Abstract
Purpose :
Retinal detachment (RD) occurs when photoreceptors (PR) are physically separated from the underlying retinal pigment epithelium, compromising PR cell survival. RD triggers an innate immune response, with microglia/macrophage activation and migration toward stressed PR. TLR2 and TLR4 are pattern recognition receptors that when activated can trigger the activation and migratory phenotype of microglia/macrophages. This study examined if TLR2 and TLR4 contribute to the response of microglia/macrophages following RD.
Methods :
C57BL/6 (B6) mice received sub-retinal injections of 1% hylaluronic acid to cause RD. Retinal samples were collected at 3-day and 7-day post-retinal detachment (dprd) and membrane proteins were enriched for the detection of TLR2 and TLR4 by Western blotting. Microglial/macrophage migration was visualized using immunofluorescence with antibodies to CD68, Iba1, and P2yr12 in 30-μm cross sections and retinal flat mounts. TUNEL was used to detect PR cell apoptosis at 3 dprd and 7 dprd. The functions of TLR2 and TLR4 were assessed using mice with germline deletion (knockout, KO) of TLR2 or TLR4.
Results :
RD resulted in 1.3-fold increased expression of TLR2 protein at 3 dprd, with a 2.6-fold elevation at 7 dprd. In contrast, TLR4 protein was not significantly increased. PR cell apoptosis peaked at 3 dprd. TLR2 KO or TLR4 KO had no affect on the number of apoptotic cells at either 3 dprd or 7 dprd. RD triggered the activation of microglia and increased microglia numbers in the inner and outer plexiform layers. In addition, activated microglia/macrophages migrated to the stressed and dying PR region. Surprisingly, compared to B6 control mice, the number of Iba1+ and CD68+ cells recruited to the inner and outer segments of PRs was diminished in both TLR2 and TLR4 KO mice at 3 dprd. Interestingly, TLR4 KO mice exhibited more Iba1+ cells at the inner and outer segments of PRs at 7 dprd.
Conclusions :
RD triggers the activation of microglia and their migration toward stressed PR. TLR2 KO or TLR4 KO affects the recruitment of microglia/macrophages to the inner and outer segments of PRs, but not the apoptosis of PR cells after RD. Further studies are needed to validate the effects of TLR2 and TLR4 on the retinal immune response and the long term survival of PR cells.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.