目的：基于Nrf2/HO-1/GPX4途径探讨和阐明葫芦巴碱(TRG)保护人视网膜色素上皮(ARPE-19)铁死亡的干预机制。

方法：用不同浓度的葫芦巴碱干预ARPE-19细胞筛选葫芦巴碱干预ARPE-19细胞的最佳浓度，随后进行分组(NC组、HG组、Fer-1组、TRG组)，收集样本进行相关指标测定。按照谷胱甘肽(GSH)、丙二醛(MDA)和铁离子检测试剂盒说明书评价各组细胞中GSH、MDA和铁离子变化水平； 流式细胞术检测各组细胞中ROS变化水平； Western blot分析各组细胞核因子E2相关因子2(Nrf2)、血红素加氧酶-1(HO-1)、谷胱甘肽过氧化物酶4(GPX4)、酰基辅酶A合成酶长链家族4(ACSL4)的表达情况。

结果：40 μg/mL葫芦巴碱的预处理干预措施可有效减轻高糖造成的细胞活性的降低。HG组ROS、MDA的水平显著高于NC组； 与HG组相比，TRG组ROS、MDA的水平显著下降，各组GSH变化情况与ROS、MDA相反，Fer-1组和TRG组中ACSL4蛋白和铁离子水平表达降低，Fer-1组和TRG组中Nrf2、HO-1、GPX4蛋白相对表达水平升高(均P<0.01)。

结论：葫芦巴碱保护ARPE-19细胞免于高糖损伤是通过靶向抑制Nrf2/HO-1/GPX4信号通路抗铁死亡实现的。

METHODS: The ARPE-19 cells were cultured and subsequently treated with varying concentrations of trigonelline to ascertain the most effective concentration for modulating the cells. Then the cells were categorized into distinct groups, including normal control(NC)group, high glucose(HG)group, Fer-1 group, TRG group based on the determined concentration. Samples from each group were then gathered to assess relevant indicators. The intracellular levels of glutathione(GSH), malondialdehyde(MDA), and Ferrion were quantified in accordance with the protocols provided by the GSH, MDA, and Ferrion detection kits. Flow cytometry was employed to measure the ROS levels within each group. Additionally, Western blot analysis was conducted to examine the expression of nuclear factor erythroid 2-related factor 2(Nrf2), heme oxygenase-1(HO-1), glutathione peroxidase(GPX4), and acyl-CoA synthetase long-chain family member 4(ACSL4)across the different groups.

RESULTS: The preconditioning intervention with 40 μg/mL TRG effectively mitigated the decline in cell activity induced by high glucose levels. The levels of reactive oxygen species(ROS)and MDA in the HG group were markedly elevated compared to the NC group; and the TRG group exhibited significantly reduced levels of ROS and MDA compared to those of the HG group, with the antioxidant stress index GSH showing opposite trends to those of ROS and MDA across all the groups. Whereas the Fer-1 and TRG groups showed decreased expression levels of ACSL4 protein and iron ions, and the expression levels of Nrf2, HO-1 and GPX4 in the Fer-1 and TRG groups were increased.

CONCLUSION: TRG protects ARPE-19 cells from the detrimental effects of high glucose by targeting the Nrf2/HO-1/GPX4 signaling pathway to counter ferroptosis.