Abstract
Multiple sclerosis (MS) is a neuroinflammatory multifocal disorder. Optic neuritis is common in MS and leads to visual disability. No current treatments repair this damage. Discerning gene expression changes within specific cell types in optic nerve (ON) may suggest new treatment targets for visual disability in MS. Astrocytes are pivotal regulators of neuroinflammation, playing either detrimental or beneficial roles. Here, we used RiboTag technology to characterize the astrocyte-specific transcriptome in ON in the experimental autoimmune encephalomyelitis (EAE) model of MS. RNA sequencing analysis showed the Complement Cascade and Cholesterol Biosynthesis Pathways as the most enriched and de-enriched pathways, respectively, in ON astrocytes in EAE. Expression of complement component 3 (C3) was confirmed to be increased in ON astrocytes at the protein level during EAE. A bigger increase in C3 expressing ON astrocytes was found in EAE females versus healthy females, as compared to that in EAE males versus healthy males. Also, there was worse retinal ganglion cell (RGC) and axonal loss in EAE females. Regression analyses showed a negative correlation between C3 expressing astrocytes and RGC density. This cell-specific and sex-specific investigation of the optic nerve provides targets for the development of therapeutic strategies tailored for optic neuritis in MS.
Highlights
The central nervous system (CNS) contains a variety of different cell types, whose composition and gene expression are altered during disease
Neuroinflammation and demyelination were assessed in EAE and nature.com/scientificreports as controls (NL) mice by double immunostaining of optic nerve (ON) sagittal sections for CD45, a marker for microglia and blood-derived inflammatory cells, and proteolipid protein (PLP), a marker for myelin (Fig. 1gi–iv)
We double immune-labelled ON sagittal sections from EAE and NL mice at the defined time points for the astrocyte reactivity marker, glial fibrillary acid protein (GFAP), alongside Lipocalin 2 (LCN2), an autocrine mediator of reactive gliosis thought to contribute to disease progression during EAE10,19,20
Summary
The central nervous system (CNS) contains a variety of different cell types, whose composition and gene expression are altered during disease. Using a cell- and region-specific approach[2,3,4], we showed regional differences in astrocyte gene expression among CNS regions in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS1. Www.nature.com/scientificreports accumulates following recurrent attacks of optic neuritis. This entails inflammation and demyelination of the ON, axonal damage and loss, and retinal ganglion cell (RGC) death. We characterized the astrocyte-specific transcriptome in ON in EAE using RiboTag technology Such technology uses the Cre-LoxP recombination to generate mice expressing hemagglutinin (HA)-tagged ribosomes in specific cell types[1,2,4]. GFAP-Cre RiboTag mice enabled us to isolate astrocyte specific transcripts from ON, overcoming limitations inherent to interpretation of gene expression analysis performed on whole tissues, characterized by variability in cell composition during disease. Sex as a biological variable was examined comparing female and male mice with optic neuritis
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