000K utf8 1100 2023$c2023-04-17 1500 eng 2050 urn:nbn:de:hbz:465-20240327-131550-6 2051 10.1186/s41232-023-00274-6 3000 Labusek, Nicole 3010 Bendix, Ivo 3010 Diesterbeck, Eva 3010 Felderhoff-Müser, Ursula 3010 Giebel, Bernd 3010 Hanenberg, Helmut 3010 Herz, Josephine 3010 Horn, Peter A. 3010 Köster, Christian 3010 Mouloud, Yanis 3010 Tertel, Tobias 3010 Wiek, Constanze 4000 Extracellular vesicles from immortalized mesenchymal stromal cells protect against neonatal hypoxic-ischemic brain injury [Labusek, Nicole] 4209 Background: Human mesenchymal stromal cell (MSC)-derived extracellular vesicles (EV) revealed neuroprotective potentials in various brain injury models, including neonatal encephalopathy caused by hypoxia–ischemia (HI). However, for clinical translation of an MSC-EV therapy, scaled manufacturing strategies are required, which is challenging with primary MSCs due to inter- and intra-donor heterogeneities. Therefore, we established a clonally expanded and immortalized human MSC line (ciMSC) and compared the neuroprotective potential of their EVs with EVs from primary MSCs in a murine model of HI-induced brain injury. In vivo activities of ciMSC-EVs were comprehensively characterized according to their proposed multimodal mechanisms of action. Methods: Nine-day-old C57BL/6 mice were exposed to HI followed by repetitive intranasal delivery of primary MSC-EVs or ciMSC-EVs 1, 3, and 5 days after HI. Sham-operated animals served as healthy controls. To compare neuroprotective effects of both EV preparations, total and regional brain atrophy was assessed by cresyl-violet-staining 7 days after HI. Immunohistochemistry, western blot, and real-time PCR were performed to investigate neuroinflammatory and regenerative processes. The amount of peripheral inflammatory mediators was evaluated by multiplex analyses in serum samples. Results: Intranasal delivery of ciMSC-EVs and primary MSC-EVs comparably protected neonatal mice from HI-induced brain tissue atrophy. Mechanistically, ciMSC-EV application reduced microglia activation and astrogliosis, endothelial activation, and leukocyte infiltration. These effects were associated with a downregulation of the pro-inflammatory cytokine IL-1 beta and an elevated expression of the anti-inflammatory cytokines IL-4 and TGF-beta in the brain, while concentrations of cytokines in the peripheral blood were not affected. ciMSC-EV-mediated anti-inflammatory effects in the brain were accompanied by an increased neural progenitor and endothelial cell proliferation, oligodendrocyte maturation, and neurotrophic growth factor expression. Conclusion: Our data demonstrate that ciMSC-EVs conserve neuroprotective effects of primary MSC-EVs via inhibition of neuroinflammation and promotion of neuroregeneration. Since ciMSCs can overcome challenges associated with MSC heterogeneity, they appear as an ideal cell source for the scaled manufacturing of EV-based therapeutics to treat neonatal and possibly also adult brain injury. 4950 https://doi.org/10.1186/s41232-023-00274-6$xR$3Volltext$534 4950 https://nbn-resolving.org/urn:nbn:de:hbz:465-20240327-131550-6$xR$3Volltext$534 4961 https://duepublico2.uni-due.de/receive/duepublico_mods_00079214 5051 610 5550 Exosomes 5550 Extracellular vesicles 5550 Hypoxia–ischemia 5550 Mesenchymal stromal cells 5550 Neonatal encephalopathy 5550 Neuroinflammation 5550 Neuroregeneration