Connexin 30 Deficiency Attenuates Chronic but Not Acute Phases of Experimental Autoimmune Encephalomyelitis Through Induction of Neuroprotective Microglia.

Mei Fang,Noriko Isobe,Katsuhisa Masaki,Ryo Yamasaki,Hiroo Yamaguchi,Jun-Ichi Kira,Guangrui Li,Atsushi Fujita

doi:10.3389/fimmu.2018.02588

Abstract

Glial connexins (Cxs) form gap junction channels through which a pan-glial network plays key roles in maintaining homeostasis of the central nervous system (CNS). In multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), expression of astrocytic Cx43 is lost in acute lesions but upregulated in chronic plaques, while astrocytic Cx30 is very low in normal white matter and changes in its expression have not been convincingly shown. In Cx30 or Cx43 single knockout (KO) mice and even in Cx30/Cx43 double KO mice, acute EAE is unaltered. However, the effects of Cx30/Cx43 deficiency on chronic EAE remains to be elucidated. We aimed to clarify the roles of Cx30 in chronic neuroinflammation by studying EAE induced by myelin oligodendrocyte glycoprotein peptide 35–55 in Cx30 KO mice. We found that Cx30 deficiency improved the clinical symptoms and demyelination of chronic but not acute EAE without influencing CD3+ T cell infiltration. Furthermore, increased ramified microglia in the naïve state and induced earlier and stronger microglial activation in the acute and chronic phases of EAE was observed. These activated microglia had an anti-inflammatory phenotype, as shown by the upregulation of arginase-1 and brain-derived neurotrophic factor and the downregulation of nitric oxide synthase 2. In the naïve state, Cx30 deficiency induced modest enlargement of astrocytic processes in the spinal cord gray matter and a partial reduction of Cx43 expression in the spinal cord white matter. These astrocytes in Cx30 KO mice showed earlier and stronger activation during the acute phase of EAE, with upregulated A2 astrocyte markers and a significant decrease in Cx43 in the chronic phases. Spinal cord neurons and axons were more preserved in Cx30 KO mice than in littermates in the chronic phase of EAE. These findings suggest that Cx30 deficiency increased ramified microglia in the CNS in the naïve state and improved chronic EAE through redirecting microglia toward an anti-inflammatory phenotype, suggesting a hitherto unknown critical role of astrocytic Cx30 in regulating microglial number and functional state.

Highlights

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) [1]
No Changes in Myelin Density In wild type (WT) littermate mice in the naïve state, Cx30 was expressed predominantly on astrocytes in the gray matter of the spinal cord, cerebellum and cerebrum, while Cx30 expression was very low in white matter astrocytes, including the optic nerve (Figure 1A; Supplementary Figure 1), which is consistent with our previous study in humans [22]
glial fibrillary acidic protein (GFAP) immunostaining revealed neither morphological nor quantitative changes in GFAP+ astrocytes in the white matter between WT and Cx30 KO mice, whereas GFAP+ astrocytes in the gray matter had thicker processes and showed a tendency to be increased in Cx30 KO mice than in WT mice (p = 0.0575, Figures 1A,D,E)

Summary

Introduction

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) [1] It initially presents as relapsing remitting MS (RRMS) but later evolves into secondary progressive MS (SPMS) in ∼20% of patients, even after disease-modifying therapies (DMTs) are introduced [2, 3]. Siponimod [7] and ozanimod [8], new functional antagonists of sphingosine1-phosphate receptor 1 (S1P1), were reported to be effective for preventing disability progression in SPMS. These drugs may directly act on glial cells harboring S1P1 such as microglia and astroglia, in addition to their inhibitory effects on lymphocyte egress from the secondary lymphoid organs [9,10,11]. In chronic MS lesions, persistent demyelination with varying degrees of remyelination and neuroaxonal degeneration are accompanied by the presence of activated microglia but few T cells [12, 13], suggesting a key role of microglia, which are not targeted by the peripherally acting DMTs, in chronic inflammation in SPMS

Objectives

Methods

Results

Conclusion