Lipids are increasingly recognized as bioactive mediators of extracellular vesicle (EV)
functions. However, while EV proteins and nucleic acids are well described, EV lipids are insufficiently
understood due to lack of adequate quantitative methods. We adapted an established
targeted and quantitative mass spectrometry (LC-MS/MS) method originally developed for analysis
of 94 eicosanoids and seven polyunsaturated fatty acids (PUFA) in human plasma. Additionally, the
influence of freeze–thaw (FT) cycles, injection volume, and extraction solvent were investigated. The
modified protocol was applied to lipidomic analysis of differently polarized macrophage-derived EVs.
We successfully quantified three PUFAs and eight eicosanoids within EVs. Lipid extraction showed
reproducible PUFA and eicosanoid patterns. We found a particularly high impact of FT cycles on EV
lipid profiles, with significant reductions of up to 70%. Thus, repeated FT will markedly influence
analytical results and may alter EV functions, emphasizing the importance of a standardized sample
pretreatment protocol for the analysis of bioactive lipids in EVs. EV lipid profiles differed largely
depending on the polarization of the originating macrophages. Particularly, we observed major
changes in the arachidonic acid pathway. We emphasize the importance of a standardized sample
pretreatment protocol for the analysis of bioactive lipids in EVs.
| Identifer | oai:union.ndltd.org:DRESDEN/oai:qucosa:de:qucosa:85940 |
| Date | 09 June 2023 |
| Creators | Reinicke, Madlen, Shamkeeva, Saikal, Hell, Max, Isermann, Berend, Ceglarek, Uta, Heinemann, Mitja L. |
| Publisher | MDPI |
| Source Sets | Hochschulschriftenserver (HSSS) der SLUB Dresden |
| Language | English |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, doc-type:article, info:eu-repo/semantics/article, doc-type:Text |
| Rights | info:eu-repo/semantics/openAccess |
| Relation | 1319 |
Page generated in 0.0024 seconds