Modifications of the endosomal compartment in peripheral blood mononuclear cells and fibroblasts from Alzheimer's disease patients

F Corlier, I Rivals, J Lagarde, L Hamelin, H Corne, L Dauphinot, K Ando, J-C Cossec, G Fontaine, G Dorothée, C Malaplate-Armand, J-L Olivier, B Dubois, M Bottlaender, C Duyckaerts, M Sarazin, M-C Potier, Clinical ImaBio3 Team, Amer Alnajjar-Carpentier, Michel Logak, Sara Leder, Dominique Marchal, Hélène Pitti-Ferandi, Hélene Brugeilles, Brigitte Roualdes, Agnes Michon, F Corlier, I Rivals, J Lagarde, L Hamelin, H Corne, L Dauphinot, K Ando, J-C Cossec, G Fontaine, G Dorothée, C Malaplate-Armand, J-L Olivier, B Dubois, M Bottlaender, C Duyckaerts, M Sarazin, M-C Potier, Clinical ImaBio3 Team, Amer Alnajjar-Carpentier, Michel Logak, Sara Leder, Dominique Marchal, Hélène Pitti-Ferandi, Hélene Brugeilles, Brigitte Roualdes, Agnes Michon

Abstract

Identification of blood-based biomarkers of Alzheimer's disease (AD) remains a challenge. Neuropathological studies have identified enlarged endosomes in post-mortem brains as the earliest cellular change associated to AD. Here the presence of enlarged endosomes was investigated in peripheral blood mononuclear cells from 48 biologically defined AD patients (25 with mild cognitive impairment and 23 with dementia (AD-D)), and 23 age-matched healthy controls using immunocytochemistry and confocal microscopy. The volume and number of endosomes were not significantly different between AD and controls. However, the percentage of cells containing enlarged endosomes was significantly higher in the AD-D group as compared with controls. Furthermore, endosomal volumes significantly correlated to [C(11)]PiB cortical index measured by positron emission tomography in the AD group, independently of the APOE genotype, but not to the levels of amyloid-beta, tau and phosphorylated tau measured in the cerebrospinal fluid. Importantly, we confirmed the presence of enlarged endosomes in fibroblasts from six unrelated AD-D patients as compared with five cognitively normal controls. This study is the first, to our knowledge, to report morphological alterations of the endosomal compartment in peripheral cells from AD patients correlated to amyloid load that will now be evaluated as a possible biomarker.

Figures

Figure 1
Figure 1
Endosomal abnormalities are present in peripheral mononuclear blood cells (PBMCs) from individuals with AD-MCI and AD-D. (a–c) Immunofluorescence confocal images showing EEA1-labelled early endosomes in representative PBMCs from cognitively healthy subjects (a), AD-MCI (b) and AD-D cases (c). Scale bar, 2 μm. Mean endosome volumes (MEVs) (d) and mean endosome numbers (MENs) (e) per PBMC are not significantly different between AD-D (455 cells), AD-MCI (497 cells) and controls (CT, 459 cells). Pgroup=0.63 for MEN and 0.61 for MEV according to analysis of variance. AD-D, Alzheimer's disease with dementia; AD-MCI, Alzheimer's disease with mild congnitive impairment.
Figure 2
Figure 2
Percentage of PBMCs containing enlarged endosomes is significantly higher in AD-D as compared with control (CT) individuals. (a) Classification of cells in three subclasses according to their MEV (small/medium/large) and (b) mean percentages and standard deviations on 1000 bootstrap samples drawn at the individual level. AD-D, Alzheimer's disease with dementia; AD-MCI, Alzheimer's disease with mild congnitive impairment; MEV, mean endosome volume; PBMC, peripheral blood mononuclear cell.
Figure 3
Figure 3
Correlation between mean endosomal volume in AD patients (AD-MCI and AD-D) and their PiB retention. Black dots correspond to non-APOE ɛ4 carriers and red dots to APOE ɛ4 carriers; r=0.682, P=0.0001 and r=0.415, P=0.008 for all AD patients (black and red dots) and APOE ɛ4 carriers only (red dots) after adjusting for age and APOE ɛ4. AD-D, Alzheimer's disease with dementia; AD-MCI, Alzheimer's disease with mild congnitive impairment; PiB, Pittsburgh Compound B.
Figure 4
Figure 4
Endosomal abnormalities are present in fibroblasts from individuals with AD-D. Immunofluorescence confocal images showing EEA1-labelled early endosomes (green) in representative fibroblasts from cognitively healthy subjects (a) and AD-D cases (b). Nuclei are labelled in blue. Scale bar, 2 μm. (c) Mean endosome volumes per fibroblast are not significantly different between control (CT) individuals (0.147±0.016 μm3) and AD-D patients (0.170±0.034 μm3; Wilcoxon P=0.18). (d) Comparison of the mean endosomal densities per fibroblast (that is, portion of the cell volume occupied by endosomes) showed no significant difference between control individuals (0.0249±0.005) and AD-D patients (0.0279±0.006; Wilcoxon P=0.46). AD-D, Alzheimer's disease with dementia; EEA1, early endosome antigen 1.
Figure 5
Figure 5
Percentage of fibroblasts containing enlarged endosomes is significantly higher in AD-D as compared with control (CT) individuals. Classification of cells in three subclasses according to their MEV (small/medium/large) showed significantly elevated percentage of cells with large MEV in AD-D patients and reduced percentage of cells with small MEV (χ2 test, P=0.0094). AD-D, Alzheimer's disease with dementia; MEV, mean endosome volume.

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