Mesenchymal Stem Cell-Derived Extracellular Vesicles as Mediators of Anti-Inflammatory Effects: Endorsement of Macrophage Polarization

TLDR

Mesenchymal stem cells promote tissue repair mainly through paracrine signaling, and recent evidence shows that extracellular vesicles they release also contribute to this therapeutic effect. This study characterizes extracellular vesicles from human adipose‑derived MSCs to determine their capacity to modulate macrophage polarization and mediate anti‑inflammatory effects. EVs were isolated by repeated ultracentrifugation of conditioned medium from MSCs cultured under normoxic or hypoxic conditions, yielding EVNormo and EVHypo preparations. Both EVNormo and EVHypo were taken up by macrophages, inducing a shift from M1 to M2 phenotype, reducing IL‑6 and Nos2 while increasing Arg1, Ym1, and CD206⁺ cells, and enhancing myogenic markers Pax7, MyoD, and eMyhc, with hypoxic EVs exerting stronger effects, confirming MSC‑EVs as potent anti‑inflammatory agents.

Abstract

Abstract Mesenchymal Stem Cells (MSCs) are effective therapeutic agents enhancing the repair of injured tissues mostly through their paracrine activity. Increasing evidences show that besides the secretion of soluble molecules, the release of extracellular vesicles (EVs) represents an alternative mechanism adopted by MSCs. Since macrophages are essential contributors toward the resolution of inflammation, which has emerged as a finely orchestrated process, the aim of the present study was to carry out a detailed characterization of EVs released by human adipose derived-MSCs to investigate their involvement as modulators of MSC anti-inflammatory effects inducing macrophage polarization. The EV-isolation method was based on repeated ultracentrifugations of the medium conditioned by MSC exposed to normoxic or hypoxic conditions (EVNormo and EVHypo). Both types of EVs were efficiently internalized by responding bone marrow-derived macrophages, eliciting their switch from a M1 to a M2 phenotype. In vivo, following cardiotoxin-induced skeletal muscle damage, EVNormo and EVHypo interacted with macrophages recruited during the initial inflammatory response. In injured and EV-treated muscles, a downregulation of IL6 and the early marker of innate and classical activation Nos2 were concurrent to a significant upregulation of Arg1 and Ym1, late markers of alternative activation, as well as an increased percentage of infiltrating CD206pos cells. These effects, accompanied by an accelerated expression of the myogenic markers Pax7, MyoD, and eMyhc, were even greater following EVHypo administration. Collectively, these data indicate that MSC-EVs possess effective anti-inflammatory properties, making them potential therapeutic agents more handy and safe than MSCs.