Functions of retinal astrocytes and Müller cells in mammalian myopia

Abstract

Background: Changes in the retina and choroid blood vessels are regularly observed in myopia. However, if the retinal glial cells, which directly contact blood vessels, play a role in mammalian myopia is unknown. We aimed to explore the potential role and mechanism of retinal glial cells in form deprived myopia.

Methods: We adapted the mice form-deprivation myopia model by covering the right eye and left the left eye open for control, measured the ocular structure with anterior segment optical coherence tomography, evaluated changes in the morphology and distribution of retinal glial cells by fluorescence staining and western blotting; we also searched the online GEO databases to obtain relative gene lists and confirmed them in the form-deprivation myopia mouse retina at mRNA and protein level.

Results: Compared with the open eye, the ocular axial length (3.54 ± 0.006 mm v.s. 3.48 ± 0.004 mm, p = 0.027) and vitreous chamber depth (3.07 ± 0.005 mm v.s. 2.98 ± 0.006 mm, p = 0.007) in the covered eye became longer. Both glial fibrillary acidic protein and excitatory amino acid transporters 4 elevated. There were 12 common pathways in human myopia and anoxic astrocytes. The key proteins were also highly relevant to atropine target proteins. In mice, two common pathways were found in myopia and anoxic Müller cells. Seven main genes and four key proteins were significantly changed in the mice form-deprivation myopia retinas.

Conclusion: Retinal astrocytes and Müller cells were activated in myopia. They may response to stimuli and secretory acting factors, and might be a valid target for atropine.

Keywords: Astrocyte; Atropine; Gene set enrichment analysis; Myopia; Müller cell.

Fig. 1

Flowchart of the experimental process

Fig. 2

Mice were induced as form deprived myopia. A Diagrammatic drawing that the right eye was covered whereas the left eye was left open of every mouse. B Measurement of the ocular structure. CCT referred to central cornea thickness, ACD referred to anterior chamber depth and VCD referred to vitreous chamber depth. The axial length was the sum of CCT, ACD and VCD. Ocular axial length (C), vitreous chamber depth (D), central cornea thickness (E) and anterior chamber depth (F) were compared between the open and the covered eye. t-test, n = 14, * p < 0.05

Fig. 3

The expression of GFAP and EAAT4 elevated in FDM mice. A GFAP (green) and EAAT4 (red) staining in retinal section (a-f) and whole-mount (g-h). Relative fluorescence area of GFAP (B) and EAAT4 (D), relative mean fluorescence intensity of GFAP (C) and EAAT4 (E), relative colocalization percentage of GFAP and EAAT4 (F) were compared between the open and the covered eye. n = 12. (G-H) GFAP and EAAT4 expression determined by WB (the cropped images), n = 4–8. t-test or Mann–Whitney test, * p < 0.05, ** p < 0.01, ***p < 0.001

Fig. 4

Myopia-related and hypoxic astrocyte-related genes in human. Venn diagram (A) and chordal graph (B) as well as GSEA analysis (C) shows the distribution and expression trend of myopia and hypoxia astrocyte genes. In partial PPI analysis (D), red nodes represent HMRGs, and thicker lines between nodes indicate a stronger relationship. (E) GO pathway analysis showing common pathways in the top 12 clusters. GO analysis of the common pathways (F), and PPI analysis of the connected nodes (G) (red: atropine target proteins, blue: myopia-related proteins, green: astrocyte-related proteins) show effect of atropine on myopia and astrocytes

Fig. 5

Myopia-related and hypoxic Müller cell-related genes in mice. A miRNA clusters in FDM mice, B the common pathways of the two conditions, and C PPI analysis and GO annotation results (nodes with a red circle represent myopia-related genes)

Fig. 6

Expression of key genes as determined by qPCR (A) and WB (B and C) (the cropped images). n = 4–8. t-test or Mann–Whitney test, * p < 0.05, ** p < 0.01

Fig. 7

Schematic diagram of possible cellular and molecular mechanism in mice FDM. Astrocytes and Müller cells suffer from retina hypoxia, and work as an injury susceptor to regular the receptors or secreted proteins, thus work as a bridge to connect vessels and neurons. Finally, retinal signals transmitter to sclera and lead to the scleral remodeling and axial elongation in FDM. A possible mechanism for atropine treatment is that atropine targets on retinal neurons or glial cells and influence GABA release, which interplay with retinal glial cells, and finally protect FDM