Background & Aims

Neuropathic pain caused by nerve injuries is a debilitating health condition with a significant global impact. Neuro-immune-glia interactions are crucial in driving neuropathic pain pathophysiology. Recently, we have shown that dendritic cells accumulate in the dorsal root/spinal cord meninges after peripheral nerve injury and mediate the development of neuropathic pain. In this study, we aimed to in-deep explore changes occurring at dorsal root/spinal cord meningeal immune cells tissues after peripheral nerve injury and how these cells contribute for the development of neuropathic pain.

Methods

The study induced neuropathic pain in male and female mice using the spared nerve injury (SNI) model. Pain behaviours were assessed by von Frey filament Lumbar (L3-L6) dorsal root/spinal cord meningeal tissues (meninges) were cirurgicaly harvested and used for whole-mount immunofluorescence, real-time PCR, scanning eletron-microscopy, single cell (sc) RNAseq, ELISA, Flow cytometry.

Results

Immunofluorescence and flow cytometry analysis revealed that CD45+ cells accumulate in the meninges in a time-dependent manner (3-28 days). The peak of leukocytes accumulation was observed 10 days after SNI. We were able to show that part of the cells that accumulate in the meninges after SNI originate and migrate from vertebrae bone marrow directly to meningeal tissue by using ossified channels in the vertebrae. ScRNAseq analyses of CD45+ cells revealed the presence of 15 different clusters of cells subtypes, neutrophils, monocytes, border macrophages, B and T/NK cells, ILC2s and dendritic cells. We combined scRNAseq data sets from different studies in the literature which analyzed the transcriptomes after SNI in the sciatic nerve, DRGs and spinal cord, and compared with our data set from meningeal cells. Among different immune mediators, Csf2 was one of the main expressed in meningeal cells. The expression of Csf2 in meningeal cells was confirmed by using Csf2 reporter animals. ScRNAseq data revealed that Csf2 expression is found in ILC2 cells. SNI-induced mechanical allodynia was reduced in Csf2 and Csf2Ra knockout mice or by killing Csf2-producing cells.

Conclusions

Our results unravel a novel role for meningeal immune cells, specialy ILC2s in the pathophysiology of neuropathic pain and also pointed out GM-CSF as a possible candidate for neuropathic pain control.

References

1. .Hamilton, J. A. . J Exp Med. 6, 217 (2020) doi: 10.1084/jem.20190945
2.Komuczki, J. et al. Immunity 50, 1289-1304 (2019) doi: 10.1016/j.immuni.2019.04.006
3.Decosterd, I. & Woolf, C. J. Pain 87, 149–158 (2000) doi: 10.1016/S0304-3959(00)00276-1
4.Guimarães, R. M. et al. J. Leukoc. Biol. 106, 541–551 (2019) doi: 10.1002/JLB.HI1118-458R
5.Fonseca, M. M. et al. Front. Immunol. 10, 1–14 (2020) doi: 10.3389/fimmu.2019.03059
6. Pulous, F. E. et al. Nature Neuroscience 25, 567–576 (2022) doi: 10.1038/s41593-022-01060-2
7. Cugurra, A. et al. Science 23; 373(6553) (2021) doi: 10.1126/science.abf7844.

Presenting Author

Glauce Regina Pigatto

Poster Authors

Glauce Pigatto

Lead Author

Andreza Urba Quadros

PhD

Lead Author

William Gonçalves

Lead Author

Thiago Cunha

Ribeirao Preto Medical School thicunha

Lead Author

Topics

  • Models: Chronic Pain - Neuropathic