A convenient model of serum-induced reactivity of human astrocytes to investigate astrocyte-derived extracellular vesicles
Journal article
Authors | White, Katherine E., Bailey, Hannah L., Shaw, Barry, Geiszler, Philippine C., Mesquita-Ribeiro, Raquel, Scott, Daniel, Layfield, Robert and Serres Sebastien |
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Abstract | Extracellular vesicles (EVs) are secreted by all cells in the CNS, including neurons and astrocytes. EVs are lipid membrane enclosed particles loaded with various bioactive cargoes reflecting the dynamic activities of cells of origin. In contrast to neurons, the specific role of EVs released by astrocytes is less well understood, partly due to the difficulty in maintaining primary astrocyte cultures in a quiescent state. The aim of this study was to establish a human serum-free astrocyte culture system that maintains primary astrocytes in a quiescent state to study the morphology, function, and protein cargoes of astrocyte-derived EVs. Serum-free medium with G5 supplement and serum-supplemented medium with 2% FBS were compared for the culture of commercially available human primary fetal astrocytes. Serum-free astrocytes displayed morphologies similar to <italic>in vivo</italic> astrocytes, and surprisingly, higher levels of astrocyte markers compared to astrocytes chronically cultured in FBS. In contrast, astrocyte and inflammatory markers in serum-free astrocytes were upregulated 24 h after either acute 2% FBS or cytokine exposure, confirming their capacity to become reactive. Importantly, this suggests that distinct signaling pathways are involved in acute and chronic astrocyte reactivity. Despite having a similar morphology, chronically serum-cultured astrocyte-derived EVs (ADEVs) were smaller in size compared to serum-free ADEVs and could reactivate serum-free astrocytes. Proteomic analysis identified distinct protein datasets for both types of ADEVs with enrichment of complement and coagulation cascades for chronically serum-cultured astrocyte-derived EVs, offering insights into their roles in the CNS. Collectively, these results suggest that human primary astrocytes cultured in serum-free medium bear similarities with <italic>in vivo</italic> quiescent astrocytes and the addition of serum induces multiple morphological and transcriptional changes that are specific to human reactive astrocytes and their ADEVs. Thus, more emphasis should be made on using multiple structural, molecular, and functional parameters when evaluating ADEVs as biomarkers of astrocyte health. |
Keywords | Extracellular vesicles (EVs); cells of origin ; human primary fetal astrocytes |
Year | 2024 |
Journal | Frontiers in Cellular Neuroscience |
Journal citation | 18, pp. 1-18 |
Publisher | Frontiers |
ISSN | 1662-5102 |
Digital Object Identifier (DOI) | https://doi.org/10.3389/fncel.2024.1414142 |
Web address (URL) | https://www.frontiersin.org/journals/cellular-neuroscience/articles/10.3389/fncel.2024.1414142/full |
Publisher's version | License File Access Level Open |
Output status | Published |
Publication dates | |
Online | 10 Jun 2024 |
Publication process dates | |
Deposited | 01 Jul 2024 |
Supplemental file | File Access Level Open |
https://repository.derby.ac.uk/item/q7098/a-convenient-model-of-serum-induced-reactivity-of-human-astrocytes-to-investigate-astrocyte-derived-extracellular-vesicles
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Publisher's version
fncel-18-1414142.pdf | ||
License: CC BY 4.0 | ||
File access level: Open |
Supplemental file
(EV_astro)_White_2024_suppl.zip | ||
File access level: Open |
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