Quercetin prevents necroptosis of oligodendrocytes by inhibiting macrophages/microglia polarization to M1 phenotype after spinal cord injury in rats
Quercetin prevents necroptosis of oligodendrocytes by inhibiting macrophages/microglia polarization to M1 phenotype after spinal cord injury in rats
复制标题
槲皮素通过抑制大鼠脊髓损伤后巨噬细胞/小胶质细胞极化为 M1 表型来预防少突胶质细胞坏死性凋亡
DOI：
10.1186/s12974-019-1613-2
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发表时间：
2019-11-07
影响因子：
9.3
通讯作者：
Hao, Ding-Jun
中科院分区：
文献类型：
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作者：
Fan, Hong;Tang, Hai-Bin;Hao, Ding-Jun
脊髓损伤（spinal cord injury，SCI）后少突胶质细胞（oligodendrocytes，OLs）的死亡导致脱髓鞘，甚至导致永久性神经功能缺损.除了凋亡，我们以前的研究表明，OLs经历受体相互作用丝氨酸-苏氨酸激酶3（RIP 3）/混合谱系激酶结构域样蛋白（MLKL）介导的坏死性凋亡。考虑到坏死性凋亡总是伴随着促炎症反应，而槲皮素长期以来被用作抗炎剂，本研究探讨了槲皮素是否可以抑制脊髓损伤后OL的坏死性凋亡，抑制M1巨噬细胞/小胶质细胞介导的免疫反应及其可能的机制。以脊髓损伤大鼠为模型，对照组注射生理盐水。采用Basso-Beattie-Bresnahan（BBB）评分和臀高指数（RHI）测定评价运动功能恢复。在体内，通过免疫组织化学和5-溴-2-脱氧尿苷（5-bromo-2-deoxyuridine，BrdU）掺入法检测OLs的坏死性凋亡、凋亡和再生。脊髓损伤后髓鞘和轴突的丢失通过Luxol坚牢蓝（LFB）染色、免疫组化和电镜研究来评估。采用定量逆转录-聚合酶链反应（qRT-PCR）和免疫组化方法检测脊髓损伤后巨噬细胞/小胶质细胞的极化及其机制。体外培养条件下，采用ATP和活性氧（ROS）水平检测、碘化丙啶（PI）标记、Western blotting等方法分析OLs的坏死性凋亡，采用qRT-PCR和Western blotting等方法检测巨噬细胞/小胶质细胞信号通路介导的极化。然后我们发现，槲皮素显着减少脊髓损伤后OL的坏死性凋亡，而不影响OL的凋亡和再生。同时，与SCI +盐水对照相比，槲皮素治疗的大鼠中髓鞘丢失和轴突丢失也显著减少。此外，我们发现，槲皮素可以通过抑制STAT 1和NF-κ B途径在体内和体外抑制极化为M1表型的巨噬细胞/小胶质细胞，这有助于减少necroptosis的OLs.ConclusionsQuercetin治疗减轻necroptosis的OLs部分通过抑制脊髓损伤后M1巨噬细胞/小胶质细胞极化。我们的研究结果表明，OLs的坏死性凋亡可能是临床SCI的潜在治疗靶点。
BackgroundOligodendrocytes (OLs) death after spinal cord injury (SCI) contributes to demyelination, even leading to a permanent neurological deficit. Besides apoptosis, our previous study demonstrated that OLs underwent receptor-interacting serine-threonine kinase 3(RIP3)/mixed lineage kinase domain-like protein (MLKL)-mediated necroptosis. Considering that necroptosis is always accompanied with pro-inflammatory response and quercetin has long been used as anti-inflammatory agent, in the present study we investigated whether quercetin could inhibit necroptosis of OLs and suppress the M1 macrophages/microglia-mediated immune response after SCI as well as the possible mechanism.MethodsIn this study, we applied quercetin, an important flavonoid component of various herbs, to treat rats with SCI and rats injected with saline were employed as the control group. Locomotor functional recovery was evaluated using Basso-Beattie-Bresnahan (BBB) scoring and rump-height Index (RHI) assay. In vivo, the necroptosis, apoptosis, and regeneration of OLs were detected by immunohistochemistry, 5-bromo-2-deoxyuridine (BrdU) incorporation. The loss of myelin and axons after SCI were evaluated by Luxol fast blue (LFB) staining, immunohistochemistry, and electron microscopic study. The polarization of macrophages/microglia after SCI and the underlying mechanisms were detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunohistochemistry. In vitro, the ATP and reactive oxygen species (ROS) level examination, propidium iodide (PI) labeling, and Western blotting were used to analyze the necroptosis of cultured OLs, while the signaling pathways-mediated polarization of cultured macrophages/microglia was detected by qRT-PCR and Western blotting.Results We demonstrated that quercetin treatment improved functional recovery in rats after SCI. We then found that quercetin significantly reduced necroptosis of OLs after SCI without influencing apoptosis and regeneration of OLs. Meanwhile, myelin loss and axon loss were also significantly reduced in quercetin-treated rats, as compared to SCI + saline control. Further, we revealed that quercetin could suppress macrophages/microglia polarized to M1 phenotype through inhibition of STAT1 and NF-kappa B pathway in vivo and in vitro, which contributes to the decreased necroptosis of OLs.ConclusionsQuercetin treatment alleviated necroptosis of OLs partially by inhibiting M1 macrophages/microglia polarization after SCI. Our findings suggest that necroptosis of OLs may be a potential therapeutic target for clinical SCI.