Journal: Acta Neuropathologica
Article Title: Retinal pathological features and proteome signatures of Alzheimer’s disease
doi: 10.1007/s00401-023-02548-2
Figure Lengend Snippet: Spatiotemporal distribution of Aβ 42 burden in retinas of MCI and AD patients and relations to brain pathology and cognition. a Illustration depicts analyzed retinal cross-sections in predefined geometrical regions including superior- and inferior temporal (ST/IT) strips (orange) extending from the optic disc (OD) to the ora serrata and separated into subregions: central (C), mid-periphery (M) and far periphery (F). Schematic flow-diagram describes human donor eyes and brains allocated for histological and protein analyses (N = subjects). b Fluorescence micrographs of retinal cross-sections from MCI and AD patients compared to normal cognition (NC) controls. Tissues were immunolabeled for GFAP + -macroglia (green), IBA1 + -microglia (red), 12F4 + -Aβ 42 (white), and DAPI + -nuclei (blue; dashed lines indicate margins of analyzed layers between the inner and outer limiting membranes–ILM/OLM). Scale bar: 50 µm. Right micrographs are from the same individuals immunolabeled with 12F4 + -Aβ 42 using peroxidase-based 3,3′diaminobenzidine (DAB) and hematoxylin counterstaining. Scale bar: 20 µm. c Violin plots display quantitative-IHC analysis of retinal (r)Aβ 42 -immunoreactive area in age- and sex-matched patients with premortem clinical diagnoses of NC ( n = 17), MCI ( n = 10), or AD ( n = 18), and paired-brain (b)Aβ-plaque severity scores in NC ( n = 6), MCI ( n = 10), and AD ( n = 17) patients. Red circle represents an ADAD patient with an A260V mutation in presenilin-1 ( PSEN1 ). d Retinal Aβ 1–42 levels determined by ELISA are shown in an additional cohort of NC and AD patients ( n = 14; ADAD patient with PSEN1- A431E mutation, red circle). e TEM-micrographs from AD patients’ retina: Left, 12F4 + -immunogold Aβ 42 -positive black puncta signals at high-magnification (red arrow) in the ILM/innermost layers. Scale bar: 200 nm. Middle: 3D-reconstruction of vertical/en face TEM images show rAβ 42 plaque ultrastructure with fibril arms emanating from its dense core and Aβ-containing deposits (red arrowheads). Scale bar: 1 μm. Right, Aβ 42 plaque (black arrow) and deposits within Müller cell (MC) endfeet (red arrows). Scale bar: 0.2 µm. f Pie charts display Aβ 42 distribution across the inner retina (IR), outer retina (OR), and C, M, and F subregions: raw data and normalized per retinal thickness (density); higher burden in darker red. g Violin plot displays rAβ 42 density for C, M, and F subregions. h Definition of inner retina (IR) and outer retina (OR) in a cross-section. Scale bar: 10 μm. i Aβ 42 burden in IR vs. OR; percentages indicate rAβ 42 area in IR of total area. Statistics: red or blue asterisks mark significance relative to NC or MCI, respectively. P d –diagnostic groups; P r –C, M, vs. F subregions; P L –IR vs. OR layers; P i –interactions. j Scatterplot presents correlations between rAβ 42 area and Aβ plaques in total brain (gray) or EC (orange). k–l Mid-sagittal brain illustration and heatmap show color-grading magnitude of Pearson’s correlation coefficient ( r ) values with multivariable Holm-Bonferroni adjusted P- values (asterisks) between rAβ 42 burden and brain pathology: Aβ-(P)laques, neuropil threads (NT), and neurofibrillary tangles (NFT) in the hippocampus (Hipp), superior (S.) frontal (F. Ctx) and temporal (temp, T. Ctx) gyrus, S. parietal lobule (P. Ctx), entorhinal (EC), primary visual (PV), and visual association (VA) cortices. m Pearson’s correlation between rAβ 42 burden and BRAAK stage. n Subjects were stratified based on high(H) or low(L) brain ATN-histopathology severity and plotted based on rAβ 42 burden; extrapolated dotted-gray line marks rAβ 42 level separating ATN H from ATN L individuals. o Pearson’s correlations between rAβ 42 area or bAβ burden and the Mini-Mental State Examination (MMSE)-cognitive scores. Data points are presented with group means ± SEMs. Filled and empty circles represent women and men, respectively. Median and lower and upper quartiles are indicated on each violin plot. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, by one-way or two-way ANOVA and Tukey’s post hoc multiple comparison test, or by two-tailed paired (parenthesis) or unpaired Student’s t test
Article Snippet: After determination of protein concentrations (Thermo Fisher Scientific), the amount of retinal Aβ 1–42 was determined using an anti-human Aβ 1–42 end-specific sandwich ELISA kit (Thermo Fisher, KHB3441).
Techniques: Fluorescence, Immunolabeling, Mutagenesis, Enzyme-linked Immunosorbent Assay, Diagnostic Assay, Histopathology, Comparison, Two Tailed Test
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