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Chen Yu, Rose Mathew, Daniel R Saban; Inner Plexiform Layer-Specific Microglia Protect the Retinal Pigment Epitheilium in Retinal Degeneration. Invest. Ophthalmol. Vis. Sci. 2019;60(9):985. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
We recently demonstrated that microglia which take up the subretinal space in retinal degeneration models function to protect RPE integrity from disease-associated damage. These subretinal microglia in degeneration originated from both inner and outer plexiform layers (IPL and OPL, respectively). However, as increasing evidence points to regional-specific heterogeneity of microglia, an emerging question is whether IPL and OPL pools both mediate RPE protection in retinal degeneration. We hypothesize that these individual pools may contribute to differential levels of RPE protection in retinal degeneration.
We addressed the role of IPL microglia using IL-34 deficient mice, as we previously showed these mice have an IPL-restricted loss in microglia numbers. We crossed F1 (BALB/c x C57BL/6) mice to C57BL/6 background for 9 generations to restore light damage susceptibility (RPE65L450/M) in IL-34 deficient mice. Dark-adapted mice were dilated and exposed to 10K lux light for 4 hours before being returned to normal housing for 5 days. We then examined the retina by OCT imaging, microglia by flow cytometry, and RPE morphology using confocal microscopy. Finally, to understand the mechanisms behind the heterogeneity, we analyzed ~5000 retinal microglia using single cell RNA-seq (scRNA-seq).
We found that microglia numbers of neuroretinas from both IL-34 deficient and sufficient mice were increased in response to light damage, but IL-34 deficient mice still had fewer microglia compared to the control. Likewise, confocal microscopy revealed subretinal microglial numbers were reduced in IL-34 deficient mice. Correspondingly, we observed that the area of RPE damage in IL-34 deficient mice was significantly (p < 0.01) greater relative to the control. Lastly, our scRNA-seq data uncovered two major microglia clusters with distinct gene markers suggesting heterogeneity possibly amongst IPL versus OPL pools.
Using IL-34 deficient mice, we demonstrate that IPL microglia contribute to the protection of RPE integrity in light damage model. Differential expressed genes from our scRNA data may help explain the IPL-restricted microglial defect of IL-34 deficient mice. Future work will specifically deplete or reduce OPL microglia to determine if the functional heterogeneity exists between two microglial pools.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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