Mesenchymal Stem Cell (MSC)–Derived Extracellular Vesicles Protect from Neonatal Stroke by Interacting with Microglial Cells

Abstract

Mesenchymal stem cell (MSC)-based therapies are beneficial in models of perinatal stroke and hypoxia-ischemia. Mounting evidence suggests that in adult injury models, including stroke, MSC-derived small extracellular vesicles (MSC-sEV) contribute to the neuroprotective and regenerative effects of MSCs. Herein, we examined if MSC-sEV protect neonatal brain from stroke and if this effect is mediated via communication with microglia. MSC-sEV derived from bone marrow MSCs were characterized by size distribution (NanoSight™) and identity (protein markers). Studies in microglial cells isolated from the injured or contralateral cortex of postnatal day 9 (P9) mice subjected to a 3-h middle cerebral artery occlusion (tMCAO) and cultured (in vitro) revealed that uptake of fluorescently labeled MSC-sEV was significantly greater by microglia from the injured cortex vs. contralateral cortex. The cell-type-specific spatiotemporal distribution of MSC-sEV was also determined in vivo after tMCAO at P9. MSC-sEV administered at reperfusion, either by intracerebroventricular (ICV) or by intranasal (IN) routes, accumulated in the hemisphere ipsilateral to the occlusion, with differing spatial distribution 2 h, 18 h, and 72 h regardless of the administration route. By 72 h, MSC-sEV in the IN group was predominantly observed in Iba1 + cells with retracted processes and in GLUT1 + blood vessels in ischemic-reperfused regions. MSC-sEV presence in Iba1 + cells was sustained. MSC-sEV administration also significantly reduced injury volume 72 h after tMCAO in part via modulatory effects on microglial cells. Together, these data establish feasibility for MSC-sEV delivery to injured neonatal brain via a clinically relevant IN route, which affords protection during sub-acute injury phase.

Switching Roles: Beneficial Effects of Adipose Tissue-Derived Mesenchymal Stem Cells on Microglia and Their Implication in Neurodegenerative Diseases

“…They even reversed memory loss in AD mouse models [ 66 ]. These effects are similar to those induced by microglia-derived exosomes in recipient microglial cells [ 93 ]; accordingly, ASC- and BM-MSC-derived exosomes are mainly accumulated in microglia and, to a lesser extent, in astrocytes and neurons [ 81 , 94 ]. This points toward a microglia-mediated reduction of neuroinflammatory parameters, supported by studies that show that ASC-EV provoke a decrease in pro-inflammatory markers, such as CD68, inducible nitric oxide synthase (iNOS) and a variety of pro-inflammatory cytokines, and an increase in arginase-1 expression, switching microglia toward a neuroprotective phenotype with a more ramified cell shape [ 75 , 79 , 81 ].…”

Pharmacokinetics and biodistribution of extracellular vesicles administered intravenously and intranasally to Macaca nemestrina

“…Previously, studies of intranasally administered recombinant vesicular stomatitis viral vectors 65 and a nanogel pneumococcal vaccine formulation 66 in macaques found no brain uptake. Certainly, numerous studies report that IN EVs or their presumed cargo enter the brain parenchyma (see, for example [67][68][69][70][71][72] ). As a result, intranasal delivery of EVs is thought to be a promising way to treat CNS disease (reviewed in 73 ).…”

“…Physiology of the recipient species or characteristics of the source cells could explain disparate results. For example, many studies relied on various brain injury models (tumors, stroke, brain injury, morphine treatment) which may enhance the capacity to take up EVs compared with healthy animals 67,69,72 . Additionally, EV uptake and signal uptake may not overlap completely depending on EV separation technique.…”

“…Our results suggest that IN delivery of EVs to the brain in large animals should not be a foregone conclusion. Certainly, numerous studies report that IN EVs or their presumed cargo enter the brain parenchyma (see, for example [60][61][62][63][64][65] ). As a result, intranasal delivery of EVs is thought to be a promising way to treat CNS disease.…”

Functional Recovery Caused by Human Adipose Tissue Mesenchymal Stem Cell-Derived Extracellular Vesicles Administered 24 h after Stroke in Rats

“…Although MSC therapy seems promising, improvements have been made to optimize their effectiveness [ 25 ]. It is currently thought that the protective effects of MSCs are due to the release of extracellular vesicles (EVs) [ 26 , 27 , 28 ]. EVs are small double-membrane vesicles (30–200 nm) released by many cell types involved in mediating cell-to-cell communication under physiological conditions [ 29 , 30 , 31 , 32 ].…”