Jason D. Ulrich

TREM2 Lipid Sensing Sustains the Microglial Response in an Alzheimer’s Disease Model

Summary Triggering receptor expressed on myeloid cells 2 (TREM2) is a microglia surface receptor that triggers intracellular protein tyrosine phosphorylation. Recent genome-wide association studies have shown that a rare R47H mutation of TREM2 correlates with a substantial increase in the risk of developing Alzheimers disease (AD). To address the basis for this genetic association, we studied TREM2 deficiency in the 5XFAD mouse model of AD. We found that TREM2 deficiency and haploinsufficiency augment β-amyloid (Aβ) accumulation due to dysfunctional response of microglia, which become apoptotic and fail to cluster around Aβ plaques. We further demonstrate that TREM2 senses a broad array of anionic and zwitterionic lipids known to associate with fibrillar Aβ in lipid membranes and to be exposed on the surface of damaged neurons. Remarkably, the R47H mutation impairs TREM2 detection of lipid ligands. Thus, TREM2 detects damage-associated lipid patterns associated with neurodegeneration, sustaining microglia response to Aβ accumulation.

TREM2-mediated early microglial response limits diffusion and toxicity of amyloid plaques

Wang et al. report that TREM2 protects mice from Alzheimers disease by enabling resident microglia to insulate and alter Aβ plaque structure, thereby limiting neuritic damage.

Altered microglial response to Aβ plaques in APPPS1-21 mice heterozygous for TREM2.

BackgroundRecent genome-wide association studies linked variants in TREM2 to a strong increase in the odds of developing Alzheimer’s disease. The mechanism by which TREM2 influences the susceptibility to Alzheimer’s disease is currently unknown. TREM2 is expressed by microglia and is thought to regulate phagocytic and inflammatory microglial responses to brain pathology. Given that a single allele of variant TREM2, likely resulting in a loss of function, conferred an increased risk of developing Alzheimer’s disease, we tested whether loss of one functional trem2 allele would affect Aβ plaque deposition or the microglial response to Aβ pathology in APPPS1-21 mice.ResultsThere was no significant difference in Aβ deposition in 3-month old or 7-month old APPPS1-21 mice expressing one or two copies of trem2. However, 3-month old mice with one copy of trem2 exhibited a marked decrease in the number and size of plaque-associated microglia. While there were no statistically significant differences in cytokine levels or markers of microglial activation in 3- or 7-month old animals, there were trends towards decreased expression of NOS2, C1qa, and IL1a in 3-month old TREM2+/− vs. TREM2+/+ mice.ConclusionsLoss of a single copy of trem2 had no effect on Aβ pathology, but altered the morphological phenotype of plaque-associated microglia. These data suggest that TREM2 is important for the microglial response to Aβ deposition but that a 50% decrease inTREM2 expression does not affect Aβ plaque burden.

In vivo measurement of apolipoprotein E from the brain interstitial fluid using microdialysis

BackgroundThe APOE4 allele variant is the strongest known genetic risk factor for developing late-onset Alzheimer’s disease. The link between apolipoprotein E (apoE) and Alzheimer’s disease is likely due in large part to the impact of apoE on the metabolism of amyloid β (Aβ) within the brain. Manipulation of apoE levels and lipidation within the brain has been proposed as a therapeutic target for the treatment of Alzheimer’s disease. However, we know little about the dynamic regulation of apoE levels and lipidation within the central nervous system. We have developed an assay to measure apoE levels in the brain interstitial fluid of awake and freely moving mice using large molecular weight cut-off microdialysis probes.ResultsWe were able to recover apoE using microdialysis from human cerebrospinal fluid (CSF) in vitro and mouse brain parenchyma in vivo. Microdialysis probes were inserted into the hippocampus of wild-type mice and interstitial fluid was collected for 36 hours. Levels of apoE within the microdialysis samples were determined by ELISA. The levels of apoE were found to be relatively stable over 36 hours. No apoE was detected in microdialysis samples from apoE KO mice. Administration of the RXR agonist bexarotene increased ISF apoE levels while ISF Aβ levels were decreased. Extrapolation to zero-flow analysis allowed us to determine the absolute recoverable concentration of apoE3 in the brain ISF of apoE3 KI mice. Furthermore, analysis of microdialysis samples by non-denaturing gel electrophoresis determined lipidated apoE particles in microdialysis samples were consistent in size with apoE particles from CSF. Finally, we found that the concentration of apoE in the brain ISF was dependent upon apoE isoform in human apoE KI mice, following the pattern apoE2>apoE3>apoE4.ConclusionsWe are able to collect lipidated apoE from the brain of awake and freely moving mice and monitor apoE levels over the course of several hours from a single mouse. Our technique enables assessment of brain apoE dynamics under physiological and pathophysiological conditions and in response to therapeutic interventions designed to affect apoE levels and lipidation within the brain.

Novel allele-dependent role for APOE in controlling the rate of synapse pruning by astrocytes

Significance Susceptibility to Alzheimer’s disease (AD) is strongly controlled by apolipoprotein E (APOE) genotype. The E4 allele greatly increases risk whereas the E2 allele decreases risk, but it is not known how the APOE allele controls AD risk. In this paper, we report a novel role for APOE by showing that APOE2 enhances and APOE4 decreases the rate of synapse pruning and turnover in the brain by astrocytes. We also show that APOE alleles control the rate of accumulation of the complement C1q protein, which we hypothesize, reflects senescent synapse accumulation during normal brain aging and vulnerability of the aging brain to neurodegenerative diseases such as AD. The strongest genetic risk factor influencing susceptibility to late-onset Alzheimer’s disease (AD) is apolipoprotein E (APOE) genotype. APOE has three common isoforms in humans, E2, E3, and E4. The presence of two copies of the E4 allele increases risk by ∼12-fold whereas E2 allele is associated with an ∼twofold decreased risk for AD. These data put APOE central to AD pathophysiology, but it is not yet clear how APOE alleles modify AD risk. Recently we found that astrocytes, a major central nervous system cell type that produces APOE, are highly phagocytic and participate in normal synapse pruning and turnover. Here, we report a novel role for APOE in controlling the phagocytic capacity of astrocytes that is highly dependent on APOE isoform. APOE2 enhances the rate of phagocytosis of synapses by astrocytes, whereas APO4 decreases it. We also found that the amount of C1q protein accumulation in hippocampus, which may represent the accumulation of senescent synapses with enhanced vulnerability to complement-mediated degeneration, is highly dependent on APOE alleles: C1q accumulation was significantly reduced in APOE2 knock-in (KI) animals and was significantly increased in APOE4 KI animals compared with APOE3 KI animals. These studies reveal a novel allele-dependent role for APOE in regulating the rate of synapse pruning by astrocytes. They also suggest the hypothesis that AD susceptibility of APOE4 may originate in part from defective phagocytic capacity of astrocytes which accelerates the rate of accumulation of C1q-coated senescent synapses, enhancing synaptic vulnerability to classical-complement-cascade mediated neurodegeneration.

Elucidating the Role of TREM2 in Alzheimer’s Disease

Alzheimers disease (AD) is the sixth leading cause of death in the United States and the most common cause of dementia in the elderly. Genetic factors, such as rare variants in the microglial-expressed gene TREM2, strongly impact the lifetime risk of developing AD. Several recent studies have described dramatic TREM2-dependent phenotypes in mouse models of amyloidosis that point to an important role for TREM2 in regulating the response of the innate immune system to Aβ pathology. Furthermore, elevations in the CSF levels of soluble TREM2 fragments implicate changes in inflammatory pathways as occurring coincident with markers of neuronal damage and the onset of clinical dementia in AD. Here, we review the rapidly developing literature surrounding TREM2 in AD that may provide novel insight into the broader role of the innate immune system in neurodegenerative disease.

TREM2 Function in Alzheimer’s Disease and Neurodegeneration

Alzheimers disease (AD), the most common cause of dementia in the elderly, is a complex neurodegenerative disease marked by the appearance of amyloid-β (Aβ) plaques and hyperphosphorylated tau tangles. Alzheimers disease has a strong genetic component, and recent advances in genome technology have unearthed novel variants in several genes, which could provide insight into the pathogenic mechanisms that contribute to AD. Particularly interesting are variants in the microglial-expressed receptor TREM2 which are associated with a 2-4-fold increased risk of developing AD. Since the discovery of a link between TREM2 and AD, multiple studies have emerged testing whether partial or complete loss of TREM2 function contributed to Aβ deposition or Aβ-associated microgliosis. Although some confounding conflicting data have emerged from these studies regarding the role of TREM2 in regulating Aβ deposition within the hippocampus, the most consistent and striking observation is a strong decrease in microgliosis surrounding Aβ plaques in TREM2 haploinsufficient and TREM2 deficient mice. Interestingly, a similar impairment in microgliosis has been reported in mouse models of prion disease, stroke, and multiple sclerosis, suggesting a critical role for TREM2 in supporting microgliosis in response to pathology in the central nervous system. In this Review, we summarize recent reports on the role of TREM2 in AD pathology and hypothesized mechanisms by which TREM2 function could influence AD-induced microgliosis.

TREM2 deficiency attenuates neuroinflammation and protects against neurodegeneration in a mouse model of tauopathy

Significance Alzheimer’s disease (AD) is the most common cause of dementia and is a major public health problem for which there is currently no disease-modifying treatment. There is an urgent need for greater understanding of the molecular mechanisms underlying neurodegeneration in patients to create better therapeutic options. Recently, genetic studies uncovered novel AD risk variants in the microglial receptor, triggering receptor expressed on myeloid cells 2 (TREM2). Previous studies suggested that loss of TREM2 function worsens amyloid-β (Aβ) plaque-related toxicity. In contrast, we observe TREM2 deficiency mitigates neuroinflammation and protects against brain atrophy in the context of tau pathology. These findings indicate dual roles for TREM2 and microglia in the context of amyloid versus tau pathology, which are important to consider for potential treatments targeting TREM2. Variants in the gene encoding the triggering receptor expressed on myeloid cells 2 (TREM2) were recently found to increase the risk for developing Alzheimer’s disease (AD). In the brain, TREM2 is predominately expressed on microglia, and its association with AD adds to increasing evidence implicating a role for the innate immune system in AD initiation and progression. Thus far, studies have found TREM2 is protective in the response to amyloid pathology while variants leading to a loss of TREM2 function impair microglial signaling and are deleterious. However, the potential role of TREM2 in the context of tau pathology has not yet been characterized. In this study, we crossed Trem2+/+ (T2+/+) and Trem2−/− (T2−/−) mice to the PS19 human tau transgenic line (PS) to investigate whether loss of TREM2 function affected tau pathology, the microglial response to tau pathology, or neurodegeneration. Strikingly, by 9 mo of age, T2−/−PS mice exhibited significantly less brain atrophy as quantified by ventricular enlargement and preserved cortical volume in the entorhinal and piriform regions compared with T2+/+PS mice. However, no TREM2-dependent differences were observed for phosphorylated tau staining or insoluble tau levels. Rather, T2−/−PS mice exhibited significantly reduced microgliosis in the hippocampus and piriform cortex compared with T2+/+PS mice. Gene expression analyses and immunostaining revealed microglial activation was significantly attenuated in T2−/−PS mice, and there were lower levels of inflammatory cytokines and astrogliosis. These unexpected findings suggest that impairing microglial TREM2 signaling reduces neuroinflammation and is protective against neurodegeneration in the setting of pure tauopathy.

Anti-tau antibody administration increases plasma tau in transgenic mice and patients with tauopathy

Administration of an anti-tau antibody to transgenic mice expressing human tau and to patients with tauopathy increased the concentration of tau in plasma. Tracking tau in mice and humans Tauopathies, such as progressive supranuclear palsy and Alzheimer’s disease, are a group of neurodegenerative diseases characterized by the accumulation of aggregated forms of tau protein in the brain. Administration of anti-tau antibodies is a new treatment approach being tested for these diseases. Tau is present at high levels in the brain and low levels in the plasma. Peripheral administration of an anti-tau antibody markedly increased tau in the plasma of both transgenic mice expressing human tau and patients with tauopathy (Yanamandra et al.). The increase in plasma tau in mice correlated with an increase in brain extracellular and soluble tau. Tauopathies are a group of disorders in which the cytosolic protein tau aggregates and accumulates in cells within the brain, resulting in neurodegeneration. A promising treatment being explored for tauopathies is passive immunization with anti-tau antibodies. We previously found that administration of an anti-tau antibody to human tau transgenic mice increased the concentration of plasma tau. We further explored the effects of administering an anti-tau antibody on plasma tau. After peripheral administration of an anti-tau antibody to human patients with tauopathy and to mice expressing human tau in the central nervous system, there was a dose-dependent increase in plasma tau. In mouse plasma, we found that tau had a short half-life of 8 min that increased to more than 3 hours after administration of anti-tau antibody. As tau transgenic mice accumulated insoluble tau in the brain, brain soluble and interstitial fluid tau decreased. Administration of anti-tau antibody to tau transgenic mice that had decreased brain soluble tau and interstitial fluid tau resulted in an increase in plasma tau, but this increase was less than that observed in tau transgenic mice without these brain changes. Tau transgenic mice subjected to acute neuronal injury using 3-nitropropionic acid showed increased interstitial fluid tau and plasma tau. These data suggest that peripheral administration of an anti-tau antibody results in increased plasma tau, which correlates with the concentration of extracellular and soluble tau in the brain.

Behavioral and transcriptomic analysis of Trem2-null mice: not all knockout mice are created equal

Abstract It is clear that innate immune system status is altered in numerous neurodegenerative diseases. Human genetic studies have demonstrated that triggering receptor expressed in myeloid cells 2 (TREM2) coding variants have a strong association with Alzheimer’s disease (AD) and other neurodegenerative diseases. To more thoroughly understand the impact of TREM2 in vivo, we studied the behavioral and cognitive functions of wild-type (WT) and Trem2−/− (KO) mice during basal conditions and brain function in the context of innate immune stimulation with peripherally administered lipopolysaccharide (LPS). Early markers of neuroinflammation preceded Aif1 and Trem2 upregulation that occurred at later stages (24–48 h post-LPS). We performed a transcriptomic study of these cohorts and found numerous transcripts and pathways that were altered in Trem2−/− mice both at baseline and 48 h after LPS challenge. Importantly, our transcriptome analysis revealed that our Trem2−/− mouse line (Velocigene allele) results in exaggerated Treml1 upregulation. In contrast, aberrantly high Treml1 expression was absent in the Trem2 knockout line generated by the Colonna lab and the Jackson Labs CRISPR/Cas9 Trem2 knockout line. Notably, removal of the floxed neomycin selection cassette ameliorated aberrant Treml1 expression, validating the artifactual nature of Treml1 expression in the original Trem2−/− Velocigene line. Clearly further studies are needed to decipher whether the Treml1 transcriptional artifact is functionally meaningful, but our data indicate that caution is warranted when interpreting functional studies with this particular line. Additionally, our results indicate that other Velocigene alleles or targeting strategies with strong heterologous promoters need to carefully consider downstream genes.