The influence of the R47H triggering receptor expressed on myeloid cells 2 variant on microglial exosome profiles

Abstract

Variants in the triggering receptor expressed on myeloid cells 2 (TREM2) gene are linked with an increased risk of dementia, in particular the R47Hhet TREM2 variant is linked to late-onset Alzheimer’s disease. Using human iPSC-derived microglia, we assessed whether variations in the dynamics of exosome secretion, including their components, from these cells might underlie some of this risk. We found exosome size was not altered between common variant controls and R47Hhet variants, but the amount and constitution of exosomes secreted were different. Exosome quantities were rescued by incubation with an ATP donor or with lipids via a phosphatidylserine TREM2 ligand. Following a lipopolysaccharide or phagocytic cell stimulus, exosomes from common variant and R47Hhet microglia were found to contain cytokines, chemokines, APOE and TREM2. Differences were observed in the expression of CCL22, IL-1β and TREM2 between common variant and R47Hhet derived exosomes. Furthermore unlike common variant-derived exosomes, R47Hhet exosomes contained additional proteins linked to negative regulation of transcription and metabolic processes. Subsequent addition of exosomes to stressed neurones showed R47Hhet-derived exosomes to be less protective. These data have ramifications for the responses of microglia in Alzheimer’s disease and may point to further targets for therapeutic intervention.

Differential stimulation of pluripotent stem cell-derived human microglia leads to exosomal proteomic changes affecting neurons

“…Whilst exosomes from basal Cv iPS-Mg contained higher levels of proteins involved in cell growth and/or maintenance, R47H het basal exosomes contained more transcription-related proteins. This is consistent with our previous finding that the most abundant proteins in exosomes from R47H het iPS-Mg were involved in the negative regulation of transcription [22].…”

“…Exosomes were extracted from supernatant with an ExoQuick kit (System Biosciences). The presence of exosomal markers and size distribution of the particles was confirmed previously [22]. Extracted exosomes were lysed in RIPA buffer and their protein content determined by BCA analysis.…”

“…previously [14,15]. In our previous study, we reported the top 200 most abundant and significantly changed exosomal proteins secreted from iPS-Mg [22], however we did not…”

“…Analysis was performed on pooled control lines and pooled R47H het cell lines. As described previously [22], proteomic data were log10 transformed prior to analysis.…”

“…We have recently shown that the R47H het TREM2 variant decreased the secretion of exosomes from patient-derived induced pluripotent stem cell-derived microglia (iPS-Mg) [22]. The R47H het TREM2 variant also affected the ability of exosomes to rescue neurons from cell death compared with exosomes secreted from common variant (Cv) TREM2 expressing iPS-Mg [22].…”