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

ABSTRACT: 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 A?-S8C dimer highlighted metabolic pathways, phagosome and immune response as the most perturbed pathways in AD neuronal cultures. Through the construction of an A?-induced gene regulatory network, we were able to identify an A? 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.

SUBMITTER: Martins S

PROVIDER: S-EPMC7350255 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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

Martins Soraia S Müller-Schiffmann Andreas A Erichsen Lars L Bohndorf Martina M Wruck Wasco W Sleegers Kristel K Van Broeckhoven Christine C Korth Carsten C Adjaye James J

International journal of molecular sciences 20200625 12

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 ca ...[more]

PMID: 32630447

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Project description:BackgroundMolecular characterization of late-onset Alzheimer's disease (LOAD), the leading cause of age-related dementia, has revealed transcripts, proteins, and pathway alterations associated with disease. Assessing these postmortem signatures of LOAD in experimental model systems can further elucidate their relevance to disease origins and progression. Model organisms engineered with human genetic factors further link these signatures to disease-associated variants, especially when studies are designed to leverage homology across species. Here we assess differential gene splicing patterns in aging mouse models carrying humanized APOE4 and/or the Trem2*R47H variant on a C57BL/6J background. We performed a differential expression of gene (DEG) and differential splicing analyses on whole brain transcriptomes at multiple ages. To better understand the difference between differentially expressed and differentially spliced genes, we evaluated enrichment of KEGG pathways and cell-type specific gene signatures of the adult brain from each alteration type. To determine LOAD relevance, we compared differential splicing results from mouse models with multiple human AD splicing studies.ResultsWe found that differentially expressed genes in Trem2*R47H mice were significantly enriched in multiple AD-related pathways, including immune response, osteoclast differentiation, and metabolism, whereas differentially spliced genes were enriched for neuronal related functions, including GABAergic synapse and glutamatergic synapse. These results were reinforced by the enrichment of microglial genes in DEGs and neuronal genes in differentially spliced genes in Trem2*R47H mice. We observed significant overlap between differentially spliced genes in Trem2*R47H mice and brains from human AD subjects. These effects were absent in APOE4 mice and suppressed in APOE4.Trem2*R47H double mutant mice relative to Trem2*R47H mice.ConclusionsThe cross-species observation that alternative splicing observed in LOAD are present in Trem2*R47H mouse models suggests a novel link between this candidate risk gene and molecular signatures of LOAD in neurons and demonstrates how deep molecular analysis of new genetic models links molecular disease outcomes to a human candidate gene.

Project description:BackgroundTREM2 is a microglial cell surface receptor, with risk mutations linked to Alzheimer's disease (AD), including R47H. TREM2 signalling via SYK aids phagocytosis, chemotaxis, survival, and changes to microglial activation state. In AD mouse models, knockout (KO) of TREM2 impairs microglial clustering around amyloid and prevents microglial activation. The R47H mutation is proposed to reduce TREM2 ligand binding. We investigated cell phenotypes of the R47H mutant and TREM2 KO in a model of human microglia, and compared their transcriptional signatures, to determine the mechanism by which R47H TREM2 disrupts function.MethodsWe generated human microglia-like iPSC-macrophages (pMac) from isogenic induced pluripotent stem cell (iPSC) lines, with homozygous R47H mutation or TREM2 knockout (KO). We firstly validated the effect of the R47H mutant on TREM2 surface and subcellular localization in pMac. To assess microglial phenotypic function, we measured phagocytosis of dead neurons, cell morphology, directed migration, survival, and LPS-induced inflammation. We performed bulk RNA-seq, comparing significant differentially expressed genes (DEGs; p < 0.05) between the R47H and KO versus WT, and bioinformatically predicted potential upstream regulators of TREM2-mediated gene expression.ResultsR47H modified surface expression and shedding of TREM2, but did not impair TREM2-mediated signalling, or gross phenotypes that were dysregulated in the TREM2 KO (phagocytosis, motility, survival). However, altered gene expression in the R47H TREM2 pMac overlapped by 90% with the TREM2 KO and was characterised by dysregulation of genes involved with immunity, proliferation, activation, chemotaxis, and adhesion. Downregulated mediators of ECM adhesion included the vitronectin receptor αVβ3, and consequently, R47H TREM2 pMac adhered weakly to vitronectin compared with WT pMac. To counteract these transcriptional defects, we investigated TGFβ1, as a candidate upstream regulator. TGFβ1 failed to rescue vitronectin adhesion of pMac, although it improved αVβ3 expression.ConclusionsThe R47H mutation is not sufficient to cause gross phenotypic defects of human pMac under standard culture conditions. However, overlapping transcriptional defects with TREM2 KO supports the hypothesised partial loss-of-function effects of the R47H mutation. Furthermore, transcriptomics can guide us to more subtle phenotypic defects in the R47H cells, such as reduced cell adhesion, and can be used to predict targets for therapeutic intervention.

Dataset Information

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

Publications

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

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