中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2018, Vol. 13 ›› Issue (5): 896-907.doi: 10.4103/1673-5374.232488

• 原著:退行性病与再生 • 上一篇    下一篇

实验性自身免疫性脑脊髓炎进展期间小胶质细胞和巨噬细胞中Nogo受体的表达

  

  • 收稿日期:2018-03-27 出版日期:2018-05-15 发布日期:2018-05-15
  • 基金资助:

    澳大利亚多发性硬化研究和Trish多发性硬化研究基金会研究生奖学金; 国家多发性硬化症协会项目Grant＃RG4398A1 / 1，国际进展多发性硬化联盟挑战奖＃PA0065，多发性硬化症研究澳大利亚和Trish多发性硬化症研究基金会＃15-022和Bethlehem Griffiths研究基金会＃BGRF1706

Nogo receptor expression in microglia/macrophages during experimental autoimmune encephalomyelitis progression

Amani A. Alrehaili1, 2, Jae Young Lee1, 3, Maha M. Bakhuraysah1, 2, Min Joung Kim1, Pei-Mun Aui1, Kylie A. Magee1, Steven Petratos1   

  1. 1 Department of Neuroscience, Central Clinical School, Monash University, Prahran Victoria, Australia
    2 Department of Clinical Laboratories, College of Applied Medical Sciences, Taif University, Taif, Kingdom of Saudi Arabia
    3 Toolgen Inc., Gasan Digital-Ro, Geumcheon, Seoul, Korea
  • Received:2018-03-27 Online:2018-05-15 Published:2018-05-15
  • Contact: Steven Petratos, Ph.D.,steven.petratos@monash.edu
  • Supported by:

    This study was supported by Multiple Sclerosis Research Australia and Trish Multiple Sclerosis Research Foundation Postgraduate Scholarship (to JYL); the National Multiple Sclerosis Society Project Grant #RG4398A1/1, International Progressive Multiple Sclerosis Alliance Challenge Award #PA0065, Multiple Sclerosis Research Australia and Trish Multiple Sclerosis Research Foundation #15-022 and Bethlehem Griffiths Research Foundation #BGRF1706 (to SP).

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摘要：

中枢神经系统（CNS）内的髓磷脂相关抑制因子被认为是疾病或损伤后轴突再生的主要障碍之一。 Nogo受体1（NgR1）在限制受损和患病哺乳动物CNS轴突再生中发挥关键作用，但nogo受体在CNS炎症过程免疫细胞激活中的作用尚未得到充分阐释。为此，实验将髓鞘少突胶质细胞糖蛋白MOG35-55肽注射于ngr1+/+ 和 ngr1-/-雌性小鼠皮下，以诱导建立实验性自身免疫性脑脊髓炎（EAE）模型。使用细胞标记物CD11b和Iba1分别在对临床评分0-3，即运动障碍越来越重的EAE小鼠脊髓中小胶质细胞和巨噬细胞的命运进行分析。采用免疫印迹法、免疫组化和流式细胞分析，检测脊髓小胶质细胞/巨噬细胞中Nogo受体及其两种同系物在EAE进展期间的表达。通过检测ngr1+/+ 和 ngr1-/-小鼠脊髓小胶质细胞/巨噬细胞中Nogo-A和MOG水平，证实了疾病进展期间髓鞘蛋白吞噬作用。流式细胞术分析来评估ngr1+/+ 和 ngr1-/-EAE小鼠脊髓中M1和M2小胶质细胞/巨噬细胞的表型。疾病临床症状进展期间在ngr1+/+ 和 ngr1-/-小鼠之间的小胶质细胞和巨噬细胞数量明显不同，且该差异也明显存在于脊髓的灰质与白质区域对比上，且这些差异与细胞上NgR的表达无关。实验证实，随着疾病进展，ngr1-/-小鼠小胶质细胞/巨噬细胞对髓磷脂碎片的吞噬作用有所增强。ngr1-/-小鼠EAE进展过程中显示出了从M1致病性细胞表型到M2-神经营养细胞表型的调节过程。这些证据表明ngr1-/-小鼠的中枢神经系统特异性巨噬细胞和小胶质细胞可表现出清除炎性病灶中隔离抑制性分子能力的增强。

orcid:0000-0003-1211-4577(Steven Petratos) 

关键词: 小胶质细胞, 实验性自身免疫性脑脊髓炎, Nogo受体, 髓鞘相关抑制因子（MAIF）, Nogo A, 神经再生

Abstract:

Myelin-associated inhibitory factors within the central nervous system (CNS) are considered to be one of the main obstacles for axonal regeneration following disease or injury. The nogo receptor 1 (NgR1) has been well documented to play a key role in limiting axonal regrowth in the injured and diseased mammalian CNS. However, the role of nogo receptor in immune cell activation during CNS inflammation is yet to be mechanistically elucidated. Microglia/macrophages are immune cells that are regarded as pathogenic contributors to inflammatory demyelinating lesions in multiple sclerosis (MS). In this study, the animal model of MS, experimental autoimmune encephalomyelitis (EAE) was induced in ngr1+/+ and ngr1–/–female mice following injection with the myelin oligodendrocyte glycoprotein (MOG35–55) peptide. A fatemap analysis of microglia/macrophages was performed throughout spinal cord sections of EAE-induced mice at clinical scores of 0, 1, 2 and 3, respectively (increasing locomotor disability) from both genotypes,using the CD11b and Iba1 cell markers. Western immunoblotting using lysates from isolated spinal cord microglia/macrophages, along with immunohistochemistry and flow cytometric analysis, was performed to demonstrate the expression of nogo receptor and its two homologs during EAE progression. Myelin protein engulfment during EAE progression in ngr1+/+ and ngr1–/– mice was demonstrated by western immunblotting of lysates from isolated spinal cord microglia/macrophages, detecting levels of Nogo-A and MOG. The numbers of M1 and M2 microglia/macrophage phenotypes present in the spinal cords of EAE-induced ngr1+/+ and ngr1–/– mice, were assessed by flow cytometric analysis using CD38 and Erg-2 markers. A significant difference in microglia/macrophage numbers between ngr1+/+ and ngr1–/– mice was identified during the progression of the clinical symptoms of EAE, in the white versus gray matter regions of the spinal cord. This difference was unrelated to the expression of NgR on these macrophage/microglial cells. We have identified that as EAE progresses, the phagocytic activity of microglia/macrophages with myelin debris, in ngr1–/– mice, was enhanced. Moreover, we show a modulation from a predominant M1-pathogenic to the M2-neurotrophic cell phenotype in the ngr1–/– mice during EAE progression. These findings suggest that CNS-specific macrophages and microglia of ngr1–/– mice may exhibit an enhanced capacity to clear inhibitory molecules that are sequestered in inflammatory lesions.

Key words: microglia, experimental autoimmune encephalomyelitis, nogo receptor, myelin-associated inhibitory factors (MAIFs), Nogo A, neural regeneration