Alzheimer's disease-associated R47H TREM2 increases, but wild-type TREM2 decreases, microglial phagocytosis of synaptosomes and neuronal loss.

Triggering receptor on myeloid cells 2 (TREM2) is an innate immune receptor, upregulated on the surface of microglia associated with amyloid plaques in Alzheimer's disease (AD). Individuals heterozygous for the R47H variant of TREM2 have greatly increased risk of developing AD. We examined the effects of wild-type (WT), R47H and knock-out (KO) of human TREM2 expression in three microglial cell systems. Addition of mouse BV-2 microglia expressing R47H TREM2 to primary mouse neuronal cultures caused neuronal loss, not observed with WT TREM2. Neuronal loss was prevented by using annexin V to block exposed phosphatidylserine, an eat-me signal and ligand of TREM2, suggesting loss was mediated by microglial phagocytosis of neurons exposing phosphatidylserine. Addition of human CHME-3 microglia expressing R47H TREM2 to LUHMES neuronal-like cells also caused loss compared to WT TREM2. Expression of R47H TREM2 in BV-2 and CHME-3 microglia increased their uptake of phosphatidylserine-beads and synaptosomes versus WT TREM2. Human iPSC-derived microglia with heterozygous R47H TREM2 had increased phagocytosis of synaptosomes vs common-variant TREM2. Additionally, phosphatidylserine liposomes increased activation of human iPSC-derived microglia expressing homozygous R47H TREM2 versus common-variant TREM2. Finally, overexpression of TREM2 in CHME-3 microglia caused increased expression of cystatin F, a cysteine protease inhibitor, and knock-down of cystatin F increased CHME-3 uptake of phosphatidylserine-beads. Together, these data suggest that R47H TREM2 may increase AD risk by increasing phagocytosis of synapses and neurons via greater activation by phosphatidylserine and that WT TREM2 may decrease microglial phagocytosis of synapses and neurons via cystatin F.

TREM2 R47H variant and risk for Alzheimer's disease: assessment in a Greek population and updated meta-analysis.

INTRODUCTION: Rare coding variants in TREM2 and their association with the susceptibility towards Alzheimer's disease (AD) were recently studied in various ethnic groups with contradictory results. The T allele of the rs75932628 (p.R47H variant) has shown a positive risk association with AD in several studies; however, neither a study in Greece nor an updated meta-analysis have been conducted. OBJECTIVE: To assess the association between TREM2 rs75932628 and late-onset (sporadic) AD in a Greek population, and perform a meta-analysis of current data. MATERIALS AND METHODS: The rs75932628 was genotyped in a total of 327 patients with AD and 700 cognitively healthy controls. A systematic search and meta-analyses of studies presenting data regarding rs75932628 in AD cases and controls were also performed. RESULTS: Three patients vs. none of the controls were found to carry the heterozygous risk allele of the rs75932628, yielding a significant association (p = 0.032), in the Greek sample. In the meta-analysis, the overall odds ratio (OR) under a fixed-effects model was 2.98 (Confidence Interval (CI):2.52-3.53) showing a significant association of the rs75932628-T allele with AD in the overall dataset, based on data from 27 studies (26200 AD cases and 142084controls). Caucasian population-only studies (n = 16) revealed a similar OR of 2.93 (CI:2.45-3.51), whereas Asian population-only studies (n = 5) had a non-significant OR of 0.84 (CI:0.19-3.74). CONCLUSION: The rs75932628 was associated with AD in the Greek sample. Our meta-analysis, covering a total population of over 168,000 people, also showed a significant association of the allele with AD in Caucasian populations.

Assessment of TREM2 rs75932628 variant's association with Parkinson's disease in a Greek population and Meta-analysis of current data.

BACKGROUND: Α number of genetic variants are considered to confer susceptibility to Parkinson's disease (PD). Rs75392628 (R47H), a rare variant of TREM2 gene, has been linked to PD, although its role on PD remains conflicting. OBJECTIVE: Detection of a possible contribution of rs75392628 variant of TREM2 gene to PD risk. METHODS: A total of 358 PD patients and 358 healthy controls genotyped for rs75392628. In addition, a meta-analysis was performed by merging our results with those from previous studies. RESULTS: The rare variant of rs75932628 (47H) of TREM2 gene was not detected on cohort. Meta-analysis of a total of 9271 PD cases and 9777 controls across 14 independent PD data sets from 9 studies, including the present study, did not show any statistically significant effect of rs75392628 on PD risk (ORFE:1.54 95% CI:0.87-2.73. ORRE: 1.54, 95%CI: 0.71-3.32). CONCLUSIONS: Rs75392628 TREM2 variant is rather unlikely to be a major genetic risk contributor of PD.

IPSC-Derived Neuronal Cultures Carrying the Alzheimer's Disease Associated TREM2 R47H Variant Enables the Construction of an Aß-Induced Gene Regulatory Network.

Genes associated with immune response and inflammation have been identified as genetic risk factors for late-onset Alzheimer´s disease (LOAD). The rare R47H variant within triggering receptor expressed on myeloid cells 2 (TREM2) has been shown to increase the risk for developing Alzheimer's disease (AD) 2-3-fold. Here, we report the generation and characterization of a model of late-onset Alzheimer's disease (LOAD) using lymphoblast-derived induced pluripotent stem cells (iPSCs) from patients carrying the TREM2 R47H mutation, as well as from control individuals without dementia. All iPSCs efficiently differentiated into mature neuronal cultures, however AD neuronal cultures showed a distinct gene expression profile. Furthermore, manipulation of the iPSC-derived neuronal cultures with an Aß-S8C dimer highlighted metabolic pathways, phagosome and immune response as the most perturbed pathways in AD neuronal cultures. Through the construction of an Aß-induced gene regulatory network, we were able to identify an Aß signature linked to protein processing in the endoplasmic reticulum (ER), which emphasized ER-stress, as a potential causal role in LOAD. Overall, this study has shown that our AD-iPSC based model can be used for in-depth studies to better understand the molecular mechanisms underlying the etiology of LOAD and provides new opportunities for screening of potential therapeutic targets.

TREM2-activating antibodies abrogate the negative pleiotropic effects of the Alzheimer's disease variant Trem2R47H on murine myeloid cell function.

Triggering receptor expressed on myeloid cells 2 (TREM2) is an orphan immune receptor expressed on cells of myeloid lineage such as macrophages and microglia. The rare variant R47H TREM2 is associated with an increased risk for Alzheimer's disease, supporting the hypothesis that TREM2 loss of function may exacerbate disease progression. However, a complete knockout of the TREM2 gene in different genetic models of neurodegenerative diseases has been reported to result in both protective and deleterious effects on disease-related end points and myeloid cell function. Here, we describe a Trem2R47H transgenic mouse model and report that even in the absence of additional genetic perturbations, this variant clearly confers a loss of function on myeloid cells. The Trem2R47H variant-containing myeloid cells exhibited subtle defects in survival and migration and displayed an unexpected dysregulation of cytokine responses in a lipopolysaccharide challenge environment. These subtle phenotypic defects with a gradation in severity across genotypes were confirmed in whole-genome RNA-Seq analyses of WT, Trem2-/-, and Trem2R47H myeloid cells under challenge conditions. Of note, TREM2-activating antibodies that boost proximal signaling abrogated survival defects conferred by the variant and also modulated migration and cytokine responses in an antibody-, ligand-, and challenge-dependent manner. In some instances, these antibodies also boosted WT myeloid cell function. Our studies provide a first glimpse into the boost in myeloid cell function that can be achieved by pharmacological modulation of TREM2 activity that can potentially be ameliorative in neurodegenerative diseases such as Alzheimer's disease.

Molecular basis for the loss-of-function effects of the Alzheimer's disease-associated R47H variant of the immune receptor TREM2.

Triggering receptor expressed on myeloid cells 2 (TREM2) is an immune receptor expressed on the surface of microglia, macrophages, dendritic cells, and osteoclasts. The R47H TREM2 variant is a significant risk factor for late-onset Alzheimer's disease (AD), and the molecular basis of R47H TREM2 loss of function is an emerging area of TREM2 biology. Here, we report three high-resolution structures of the extracellular ligand-binding domains (ECDs) of R47H TREM2, apo-WT, and phosphatidylserine (PS)-bound WT TREM2 at 1.8, 2.2, and 2.2 Å, respectively. The structures reveal that Arg47 plays a critical role in maintaining the structural features of the complementarity-determining region 2 (CDR2) loop and the putative positive ligand-interacting surface (PLIS), stabilizing conformations capable of ligand interaction. This is exemplified in the PS-bound structure, in which the CDR2 loop and PLIS drive critical interactions with PS via surfaces that are disrupted in the variant. Together with in vitro and in vivo characterization, our structural findings elucidate the molecular mechanism underlying loss of ligand binding, putative oligomerization, and functional activity of R47H TREM2. They also help unravel how decreased in vitro and in vivo stability of TREM2 contribute to loss of function in disease.

The role of TREM2 R47H as a risk factor for Alzheimer's disease, frontotemporal lobar degeneration, amyotrophic lateral sclerosis, and Parkinson's disease.

A rare variant in TREM2 (p.R47H, rs75932628) was recently reported to increase the risk of Alzheimer's disease (AD) and, subsequently, other neurodegenerative diseases, i.e. frontotemporal lobar degeneration (FTLD), amyotrophic lateral sclerosis (ALS), and Parkinson's disease (PD). Here we comprehensively assessed TREM2 rs75932628 for association with these diseases in a total of 19,940 previously untyped subjects of European descent. These data were combined with those from 28 published data sets by meta-analysis. Furthermore, we tested whether rs75932628 shows association with amyloid beta (Aß42) and total-tau protein levels in the cerebrospinal fluid (CSF) of 828 individuals with AD or mild cognitive impairment. Our data show that rs75932628 is highly significantly associated with the risk of AD across 24,086 AD cases and 148,993 controls of European descent (odds ratio or OR = 2.71, P = 4.67 × 10(-25)). No consistent evidence for association was found between this marker and the risk of FTLD (OR = 2.24, P = .0113 across 2673 cases/9283 controls), PD (OR = 1.36, P = .0767 across 8311 cases/79,938 controls) and ALS (OR = 1.41, P = .198 across 5544 cases/7072 controls). Furthermore, carriers of the rs75932628 risk allele showed significantly increased levels of CSF-total-tau (P = .0110) but not Aß42 suggesting that TREM2's role in AD may involve tau dysfunction.

Alzheimer's disease-linked risk alleles elevate microglial cGAS-associated senescence and neurodegeneration in a tauopathy model.

The strongest risk factors for late-onset sporadic Alzheimer's disease (AD) include the ε4 allele of apolipoprotein E (APOE), the R47H variant of triggering receptor expressed on myeloid cells 2 (TREM2), and female sex. Here, we combine APOE4 and TREM2R47H (R47H) in female P301S tauopathy mice to identify the pathways activated when AD risk is the strongest, thereby highlighting detrimental disease mechanisms. We find that R47H induces neurodegeneration in 9- to 10-month-old female APOE4 tauopathy mice. The combination of APOE4 and R47H (APOE4-R47H) worsened hyperphosphorylated tau pathology in the frontal cortex and amplified tauopathy-induced microglial cyclic guanosine monophosphate (GMP)-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling and downstream interferon response. APOE4-R47H microglia displayed cGAS- and BAX-dependent upregulation of senescence, showing association between neurotoxic signatures and implicating mitochondrial permeabilization in pathogenesis. By uncovering pathways enhanced by the strongest AD risk factors, our study points to cGAS-STING signaling and associated microglial senescence as potential drivers of AD risk.

A Trem2R47H mouse model without cryptic splicing drives age- and disease-dependent tissue damage and synaptic loss in response to plaques.

BACKGROUND: The TREM2 R47H variant is one of the strongest genetic risk factors for late-onset Alzheimer's Disease (AD). Unfortunately, many current Trem2 R47H mouse models are associated with cryptic mRNA splicing of the mutant allele that produces a confounding reduction in protein product. To overcome this issue, we developed the Trem2R47H NSS (Normal Splice Site) mouse model in which the Trem2 allele is expressed at a similar level to the wild-type Trem2 allele without evidence of cryptic splicing products. METHODS: Trem2R47H NSS mice were treated with the demyelinating agent cuprizone, or crossed with the 5xFAD mouse model of amyloidosis, to explore the impact of the TREM2 R47H variant on inflammatory responses to demyelination, plaque development, and the brain's response to plaques. RESULTS: Trem2R47H NSS mice display an appropriate inflammatory response to cuprizone challenge, and do not recapitulate the null allele in terms of impeded inflammatory responses to demyelination. Utilizing the 5xFAD mouse model, we report age- and disease-dependent changes in Trem2R47H NSS mice in response to development of AD-like pathology. At an early (4-month-old) disease stage, hemizygous 5xFAD/homozygous Trem2R47H NSS (5xFAD/Trem2R47H NSS) mice have reduced size and number of microglia that display impaired interaction with plaques compared to microglia in age-matched 5xFAD hemizygous controls. This is associated with a suppressed inflammatory response but increased dystrophic neurites and axonal damage as measured by plasma neurofilament light chain (NfL) level. Homozygosity for Trem2R47H NSS suppressed LTP deficits and loss of presynaptic puncta caused by the 5xFAD transgene array in 4-month-old mice. At a more advanced (12-month-old) disease stage 5xFAD/Trem2R47H NSS mice no longer display impaired plaque-microglia interaction or suppressed inflammatory gene expression, although NfL levels remain elevated, and a unique interferon-related gene expression signature is seen. Twelve-month old Trem2R47H NSS mice also display LTP deficits and postsynaptic loss. CONCLUSIONS: The Trem2R47H NSS mouse is a valuable model that can be used to investigate age-dependent effects of the AD-risk R47H mutation on TREM2 and microglial function including its effects on plaque development, microglial-plaque interaction, production of a unique interferon signature and associated tissue damage.

Untangling the Role of TREM2 in Conjugation with Microglia in Neuronal Dysfunction: A Hypothesis on a Novel Pathway in the Pathophysiology of Alzheimer's Disease.

Alzheimer's disease (AD) is a complex neurodegenerative disorder involving heterogenous pathophysiological characteristics, which has become a challenge to therapeutics. The major pathophysiology of AD comprises amyloid-ß (Aß), tau, oxidative stress, and apoptosis. Recent studies indicate the significance of Triggering receptor expressed on myeloid cells 2 (TREM2) and its mutant variants in AD. TREM2 are the transmembrane receptors of microglial cells that performs a broad range of physiological cell processes. Phagocytosis of Aß is one of the physiological roles of TREM2, which plays a pivotal role in AD progression. R47H, a mutant variant of TREM2, increases the risk of AD by impairing TREM2-Aß binding. Inconclusive evidence regarding the TREM2 signaling cascade mechanism of Aß phagocytosis motivates the current review to propose a new hypothesis. The review systematically assesses the cross talk between TREM2 and other AD pathological domains and the influence of TREM2 on amyloid and tau seeding. Disease associated microglia (DAM), a novel state of microglia with unique transcriptional and functional signatures reported in neurodegenerative conditions, also depend on the TREM2 pathway for its differentiation. DAM is suggested to have a neuroprotective role. We hypothesize that TREM2, along with its signaling adaptors and endogenous proteins, play a key role in ameliorating Aß clearance. We indicate that TREM2 has the potential to ameliorate the Aß burden, though with differential clearance ability and may act as a potential therapeutic target.