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Updated: Jan 1, 2026
Isolation of Region-specific Microglia from One Adult Mouse Brain Hemisphere for Deep Single-cell RNA Sequencing
Published on: December 3, 2019
Identification of a Unique Subretinal Microglia Type in Retinal Degeneration Using Single Cell RNA-Seq
Chen Yu1, Daniel R Saban2,3
1Departments of Ophthalmology, Duke University School of Medicine, Durham, NC, USA.
Abstract:
As the resident macrophages of central nervous system, microglia reside in the plexiform and nerve fiber layers of the retina. In degenerative diseases, monocyte-derived macrophages can be recruited to the retina, and histopathology shows abnormal accumulation of macrophages subretinally. However, due to lack of known markers, recruited cells and resident microglia are phenotypically indistinguishable, leaving a major knowledge gap about their potentially independent roles. Here, we used single cell RNA-seq and analyzed over 10,000 immune cells of mouse retinas from normal control and light damage-induced retinal degeneration. We observed ten major macrophage clusters. Moreover, combining trajectory analysis and in situ validation allowed us to pinpoint that subretinal phagocytes are microglia-derived and express high levels of Gal3, Cd68, and Lpl but not P2ry12. Hence, we have identified novel subretinal macrophage markers indicative of their origin and phenotype, which may be useful in other degeneration models and human specimens.
Insights
Researchers identified new markers to distinguish between resident microglia and infiltrating macrophages in retinal degeneration. This finding helps understand their distinct roles in eye diseases.
Area of Science:
- Neuroscience
- Immunology
- Ophthalmology
Background:
- Microglia are the primary immune cells in the retina, while monocyte-derived macrophages can infiltrate during disease.
- Distinguishing between these cell types is crucial for understanding their roles in retinal degeneration.
- Current markers do not reliably differentiate between resident microglia and recruited macrophages.
Purpose of the Study:
- To identify novel markers for distinguishing resident microglia from recruited macrophages in the retina.
- To investigate the origin and phenotype of subretinal macrophages during retinal degeneration.
Main Methods:
- Single-cell RNA sequencing of over 10,000 immune cells from normal and light-damaged mouse retinas.
- Analysis of ten distinct macrophage clusters.
- Trajectory analysis and in situ validation to determine cell origins and markers.
Main Results:
- Identified ten major macrophage clusters in the mouse retina.
- Subretinal phagocytes were confirmed to be microglia-derived.
- Novel markers, including Gal3, Cd68, and Lpl (but not P2ry12), were identified for subretinal macrophages.
Conclusions:
- Established novel markers to differentiate microglia-derived subretinal macrophages from other retinal immune cells.
- These markers provide insights into the origin and phenotype of macrophages in retinal degeneration.
- The identified markers may be valuable for studying other degenerative conditions and human retinal tissues.
