Roles of blood monocytes carrying TREM2R47H mutation in pathogenesis of Alzheimer's disease and its therapeutic potential in APP/PS1 mice

Abstract

Introduction: The triggering receptor expressed on myeloid cells 2 (TREM2) arginine-47-histidine (R47H) mutation is a significant risk for Alzheimer's disease (AD) with unclear mechanisms. Previous studies focused on microglial amyloid-β (Aβ) phagocytosis with less attention on the impact of TREM2^R47H mutation on blood monocytes.

Methods: Bone marrow transplantation (BMT) models were used to assess the contribution of blood monocytes carrying TREM2^R47H mutation to AD.

Results: Aβ phagocytosis was compromised in mouse monocytes carrying the TREM2^R47H mutation. Transplantation of bone marrow cells (BMCs) carrying TREM2^R47H mutation increased cerebral Aβ burden and aggravated AD-type pathologies. Moreover, the replacement of TREM2^R47H-BMCs restored monocytic Aβ phagocytosis, lowered Aβ levels in the blood and brain, and improved cognitive function.

Discussion: Our study reveals that blood monocytes carrying the TREM2^R47H mutation substantially contribute to the pathogenesis of AD, and correcting the TREM2^R47H mutation in BMCs would be a potential therapeutic approach for those carrying this mutation.

Highlights: TREM2^R47H mutation compromises the Aβ phagocytosis of blood monocytes. Blood monocytes carrying TREM2^R47H mutation contribute substantially to AD pathogenesis. Correction of the TREM2^R47H mutation in bone marrow cells ameliorates AD pathologies and cognitive impairments.

Keywords: Alzheimer's disease; Aβ; TREM2R47H mutation; bone marrow transplantation; monocytes; phagocytosis.

FIGURE 1

Effect of the TREM2^R47H mutation on monocytic Aβ phagocytosis. (A) Schematic diagram of investigation of the Aβ uptake by blood monocytes in the context of mixing CD45.2‐TREM2^R47H and CD45.1‐TREM2^Wt PBMCs. (B) Representative flow‐cytometric images of CD115+ and CD11b+ monocytes scavenging Aβ. (C) Statistics of the proportion of CD115⁺ and CD11b⁺ cells in response to Aβ and mean fluorescence intensity. (D) Schematic diagram of investigation of the Aβ uptake by CD115+ monocytes in the context of mixing CD45.2‐ TREM2^R47H‐and TREM2^Wt‐CD115+ cells. (E) Representative flow‐cytometric images of CD115+ and CD11b+ monocytes scavenging Aβ. F. Statistics of the proportion of CD115⁺ and CD11b⁺ cells responding to Aβ and mean fluorescence intensity. n = 6 per group, ***p < 0.001, ****p < 0.0001. The error bars are the SEMs. Aβ, amyloid‐β; PBMCs, peripheral blood mononuclear cells; TREM2^Wt, Wt TREM2 gene; TREM2^R47H, R47H mutation in the TREM2 gene; Wt, wild‐type.

FIGURE 2

TREM2^R47H‐BMC transplantation impaired monocytic Aβ phagocytosis in AD mice. (A) Schematic diagram of generating a specific mouse model and results of chimerism. (B) Statistics of the number of CD115⁺ and CD11b⁺ cells responding to Aβ and mean fluorescence intensity. (C) Blood Aβ40 and Aβ42 levels of AD‐T^Wt→Wt, AD‐T^Wt→Wt, and AD‐T^R47H→Wt mice. (D) Correlation between blood total Aβ levels and the ability of monocytic Aβ phagocytosis. n = 8 per group for Figure A–C; n = 24 for Figure D. *p < 0.05, **p < 0.01. The error bar in C is the SEMs. Aβ, amyloid‐β; AD‐T^Wt, AD mice carrying the Wt TREM2 gene; AD‐T^Wt→W, AD mice carrying the Wt TREM2 gene that received BMCs from AD mice carrying the Wt TREM2 gene; AD‐T^R47H→Wt, AD mice carrying the Wt TREM2 gene that received BMCs from AD mice carrying the R47H mutation in the TREM2 gene; ns, no statistical significance; BMC, bone marrow cells; Wt, wild‐type.

FIGURE 3

TREM2^R47H‐BMC transplantation aggravated the Aβ burden in AD mice. (A) Immunostaining and quantification of Aβ plaques stained with the 6E10 antibody and Congo red in the neocortex and hippocampus. (B) Comparison of Aβ40 and Aβ42 levels in the TBS, SDS and FA fractions of brain. (C–E) Representative western‐blotting images and quantitative analysis of the levels of APP‐metabolizing enzymes, Aβ‐degrading enzymes and Aβ‐transporting receptors in brain homogenates. n = 8 per group; *p < 0.05. The scale bar in A is 500 #x000B5;m. The error bars are the SEMs. Aβ, amyloid‐β; AD‐T^Wt, AD mice carrying the Wt TREM2 gene; AD‐T^Wt→Wt, AD mice carrying the Wt TREM2 gene that received BMCs from AD mice carrying the Wt TREM2 gene; AD‐T^R47H→Wt, AD mice carrying the Wt TREM2 gene that received BMCs from AD mice carrying the R47H mutation in the TREM2 gene; ns, no statistical significance; BMC, bone marrow cells; Wt, wild‐type.

FIGURE 4

TREM2^R47H‐BMC transplantation exacerbated AD‐type pathologies in AD mice. (A) Immunostaining and quantification of astrocytes and microglia with the GFAP antibody and the CD68 antibody in the neocortex and hippocampus. (B–C) Representative immunofluorescent images and quantification of neurons (NeuN, red) and dendrites (Map2, green) in the CA1 region of the hippocampus and neural apoptosis (caspase 3, green) in the CA3 region of the hippocampus. (D) Representative immunostaining images and quantification of the levels of PT231, PS199, and Tau46 expression. (E) Representative western‐blotting images and quantification of synaptic proteins (PSD95, SYN, SNAP25, and VAMP1). n = 8 per group; *p < 0.05; ***p < 0.001. The scale bar in A is 500 #x000B5;m, and the scale bar in B is 50 #x000B5;m. The error bars are the SEMs. AD, Alzheimer's disease; AD‐T^Wt, AD mice carrying the Wt TREM2 gene; AD‐T^Wt→W, AD mice carrying the Wt TREM2 gene that received BMCs from AD mice carrying the Wt TREM2 gene; AD‐T^R47H→Wt, AD mice carrying the Wt TREM2 gene that received BMCs from AD mice carrying the R47H mutation in the TREM2 gene; BMC, bone marrow cells; PSD95, postsynaptic protein‐95; SNAP25, anti‐ synaptosomal‐associated protein of 25 kDa; Wt, wild‐type.

FIGURE 5

Replacement of TREM2^R47H‐BMCs with TREM2^Wt‐BMCs recovered Aβ phagocytosis and enhanced blood Aβ clearance. (A) Schematic diagram of analyzing in vivo monocytic Aβ phagocytosis. (B) Confocal images of monocytic Aβ phagocytosis (red for CD11b, green for Aβ). (C) The changes in FITC fluorescence intensity in blood post‐FITC‐labelled Aβ injection (n = 4 per group). (D) Representative images of monocytic Aβ phagocytosis taken by imaging flow cytometry. (E) Statistics of mean fluorescence intensity (MFI) in the double CD115+ and CD11b+ cells. *p < 0.05. The scale bar in B is 10 #x000B5;m. Aβ, amyloid‐β; AD‐T^R47H→R47H, AD mice carrying the R47H mutation in the TREM2 gene that received BMCs from AD mice carrying the R47H mutation in the TREM2 gene; AD‐T^Wt→R47H, AD mice carrying the R47H mutation in the TREM2 gene that received BMCs from AD mice carrying the Wt TREM2 gene; BMC, bone marrow cells; MFI, mean fluorescence intensity; Wt, wild‐type.

FIGURE 6

Replacement of TREM2^R47H‐BMCs with TREM2^Wt‐BMCs alleviated brain Aβ burden. (A) Immunostaining and quantification of Aβ plaques stained with the 6E10 antibody and Congo red in the neocortex and hippocampus. (B) Comparison of Aβ40 and Aβ42 levels in the TBS, SDS and FA fractions of brain. (C–E) Representative western‐blotting images and quantitative analysis of the levels of APP‐metabolizing enzymes, Aβ‐degrading enzymes, and Aβ‐transporting receptors in brain homogenates. n = 8 per group; *p < 0.05, ns denotes no statistical significance. The scale bar in A is 500 #x000B5;m. The error bars are the SEMs. Aβ, amyloid‐β; AD‐T^R47H, AD mice carrying the R47H mutation in the TREM2 gene; AD‐T^R47H→R47H, AD mice carrying the R47H mutation in the TREM2 gene that received BMCs from AD mice carrying the R47H mutation in the TREM2 gene; AD‐T^Wt→R47H, AD mice carrying the R47H mutation in the TREM2 gene that received BMCs from AD mice carrying the Wt TREM2 gene.

FIGURE 7

Replacement of TREM2^R47H‐BMCs with TREM2^Wt‐BMCs ameliorated AD‐type pathologies. (A) Immunostaining and quantification of astrocytes and microglia with the GFAP antibody and the CD68 antibody in the neocortex and hippocampus. (B–C) Representative immunofluorescent images and quantification of neurons (NeuN, red) and dendrites (Map2, green) in the CA1 region of the hippocampus and neural apoptosis (caspase 3, green) in the CA3 region of the hippocampus. (D) Representative immunostaining images and quantification of the levels of PT231, PS396, and Tau46 expression. E. Representative images and quantification of synaptic proteins (PSD95, SNAP25, and VAMP1) and NeuN expression. n = 8 per group; *p < 0.05; ***p < 0.001; ns denotes no statistical significance. The scale bar in A is 500 #x000B5;m, and the scale bar in B is 50 #x000B5;m. The error bars are the SEMs. AD‐T^R47H, AD mice carrying the R47H mutation in the TREM2 gene; AD‐T^R47H→R47H, AD mice carrying the R47H mutation in the TREM2 gene that received BMCs from AD mice carrying the R47H mutation in the TREM2 gene; AD‐T^Wt→R47H, AD mice carrying the R47H mutation in the TREM2 gene that received BMCs from AD mice carrying the Wt TREM2 gene; ns, no statistical significance.; BMC, bone marrow cells; PSD95, postsynaptic protein‐95; SNAP25, synaptosomal‐associated protein of 25 kDa; Wt, wild‐type.