Species: Mouse
Genes: Trem2, TREM2, APP, PSEN1
Mutations: TREM2 R47H, APP K670_M671delinsNL (Swedish), APP I716V (Florida), APP V717I (London), PSEN1 M146L (A>C), PSEN1 L286V
Modification: Trem2: Knock-Out; TREM2: Transgenic; APP: Transgenic; PSEN1: Transgenic
Disease Relevance: Alzheimer's Disease
Strain Name: N/A
Genetic Background: C57BL/6 X CBA, back-crossed for at least 4 generations to C57BL/6
Availability: TREM2 mice: available through Marco Colonna; 5XFAD: The Jackson Lab; available through the JAX MMRRC Stock# 034848; Live

Summary

The R47H variant of TREM2 has been found to confer an approximately threefold greater risk for Alzheimer’s disease in humans heterozygous for this allele. To study the effects of the R47H variant in the context of amyloidosis, mice carrying a human TREM2 gene were crossed with 5XFAD mice. Compared with mice expressing the common variant of human TREM2, mice expressing the R47H variant have fewer plaque-associated microglia, express fewer TREM2-dependent activation markers, and show less binding of soluble TREM2 to neurons and amyloid plaques (Song et al., 2018).

Generation of mice

Humanized TREM2 mice were created in a two-step process: First, researchers generated transgenic mice that carry BACs containing the R47H variant of human TREM2 (R47H). Then these BAC transgenic mice were back-crossed to Trem2^-/- mice (Turnbull et al., 2006) to yield mice that express the R47H variant of human TREM2 in the absence of mouse Trem2 (R47H⁺mTrem2^−/−, or R47H-KO). For comparison, BAC transgenic mice carrying the common variant of human TREM2 in the absence of mouse Trem2 were also generated (CV⁺mTrem2^−/−, or CV-KO).

5XFAD mice express human APP and human presenilin-1, with a total of five AD-linked mutations, and aggressively deposit amyloid plaques starting at 6 weeks of age.

Humanized TREM2 mice were crossed with 5XFAD mice to obtain 5XFAD expressing the R47H variant (R47H⁺mTrem2^−/−5XFAD, or R47H-KO-5XFAD) or the common variant (CV⁺mTrem2^−/−5XFAD, or CV-KO-5XFAD). Additionally, mTrem2^−/−5XFAD (KO-5XFAD) were available for comparison.

Thus far, 8.5-month-old mice have been studied.

Expression of human TREMs

Transgene copy numbers are six to eight times greater in CV-KO than in R47H-KO mice, but TREM2 mRNA levels only differ by approximately 20 percent, and levels of TREM2 protein in hippocampus are similar in CV-KO and R47H-KO. Three hTREM2 mRNA isoforms were detected in cortex by RT-qPCR in animals expressing the common variant, while only two isoforms were found in mice expressing the R47H variant. In the non-5XFAD background, CV-KO brains have slightly higher levels of isoforms 1 and 2 than R47H-KO brains do; these differences are magnified in the 5XFAD background. Levels of Isoform 3, found only in CV-KO brains, do not differ between non-5XFAD and 5XFAD.

The BAC used to create these mice also contains TREML1 and TREML2 genes. The low levels of TREML1 transcript in CV-KO and R47H-KO are similar to levels in Trem2^-/- (KO) brain, regardless of 5XFAD background, suggesting that there is little basal expression of the human TREML1 gene contained in the BAC. TREML2 transcripts, absent from KO brains, are elevated in both CV-KO and R47H-KO. However, the TREML2 mRNA levels do not differ between CV-KO and R47H-KO, regardless of 5XFAD background.

Phenotype

Microglia cluster around amyloid plaques in 5XFAD mice, but genetic ablation of mTrem2 reduces the number of microglia surrounding plaques (Wang et al., 2016). Replacement of mTrem2 with CV, but not with R47H, hTREM2 rescues this deficit: CV-KO-5XFAD mice have more plaque-associated microglia than R47H-KO-5XFAD or KO-5XFAD mice do (Song et al., 2018). Despite differing in plaque-associated microglial density, the coverage of amyloid plaques by microglial processes was similar in CV-KO-5XFAD and R47H-KO-5XFAD mice, and exceeded that in KO-5XFAD mice.

Amyloid plaque burdens in cortex, hippocampus, and subiculum were similar in CV-KO-5XFAD, R47H-KO-5XFAD, and KO-5XFAD. Genotype did not affect levels of PBS- or guanidine-extractable Aβ40 or Aβ42, with the exception of a modest decrease in PBS-soluble Aβ in hippocampus in CV-KO-5XFAD compared with R47H-KO-5XFAD or KO-5XFAD.

In terms of mRNA markers of microglial activation, R47H-KO-5XFAD mice more closely resemble KO-5XFAD than CV-KO-5XFAD. The neurodegeneration-associated microglial activation markers Cst7, Spp1, and Gpnmb were elevated in CV-KO-5XFAD compared with KO-5XFAD and R47H-KO-5XFAD. Further, KO-5XFAD and R47H-KO-5XFAD grouped together, apart from CV-KO-5XFAD, in an unbiased cluster analysis.

An antibody directed against an intracellular epitope on TREM2 stained only microglia, but an antibody directed against the extracellular domain of TREM2 stained plaques and neurons as well. The labeling of neurons and plaques is consistent with binding soluble TREM2 (sTREM2) to these elements. Anti-TREM2 staining of plaques and neurons was greater in the brains of CV-KO-5XFAD than R47H-KO-5XFAD mice.

Modification Details

BAC-transgenic mice carrying human TREM2 (R47H variant), TREML1, and TREML2 were back-crossed to Trem2 KO mice (Colonna) to yield mice that express the R47H variant of human TREM2 in the absence of mouse Trem2. These mice were then crossed with 5XFAD mice.

Phenotype Characterization

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References

Research Models Citations

1. 5xFAD (C57BL6)
2. Trem2 KO (Colonna)
3. Trem2 KO (Colonna) x 5XFAD

Paper Citations

1. Song WM, Joshita S, Zhou Y, Ulland TK, Gilfillan S, Colonna M. Humanized TREM2 mice reveal microglia-intrinsic and -extrinsic effects of R47H polymorphism. J Exp Med. 2018 Mar 5;215(3):745-760. Epub 2018 Jan 10 PubMed.
2. Turnbull IR, Gilfillan S, Cella M, Aoshi T, Miller M, Piccio L, Hernandez M, Colonna M. Cutting edge: TREM-2 attenuates macrophage activation. J Immunol. 2006 Sep 15;177(6):3520-4. PubMed.
3. Wang Y, Ulland TK, Ulrich JD, Song W, Tzaferis JA, Hole JT, Yuan P, Mahan TE, Shi Y, Gilfillan S, Cella M, Grutzendler J, DeMattos RB, Cirrito JR, Holtzman DM, Colonna M. TREM2-mediated early microglial response limits diffusion and toxicity of amyloid plaques. J Exp Med. 2016 May 2;213(5):667-75. Epub 2016 Apr 18 PubMed.

Other Citations

1. R47H

External Citations

1. Marco Colonna
2. JAX MMRRC Stock# 034848

Further Reading