Journal: Molecular Neurodegeneration

Article Title: Astrocytic APOE4 removal confers cerebrovascular protection despite increased cerebral amyloid angiopathy

doi: 10.1186/s13024-023-00610-x

Figure Lengend Snippet: Depletion of APOE in astrocytes diminishes certain disease-associated glial signatures. a – e , GFAP + astrocytic staining ( a ) and quantification of area coverage ( b ) in the cortex of 10-month-old 5X+AL- or 5X+AL+ mice. Scalebar: 300 µm. Representative images ( c ) and quantification of the number of astrocytes surrounding CAA ( d ) or plaques ( e ). Scalebar: 20 µm. Each point represents the number of astrocytes surrounding a single CAA or plaque normalized to that CAA or plaque area from n = 8–10 mice per group. f–j , IBA1 + microglial coverage ( f ) and quantification of area coverage ( g ) in the cortex overlying the hippocampus. Scalebar: 300 µm. Per plaque analysis of number of microglia clustering CAA ( h , i ) or plaques ( j ). Scalebar: 20 µm. Each point represents the number of microglia surrounding a single CAA or plaque normalized to that CAA or plaque area from n = 8–10 mice per group. k, l , Relative mRNA expression of homeostatic and disease-associated astrocytic ( k ) and microglial genes ( l ). Data imputed for Clec7a mRNA (column 10) but not used in statistical analysis. Each column represents an individual mouse. Data expressed as mean ± SD, student’s t -test (two-sided) performed for all statistical analyses except ( d ), ( e ), ( k – S100β ), and ( l – Cst7 , Cst3 , Itgax , Spp1 ) where Welch’s t -test was performed. ∆ = males, ○ = females. * P < 0.05, ** P < 0.01. No other statistical comparisons are significant unless indicated

Article Snippet: Primary antibodies include: biotinylated antibody HJ3.4 for human Aβ 1–13 (produced in-house, 2 µg/mL), rabbit Aβ 40 (Thermo Fisher, 44,136, 1:500), rabbit Aβ 42 (Thermo Fisher, 700,254, 1:500), rabbit APOE antibody (Cell signaling, #13,366, 1:500), biotinylated GFAP for astrocytes (Sigma-Aldrich, MAB3402B, 1:1000), rabbit Iba1 (Wako, 019–19,741, 1:5000), rabbit fibrinogen (Abcam, ab34269, 1:1000), and rat LAMP1 (Developmental Studies Hybridoma Bank, #1D4B, 1:400).

Techniques: Staining, Expressing

Journal: Nature neuroscience

Article Title: Peripheral apoE4 enhances Alzheimer’s pathology and impairs cognition by compromising cerebrovascular function

doi: 10.1038/s41593-022-01127-0

Figure Lengend Snippet: a, Schematic illustration of the experimental paradigm. 5xFAD amyloid mice at 1-1.5 month of age were transduced with AAV-Alb-apoE3 or AAV-Alb-apoE4 virus via intravenous injection. b, c, The amyloid deposition in the brain of experimental mice at 4 months of age was examined by immunostaining for Aβ. Scale bar, 1 mm. The amyloid plaque burdens in the cortex and hippocampus (E3, n=14; E4, n=16) were quantified. **, P = 0.007. d, e, ApoE levels in the plasma and brain of experimental mice (E3, n=15; E4, n=16) were measured by ELISAs. **, P = 0.0098. f, TBSX-soluble and -insoluble (guanidine; GDN) Aβ40 and Aβ42 levels in the cortex of 4-month-old 5xFAD mice transduced with AAV-Alb-apoE3 (n=15) or AAV-Alb-apoE4 (n=16) were examined by specific Aβ ELISA. TBSX-Aβ40 (*, P = 0.012); TBSX-Aβ42 (*, P = 0.031). GDN-Aβ40 (*, P = 0.049); GDN-Aβ42 (*, P = 0.033). g, Thio S-positive plaques in the cortex of experimental mice were shown and quantified. Scale bar, 100 μm. h, i, Brain sections from experimental mice (n=12/genotype) were immunostained with GFAP or Iba1 antibody. Scale bar, 100 μm. The immunoreactivity of GFAP and Iba1 in cortex and hippocampus were quantified. j, Representative images of plaque-associated microglia in mice expressing apoE3 or apoE4 in the liver are shown. Scale bar, 50 μm. The number of Iba1-positive microglia (green) surrounding Aβ plaque (red) between 50-300 μm2 plaque sizes were quantified. Each dot represents the average value from an individual mouse (E3, n=10; E4, n=11). *, P = 0.049. k, Co-immunofluorescence staining of LAMP1 (green) and Aβ plaques (red) was used to examine plaque-associated neuritic dystrophy. Scale bar, 50 μm. The LAMP1 immunoreactivity was quantified. *, P = 0.027. l, LAMP1 immunoreactivity was positively correlated with Thio S-positive fibrillar plaques. c-l, Data represent mean ± s.e.m., two-tailed Student's t-test.

Article Snippet: The following antibodies were used in this study: anti-GFAP (Millipore; MAB360), anti-apoE (WUE4; NB110-60531) anti-PSD-95 (Cell Signaling; #3450), anti-synaptophysin (Millipore; MAB5258), and anti-β-actin (Sigma; A2228) antibodies.

Techniques: Expressing, Transduction, Injection, Immunostaining, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Two Tailed Test

Journal: Nature neuroscience

Article Title: Peripheral apoE4 enhances Alzheimer’s pathology and impairs cognition by compromising cerebrovascular function

doi: 10.1038/s41593-022-01127-0

Figure Lengend Snippet: a, Brain sections from 9-month-old APP/PS1 mice expressing apoE3 (Cre− , n=21; Cre+, n=22) or apoE4 (Cre−, n=19; Cre+, n=15) in the liver were immunostained with a pan-Aβ antibody. The Aβ plaque burden in the hippocampus was quantified. Black circle: male; Grey circle: female. E3: *, P=0.041; E4: **, P=0.009, two-tailed Student's t-test. b, Representative images of Aβ staining in the cortex of APP/Alb/iE3 or APP/Alb/iE4 mice (murine Apoe−/− background) are shown. Scale bar, 200 μm. Images from APP/iE mice (murine Apoe+/+ background) were included as visual representation. Note that only diffused plaques were observed in APP/Alb/iE3 or iE4 mice due to the absence of murine apoE in the brain. c, d, TBS- and TBSX-soluble Aβ40 and Aβ42 levels in the cortex of 9-month-old APP/iE3/Cre mice (Cre−, n=18; Cre+, n=19) or APP/iE4/Cre mice (Cre−, n=22; Cre+, n=25) were examined by specific Aβ ELISA. c, TBS-E3_Aβ40: *, P=0.036; Aβ42: *, P=0.045. TBS-E4_Aβ40: **, P=0.008; Aβ42: **, P=0.001. d, TBSX-E3_Aβ40: *, P=0.024; Aβ42: *, P=0.030. TBSX-E4_Aβ40: **, P=0.002; Aβ42: **, P=0.0003. e, f, Brain sections from APP/PS1 mice expressing apoE3 (Cre−, n=8; Cre+, n=8) or apoE4 (Cre−, n=6; Cre+, n=6) in the liver (murine Apoe−/− background) were labeled for fibrillar Aβ using Thioflavin S (Thio S). Scale bar (upper panels), 1 mm; Scale bar (bottom panels), 100 μm. Images from APP/iE mice (murine Apoe+/+ background) were included for comparison. The amount of fibrillar plaques in the APP/Alb/iE3 or APP/Alb/iE4 mice was minimal due to the absence of murine apoE in the brain. The percentage of area covered by Thio S-positive plaques in the cortex and hippocampus of experimental mice was quantified. g, h, Brain sections from 9-month-old APP/PS1 mice expressing apoE3 (Cre−, n=27; Cre+, n=28) or apoE4 (Cre−, n=19; Cre+, n=19) in the liver were immunostained with an Iba1 antibody. Scale bar, 100 μm. The immunoreactivity of Iba1 in cortex and hippocampus were quantified. Data expressed as mean ± s.e.m. Cortex: *, P = 0.011; Hippo: *, P = 0.030. N.S., not significant, two-tailed Student's t-test.

Article Snippet: The following antibodies were used in this study: anti-GFAP (Millipore; MAB360), anti-apoE (WUE4; NB110-60531) anti-PSD-95 (Cell Signaling; #3450), anti-synaptophysin (Millipore; MAB5258), and anti-β-actin (Sigma; A2228) antibodies.

Techniques: Expressing, Two Tailed Test, Staining, Enzyme-linked Immunosorbent Assay, Labeling

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Protective Effect of Buyang Huanwu Decoction on Neurovascular Unit in Alzheimer’s Disease Cell Model via Inflammation and RAGE/LRP1 Pathway

doi: 10.12659/MSM.917020

Figure Lengend Snippet: Protein levels of RAGE, LRP1, NFκB-p65, and ApoE in 6 groups. * p<0.05, compared with the control group; & P <0.05, compared with the model group. A – control group; B – model group; C – Donepezil group; D – BYHWDL group; E – BYHWDM group; F – BYHWDH group.

Article Snippet: The primary antibodies used in this study were: rabbit anti-RAGE antibody (CST, Danvers, MA, USA), rabbit anti-LRP1 antibody (CST, Danvers, MA, USA), rabbit anti-ICAM-1 antibody (CST, Danvers, MA, USA), rabbit anti-VCAM-1 antibody (CST, Danvers, MA, USA), rabbit anti-ApoJ antibody (CST, Danvers, MA, USA), rabbit anti-ApoE antibody (CST, Danvers, MA, USA), and rabbit anti-NFκB-p65 antibody (CST, Danvers, MA, USA), with β-actin (CST, Danvers, MA, USA) as the internal reference.

Techniques:

Journal: Antioxidants

Article Title: ApoE3 vs. ApoE4 Astrocytes: A Detailed Analysis Provides New Insights into Differences in Cholesterol Homeostasis

doi: 10.3390/antiox11112168

Figure Lengend Snippet: Analysis of the levels of key proteins involved in lipid homeostasis. The protein levels of the ATP binding cassette subfamily A member 1 (ABCA1), ApoE receptor 2 (ApoER2), low-density lipoprotein receptor (LDLR), LDLR-related protein 1 (LRP1), and peroxisome proliferator activated receptor γ (PPARγ) were determined by Western blotting in ApoE3 and ApoE4 immortalized mouse astrocytes. Data were normalized to the corresponding β-actin levels and are shown as percentage of average ApoE3 values. Data are expressed as mean values ± SD of three different experiments ( n = 9, Student’s t -test). **** p < 0.0001 and *** p < 0.001 vs. ApoE3 astrocytes.

Article Snippet: The following primary antibodies (1:1000 dilution) were used: anti-ApoE (D7I9N, Cell Signaling Technology, Danvers, MA, USA), anti-PPARγ (C26H12, Cell Signaling Technology), anti-ABCA1 (MA5-16026, Invitrogen), anti-ApoER2 (LRP8, PA5-109269, Invitrogen), anti-LDLR (MA5-32075, Invitrogen), anti-LRP1 (37-7600, Invitrogen), and anti-β-actin (sc-47778, Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Binding Assay, Western Blot