Activated myeloid cells and astrocytes are the predominant cell types in active multiple sclerosis (MS) lesions. Both cell types can adopt diverse functional states that play critical roles in lesion formation and resolution. In order to identify phenotypic subsets of myeloid cells and astrocytes, we profiled two active MS lesions with thirteen glial activation markers using imaging mass cytometry (IMC), a method for multiplexed labeling of histological sections. In the acutely demyelinating lesion, we found multiple distinct myeloid and astrocyte phenotypes that populated separate lesion zones. In the post-demyelinating lesion, phenotypes were less distinct and more uniformly distributed. In both lesions cell-to-cell interactions were not random, but occurred between specific glial subpopulations and lymphocytes. Finally, we demonstrated that myeloid, but not astrocyte phenotypes were activated along a lesion rim-to-center gradient, and that marker expression in glial cells at the lesion rim was driven more by cell-extrinsic factors than in cells at the center. This proof-of-concept study demonstrates that highly multiplexed tissue imaging, combined with the appropriate computational tools, is a powerful approach to study heterogeneity, spatial distribution and cellular interactions in the context of MS lesions. Identifying glial phenotypes and their interactions at different lesion stages may provide novel therapeutic targets for inhibiting acute demyelination and low-grade, chronic inflammation. Electronic supplementary material The online version of this article (10.1186/s40478-019-0779-2) contains supplementary material, which is available to authorized users.

There is a gap in understanding the effect of the essential ω-3 and ω-6 long-chain polyunsaturated fatty acids (LCPUFA) on Phase I retinopathy of prematurity (ROP), which precipitates proliferative ROP. Postnatal hyperglycemia contributes to Phase I ROP by delaying retinal vascularization. In mouse neonates with hyperglycemia-associated Phase I retinopathy, dietary ω-3 (vs. ω-6 LCPUFA) supplementation promoted retinal vessel development. However, ω-6 (vs. ω-3 LCPUFA) was also developmentally essential, promoting neuronal growth and metabolism as suggested by a strong metabolic shift in almost all types of retinal neuronal and glial cells identified with single-cell transcriptomics. Loss of adiponectin (APN) in mice (mimicking the low APN levels in Phase I ROP) decreased LCPUFA levels (including ω-3 and ω-6) in retinas under normoglycemic and hyperglycemic conditions. ω-3 (vs. ω-6) LCPUFA activated the APN pathway by increasing the circulating APN levels and inducing expression of the retinal APN receptor. Our findings suggested that both ω-3 and ω-6 LCPUFA are crucial in protecting against retinal neurovascular dysfunction in a Phase I ROP model; adequate ω-6 LCPUFA levels must be maintained in addition to ω-3 supplementation to prevent retinopathy. Activation of the APN pathway may further enhance the ω-3 and ω-6 LCPUFA’s protection against ROP.

Photoreceptor degeneration caused by genetic defects leads to retinitis pigmentosa, a rare disease typically diagnosed in adolescents and young adults. In most cases, rod loss occurs first, followed by cone loss as well as altered function in cells connected to photoreceptors directly or indirectly. There remains a gap in our understanding of retinal cellular responses to photoreceptor abnormalities. Here, we utilized single-cell transcriptomics to investigate cellular responses in each major retinal cell type in retinitis pigmentosa model (P23H) mice vs. wild-type littermate mice. We found a significant decrease in the expression of genes associated with phototransduction, the inner/outer segment, photoreceptor cell cilium, and photoreceptor development in both rod and cone clusters, in line with the structural changes seen with immunohistochemistry. Accompanying this loss was a significant decrease in the expression of genes involved in metabolic pathways and energy production in both rods and cones. We found that in the Müller glia/astrocyte cluster, there was a significant increase in gene expression in pathways involving photoreceptor maintenance, while concomitant decreases were observed in rods and cones. Additionally, the expression of genes involved in mitochondrial localization and transport was increased in the Müller glia/astrocyte cluster. The Müller glial compensatory increase in the expression of genes downregulated in photoreceptors suggests that Müller glia adapt their transcriptome to support photoreceptors and could be thought of as general therapeutic targets to protect against retinal degeneration.