RGMa Signal in Macrophages Induces Neutrophil-Related Astrocytopathy in NMO.

OBJECTIVE: Repulsive guidance molecule-a (RGMa) is a glycosylphosphatidylinositol-linked glycoprotein which has multiple functions including axon growth inhibition and immune regulation. However, its role in the pathophysiology of neuromyelitis optica (NMO) is poorly understood. Perivascular astrocytopathy, which is induced by the leakage of aquaporin-4 (AQP4)-specific IgG into the central nervous system parenchyma, is a key feature of NMO pathology. We investigated the RGMa involvement in the pathology of NMO astrocytopathy, and tested a therapeutic potential of humanized anti-RGMa monoclonal antibody (RGMa-mAb). METHODS: Using a clinically relevant NMO rat model, we evaluated the therapeutic effect of a RGMa-mAb by behavioral testing, immunohistochemistry, and gene expression assay. We further performed in vitro experiments to address the RGMa-signaling in macrophages. RESULTS: In both NMO rats and an NMO-autopsied sample, RGMa was expressed by the spared neurons and astrocytes, whereas its receptor neogenin was expressed by infiltrating macrophages. AQP4-IgG-induced astrocytopathy and clinical exacerbation in NMO rats were ameliorated by RGMa-mAb treatment. RGMa-mAb treatment significantly suppressed neutrophil infiltration, and decreased the expression of neutrophil chemoattractants. Interestingly, neogenin-expressing macrophages accumulated in the lesion expressed CXCL2, a strong neutrophil chemoattractant, and further analysis revealed that RGMa directly regulated CXCL2 expression in macrophages. Finally, we found that our NMO rats developed neuropathic pain, and RGMa-mAb treatment effectively ameliorated the severity of neuropathic pain. INTERPRETATION: RGMa signaling in infiltrated macrophages is a critical driver of neutrophil-related astrocytopathy in NMO lesions, and RGMa-mAb may provide an efficient therapeutic strategy for NMO-associated neuropathic pain and motor deficits in patients with NMO. ANN NEUROL 2022;91:532-547.

Dynamics of neuronal oscillations underlying nociceptive response in the mouse primary somatosensory cortex.

Pain is caused by tissue injury, inflammatory disease, pathogen invasion, or neuropathy. The perception of pain is attributed to the neuronal activity in the brain. However, the dynamics of neuronal activity underlying pain perception are not fully known. Herein, we examined theta-oscillation dynamics of local field potentials in the primary somatosensory cortex of a mouse model of formalin-induced pain, which usually shows a bimodal behavioral response interposed between pain-free periods. We found that formalin injection exerted a reversible shift in the theta-peak frequency toward a slower frequency. This shift was observed during nociceptive phases but not during the pain-free period and was inversely correlated with instantaneous pain intensity. Furthermore, instantaneous oscillatory analysis indicated that the probability of slow theta oscillations increased during nociceptive phases with an association of augmented slow theta power. Finally, cross-frequency coupling between theta and gamma oscillations indicated that the coupling peak frequency of theta oscillations was also shifted toward slower oscillations without affecting coupling strength or gamma power. Together, these results suggest that the dynamic changes in theta oscillations in the mouse primary somatosensory cortex represent the ongoing status of pain sensation.

Transcription factor IRF1 is responsible for IRF8-mediated IL-1ß expression in reactive microglia.

Interferon regulatory factor-8 (IRF8) plays a crucial role in the transformation of microglia to a reactive state by regulating the expression of various genes. In the present study, we show that IRF1 is required for IRF8-induced gene expression in microglia. Peripheral nerve injury induced IRF1 gene upregulation in the spinal microglia in an IRF8-dependent manner. IRF8 transduction in cultured microglia induced de novo gene expression of IRF1. Importantly, knockdown of the IRF1 gene in IRF8-transduced microglia prevented upregulation of interleukin-1ß (IL-1ß). Therefore, our findings suggest that expression of IL-1ß is dependent on IRF1 in IRF8-expressing reactive microglia.

Transcription factor IRF5 drives P2X4R+-reactive microglia gating neuropathic pain.

In response to neuronal injury or disease, microglia adopt distinct reactive phenotypes via the expression of different sets of genes. Spinal microglia expressing the purinergic P2X4 receptor (P2X4R) after peripheral nerve injury (PNI) are implicated in neuropathic pain. Here we show that interferon regulatory factor-5 (IRF5), which is induced in spinal microglia after PNI, is responsible for direct transcriptional control of P2X4R. Upon stimulation of microglia by fibronectin, IRF5 induced de novo expression of P2X4R by directly binding to the promoter region of the P2rx4 gene. Mice lacking Irf5 did not upregulate spinal P2X4R after PNI, and also exhibited substantial resistance to pain hypersensitivity. Furthermore, we found that expression of IRF5 in microglia is regulated by IRF8. Thus, an IRF8-IRF5 transcriptional axis may contribute to shifting spinal microglia toward a P2X4R-expressing reactive state after PNI. These results may provide a new target for treating neuropathic pain.