Targeting post-ischemic thromboinflammation by mesenchymal stromal cell (MSC)-derived small extracellular vesicles

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As a consequence of their recovery-promoting actions in animal models of ischemic stroke, small extracellular vesicles (sEV) obtained from mesenchymal stromal cells (MSCs) are rapidly approaching clinical proof-of-concept studies in human stroke patients. Despite increasing efficacy data in animal models, our basic knowledge about the mechanisms of action and sites of action of MSC-sEVs remains sparse. Using mice exhibiting normolipidemia or dietary-induced hyperlipidemia, which is a clinically relevant vascular risk factor that exacerbates post-ischemic neuroinflammation, we plan to characterize immunomodulatory mechanisms mediating post-ischemic brain tissue recovery induced by MSC-sEVs in an intraluminal middle cerebral artery occlusion model. Immune responses will be characterized in the blood, cerebral microvessels and brain by flow cytometry, histochemistry and 3D light sheet microscopy. Target cells will be identified in vitro and in vivo via the uptake of dye labelled sEVs or sEVs loaded with CD63-mCherry or CRE recombinase via genetic engineering approaches. For unravelling the mechanisms of action of MSC-EVs, global gene expression profiles of isolated target cells and sEV proteome profiling will be performed. Using antibody-mediated or pharmacological studies, we will functionally deactivate platelet aggregation and/or associated immune responses, dissecting thromboinflammatory events targeted by MSC-sEVs.