Abstract
Purpose :
Microglia and macrophages play roles in AMD-relevant processes of immune surveillance, neurodegeneration, and neovascularization. We analyzed the distribution of cells expressing Iba1 and TMEM119, specific markers for microglia/ macrophage and brain microglia, respectively, in human donor eyes at different AMD stages.
Methods :
Twelve μm-thick sections of maculae from 20 eyes of white donors were immuno-stained with Iba1 (Wako 019-19741) and TMEM119 (Abcam ab185333), using red-substrate enzymatic detection and scanned with a 40X objective. Immunoreactivity presence was assessed by one observer as 0 (none), 1 (a few cells), 2 (some cells) and 3 (all over or all cells). Positive controls were resident microglia cells in the inner retina, choroidal macrophages, and Kupffer cells in surgically excised human liver. Negative control experiments omitted primary antibody. RPE cells were phenotyped by published methods (PMID 25813989, 26024109).
Results :
In all eyes (4 normal,16 AMD; 7 early-or-intermediate (e-i)AMD, 6 geographic atrophy (GA) AMD, 3 neovascular (nv)AMD), elongated and dendritic cells in inner retina and in choroid were Iba1+, and only in these locations, in normal eyes. Iba1+ cells appeared in outer retina in e-iAMD eyes. In advanced AMD, Iba1+ cells were amoeboid and found in GA areas and in fibrotic scar (nvAMD), where photoreceptors (PR) and RPE layer are absent (both stages) or scarred (nvAMD). Fewer Iba1+ cells were found in inner retina and choroid compared normal and e-iAMD eyes. A few RPE cells were stained (sloughed, intraretinal and subducted phenotypes); most were not.
Like Iba1, TMEM119+ cells were found in inner retina and choroid in normal eyes and additionally in outer retina of AMD eyes. Unlike Iba1, TMEM119 signal was found in PR inner segment ellipsoids, ganglion cells, and Müller cells, as seen in mouse (PMID 31853425).
Conclusions :
Iba1+ and TMEM119+ cells in outer retina of advanced AMD eyes are most likely activated microglia; we cannot exclude choroidal cells. Some RPE phenotypes that express immune markers in AMD (Cao ARVO 2020) were Iba1+ and TMEM119+ but most RPE cells were unlabeled. TMEM119 signal in PR ellipsoids possibly localizes to mitochondria. Results will assist interpretation of planned single-cell RNA sequencing studies.
This is a 2021 ARVO Annual Meeting abstract.