萝卜硫素通过增强抗氧化能力抑制髓核细胞铁死亡

王泽; 于涵; 焦磊; 王琦; 郑国权; 孙建华

doi:10.12435/j.issn.2095-5227.25021904

萝卜硫素通过增强抗氧化能力抑制髓核细胞铁死亡

Sulforaphane inhibits ferroptosis in nucleus pulposus cells by enhancing antioxidant capacity

摘要

摘要:
背景椎间盘退变过程中，髓核细胞铁死亡的发生与氧化应激失衡密切相关。萝卜硫素作为天然抗氧化剂，其对髓核细胞铁死亡的调控作用及分子机制尚不明确。
目的分析萝卜硫素对Erastin诱导的髓核细胞铁死亡的影响，并评估萝卜硫素对细胞活力及抗氧化能力的调节作用。
方法以人髓核细胞为研究对象，分为对照组、Erastin处理组以及不同浓度的萝卜硫素干预组(2.5 ~ 40 μmol/L)。通过光学显微镜、流式细胞术、CCK-8试剂盒、总抗氧化能力(T-AOC)检测试剂盒、Western blot等方法，检测细胞形态、病死率(PI⁺率)、活性氧(reactive oxygen species，ROS)水平、细胞活力和抗氧化能力以及细胞内Collagen Ⅱ、Nrf2、GPX4、SLC7A11蛋白的表达情况。
结果 Erastin处理组中，髓核细胞体积增大、形态改变，并伴随细胞死亡和细胞内ROS水平升高，提示Erastin成功诱导了髓核细胞的铁死亡(P＜0.001)。流式细胞术分析结果显示，与对照组比较，Erastin处理可以使PI⁺细胞比例增加，从对照组的5.11% ± 0.04%升至10.11% ± 0.58%。而萝卜硫素5 μmol/L和10 μmol/L处理组可以降低PI⁺细胞比例，分别为5.08% ± 0.44%和3.38% ± 0.32%，10 μmol/L萝卜硫素组的PI⁺细胞比例低于5 μmol/L萝卜硫素组(P＜0.001)。以上结果表明，萝卜硫素在一定浓度下能够显著抑制Erastin诱导的髓核细胞铁死亡，10 μmol/L萝卜硫素对PI⁺细胞比例的抑制作用更强。Western blot结果进一步证实，10 μmol/L萝卜硫素处理能够有效增强Nrf2的表达，并恢复Collagen Ⅱ、GPX4、SLC7A11蛋白的表达，逆转Erastin处理引起的蛋白表达下降。
结论萝卜硫素能够通过增强髓核细胞的抗氧化能力，抑制Erastin诱导的铁死亡，进而维持细胞活力。

Abstract:
Background Ferroptosis in nucleus pulposus cells plays a crucial role in intervertebral disc degeneration, and sulforaphane (SFN), a natural compound, is known for its antioxidant properties.
Objective To analyze the effects of SFN on Erastin-induced ferroptosis in nucleus pulposus cells and evaluate its regulation of cell viability and antioxidant capacity.
Methods Human nucleus pulposus cells were used as the study subjects and treated with different groups, including control group, Erastin-treated group, and SFN intervention groups (2.5 μmol/L to 40 μmol/L). Cell morphology, death rate, ROS levels, cell viability, and antioxidant capacity were assessed using optical microscopy, flow cytometry, CCK-8 assay, and Western blotting.
Results In the Erastin-treated group, cells exhibited increased volume, morphological changes, and significant cell death with elevated ROS levels, indicating successful induction of ferroptosis in nucleus pulposus cells. Flow cytometry analysis showed a significant increase in PI⁺ cell proportion after Erastin treatment, rising from (5.11 ± 0.04)% in the control group to (10.11 ± 0.58)%. However, the PI⁺ cell proportion in the SFN 5 μmol/L and 10 μmol/L groups significantly decreased to (5.08 ± 0.44)% and (3.38 ± 0.32)%, respectively, with the 10 μmol/L SFN group showing a significantly lower proportion compared to the 5 μmol/L SFN group (P < 0.001). These results indicated that SFN at certain concentrations could significantly inhibit Erastin-induced ferroptosis, with 10 μmol/L SFN showing the most significant effect in reducing PI⁺ cells. Western blot analysis further confirmed that 10 μmol/L SFN treatment effectively enhanced the expression of Nrf2 and restored the expression of Collagen Ⅱ, GPX4, and SLC7A11 proteins, reversing the protein expression decline caused by Erastin treatment.
Conclusion Sulforaphane can enhance the antioxidant capacity of nucleus pulposus cells and inhibit Erastin-induced ferroptosis, thereby maintaining cell viability, which provides experimental evidence for sulforaphane as a potential therapeutic strategy for intervertebral disc degeneration.

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