Department of Neurology, Cangzhou Central Hospital, No. 16, Xinhua West Road, Yunhe District, Hebei, Cangzhou, 061014, China.
Metab Brain Dis. 2022 Jun;37(5):1517-1526. doi: 10.1007/s11011-022-00950-5. Epub 2022 Mar 25.
Ischemic stroke is one of the most common causes of death worldwide. The transformation of microglia from the classic M1 to the alternative M2 state has been shown to have both deleterious and immunosuppressive roles in neuroinflammation. Microglial polarization toward the M2 phase is currently proposed to be a beneficial phenotype in brain ischemic injury. Phoenixin-20 is a newly identified pleiotropic neuropeptide expressed abundantly in different brain regions. In this study, we found that administration of Phoenixin-20 in ischemic stroke middle cerebral artery occlusion (MCAO) mice significantly reduced the brain infarction area but improved the neurological deficit score. Gene expression analysis showed Phoenixin-20 treatment inhibited pro-inflammatory M1 phase microglial markers: a cluster of differentiation molecule 11b (CD11b), cluster of differentiation molecule 86 (CD86), inducible nitric oxide synthase (iNOS), tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and increased anti-inflammatory M2 phase markers (found in Inflammatory Zone 1 (FIZZ1), Arginase 1 (Arg-1), Chitinase 3-like 3 (YM1), and interleukin-10 (IL-10)) in the infarcted brain. We further investigated the molecular mechanism of Phoenixin-20 in cultured microglia. We found that treatment with it induced signature genes expression in microglial M2 state, including Fizz1, Arg-1, YM1, and IL-10, indicating the promotion of microglial polarization toward the M2 state. Furthermore, we found that treatment with the M2 phase cytokine interleukin 4 (IL-4) induced the expression of microglial G Protein-Coupled Receptor (GPR173), which is the receptor of Phoenixin-20. Silencing of the microglial signal transducer and activator of transcription 6 (STAT6) partially blocked the effect of IL-4 on GPR173, suggesting that STAT6 is the upstream regulator of GPR173. Finally, we showed that the silencing of GPR173 completely abolished the effect of Phoenixin-20 in microglia, indicating the dependency of its regulatory role on GPR173. Collectively, our study demonstrates that Phoenixin-20 has a protective role in the acute stroke model. Our cell-based study demonstrates Phoenixin-20 promotes microglia toward M2 transformation, which could be the mechanism of its neuroprotection.
缺血性中风是全球最常见的死亡原因之一。小胶质细胞从经典的 M1 向替代的 M2 状态的转化，在神经炎症中既有有害作用，也有免疫抑制作用。小胶质细胞向 M2 阶段的极化目前被认为是脑缺血损伤中的有益表型。凤凰素-20 是一种新发现的多效神经肽，在不同的脑区大量表达。在这项研究中，我们发现，给予缺血性中风大脑中动脉闭塞 (MCAO) 小鼠凤凰素-20 治疗显著减少了脑梗死面积，但改善了神经功能缺损评分。基因表达分析显示，凤凰素-20 治疗抑制了促炎 M1 期小胶质细胞标志物：分化抗原分子 11b (CD11b)、分化抗原分子 86 (CD86)、诱导型一氧化氮合酶 (iNOS)、肿瘤坏死因子-α (TNF-α)、白细胞介素 6 (IL-6)，并增加了抗炎 M2 期标志物（在炎症区 1 (FIZZ1)、精氨酸酶 1 (Arg-1)、几丁质酶 3 样 3 (YM1) 和白细胞介素 10 (IL-10)）在梗死的大脑中。我们进一步研究了凤凰素-20 在培养的小胶质细胞中的分子机制。我们发现，用它处理诱导了小胶质细胞 M2 状态的特征基因表达，包括 Fizz1、Arg-1、YM1 和 IL-10，表明促进了小胶质细胞向 M2 状态的极化。此外，我们发现，用 M2 期细胞因子白细胞介素 4 (IL-4) 处理诱导了小胶质细胞 G 蛋白偶联受体 (GPR173) 的表达，GPR173 是凤凰素-20 的受体。沉默小胶质细胞信号转导和转录激活因子 6 (STAT6) 部分阻断了 IL-4 对 GPR173 的作用，表明 STAT6 是 GPR173 的上游调节因子。最后，我们发现沉默 GPR173 完全消除了凤凰素-20 在小胶质细胞中的作用，表明其调节作用依赖于 GPR173。总的来说，我们的研究表明凤凰素-20 在急性中风模型中具有保护作用。我们的细胞研究表明，凤凰素-20 促进小胶质细胞向 M2 转化，这可能是其神经保护作用的机制。
