目的 探讨粗糠柴苦素（RL）对人结肠癌HCT116细胞自噬和凋亡水平的影响，揭示其潜在的抗肿瘤机制。方法 采用体外培养的HCT116细胞，设置对照组和不同浓度的RL (3.125、6.250、12.500、25.000、50.000μmol·L^-1)组。通过CCK-8法检测细胞存活率；通过流式细胞术分析细胞周期和凋亡率；通过Western blotting法检测细胞自噬相关蛋白的表达水平。设置对照组和不同时间（3、6、12、24 h）的RL (12.5μmol·L^-1)处理组，通过Western blotting法检测细胞自噬相关蛋白与凋亡相关蛋白的表达水平；通过透射电镜检测细胞自噬小体、自噬溶酶体、凋亡小体。进一步设置对照组、RL (12.5μmol·L^-1)组、3-甲基腺嘌呤(3-MA，自噬起始阶段抑制剂，5 mmol·L^-1)组、RL (12.5μmol·L^-1)+3-MA (5 mmol·L^-1)组，以及对照组、RL (12.5μmol·L^-1)组、氯喹(CQ，自噬晚期抑制剂，20μmol·L^-1)组、RL (12.5μmol·L^-1)+CQ (20μmol·L^-1)组，通过Western blotting法检测自噬相关蛋白与凋亡相关蛋白的表达水平；通过流式细胞术分析细胞凋亡率。结果 CCK-8法结果显示，与对照组相比，RL能够显著抑制HCT116细胞的存活率（P<0.01）。流式细胞术结果显示，RL能够显著增加细胞S期比例和细胞凋亡率（P<0.05、0.01）。Western blotting结果显示，RL能够显著提高细胞中LC3II/LC3I、cleaved Caspase-3/Caspase-3比值和Atg5、p62、Caspase-3的蛋白水平（P<0.05、0.01），同时显著降低Beclin-1、Bcl-2的蛋白水平（P<0.05、0.01）。透射电镜结果显示，RL能够显著增加细胞自噬小体、自噬溶酶体、凋亡小体的数量。联合自噬抑制剂的实验中，联用3-MA能够逆转RL单独处理对自噬、凋亡蛋白的影响（P<0.05、0.01），同时其能够逆转RL单独处理对细胞凋亡率的显著增加作用（P<0.01）；而联用CQ则进一步增强RL单独处理的上述效应（P<0.05、0.01）。结论 RL通过诱导HCT116细胞自噬体生成的同时抑制其降解，破坏自噬稳态诱导细胞凋亡，发挥抗肿瘤作用。

Objective To explore the effects of rottlerin（RL） on the autophagy and apoptosis levels of human colon cancer HCT116 cells, and to reveal its potential anti-tumor mechanism. Methods HCT116 cells were cultured in vitro, and divided into a blank group and groups treated with RL at increasing concentrations(3.125, 6.250, 12.500, 25.000, 50.000 μmol·L^-1). Then the cell viability was measured by CCK-8 assay; Cell cycle distribution and apoptosis rate were analyzed by flow cytometry; Cell autophagy-related proteins were assessed by Western blotting. Blank groups and RL treated groups(12.5 μmol·L^-1) at different times（3, 6, 12, 24 h） were set up, and the expression of cell autophagy-related proteins and apoptosis-related proteins were assessed by Western blotting levels; Detection of cellular autophagic vesicles, autophagic lysosomes and apoptotic vesicles by transmission electron microscopy. Additional groups included: blank control, RL alone, 3-methyladenine(3-MA, 5 mmol·L^-1)/chloroquine(CQ, 20 μmol·L^-1) alone, RL combined with 3-MA(5 mmol·L^-1) or CQ(20 μmol·L^-1). Autophagy-related proteins and apoptosis-related proteins were assessed by Western blotting, while the apoptosis rate was analyzed by flow cytometry. Results The CCK-8 assay showed that RL dose-dependently inhibited cell viability of HCT116 cells（P ＜ 0.05, 0.01）. Flow cytometry revealed that RL induced S phase arrest and significantly increased the rate of cellular apoptosis（P ＜ 0.05, 0.01）. Western blotting analysis showed that RL treatment significantly increased the LC3 II/I ratio and upregulated the levels of Atg5, p62, Caspase3 and its cleaved form（P ＜ 0.05, 0.01）, while suppressing Beclin-1 and Bcl-2 proteins（P ＜ 0.05, 0.01）. Transmission electron microscopy results showed that RL significantly increased the number of cellular autophagic vesicles, autophagic lysosomes and apoptotic vesicles. In experiments with combined autophagy inhibitors, 3-MA cotreatmen was able to reverse the effects of RL alone on autophagy and apoptosis proteins（P ＜ 0.05, 0.01）. Additionally, it was able to reverse the significant increase in the rate of cellular apoptosis by RL alone（P ＜ 0.01）. In contrast, CQ synergistically enhanced these effects of RL treatment alone（P ＜ 0.05, 0.01）. Conclusion RL induces autophagy in HCT116 cells while inhibiting its degradation, disrupting autophagy homeostasis and inducing apoptosis, thereby exerting an antitumor effect.

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国家自然科学基金资助项目（81903855）；福建省自然科学基金项目（2023J01860）；福建省科技厅项目（2024J01130，2024Y9510）