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Matthew Zabel, Lian Zhao, Robert N Fariss, Wai T Wong; Mechanisms Underlying Microglial Phagocytosis of Photoreceptors During Photoreceptor Degeneration in the rd10 Mouse. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4385.
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© ARVO (1962-2015); The Authors (2016-present)
Microglia, the resident immune cells of the retina, have been implicated in the progression of photoreceptor degeneration in human and mouse models of retinal disease. However, the molecular mechanisms guiding the activity of microglia engaged in photoreceptor phagocytosis have not been well characterized. Mice with the rd10 mutation undergo a progressive photoreceptor degeneration beginning around postnatal day 16. These mice were used to investigate the molecular cues guiding recruitment and phagocytic activity of retinal microglia.
Eyes from rd10 homozygous mice were collected at P18 - P30, spanning the period of rod degeneration. Vibratome sections were prepared and labeled with antibodies to Iba1 and/or CD11b to identify microglia infiltrating the outer nuclear layer (ONL). The spatiotemporal expression patterns of phosphatidylserine (PS), a phagocytic “eat-me” ligand, milk fat globule-EGF 8 (MFG-E8) a phagocytic bridging molecule, and CD68, a phagocytic marker, were also assessed by confocal microscopy.
At P18, prior to microglial infiltration into the ONL, PRs were negative for the “eat-me” signal, PS. At P23, multiple PRs in the ONL developed PS positivity transiently in their somata cell membrane. This signal diminishes by P30. The transient increase in PS labeling corresponds to the peak and decline in rod phagocytosis by infiltrating microglia. At P18, just prior to microglial infiltration, positivity for MFG-E8 emerged within and around inner retinal microglia. At P22, following microglial infiltration, positivity for MFG-E8 developed around PS+ PR nuclei and ONL microglia, consistent with the secretion of this extracellular bridging molecule from microglia onto target PRs. Correspondingly, ramified inner retinal microglia developed low levels of punctate positivity for CD68 at P18. As microglia infiltrated into the ONL at P23, CD68 expression increased in amoeboid microglia, particularly around phagosomes containing phagocytosed PRs.
Primary phagocytosis by infiltrating microglia appears to contribute significantly to rod degeneration in the rd10 mouse. The spatiotemporal patterns of expression of PS, MFG-E8, and CD68 in PR/microglia provide a molecular basis for this process and present potential targets for intervention in the treatment of inherited retinopathies through targeted inhibition of microglial phagocytosis.
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