SW620细胞24 h后对细胞增殖活性的影响，计算两种药物的IC₅₀值，均选择IC₅₀浓度进行后续联合处理。然后将SW620细胞分为8组，包括对照组、5-FU组、L-OHP组、5-FU与L-OHP组、自噬抑制剂3-甲基腺嘌呤(3-MA)组、3-MA与5-FU组、3-MA与L-OHP组以及3-MA、5-FU、L-OHP三药联合组。采用试剂盒检测各组细胞的活性氧(ROS)和丙二醛(MDA)水平，Western blot法检测铁死亡相关蛋白SLC7A11、GPX4和ACSL4及自噬相关蛋白LC3B的表达，使用透射电子显微镜观察线粒体形态。结果：与对照组相比，5-FU和L-OHP分别单独作用时SW620细胞的存活率均降低(P<0.01)，计算得到5-FU和L-OHP单独作用时细胞存活的IC₅₀值分别为2 971和185 μmol/L，将两种药物按IC₅₀值浓度进行联合处理后进一步抑制了SW620细胞的增殖(P<0.01)；5-FU、L-OHP单独及联合处理可显著提高细胞内ROS和MDA水平(P<0.01)，使细胞铁死亡相关蛋白SLC7A11、GPX4表达水平降低(P<0.01)，ACSL4蛋白表达水平升高(P<0.05)。使用自噬抑制剂3-MA干预后，与对照组相比，三药联合处理组铁死亡相关蛋白SLC7A11和GPX4表达降低(P<0.01)，ACSL4表达升高(P<0.01)；L-OHP处理组、5-FU联合L-OHP组细胞内自噬相关蛋白LC3B-II/I表达水平上调(P<0.05或P<0.01)。与5-FU和/或L-OHP组相比，三药联合处理组细胞存活率进一步降低(P<0.01)、MDA和ROS水平显著升高(P<0.01)；添加了3-MA的5-FU和/或L-OHP干预组ACSL4蛋白表达水平升高(P<0.05或P<0.01)；而SLC7A11和GPX4蛋白表达水平降低(P<0.05或P<0.01)；透射电镜下5-FU和/或L-OHP使SW620细胞线粒体损伤，联合处理组线粒体形态损伤更加明显。结论： 5-FU和L-OHP可抑制结肠癌SW620细胞增殖，诱导铁死亡发生，共同处理进一步提高癌细胞铁死亡水平；抑制自噬可加强5-FU和L-OHP联合处理的效果。

关键词: 结直肠癌, 自噬, 铁死亡, 奥沙利铂, 5-氟尿嘧啶

Abstract: OBJECTIVE： To investigate the role of autophagy in inducing ferroptosis in human colon cancer SW620 cells treated with 5-fluorouracil (5-FU) combined with oxaliplatin (L-OHP). METHODS： The CCK-8 assay was used to assess the effects of different concentrations of 5-FU and L-OHP acting alone on SW620 cell proliferation after 24 h. The IC₅₀ values for both drugs were calculated，and the IC₅₀ concentrations were selected for subsequent combination treatment. SW620 cells were then divided into eight groups： control， 5-FU，L-OHP，5-FU +L-OHP，autophagy inhibitor 3-methyladenine (3-MA)，3-MA+5-FU，3-MA+L-OHP， and the triple combination of 3-MA， 5-FU and L-OHP. Reagent kits were used to detect reactive oxygen species (ROS) and malondialdehyde (MDA) levels in each cell group. Western blot analysis was employed to assess the expression of ferroptosis-related proteins SLC7A11， GPX4， and ACSL4， as well as autophagy-related protein LC3B. Transmission electron microscopy was utilized to observe mitochondrial morphology. RESULTS： Compared with the control group， both 5-FU and L-OHP individually inhibited SW620 cell proliferation (P<0.01). The calculated IC₅₀ values for cell survival under single treatment with 5-FU and L-OHP were 2 971 and 185 μmol/L，respectively. Combined treatment at the IC₅₀ concentrations of both drugs further inhibited SW620 cell proliferation (P<0.01). Both single and combined treatments with 5-FU and L-OHP significantly increased ROS and MDA levels (P<0.01)，while decreasing the ferroptosis-related proteins SLC7A11 and GPX4 (P<0.01)， and increased ACSL4 expression (P<0.05). Following intervention with the autophagy inhibitor 3-MA， compared to the control group， the triple combination group exhibited decreased expression of ferroptosis-related proteins SLC7A11 and GPX4 (P<0.01) and increased ACSL4 expression (P<0.01). In the L-OHP-treated group and the 5-FU and L-OHP group，intracellular autophagy-related protein LC3B-II/I expression was upregulated (P<0.05 or P<0.01). Compared with the 5-FU and/or L-OHP groups，the triple-drug combination group exhibited further reduced cell viability (P<0.01) and significantly elevated MDA and ROS levels (P<0.01). In the 5-FU and/or L-OHP intervention groups supplemented with 3-MA，ACSL4 protein expression increased (P<0.05 or P<0.01)，while SLC7A11 and GPX4 proteins expression decreased (P<0.05 or P<0.01). Transmission electron microscopy revealed that 5-FU and/or L-OHP induced mitochondrial damage in SW620 cells，with more pronounced morphological alterations observed in the combination treatment group.CONCLUSION： 5-FU and L-OHP inhibited proliferation of SW620 colon cancer cells and induced ferroptosis. Co-treatment further enhanced the level of ferroptosis in cancer cells. Inhibition of autophagy potentiated the effect of combined 5-FU and L-OHP treatment.

Key words: colorectal cancer, autophagy, ferroptosis, oxaliplatin, 5-fluorouracil

中图分类号:

R735.3+5

吉佳音, 罗梦雨, 李红霞, 钟照玥, 康玲. 自噬在5-氟尿嘧啶联合奥沙利铂诱导结肠癌细胞铁死亡中的作用[J]. 癌变·畸变·突变, 2026, 38(1): 7-14.

JI Jiayin, LUO Mengyu, LI Hongxia, ZHONG Zhaoyue, KANG Ling. Effect of autophagy on ferroptosis in colon cancer cells treated with the combination of 5-fluorouracil and oxaliplatin[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2026, 38(1): 7-14.

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