Peripheral immune profiling highlights a dynamic role of low-density granulocytes in myasthenia gravis

Abstract

Background

Myasthenia gravis (MG) is an autoimmune neuromuscular disease marked by dysregulation of several immune cell populations. Here we explored peripheral immune landscape, particularly the role of low-density granulocytes (LDGs).

Methods

Single-cell and bulk RNA sequencing analyzed peripheral immune cells from MG patients pre- (n = 4) and after treatment (n = 2), as well as healthy controls (n = 3). Flow cytometry was employed for validating LDG subsets, and various functional assays were conducted to assess their impact on T cell proliferation and differentiation, NET formation, and ROS production.

Results

Single-cell analysis highlighted a shift towards inflammatory Th1/Th17/Tfh subsets, an intense interferon-mediated immune response, and an expansion of immature myeloid subsets in MG. Flow cytometry showed increased LDGs correlated with disease severity. Unlike myeloid-derived suppressor cells, MG LDGs do not restrict T cell proliferation but induce a pro-inflammatory Th1/Th17 response. They also display enhanced spontaneous neutrophil extracellular traps (NETs) formation and basal reactive oxygen species (ROS) production. LDGs decreased after intravenous immunoglobulin and increased after prolonged immunotherapy in minimal manifestation status (MM), with reduced pro-inflammatory activity. Bulk RNA sequencing revealed significant transcriptional differences in LDGs, especially in cell cycle and granule protein genes.

Conclusion

Peripheral immune profiling sheds light on the intricate role of LDGs in MG. These cells, as a distinct subtype of neutrophils with a proinflammatory phenotype, are notable increased in MG, exacerbating chronic inflammation. Furthermore, immunotherapy expanded LDGs but reduced their proinflammatory capacities. The complex interplay of LDGs in MG underscores their potential as biomarkers and therapeutic targets.