Neuromyelitis optica, a rare neuroinflammatory demyelinating disease of the CNS, is characterized by the presence of specific pathogenic autoantibodies directed against the astrocytic water channel aquaporin 4 (AQP4) and is now considered as an astrocytopathy associated either with complement-dependent astrocyte death or with astrocyte dysfunction. However, the link between astrocyte dysfunction and demyelination remains unclear. We propose glial intercellular communication, supported by connexin hemichannels and gap junctions, to be involved in demyelination process in neuromyelitis optica. Using mature myelinated cultures, we demonstrate that a treatment of 1 h to 48 h with immunoglobulins purified from patients with neuromyelitis optica (NMO-IgG) is responsible for a complement independent demyelination, compared to healthy donors’ immunoglobulins (P < 0.001). In parallel, patients’ immunoglobulins induce an alteration of connexin expression characterized by a rapid loss of astrocytic connexins at the membrane followed by an increased size of gap junction plaques (+60%; P < 0.01). This was co-observed with connexin dysfunction with gap junction disruption (−57%; P < 0.001) and increased hemichannel opening (+17%; P < 0.001), associated with glutamate release. Blocking connexin 43 hemichannels with a specific peptide was able to prevent demyelination in co-treatment with patients compared to healthy donors’ immunoglobulins. By contrast, the blockade of connexin 43 gap junctions with another peptide was detrimental for myelin (myelin density −48%; P < 0.001). Overall, our results suggest that dysregulation of connexins would play a pathogenetic role in neuromyelitis optica. The further identification of mechanisms leading to connexin dysfunction and soluble factors implicated, would provide interesting therapeutic strategies for demyelinating disorders.

Neuromyelitis Optica (NMO) is an autoimmune demyelinating disease of the central nervous system characterized by the presence of autoantibodies (called NMO-IgG) targeting aquaporin-4. Aquaporin-4 is expressed at the perivascular foot processes of astrocytes, in the glia limitans but also at the ependyma. Most studies have focus on studying the pathogenicity of NMO-IgG on astrocytes and NMO is now considered an astrocytopathy. However, periependymal lesions are observed in NMO suggesting that ependymal cells could also be targeted by NMO-IgG. Ependymal cells regulate CSF-parenchyma molecular exchanges, CSF flow and is a niche for subventricular neural stem cells. Our aim was to examine the effect of antibodies from NMO patients on ependymal cells. We exposed two models, i.e., primary culture of rat ependymal cells and explant cultures of rat lateral ventricular wall wholemounts, to purified IgG of NMO patients (NMO-IgG) for 24 hours. We then evaluated the treatment effect using immunolabeling, functional assays, ependymal flow analysis and bulk RNA sequencing. For each experiment, the effects were compared to purified IgG of healthy donor and to non-treated cells. We found that: i) NMO-IgG patients induced AQP4 agglomeration at the surface of ependymal cells, and induced cell enlargement in comparison to controls. In parallel, it induced an increase in gap junction connexin-43 plaque size; ii) NMO-IgG altered the orientation of ciliary basal bodies and functionally impaired cilia motility; iii) NMO-IgG activated the proliferation of subventricular neural stem cells; and iv) treatment with NMO-IgG upregulated the expression of pro-inflammatory cytokines and chemokines in the transcriptomic analysis. Our study showed that NMO-IgG can trigger an early and specific reactive phenotype in ependymal cells, with functional alterations of intercellular communication and cilia, activation of the subventricular stem cell proliferation and the secretion of pro-inflammatory cytokines. These findings suggest a key role for ependymal cells in the early phase of NMO lesion formation.