多巴胺通过抑制髓核细胞的铁死亡减缓椎间盘退行性变

关键词：椎间盘退行性变  多巴胺  铁死亡  脂质代谢

Keywords:Intervertebral disc degeneration  Dopamine  Ferroptosis  Lipid metabolism

摘要：

      目的 探讨多巴胺（DA）通过抑制铁死亡通路对脂多糖（LPS）诱导的髓核细胞（NPCs）损伤的保护作用及机制。方法 采用大鼠原代髓核细胞，通过LPS（50 ng/mL）构建炎性损伤模型。检测DA（0.1~10.0 μmol）的细胞毒性，测定氧化应激指标［活性氧（ROS）、丙二醛（MDA）、还原型谷胱甘肽（GSH）］、炎性因子［白细胞介素-1β（IL-1β）、IL-6、肿瘤坏死因子-α（TNF-α）］、细胞外基质（ECM）相关分子［Ⅱ型胶原蛋白A1（COL2A1）、聚集蛋白聚糖（ACAN）、基质金属蛋白酶1（MMP1）、MMP13］、铁死亡标志物［谷胱甘肽过氧化物酶4（GPX4）、溶质运载家族7成员11（SLC7A11）、酰基-CoA-合成酶长链家族4（ACSL4）、溶血卵磷脂酰基转移酶3（LPCAT3）］的表达。采用铁死亡诱导剂和抑制剂验证DA的作用机制，测定细胞内铁离子含量，采用透射电镜观察线粒体超微结构。结果 DA（0.1~10.0 μmol）对NPCs无毒性，可剂量依赖性减轻LPS诱导的氧化应激反应（提升GSH表达，降低ROS、MDA表达），抑制炎性因子表达（IL-1β、IL-6、TNF-α），恢复ECM稳态（上调COL2A1、ACAN表达，下调MMP1、MMP13表达），逆转铁死亡标志物表达（上调GPX4、SLC7A11，下调ACSL4、LPCAT3），降低细胞内铁离子蓄积，修复线粒体损伤。且以上保护作用可被铁死亡诱导剂拮抗、被铁死亡抑制剂增强。结论 DA通过抑制铁死亡通路，有效缓解LPS诱导的髓核细胞氧化应激、炎性反应及ECM降解，为椎间盘退行性变的靶向治疗提供了新策略。

Abstract:

      Objective To investigate the protective effect and mechanism of dopamine（DA） against lipopolysaccharide（LPS）-induced injury in nucleus pulposus cells（NPCs） through suppression of the ferroptosis pathway. Methods Rat primary NPCs were treated with LPS（50 ng/mL） to establish an inflammatory injury model. Cytotoxicity of DA（0.1-10.0 μmol） was assessed. The oxidative stress indicators（reactive oxygen species［ROS］，malondialdehyde［MDA］，glutathione［GSH］），inflammatory factors（interleukin-1β［IL-1β］，IL-6，tumor necrosis factor-α［TNF-α］），extracellular matrix（ECM）-related molecules（collagenⅡA1［COL2A1］，aggrecan［ACAN］，matrix metalloproteinase-1［MMP1］，MMP13） and ferroptosis markers（glutathione peroxidase 4［GPX4］，solute carrier family 7 member 11［SLC7A11］，acyl-CoA-synthetase long-chain family member 4［ACSL4］，lysophosphatidylcholine acyltransferase 3［LPCAT3］）were measured. The mechanism of DA was validated using ferroptosis inducer and inhibitor，and the intracellular iron ion content were measured. The mitochondrial ultrastructure was observed using transmission electron microscopy. Results DA（0.1-10.0 μmol） exhibited no cytotoxicity toward NPCs. It dose-dependently attenuated LPS-induced oxidative stress（increased GSH，decreased ROS and MDA），suppressed expression of inflammatory factors（IL-1β，IL-6，TNF-α），restored ECM homeostasis（upregulated COL2A1 and ACAN，downregulated MMP1 and MMP13），reversed ferroptosis marker expression（upregulated GPX4 and SLC7A11，downregulated ACSL4 and LPCAT3），and reduced intracellular iron accumulation，repaired mitochondrial damage. These protective effects were antagonized by ferroptosis inducer and enhanced by ferroptosis inhibitor. Conclusion DA alleviates LPS-induced oxidative stress，inflammatory response，and ECM degradation in NPCs by inhibiting the ferroptosis pathway，providing a novel therapeutic strategy for targeted treatment of intervertebral disc degeneration.