Alzheimer’s disease (AD) is a brain disorder that causes progressive neurological atrophy and is the most common cause of dementia. The majority of AD cases are late onset and a clear cause is still unknown. However, there are likely different contributors including age, environment, biology, and genetics which can increase a person’s risk for developing AD.
Research advances in recent years have furthered our understanding of the pathophysiology of AD, and it has been suggested that genetic factors, such as rare variants of triggering receptor expressed on myeloid cells 2 (TREM2), strongly increase a person’s risk of developing AD.
What is TREM2?
TREM2 is a membrane protein that forms part of the TREM protein family of cell surface receptors that participate in diverse cellular processes, including inflammation, bone homeostasis, neurological development, and coagulation (1). The TREM2 gene is expressed in a subgroup of myeloid cells such as dendritic cells, granulocytes, and tissue-specific macrophages. In the brain, TREM2 is exclusively expressed by microglia. TREM2 expression in the central nervous system (CNS) is highest in the hippocampus, spinal cord, and white matter (2).
TREM2 is a receptor that recognizes a variety of ligands and can be activated by neuronal debris, anionic bacterial ligands, mammalian ligands (phosphatidylethanolamine, phosphatidylserine, or DNA, for example), and cellular proteins such as HSP60 (5).
Several functions of TREM2 have been well characterized in the last decade. For example, in vitro loss of TREM2 in microglia and macrophages leads to decreased phagocytosis of apoptotic neurons, cellular debris, and bacteria or bacterial products. Conversely, increasing TREM2 expression improves the phagocytosis rate of these substrates (2).
Figure 1. Activation of the TREM2 pathway. TREM-2 requires the adaptor protein DAP12 for downstream signaling. The TREM family of receptors regulates the activity of various cell types in the immune system, including neutrophils, monocyte/macrophages, microglia, and dendritic cells.
Disease implications
TREM2 is a mediator of neuroprotection and is associated with various neurodegenerative diseases. For example, upregulation of TREM2 expression has been observed in Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS), stroke, traumatic brain injury, and AD (2).
TREM2 mediates response to phospholipids, apolipoprotein E (APOE), and several other potential stimuli. APOE is the largest genetic risk factor for AD; however, it remains unclear whether APOE-mediated disease risk is specifically related to its interactions with TREM2.
Microglia are strongly implicated in the genetics and neuropathology of late-onset AD and several AD-risk loci near immune genes have recently been identified. Among these microglial-specific AD risk loci, variants in TREM2 confer the largest effect on disease risk, causing a similar increase in risk as one APOE ε4 allele (5).
TREM2 undergoes proteolytic cleavage by ADAM10 and ADAM17 proteins, releasing soluble TREM2 (sTREM2). sTREM2 has been detected in human cerebrospinal fluid (CSF), and its high level in individuals with AD is modulated throughout disease progression. The level of sTREM2 has been shown to correlate with the level of neuronal injury (3, 4).
Deep dive into TREM2 and Alzheimer’s
The discovery of TREM2 as a myeloid-specific AD risk gene has accelerated research into the role of microglia in AD. While TREM2 mouse models have provided some insights, the normal and disease-associated functions of TREM2 in human microglia still remain unclear.
To address this, a team of researchers led by Mathew Blurton-Jones, from the University of California Irvine in the US, generated three isogenic TREM2 knockout induced pluripotent stem cell (iPSC) lines, differentiated these cells into microglia, and examined the transcriptional and functional effects of TREM2 deletion in human microglia.
Following microglial differentiation, the loss of TREM2 expression at the protein level in all knockout lines were confirmed with western blot and Homogenous Time Resolved Fluorescence (HTRF™) analysis (Figures 2 and 3). HTRF analysis of conditioned culture medium further demonstrated secretion of sTREM2 exclusively in wild type but not in TREM2 knockout cell lines.
Results
The team reports that TREM2 knockout reduced microglial survival, impaired phagocytosis of key substrates including APOE, and inhibited SDF-1α/CXCR4-mediated chemotaxis. TREM2 deletion also reduced clustering around amyloid plaques and impaired migration toward beta-amyloid producing cultures.
Single-cell sequencing of xenotransplanted human microglia further highlighted a loss of disease-associated microglial (DAM) responses in human TREM2 knockout microglia. This was also verified by flow cytometry and immunohistochemistry.
Taken together, the results of this study provide an improved understanding of TREM2 and its downstream genetic targets. Furthermore, both conserved and novel aspects of human TREM2 biology have been identified, which likely play critical roles in the development and progression of AD.
HTRF TREM2 Assay – how does it work?
1. Assay principle
Human TREM2 is measured using a sandwich immunoassay involving two specific anti-human TREM2 antibodies, respectively labeled with Europium Cryptate (donor) and d2 (acceptor). The intensity of the signal is proportional to the concentration of the TREM2 present in the sample.
Figure 2. HTRF Assay Principle.
2. Assay protocol
The simple HTRF TREM2 assay protocol, using a 384-well small volume white plate (20 µL final), is described on the right. Cell supernatant, sample, or standard is dispensed directly into the assay plate for the detection of human TREM2. The antibodies labeled with HTRF donor and acceptor may be pre-mixed and added in a single dispensing step to further streamline the assay procedure. The assay can be run in 96- to 384-well plates by simply resizing each addition volume proportionally.
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References:
- Klesney-Tait J, Turnbull IR, Colonna M. The TREM receptor family and signal integration. Nat Immunol. 2006 Dec;7(12):1266-73.
- Gratuze M, Leyns CEG, Holtzman DM. New insights into the role of TREM2 in Alzheimer’s disease. Mol Neurodegener. 2018 Dec 20;13(1):66.
- Hammond TR, Marsh SE, Stevens B. Immune Signaling in Neurodegeneration. Immunity 2019;50(4):955–74.
- Yeh FL, Hansen DV, Sheng M. TREM2, Microglia, and Neurodegenerative Diseases. Trends in Molecular Medicine 2017;23(6):512–33.
- Kober DL, Brett TJ. TREM2-Ligand Interactions in Health and Disease. Journal of molecular biology 2017;429(11):1607–29.
- Shi Y, Holtzman DM. Interplay between innate immunity and Alzheimer disease: APOE and TREM2 in the spotlight. Nat Rev Immunol. 2018 Dec;18(12):759-772
