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Cited by 210Alzheimer's disease research and treatmentsNeuroinflammation and Neurodegeneration MechanismsDementia and Cognitive Impairment Research

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Trem2 Deletion Reduces Late-Stage Amyloid Plaque Accumulation, Elevates the Aβ42:Aβ40 Ratio, and Exacerbates Axonal Dystrophy and Dendritic Spine Loss in the PS2APP Alzheimer's Mouse Model

Open access
Meilandt, William J., Ngu, Hai, Gogineni, Alvin, Lalehzadeh, Guita, Lee, Seung-Hye, Srinivasan, Karpagam, Imperio, Jose, Wu, Tiffany, Weber, Martin, Kruse, Agatha J., Stark, Kimberly L., Chan, Pamela, Kwong, Mandy, Modrusan, Zora, Friedman, Brad A., Elstrott, Justin, Foreman, Oded, Easton, Amy, Sheng, Morgan, Hansen, David V.·2020·The Journal of Neuroscience
DOI 10.1523/jneurosci.1871-19.2019PMC PMC7046459
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Abstract

TREM2 is an Alzheimer's disease (AD) risk gene expressed in microglia. To study the role of Trem2 in a mouse model of β-amyloidosis, we compared PS2APP transgenic mice versus PS2APP mice lacking Trem2 (PS2APP;Trem2 ko ) at ages ranging from 4 to 22 months. Microgliosis was impaired in PS2APP;Trem2 ko mice, with Trem2 -deficient microglia showing compromised expression of proliferation/Wnt-related genes and marked accumulation of ApoE. Plaque abundance was elevated in PS2APP;Trem2 ko females at 6–7 months; but by 12 or 19–22 months of age, it was notably diminished in female and male PS2APP;Trem2 ko mice, respectively. Across all ages, plaque morphology was more diffuse in PS2APP;Trem2 ko brains, and the Aβ42:Aβ40 ratio was elevated. The amount of soluble, fibrillar Aβ oligomers also increased in PS2APP;Trem2 ko hippocampi. Associated with these changes, axonal dystrophy was exacerbated from 6 to 7 months onward in PS2APP;Trem2 ko mice, notwithstanding the reduced plaque load at later ages. PS2APP;Trem2 ko mice also exhibited more dendritic spine loss around plaque and more neurofilament light chain in CSF. Thus, aggravated neuritic dystrophy is a more consistent outcome of Trem2 deficiency than amyloid plaque load, suggesting that the microglial packing of Aβ into dense plaque is an important neuroprotective activity. SIGNIFICANCE STATEMENT Genetic studies indicate that TREM2 gene mutations confer increased Alzheimer's disease (AD) risk. We studied the effects of Trem2 deletion in the PS2APP mouse AD model, in which overproduction of Aβ peptide leads to amyloid plaque formation and associated neuritic dystrophy. Interestingly, neuritic dystrophies were intensified in the brains of Trem2 -deficient mice, despite these mice displaying reduced plaque accumulation at later ages (12–22 months). Microglial clustering around plaques was impaired, plaques were more diffuse, and the Aβ42:Aβ40 ratio and amount of soluble, fibrillar Aβ oligomers were elevated in Trem2 -deficient brains. These results suggest that the Trem2-dependent compaction of Aβ into dense plaques is a protective microglial activity, limiting the exposure of neurons to toxic Aβ species.

Overview

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TL;DR

In PS2APP mice, Trem2 deletion impairs plaque-associated microgliosis and shifts amyloid pathology toward a more diffuse, Aβ42-enriched form, with reduced late-stage plaque load but worse axonal dystrophy and spine loss. The authors conclude that Trem2-dependent microglial activation is mainly neuroprotective in β-amyloidosis and that neuronal pathology is a better readout than plaque load.

Key findings

  • Trem2 deletion impaired microglial responses to plaque: reduced Iba1+ clusters, CD68 staining, CD45high microglia, and methoxy-X04+ amyloid-containing microglia across ages; RNAseq found 144 transcripts reduced and only 7 increased in PS2APP;Trem2ko microglia (≥2-fold, adjusted p ≤ 0.05), including impaired proliferation/Wnt-related genes.
  • Plaque effects were age- and sex-dependent: plaque load was increased in 6–7 month PS2APP;Trem2ko females, but reduced in 12 month females and 19–22 month males; plaques were more diffuse in Trem2ko brains at all ages tested, and amber cores were reduced in Trem2het and nearly absent in Trem2ko aged males.
  • Aβ42:Aβ40 ratio was elevated in PS2APP;Trem2ko brains in both TBS and GuHCl fractions across ages, with increased soluble fibrillar oligomers by OC dot blot; ApoE accumulated in Trem2ko microglia in 12 month females despite no difference in microglial Apoe mRNA.
  • Neuronal pathology worsened: plaque-associated Lamp1+ axonal dystrophy and total Lamp1+ area increased from 7 months onward in Trem2ko mice; amino cupric silver degeneration was greater at 12 months in females and 19–22 months in males; CSF NfL was significantly higher in Trem2ko mice, while plasma NfL did not differ; plaque-proximal dendritic spine density was lower in Trem2ko than Trem2wt or Trem2het mice, with spine density ~50% reduced near plaque versus distal segments.

Methods

  • Compared PS2APP transgenic mice with PS2APP;Trem2ko mice across ages 4 to 22 months, using single-sex cohorts: females at 4, 6–7, and 12 months; males at 6–7 and 19–22 months, plus a 19–22 month male cohort including Trem2het mice.
  • Used histology and whole-slide imaging for plaque, Iba1, CD68, Gfap, Lamp1, ApoE, and silver stains; flow cytometry/FACS for brain-resident myeloid cells; RNAseq of sorted microglia; Aβ40/Aβ42 ELISA; nondenaturing dot blots; two-photon imaging of plaque and dendritic spines; and Simoa NfL measurements in CSF/plasma.
  • Main analyses included unpaired t tests, one-way ANOVA with Tukey’s multiple-comparisons, differential expression with voom+limma, GO overrepresentation, and gene set scoring.

Limitations

  • Stated in the paper: RNAseq was from bulk sorted brain myeloid cells, not single-cell resolution; the CSF/plasma NfL cohort was not littermates and was assembled specifically for NfL measurements after a peer-review recommendation; aged male numbers were depleted, so available Trem2het mice were included to round out that analysis.
  • Evident from the text: findings are from a mouse β-amyloidosis model and may not translate directly to human AD; several cohorts were processed as separate batches, and some flow cytometry/methoxy-X04 analyses used very small n (e.g., n = 3 Trem2wt and n = 2 Trem2ko).
  • Evident from the text: the study reports histopathological and biochemical outcomes, not cognitive/behavioral measures, so the functional consequences of the observed neuronal pathology are not directly tested.

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Registry

Crossref · 10.1523/jneurosci.1871-19.2019

Cites

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Statistics

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Figures

Figure 1. Trem2 deletion impairs microglial clustering, activation, and plaque uptake in PS2APP β-amyloidosis model. A, Immunohistochemical detection of microglial clustering in transgenic PS2APP mice or nontransgenic (Ntg) controls with WT (wt), heterozygous (het), or homozygous KO (ko) Trem2 alleles. For analysis of total, rather than clustered, Iba1 signal, see Figure 1-1. Quantification of the percent area covered by clusters of Iba1+ microglia was measured from coronal sections of female mice at 4, 6–7, and 12 months of age, and from male mice at 6–7 and 19–22 months of age. Each data point represents the composite (average) histological score from several sections of an individual mouse. Bars and lines represent mean ± SEM. Unpaired t test for most cohorts, or by ANOVA followed by Tukey's multiple-comparison test for the 19–22 month cohort with three genotypes: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated. B, Representative low (left, scale bar, 400 μm) and high (right, scale bar, 100 μm) magnification images of Iba1+ staining in 12 month female mice are shown. Iba1+ clusters (red arrowheads) showed reduced presence across Trem2ko mice at all ages. C, D, Same as in A and B, except analyzing active microglial lysosomes as indicated by CD68 protein expression. E, Representative flow cytometry plots measuring CD45 immunoreactivity (low or high) of CD11b+ brain-resident myeloid cells from 14 month PS2APP mice with or without Trem2 and from Ntg mice. F, Percent of brain-resident myeloid cells with high CD45 expression measured from several mice of each genotype. Bars and lines represent mean ± SEM. G, H, Same as in E and F, except analyzing plaque content in brain CD11b+ cells from ∼12 months mice injected with methoxy-X04 dye to stain amyloid material. Ntg mice are not plotted in H since they have zero methoxy labeling. n = 3 in Trem2wt; n = 2 in Trem2ko. For histological analysis of methoxy-X04 and Iba1+ microglia colocalization in sections, see Figure 1-1.
Figure 1. Trem2 deletion impairs microglial clustering, activation, and plaque uptake in PS2APP β-amyloidosis model. A, Immunohistochemical detection of microglial clustering in transgenic PS2APP mice or nontransgenic (Ntg) controls with WT (wt), heterozygous (het), or homozygous KO (ko) Trem2 alleles. For analysis of total, rather than clustered, Iba1 signal, see Figure 1-1. Quantification of the percent area covered by clusters of Iba1+ microglia was measured from coronal sections of female mice at 4, 6–7, and 12 months of age, and from male mice at 6–7 and 19–22 months of age. Each data point represents the composite (average) histological score from several sections of an individual mouse. Bars and lines represent mean ± SEM. Unpaired t test for most cohorts, or by ANOVA followed by Tukey's multiple-comparison test for the 19–22 month cohort with three genotypes: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated. B, Representative low (left, scale bar, 400 μm) and high (right, scale bar, 100 μm) magnification images of Iba1+ staining in 12 month female mice are shown. Iba1+ clusters (red arrowheads) showed reduced presence across Trem2ko mice at all ages. C, D, Same as in A and B, except analyzing active microglial lysosomes as indicated by CD68 protein expression. E, Representative flow cytometry plots measuring CD45 immunoreactivity (low or high) of CD11b+ brain-resident myeloid cells from 14 month PS2APP mice with or without Trem2 and from Ntg mice. F, Percent of brain-resident myeloid cells with high CD45 expression measured from several mice of each genotype. Bars and lines represent mean ± SEM. G, H, Same as in E and F, except analyzing plaque content in brain CD11b+ cells from ∼12 months mice injected with methoxy-X04 dye to stain amyloid material. Ntg mice are not plotted in H since they have zero methoxy labeling. n = 3 in Trem2wt; n = 2 in Trem2ko. For histological analysis of methoxy-X04 and Iba1+ microglia colocalization in sections, see Figure 1-1.CC BY · PubMed Central
Figure 2. Induction of Neurodegeneration-related and Proliferation gene modules and Wnt-related signaling components is impaired in Trem2ko microglia. A, Heat map of differentially expressed genes (DEGs) between Trem2wt and Trem2ko microglia from 14 month PS2APP mouse brains (fold change ≥ 2, adjusted p ≤ 0.05), cross-referenced against previously published datasets from the PS2APP (GSE89482), APPswe/PS1de9 (GSE74615), and 5xFAD (GSE65067) models. Each row is a DEG, and each column is a microglial sort from a different mouse. z-score coloring represents a sample's distance in SDs from the mean expression value for a given gene across samples within a dataset. Most of the downregulated DEGs in Trem2ko microglia were typically induced in models of β-amyloid pathology, and the majority also showed impaired microglial induction in Trem2ko 5xFAD mice. For a complete list of genome-wide expression values in PS2APP;Trem2wt and PS2APP;Trem2ko microglia, see Figure 2-1). B, Induction of the Wnt/proliferation regulators Dkk2, Wif1, Ctnna3, and Asb11 was completely impaired in Trem2ko microglia of PS2APP mice. C, DEGs identified by GO analysis as “positive regulators of Wnt signaling” with reduced expression in PS2APP;Trem2ko microglia included Fzd9, Sulf2, Bambi, Ptk7, Aspm, and Dkk2. B, C, Bars and lines represent mean ± SEM, with each data point representing microglial gene expression level from a given mouse. For genes in B and C, see also Figure 2-2, showing microglial expression in the 5xFAD model with or without Trem2 (data from independent investigators). D, Top, Analysis of previously published expression profiles from the PS2APP model (GSE89482) indicated that six gene expression modules for brain myeloid cells (defined by Friedman et al., 2018) showed altered expression in microglia from brains with β-amyloid pathology. Bottom, Analysis of Trem2ko and Trem2wt microglia expression profiles from PS2APP mice showed that the Neurodegeneration-related and Proliferation gene sets showed significant Trem2 dependence. For data from independent investigators showing enriched expression of Proliferation module in 5xFAD model microglia and its dependence on Trem2 for full induction, see also Figure 2-2). For heat map displays of all individual genes in each module, see also Figure 2-3. Each data point represents a gene set expression score for microglia isolated from a given mouse. Lines indicate mean ± SEM. Two-tailed t tests assuming unequal variance between groups: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. E, Four-way plot of Neurodegeneration-related gene set, with each point representing a gene's fold change in expression between PS2APP versus nontransgenic microglia on the x axis (red and blue points, adjusted p < 0.05) and between PS2APP;Trem2ko versus PS2APP;Trem2wt microglia on the y axis (green and blue points, adjusted p < 0.05). Blue points represent significant differential expression in both datasets. Tiny black points represent genes not differentially expressed in either dataset.
Figure 2. Induction of Neurodegeneration-related and Proliferation gene modules and Wnt-related signaling components is impaired in Trem2ko microglia. A, Heat map of differentially expressed genes (DEGs) between Trem2wt and Trem2ko microglia from 14 month PS2APP mouse brains (fold change ≥ 2, adjusted p ≤ 0.05), cross-referenced against previously published datasets from the PS2APP (GSE89482), APPswe/PS1de9 (GSE74615), and 5xFAD (GSE65067) models. Each row is a DEG, and each column is a microglial sort from a different mouse. z-score coloring represents a sample's distance in SDs from the mean expression value for a given gene across samples within a dataset. Most of the downregulated DEGs in Trem2ko microglia were typically induced in models of β-amyloid pathology, and the majority also showed impaired microglial induction in Trem2ko 5xFAD mice. For a complete list of genome-wide expression values in PS2APP;Trem2wt and PS2APP;Trem2ko microglia, see Figure 2-1). B, Induction of the Wnt/proliferation regulators Dkk2, Wif1, Ctnna3, and Asb11 was completely impaired in Trem2ko microglia of PS2APP mice. C, DEGs identified by GO analysis as “positive regulators of Wnt signaling” with reduced expression in PS2APP;Trem2ko microglia included Fzd9, Sulf2, Bambi, Ptk7, Aspm, and Dkk2. B, C, Bars and lines represent mean ± SEM, with each data point representing microglial gene expression level from a given mouse. For genes in B and C, see also Figure 2-2, showing microglial expression in the 5xFAD model with or without Trem2 (data from independent investigators). D, Top, Analysis of previously published expression profiles from the PS2APP model (GSE89482) indicated that six gene expression modules for brain myeloid cells (defined by Friedman et al., 2018) showed altered expression in microglia from brains with β-amyloid pathology. Bottom, Analysis of Trem2ko and Trem2wt microglia expression profiles from PS2APP mice showed that the Neurodegeneration-related and Proliferation gene sets showed significant Trem2 dependence. For data from independent investigators showing enriched expression of Proliferation module in 5xFAD model microglia and its dependence on Trem2 for full induction, see also Figure 2-2). For heat map displays of all individual genes in each module, see also Figure 2-3. Each data point represents a gene set expression score for microglia isolated from a given mouse. Lines indicate mean ± SEM. Two-tailed t tests assuming unequal variance between groups: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. E, Four-way plot of Neurodegeneration-related gene set, with each point representing a gene's fold change in expression between PS2APP versus nontransgenic microglia on the x axis (red and blue points, adjusted p < 0.05) and between PS2APP;Trem2ko versus PS2APP;Trem2wt microglia on the y axis (green and blue points, adjusted p < 0.05). Blue points represent significant differential expression in both datasets. Tiny black points represent genes not differentially expressed in either dataset.CC BY · PubMed Central
Figure 3. Trem2 deletion shows age- and sex-dependent effects on amyloid plaque pathology, with reduced plaque accumulation at later ages. A, B, Amyloid plaque was visualized using the Campbell-Switzer silver stain method in nontransgenic (Ntg) controls or transgenic PS2APP mice with WT (wt), heterozygous (het), or homozygous KO (ko) Trem2 alleles. Quantification of the percent area covered by amyloid plaque was measured from coronal sections of indicated sex and age. Representative low (left, scale bar, 400 μm) and high (right, scale bar, 100 μm) magnification images of amyloid stains are shown. In the absence of Trem2, plaque loads were elevated in 6–7 month females but reduced in 12 month females and in 19–22 month males. For two-photon imaging of methoxy-X04-labeled plaque in somatosensory cortex of 6 month females and 8 month males, see also Figure 3-1. C, The Campbell-Switzer silver stain turns highly mature plaque cores amber (red arrowheads). Quantification plot (left) and representative images (right, scale bar, 20 μm) of amber core frequency in 19–22 month male mice. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: *p < 0.05, **p < 0.01, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.
Figure 3. Trem2 deletion shows age- and sex-dependent effects on amyloid plaque pathology, with reduced plaque accumulation at later ages. A, B, Amyloid plaque was visualized using the Campbell-Switzer silver stain method in nontransgenic (Ntg) controls or transgenic PS2APP mice with WT (wt), heterozygous (het), or homozygous KO (ko) Trem2 alleles. Quantification of the percent area covered by amyloid plaque was measured from coronal sections of indicated sex and age. Representative low (left, scale bar, 400 μm) and high (right, scale bar, 100 μm) magnification images of amyloid stains are shown. In the absence of Trem2, plaque loads were elevated in 6–7 month females but reduced in 12 month females and in 19–22 month males. For two-photon imaging of methoxy-X04-labeled plaque in somatosensory cortex of 6 month females and 8 month males, see also Figure 3-1. C, The Campbell-Switzer silver stain turns highly mature plaque cores amber (red arrowheads). Quantification plot (left) and representative images (right, scale bar, 20 μm) of amber core frequency in 19–22 month male mice. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: *p < 0.05, **p < 0.01, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.CC BY · PubMed Central
Figure 4. Trem2 deletion impairs the consolidation of β-amyloid into dense plaque. A, β-amyloid plaques were visualized by X-34 staining and confocal z-stack imaging. Representative maximum projection images of X-34+ plaques from 12 month PS2APP females of different Trem2 genotypes are shown (top). High-magnification images show a representative plaque from each genotype with the outlined masks used to delineate the compact core (green) and compact+diffuse (blue) areas of the plaque (bottom). Scale bars, 20 μm. B, The diffuseness index (see Materials and Methods) of the X-34+ plaques in cohorts of the indicated age and sex was quantified. Each data point represents 1 animal's plaque diffuseness index averaged from 3 to 4 FOV. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt.
Figure 4. Trem2 deletion impairs the consolidation of β-amyloid into dense plaque. A, β-amyloid plaques were visualized by X-34 staining and confocal z-stack imaging. Representative maximum projection images of X-34+ plaques from 12 month PS2APP females of different Trem2 genotypes are shown (top). High-magnification images show a representative plaque from each genotype with the outlined masks used to delineate the compact core (green) and compact+diffuse (blue) areas of the plaque (bottom). Scale bars, 20 μm. B, The diffuseness index (see Materials and Methods) of the X-34+ plaques in cohorts of the indicated age and sex was quantified. Each data point represents 1 animal's plaque diffuseness index averaged from 3 to 4 FOV. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt.CC BY · PubMed Central
Figure 5. Trem2 deletion increases the Aβ42:Aβ40 ratio and soluble fibrillar Aβ oligomers in PS2APP brains. A, B, Frozen hippocampal tissues were homogenized and processed for measuring the abundance of Aβ40 and Aβ42 peptides by ELISA in the soluble (TBS) and insoluble (GuHCl) fractions. The ratio of Aβ42:Aβ40 in the TBS (A) and GuHCl (B) fractions is shown. Each data point represents 1 animal's Aβ42:Aβ40 ratio. For individual Aβ40 and Aβ42 peptide measurements, see Figure 5-1. C, Image of nondenaturing dot blot of hippocampal soluble TBS homogenates from 12 month female animals immunostained with the fibrillar oligomeric Aβ antibody OC (green) and control β-actin antibody (red). Squares outline PS2APP;Trem2wt samples. Circles outline PS2APP;Trem2ko samples. #, Control sample from a PS2APP;Bace1ko mouse. D, Signal intensity ratios of OC antibody to actin antibody are plotted for the dot blot shown in C. Dotted line indicates the OC:actin ratio for a control sample from a PS2APP;Bace1ko mouse. E, The Aβ42:Aβ40 ratio (from A) and normalized OC+ dot blot signal (from D) in the TBS soluble fraction showed a significant positive correlation (linear regression; df(1,28), F = 8.63, p < 0.01). F, Same as in C, except immunostained with pan-Aβ antibody 6E10 (green) and control GAPDH antibody (red). G, Signal intensity ratios of Aβ 6E10 antibody to control GAPDH antibody are plotted for the dot blot shown in F. The PS2APP;Bace1ko control sample still has substantial 6E10 signal (see dotted line) since the N-terminal Aβ residues recognized by 6E10 are present in soluble APP when α-secretase is the responsible enzyme. H, The Aβ42:Aβ40 ratio (from A) and normalized Aβ 6E10 dot blot signal (from G) in the TBS soluble fraction showed a significant negative correlation (linear regression; df(1,28), F = 4.98, p < 0.05). Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.
Figure 5. Trem2 deletion increases the Aβ42:Aβ40 ratio and soluble fibrillar Aβ oligomers in PS2APP brains. A, B, Frozen hippocampal tissues were homogenized and processed for measuring the abundance of Aβ40 and Aβ42 peptides by ELISA in the soluble (TBS) and insoluble (GuHCl) fractions. The ratio of Aβ42:Aβ40 in the TBS (A) and GuHCl (B) fractions is shown. Each data point represents 1 animal's Aβ42:Aβ40 ratio. For individual Aβ40 and Aβ42 peptide measurements, see Figure 5-1. C, Image of nondenaturing dot blot of hippocampal soluble TBS homogenates from 12 month female animals immunostained with the fibrillar oligomeric Aβ antibody OC (green) and control β-actin antibody (red). Squares outline PS2APP;Trem2wt samples. Circles outline PS2APP;Trem2ko samples. #, Control sample from a PS2APP;Bace1ko mouse. D, Signal intensity ratios of OC antibody to actin antibody are plotted for the dot blot shown in C. Dotted line indicates the OC:actin ratio for a control sample from a PS2APP;Bace1ko mouse. E, The Aβ42:Aβ40 ratio (from A) and normalized OC+ dot blot signal (from D) in the TBS soluble fraction showed a significant positive correlation (linear regression; df(1,28), F = 8.63, p < 0.01). F, Same as in C, except immunostained with pan-Aβ antibody 6E10 (green) and control GAPDH antibody (red). G, Signal intensity ratios of Aβ 6E10 antibody to control GAPDH antibody are plotted for the dot blot shown in F. The PS2APP;Bace1ko control sample still has substantial 6E10 signal (see dotted line) since the N-terminal Aβ residues recognized by 6E10 are present in soluble APP when α-secretase is the responsible enzyme. H, The Aβ42:Aβ40 ratio (from A) and normalized Aβ 6E10 dot blot signal (from G) in the TBS soluble fraction showed a significant negative correlation (linear regression; df(1,28), F = 4.98, p < 0.05). Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.CC BY · PubMed Central
Figure 6. ApoE markedly accumulates on plaques and in microglia in PS2APP;Trem2ko females. A, Plaque-associated ApoE signal was quantified for each animal in each cohort using 2 or 3 sections per animal, with each section having a range of ∼50–800 plaques contributing to the analysis. Significant increases were observed in 6–7 month and 12 month female (f) PS2APP;Trem2ko mice, but not in males (m). B, Representative low-magnification images of the hippocampus (top row) and high-magnification images of the subiculum (enlarged from the boxed regions) from 12 month female brain sections stained with methoxy-X04 to label plaque (blue), anti-Iba1 to label microglia (green), and anti-ApoE (red). Arrows point to examples of plaques with intense ApoE staining, which are atypical in Trem2wt females but typical in Trem2ko females at this age. C, Representative 20× maximum projection of confocal z-stacks imaged from cortex, highlighting ApoE (red) localization in microglia (Iba1, green) and plaque (blue). Microglia with small ApoE puncta (arrows) are typical in Trem2wt, whereas microglia with enlarged ApoE structures (arrowheads) are frequent in Trem2ko. Plaques with strong ApoE labeling are more common in Trem2ko (asterisks). D, Analysis of the fraction of Iba1 signal that overlaps with ApoE staining (Mander's colocalization coefficient) in 12 month PS2APP females revealed increased colocalization in Trem2ko microglia. E, RNA-Seq profiles of microglia FACS-purified from brains of 14 month PS2APP mice showed no difference in Apoe expression between Trem2wt and Trem2ko microglia (MG). F, Expression profiles of microglia FACS-purified from brains of 8 month nontransgenic or 5xFAD mice revealed strong Apoe induction by β-amyloid pathology in both Trem2wt and Trem2ko microglia, with induction in 5xFAD;Trem2ko relative to 5xFAD;Trem2wt microglia being twofold reduced in one dataset (GSE132508, RNA-Seq) and not significantly different in another (GSE65067, microarray). Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: *p < 0.05, **p < 0.01, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated. n.s. - not significant.
Figure 6. ApoE markedly accumulates on plaques and in microglia in PS2APP;Trem2ko females. A, Plaque-associated ApoE signal was quantified for each animal in each cohort using 2 or 3 sections per animal, with each section having a range of ∼50–800 plaques contributing to the analysis. Significant increases were observed in 6–7 month and 12 month female (f) PS2APP;Trem2ko mice, but not in males (m). B, Representative low-magnification images of the hippocampus (top row) and high-magnification images of the subiculum (enlarged from the boxed regions) from 12 month female brain sections stained with methoxy-X04 to label plaque (blue), anti-Iba1 to label microglia (green), and anti-ApoE (red). Arrows point to examples of plaques with intense ApoE staining, which are atypical in Trem2wt females but typical in Trem2ko females at this age. C, Representative 20× maximum projection of confocal z-stacks imaged from cortex, highlighting ApoE (red) localization in microglia (Iba1, green) and plaque (blue). Microglia with small ApoE puncta (arrows) are typical in Trem2wt, whereas microglia with enlarged ApoE structures (arrowheads) are frequent in Trem2ko. Plaques with strong ApoE labeling are more common in Trem2ko (asterisks). D, Analysis of the fraction of Iba1 signal that overlaps with ApoE staining (Mander's colocalization coefficient) in 12 month PS2APP females revealed increased colocalization in Trem2ko microglia. E, RNA-Seq profiles of microglia FACS-purified from brains of 14 month PS2APP mice showed no difference in Apoe expression between Trem2wt and Trem2ko microglia (MG). F, Expression profiles of microglia FACS-purified from brains of 8 month nontransgenic or 5xFAD mice revealed strong Apoe induction by β-amyloid pathology in both Trem2wt and Trem2ko microglia, with induction in 5xFAD;Trem2ko relative to 5xFAD;Trem2wt microglia being twofold reduced in one dataset (GSE132508, RNA-Seq) and not significantly different in another (GSE65067, microarray). Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: *p < 0.05, **p < 0.01, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated. n.s. - not significant.CC BY · PubMed Central
Figure 7. Trem2 deletion exacerbates plaque-proximal axonal dystrophy. A, Representative images from 12 month brain sections stained with methoxy-X04 to label plaque, anti-Iba1 to label microglia, and anti-Lamp1 to label dystrophic axons around plaque. B, Microglial clustering around plaque is impaired in Trem2-deficient mice. Plaque-associated Iba1 signal was quantified for each animal in each cohort using 2 or 3 sections per animal, with each section having hundreds or thousands of plaques contributing to the analysis. C, Axonal dystrophy per plaque is exacerbated in Trem2-deficient mice. Plaque-associated Lamp1 signal was quantified for each animal in a similar manner as Iba1 signal in B. D, Total axonal dystrophy is exacerbated in Trem2-deficient mice. Total Lamp1 signal in each section was quantified, and each data point represents the average score from 2 or 3 sections per animal. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.
Figure 7. Trem2 deletion exacerbates plaque-proximal axonal dystrophy. A, Representative images from 12 month brain sections stained with methoxy-X04 to label plaque, anti-Iba1 to label microglia, and anti-Lamp1 to label dystrophic axons around plaque. B, Microglial clustering around plaque is impaired in Trem2-deficient mice. Plaque-associated Iba1 signal was quantified for each animal in each cohort using 2 or 3 sections per animal, with each section having hundreds or thousands of plaques contributing to the analysis. C, Axonal dystrophy per plaque is exacerbated in Trem2-deficient mice. Plaque-associated Lamp1 signal was quantified for each animal in a similar manner as Iba1 signal in B. D, Total axonal dystrophy is exacerbated in Trem2-deficient mice. Total Lamp1 signal in each section was quantified, and each data point represents the average score from 2 or 3 sections per animal. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: **p < 0.01, ***p < 0.001, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.CC BY · PubMed Central
Figure 8. Disintegrative degeneration staining and CSF NfL measurements reveal exacerbated neuronal damage in Trem2-deficient mice at later ages. A, Quantification of sections from nontransgenic (Ntg) and PS2APP mice with indicated Trem2 genotypes stained using an amino cupric silver staining method that labels degenerative neuronal processes. Each data point represents the average percent area covered/section for ∼10 sections per animal. B, Representative low (left, scale bar, 400 μm) and high (right, scale bar, 100 μm) magnification images from the 12 month female and 19–22 month male cohorts are shown. Degenerative signal is apparent in plaque-accompanying foci (red arrowheads) throughout the cortex and hippocampus and in certain white matter tracts, including the corpus callosum (cc). C, Plasma (left) and CSF (right) NfL were measured from a separate, mixed sex cohort of 12 months PS2APP;Trem2wt and PS2APP;Trem2ko mice. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: **p < 0.01, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.
Figure 8. Disintegrative degeneration staining and CSF NfL measurements reveal exacerbated neuronal damage in Trem2-deficient mice at later ages. A, Quantification of sections from nontransgenic (Ntg) and PS2APP mice with indicated Trem2 genotypes stained using an amino cupric silver staining method that labels degenerative neuronal processes. Each data point represents the average percent area covered/section for ∼10 sections per animal. B, Representative low (left, scale bar, 400 μm) and high (right, scale bar, 100 μm) magnification images from the 12 month female and 19–22 month male cohorts are shown. Degenerative signal is apparent in plaque-accompanying foci (red arrowheads) throughout the cortex and hippocampus and in certain white matter tracts, including the corpus callosum (cc). C, Plasma (left) and CSF (right) NfL were measured from a separate, mixed sex cohort of 12 months PS2APP;Trem2wt and PS2APP;Trem2ko mice. Error bars indicate mean ± SEM. Unpaired t test or ANOVA followed by Tukey's multiple-comparison test: **p < 0.01, ****p < 0.0001 versus PS2APP;Trem2wt or as indicated.CC BY · PubMed Central
Figure 9. Trem2 deletion exacerbates dendritic spine loss near plaque. A, Intact brains from 6 month female PS2APP mice with WT (wt), KO (ko), or heterozygous (het) Trem2 genotypes and carrying the Thy1::GFP-M transgene, which sparsely labels excitatory cortical neurons, were imaged in the somatosensory cortex (SS Cx) using two-photon microscopy. Dendritic shafts proximal to plaque (within 20 μm) or distally located (≥100 μm from any plaque) were imaged, with representative images shown. B, Each data point represents the average of five spine density measurements from 1 animal. Spine density was ∼50% reduced on plaque-proximal dendrite segments relative to distal dendrite segments from the same animals. Spine density near plaque was lower in PS2APP;Trem2ko mice than in PS2APP;Trem2wt or PS2APP;Trem2het mice. Error bars indicate mean ± SEM. ANOVA: *p < 0.05, **p < 0.01. Spine volume was unaffected by either plaque proximity or Trem2 genotype (data not shown).
Figure 9. Trem2 deletion exacerbates dendritic spine loss near plaque. A, Intact brains from 6 month female PS2APP mice with WT (wt), KO (ko), or heterozygous (het) Trem2 genotypes and carrying the Thy1::GFP-M transgene, which sparsely labels excitatory cortical neurons, were imaged in the somatosensory cortex (SS Cx) using two-photon microscopy. Dendritic shafts proximal to plaque (within 20 μm) or distally located (≥100 μm from any plaque) were imaged, with representative images shown. B, Each data point represents the average of five spine density measurements from 1 animal. Spine density was ∼50% reduced on plaque-proximal dendrite segments relative to distal dendrite segments from the same animals. Spine density near plaque was lower in PS2APP;Trem2ko mice than in PS2APP;Trem2wt or PS2APP;Trem2het mice. Error bars indicate mean ± SEM. ANOVA: *p < 0.05, **p < 0.01. Spine volume was unaffected by either plaque proximity or Trem2 genotype (data not shown).CC BY · PubMed Central

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  • It may state two separate claims as one. 42
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  • The paper relies on this without support.

    The effects of Trem2 deletion in PS2APP mice are mediated specifically by microglial Trem2 rather than by other Trem2-expressing myeloid cells or non-myeloid cells.

    “TREM2 is an Alzheimer's disease (AD) risk gene expressed in microglia.”

  • The paper relies on this without support.

    The Trem2tm1(KOMP)Vlcg allele completely abolishes Trem2 function in PS2APP;Trem2ko mice.

    “The Trem2tm1(KOMP)Vlcg null allele (C57BL/6N background) was crossed into the PS2APP model (C57BL/6J background).”

  • The paper relies on this without support.

    Amber core amyloid staining represents a mature form of highly condensed plaque.

    “At this age, the Campbell-Switzer stain also revealed a distinctive pattern of “amber core” amyloid staining in PS2APP brains (Fig. 3C), thought to represent a mature form of highly condensed plaque.”

  • The paper relies on this without support.

    Cognitive decline correlates better with tau pathology and synapse loss than with brain amyloid detection.

    “helps explain why cognitive decline correlates better with tau pathology and synapse loss than with brain amyloid detection.”
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  1. Campbell, 1987, Alzheimer's plaques and tangles: a controlled and enhanced silver staining method, Soc Neuroscience Abstr, 13, 678

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Metadata

Authors
Meilandt, W. J., Ngu, H., Gogineni, A., Lalehzadeh, G., Lee, S.-H., Srinivasan, K., Imperio, J., Wu, T., Weber, M., Kruse, A. J., Stark, K. L., Chan, P., Kwong, M., Modrusan, Z., Friedman, B. A., Elstrott, J., Foreman, O., Easton, A., Sheng, M., & Hansen, D. V.
Affiliations
Genentech (US)
Corresponding author
  • Meilandt, William J.
  • Hansen, David V.
ORCID
Venue
The Journal of Neuroscience
Publisher
Society for Neuroscience
Published
2020-01-24
Volume
40
Issue
9
Pages
1956-1974
ISSN
  • 0270-6474 (ISSN-L)
  • 1529-2401
DOI
10.1523/jneurosci.1871-19.2019
PMC
PMC7046459
OpenAlex
W3003126983
Semantic Scholar
24b2107485715864dc70c72f73ce957a07b3f7a9
Cited by
210
References
98
Citations by year
2026: 16 · 2025: 30 · 2024: 44 · 2023: 48 · 2022: 33 · 2021: 25 · 2020: 14
FWCI
18.04
Type
journal article
Language
en
Open access
Licence
Locations
  • Journal of Neuroscience (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc)
Topics
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Alzheimer's disease research and treatments
  • Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
  • Health Sciences › Medicine › Psychiatry and Mental health › Dementia and Cognitive Impairment Research
Keywords
TREM2, Alzheimer's disease, amyloid plaques, amyloid-beta 42/40 ratio, axonal dystrophy, dendritic spine loss, neuritic dystrophy, microglial activation, soluble amyloid-beta oligomers, neurofilament light chain, apolipoprotein E, Wnt signaling, sex differences
MeSH terms
Trefoil Factor-1, Alzheimer Disease, Animals, Axons, Female, Male, Membrane Glycoproteins, Mice, Inbred C57BL, Peptide Fragments, Receptors, Immunologic, Amyloid beta-Peptides, Neurites, Amyloid beta-Protein Precursor, Neurofilament Proteins, Microglia, Mice, Knockout, Dendritic Spines, Mice, Plaque, Amyloid

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