Hypoxic preconditioning (HPC) has been reported to alleviate neuronal damage and microglial activation in hippocampal CA1 after transient global cerebral ischemia (tGCI). However, the molecular mechanism is unclear. Recent studies identified that nuclear factor-kappa-B (NF-κB)/oligomerization domain-like receptors protein (NLRP) 3 inflammasome pathway is mainly involved in the activation of microglia and that phosphorylated (p)-mixed lineage kinase domain-like (MLKL) is related to the regulation of NF-κB/NLRP3 axis. Hence, in this study, we set out to investigate whether HPC attenuates neuronal damage and microglial activation through inhibiting NF-κB/NLRP3 axis mediated by p-MLKL after tGCI in CA1 of male rats. We found that HPC decreased NLRP3 inflammasome in microglia and inhibited M1 polarization of microglia in CA1 after tGCI. Mechanistically, HPC inhibited the activation of NF-κB signaling pathway and reduced the mRNA and protein levels of NLRP3 inflammasome after tGCI. Additionally, the knockdown of p-MLKL by short hairpin RNA (shRNA) administration inhibited the activation of the NF-κB signaling pathway and reduced the formation of NLRP3 inflammasome, thus attenuating M1 polarization of microglia and decreasing the release of interleukin 1 beta (IL-1β) and necrosis factor alpha (TNF-α) in CA1 post ischemia. We consider that p-MLKL in microglia may be derived from necroptotic neurons after tGCI. In conclusion, the new finding in this study is that HPC-induced neuroprotection against tGCI through inhibiting NF-κB/NLRP3 pathway mediated by p-MLKL.

据报道，低氧预处理 (HPC) 可减轻短暂性全脑缺血 (tGCI) 后海马 CA1 区的神经元损伤和小胶质细胞活化。然而，其分子机制尚不清楚。最近的研究发现，核因子-κ-B（NF-κB）/寡聚化结构域样受体蛋白（NLRP）3炎症小体通路主要参与小胶质细胞的激活，并且磷酸化（p）-混合谱系激酶结构域样（ MLKL）与NF-κB/NLRP3轴的调节有关。因此，在本研究中，我们着手研究HPC是否通过抑制雄性大鼠CA1区tGCI后p-MLKL介导的NF-κB/NLRP3轴来减轻神经元损伤和小胶质细胞活化。我们发现，HPC 减少了小胶质细胞中的 NLRP3 炎性体，并抑制了 tGCI 后 CA1 区小胶质细胞的 M1 极化。从机制上讲，HPC在tGCI后抑制NF-κB信号通路的激活并降低NLRP3炎症小体的mRNA和蛋白水平。此外，通过短发夹 RNA (shRNA) 敲低 p-MLKL 可抑制 NF-κB 信号通路的激活，并减少 NLRP3 炎症小体的形成，从而减弱小胶质细胞的 M1 极化并减少白细胞介素 1 β (IL) 的释放。 -1β）和坏死因子α（TNF-α）在CA1缺血后。我们认为小胶质细胞中的 p-MLKL 可能源自 tGCI 后坏死性凋亡的神经元。总之，本研究的新发现是HPC通过抑制p-MLKL介导的NF-κB/NLRP3通路而诱导针对tGCI的神经保护作用。