Department of Neurology, Neurological Institute, Graduate School of Medical Sciences, Kyushu University, 3-1-1, Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan.
Physiological Genomics, Biomedical Center, Ludwig-Maximilians University Munich, Munich, Germany.
J Neuroinflammation. 2021 Jun 5;18(1):126. doi: 10.1186/s12974-021-02176-1.
Brain astroglia are activated preceding the onset of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). We characterized the effects of brain astroglia on spinal cord inflammation, focusing on astroglial connexin (Cx)43, because we recently reported that Cx43 has a critical role in regulating neuroinflammation.
Because glutamate aspartate transporter (GLAST) astroglia are enriched in the brain gray matter, we generated Cx43;GLAST-CreER mice that were brain gray matter astroglia-specific Cx43 conditional knockouts (Cx43 icKO). EAE was induced by immunization with myelin oligodendroglia glycoprotein (MOG) peptide 10 days after tamoxifen injection. Cx43 mice were used as controls.
Acute and chronic EAE signs were significantly milder in Cx43 icKO mice than in controls whereas splenocyte MOG-specific responses were unaltered. Histologically, Cx43 icKO mice showed significantly less demyelination and fewer CD45 infiltrating immunocytes, including F4/80 macrophages, and Iba1 microglia in the spinal cord than controls. Microarray analysis of the whole cerebellum revealed marked upregulation of anti-inflammatory A2-specific astroglia gene sets in the pre-immunized phase and decreased proinflammatory A1-specific and pan-reactive astroglial gene expression in the onset phase in Cx43 icKO mice compared with controls. Astroglia expressing C3, a representative A1 marker, were significantly decreased in the cerebrum, cerebellum, and spinal cord of Cx43 icKO mice compared with controls in the peak phase. Isolated Cx43 icKO spinal microglia showed more anti-inflammatory and less proinflammatory gene expression than control microglia in the pre-immunized phase. In particular, microglial expression of Ccl2, Ccl5, Ccl7, and Ccl8 in the pre-immunized phase and of Cxcl9 at the peak phase was lower in Cx43 icKO than in controls. Spinal microglia circularity was significantly lower in Cx43 icKO than in controls in the peak phase. Significantly lower interleukin (IL)-6, interferon-γ, and IL-10 levels were present in cerebrospinal fluid from Cx43 icKO mice in the onset phase compared with controls.
The ablation of Cx43 in brain gray matter astroglia attenuates EAE by promoting astroglia toward an anti-inflammatory phenotype and suppressing proinflammatory activation of spinal microglia partly through depressed cerebrospinal fluid proinflammatory cytokine/chemokine levels. Brain astroglial Cx43 might be a novel therapeutic target for MS.
在实验性自身免疫性脑脊髓炎（EAE）发作之前，大脑神经胶质细胞被激活，EAE 是多发性硬化症（MS）的动物模型。我们描述了大脑神经胶质细胞对脊髓炎症的影响，重点研究了神经胶质细胞缝隙连接蛋白（Cx）43，因为我们最近的研究表明 Cx43 在调节神经炎症中具有关键作用。
由于谷氨酸天冬氨酸转运蛋白（GLAST）神经胶质细胞在大脑灰质中丰富，我们生成了 Cx43；GLAST-CreER 小鼠，这是大脑灰质神经胶质细胞特异性 Cx43 条件性敲除（Cx43icKO）。用髓鞘少突胶质糖蛋白（MOG）肽免疫后 10 天，通过给予他莫昔芬注射来诱导 EAE。Cx43 小鼠用作对照。
与对照相比，Cx43icKO 小鼠的急性和慢性 EAE 症状明显较轻，而脾细胞 MOG 特异性反应未改变。组织学上，与对照相比，Cx43icKO 小鼠脊髓中的脱髓鞘和 CD45 浸润免疫细胞（包括 F4/80 巨噬细胞和 Iba1 小胶质细胞）明显减少。在 Cx43icKO 小鼠的预免疫阶段，小脑的全基因芯片分析显示抗炎 A2 特异性神经胶质细胞基因集明显上调，在发病阶段促炎 A1 特异性和全反应性神经胶质细胞基因表达下调，与对照相比。与对照相比，在高峰期，Cx43icKO 小鼠大脑、小脑和脊髓中表达 C3 的 A1 标志物的神经胶质细胞明显减少。与对照相比，在预免疫阶段，分离的 Cx43icKO 脊髓小胶质细胞表现出更多的抗炎和更少的促炎基因表达。特别是，在预免疫阶段，Cx43icKO 中的小胶质细胞表达 Ccl2、Ccl5、Ccl7 和 Ccl8，在高峰期表达 Cxcl9 的水平低于对照。在高峰期，Cx43icKO 中的小胶质细胞圆形度明显低于对照。与对照相比，在发病阶段，Cx43icKO 小鼠的脑脊液中白细胞介素（IL）-6、干扰素-γ和 IL-10 水平明显降低。
大脑灰质神经胶质细胞中 Cx43 的缺失通过促进神经胶质细胞向抗炎表型和抑制脊髓小胶质细胞的促炎激活来减轻 EAE，部分通过降低脑脊液促炎细胞因子/趋化因子水平。大脑神经胶质细胞 Cx43 可能是治疗多发性硬化症的一个新的治疗靶点。
