Multiple sclerosis (MS) is a chronic autoimmune disorder that impacts the central nervous system (CNS). It is characterized by persistent inflammation, demyelination, and neurodegeneration [1], [2]. Despite immunotherapy, MS patients experience disability due to damage to myelin and neurons [3]. Microglia, the resident immune cells of the CNS, have a significant impact on the development and progression of MS. Their involvement in the pathogenesis of MS is crucial and contributes to the complex immune responses and neuroinflammation observed in this chronic autoimmune disorder [4]. Recently, mesenchymal stem cells (MSCs) hold great promise as a potential therapeutic option for MS. MSCs have shown the ability to promote remyelination and reduce neuroinflammation, making them attractive candidates for addressing the pathophysiology of MS. Additionally, MSCs possess immunomodulatory properties and can suppress immune cell activation, creating a favorable environment within the CNS [5].

Extracellular vesicles (EVs) released by MSCs have emerged as important mediators of intercellular communication [6]. These EVs possess the remarkable ability to modulate the functions of recipient cells, thereby exerting a significant impact on various biological processes [7]. The intricate interplay between MSC-EVs and recipient cells holds great promise for advancing our understanding of cell-to-cell communication mechanisms and exploring novel therapeutic avenues for a wide range of diseases, including neuroinflammatory disorders like MS [8], [9]. Furthermore, MSC-EVs exhibit potent immunomodulatory properties, expanding their therapeutic potential beyond tissue repair and regeneration [10]. They can modulate the immune response and reduce inflammation by activating immune cells and facilitating the expansion of regulatory T cells, which play a pivotal role in suppressing excessive immune responses [12]. These findings highlight the multifaceted immunomodulatory effects of MSC-EVs and their potential as a therapeutic strategy for mitigating inflammation in MS. They can modulate the immune response and reduce inflammation by the activation of immune cells and facilitate the expansion of regulatory T cells, which play a pivotal role in suppressing excessive immune responses [11]. Also, MSC-EVs offer a safer therapeutic option compared to stem cell transplantation, as they have a lower risk of immune rejection and tumorigenesis [12], [13]. These findings highlight the multifaceted immunomodulatory effects of MSC-EVs and their potential as a therapeutic strategy for mitigating inflammation in MS and other neuroinflammatory conditions [11], [13], [14], [15].

Abnormal expression of microRNA-181 (miR-181) family members, specifically miR-181a and miR-181b, has been observed in experimental autoimmune encephalomyelitis (EAE), a well-established animal model for studying MS [16]. MiR-181 plays a crucial role in regulating immune responses and inflammation, and its dysregulated expression has been observed in various autoimmune and inflammatory diseases, including MS and EAE [17]. A recent study shows that MSC-EVs containing miR-181a trigger remyelination and reduce neuroinflammation in ulcerative colitis [18]. This can be attributed to the capacity of miR-181a to modulate the expression of target genes involved in inflammation and immune regulation. The therapeutic effects of MSC-EVs are attributed to their ability to deliver functional miRNAs to recipient cells, influencing their cellular functions [19]. EVs-enriched miR-181a holds promise for treating immune-mediated inflammatory diseases [18], [20]. In addition, the role of miR-181a levels in modulating inflammatory cytokine secretion in MS has been suggested [18], [20]. However, the specific anti-inflammatory and immunomodulatory effects of MSC-EVs containing miR-181a-5p in MS remain unclear. Further ongoing research is needed to optimize the therapeutic potential of these EVs and unravel the underlying mechanisms involved [21].

In the intricate molecular interplay of microglial responses, the USP15/RelA/NEK7/NLRP3 axis orchestrates dynamic connections between key pathways [22]. Ubiquitin-specific protease 15 (USP15) acts as a regulatory hub, interacting with nuclear factor kappa B (NF-κB) p65 subunit (RelA), NIMA-related kinase 7 (NEK7), and nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain containing 3 (NLRP3), finely orchestrating a network of inflammasome [23]. Crosstalk between these elements, alongside the involvement of apoptosis-associated speck-like protein containing a CARD (ASC) and Caspase-1, paints a comprehensive picture of the molecular landscape driving microglial responses and pyroptosis in the EAE neuroinflammation [24], [25]. NLRP3, integral to the NLRP3 inflammasome, initiates the cascade of microglial activation, while ASC serves as a crucial bridge between sensors and effectors in the inflammatory cascade [26]. Caspase-1, as a central executor, propels microglial pyroptosis, dynamically contributing to the equilibrium of the neuroinflammation [24]. Recognizing the central roles played by these proteins in the microglial landscape, our study aims to unravel the intricate molecular events governing microglial pyroptosis in neuroinflammatory disorders [27].

Our study is intricately designed to unravel the molecular mechanisms underlying the protective effects of MSC-EVs containing miR-181a-5p in neuroinflammation and demyelination, employing both in vitro and in vivo models. Specifically, we explore the therapeutic potential of these EVs in the context of EAE, a well-established animal model for MS. Significantly, we establish a link between miR-181a-5p and USP15, a molecule implicated in EAE pathogenesis, neuroinflammation, and the interferon response [28]. This groundbreaking study provides compelling evidence that MSC-EVs with miR-181a-5p can modulate the USP15/RelA/NEK7/NLRP3 axis, exerting protective effects in EAE. These findings offer crucial insights for the development of innovative therapies targeting MS and related disorders.