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Characterization of Immune Cell-derived Extracellular Vesicles and Studying Functional Impact on Cell Environment
Published on: June 2, 2020
Identification of State-Specific Proteomic and Transcriptomic Signatures of Microglia-Derived Extracellular Vesicles
Juliet V Santiago1, Aditya Natu1, Christina C Ramelow1
1Department of Neurology, Emory University, Atlanta, Georgia, USA; Center for Neurodegenerative Diseases, Emory University, Atlanta, Georgia, USA.
Abstract:
Microglia are resident immune cells of the brain that play important roles in mediating inflammatory responses in several neurological diseases via direct and indirect mechanisms. One indirect mechanism may involve extracellular vesicle (EV) release, so that the molecular cargo transported by microglia-derived EVs can have functional effects by facilitating intercellular communication. The molecular composition of microglia-derived EVs, and how microglial activation states impact EV composition and EV-mediated effects in neuroinflammation, remain poorly understood. We hypothesize that microglia-derived EVs have unique molecular profiles that are determined by microglial activation state. Using size-exclusion chromatography to purify EVs from BV2 microglia, combined with proteomic (label-free quantitative mass spectrometry or LFQ-MS) and transcriptomic (mRNA and noncoding RNA seq) methods, we obtained comprehensive molecular profiles of microglia-derived EVs. LFQ-MS identified several classic EV proteins (tetraspanins, ESCRT machinery, and heat shock proteins), in addition to over 200 proteins not previously reported in the literature. Unique mRNA and microRNA signatures of microglia-derived EVs were also identified. After treating BV2 microglia with lipopolysaccharide (LPS), interleukin-10, or transforming growth factor beta, to mimic pro-inflammatory, anti-inflammatory, or homeostatic states, respectively, LFQ-MS and RNA seq revealed novel state-specific proteomic and transcriptomic signatures of microglia-derived EVs. Particularly, LPS treatment had the most profound impact on proteomic and transcriptomic compositions of microglia-derived EVs. Furthermore, we found that EVs derived from LPS-activated microglia were able to induce pro-inflammatory transcriptomic changes in resting responder microglia, confirming the ability of microglia-derived EVs to relay functionally relevant inflammatory signals. These comprehensive microglia-EV molecular datasets represent important resources for the neuroscience and omics communities and provide novel insights into the role of microglia-derived EVs in neuroinflammation.
Insights
Microglia release extracellular vesicles (EVs) carrying molecular cargo that influences brain inflammation. Their composition changes with microglial activation state, impacting intercellular communication in neurological diseases.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglia, the brain's immune cells, mediate neuroinflammation through various mechanisms.
- Extracellular vesicles (EVs) released by microglia are implicated in intercellular communication and disease pathogenesis.
- The molecular makeup of microglia-derived EVs and its dependence on microglial activation states are not well understood.
Purpose of the Study:
- To comprehensively profile the molecular composition of microglia-derived EVs.
- To investigate how different microglial activation states alter EV molecular signatures.
- To determine if microglia-derived EVs can transmit functional inflammatory signals.
Main Methods:
- Purification of EVs from BV2 microglia using size-exclusion chromatography.
- Proteomic analysis via label-free quantitative mass spectrometry (LFQ-MS).
- Transcriptomic analysis of mRNA and noncoding RNA sequencing.
- Treatment of microglia with lipopolysaccharide (LPS), interleukin-10, or transforming growth factor beta to induce activation states.
Main Results:
- Identification of numerous EV proteins, including novel ones not previously reported.
- Discovery of unique mRNA and microRNA signatures in microglia-derived EVs.
- Demonstration that microglial activation states, particularly LPS-induced inflammation, significantly alter EV proteomic and transcriptomic profiles.
- Evidence that EVs from activated microglia can induce pro-inflammatory changes in recipient microglia.
Conclusions:
- Microglia-derived EVs possess distinct molecular profiles influenced by microglial activation states.
- These EVs play a role in intercellular communication by transmitting functional inflammatory signals.
- The findings provide valuable molecular datasets and insights into the role of microglia-derived EVs in neuroinflammation.
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