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Advanced Neurology Cytokine response to EV therapy in SCI

1. Introduction SCI, offering targeted effect at the site of inflammation
and reducing systemic side effects. However, their use
Spinal cord injury (SCI) is one of the most serious requires the development of an optimal delivery system
public health concerns, as it often results in permanent that ensures prolonged and controlled release of EVs at the
disability. Primary mechanical damage to nerve tissue injury site. One promising approach is the use of a fibrin
triggers a cascade of secondary pathological reactions, matrix (FM), which is widely employed in clinical practice
with neuroinflammation playing a key role. A crucial due to its biocompatibility and natural biodegradability.
component of this process is the excessive production of FM serves as a biocompatible and biodegradable scaffold
pro-inflammatory cytokines, which exacerbates neural that enables sustained release of encapsulated MSCs and
tissue damage. Analyses of cerebrospinal fluid and blood their exosomes. 8-11 In addition, its intrinsic bioactivity may
samples from patients with SCI have revealed pronounced synergistically contribute to inflammation modulation,
alterations in the cytokine profile, which are important for thereby improving the therapeutic efficacy of MSC-EVs.
predicting clinical outcomes and evaluating therapeutic
efficacy. Therefore, approaches to post-traumatic spinal Our previous study demonstrated that the application
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cord repair should incorporate anti-inflammatory effects. of MSC-EVs promoted the preservation of mature
oligodendrocytes and improved functional outcomes in rats
The use of mesenchymal stem cells (MSCs) and/or with SCI. However, no studies have thoroughly evaluated
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their paracrine mediators encapsulated in extracellular the effects of combining MSC-EVs with FM on both pro-
vesicles (MSC-EVs) is considered a promising therapeutic inflammatory and anti-inflammatory cytokine levels in
strategy. Studies using a rat model of SCI have shown that SCI. Chronic inflammation persisting beyond the acute
systemic administration of MSC-EVs reduces levels of pro- injury phase contributes to ongoing tissue damage and
inflammatory cytokines such as tumor necrosis factor (TNF)-α impedes functional recovery. 13-15 Targeting inflammatory
and interleukin (IL)-1β, while increasing the production of processes at the chronic stage is therefore critical for
anti-inflammatory molecules like IL-10. In this context, EVs promoting regenerative mechanisms and improving long-
regulate inflammatory factors through multiple mechanisms, term outcomes. Understanding how therapies modulate
such as the delivery of miRNAs and proteins. In addition, inflammation during this phase is essential for developing
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beyond influencing apoptosis and neuroinflammation, effective treatments for SCI. The present study aimed to
systemic administration of MSC-EVs has also been shown to investigate the impact of co-administering MSC-EVs with
modulate angiogenesis, thereby promoting tissue healing by FM on cytokine levels in the rat spinal cord during the
accelerating the elimination of inflammatory mediators and chronic phase of SCI.
attracting immune cells to the injury site. 4
A study by Romanelli et al. (2019) examined the long- 2. Materials and methods
term effects of intravenously administered EVs in a rat All procedures were designed to minimize animal use and
model of spinal contusion. The study found that during the reduce the severity of interventions. The experimental
chronic phase of SCI, both the experimental and control design is summarized in Figure 1.
groups continued to lose neural tissue. However, by the end
of the experiment, significantly more tissue was preserved 2.1. Isolation and cultivation of MSCs
in the EV-treated group compared to the control group. In MSCs were obtained from the adipose tissue of female
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a follow-up study, intraspinal administration of EVs led to a Wistar rats weighing 250–300 g (Pushchino Laboratory,
more pronounced suppression of inflammatory responses, Russia). The rats were anesthetized using isoflurane (1.3%;
a reduction in pro-inflammatory cytokine release (TNF-α Laboratories Karizoo, Spain) and Zoletil (20 mg/kg; Virbac,
and IL-6), and an increase in anti-inflammatory cytokine France) before undergoing surgery. Adipose tissue was
production (IL-10). In addition, this approach reduced carefully collected in a sterile environment and placed into
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scar tissue formation and improved motor function a container with 0.9% sodium chloride (NaCl) solution
compared to systemic EV administration. This difference (PanEco, Russia). The tissue was homogenized, centrifuged
may be due to localized administration providing a more in 0.9% NaCl, and finely minced for 10 min at 1,500 rpm.
rapid and targeted effect on inflammatory processes at the Afterward, it was treated with a 0.5% collagenase solution
injury site. Despite being more invasive than intravenous derived from crab pancreas (Biolot, Russia) at 37°C for
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injection, local delivery of cells or EVs near the spinal cord 1 h with constant shaking at 180 rpm. Following this, the
lesion has generally been shown to be safe when performed homogenate was centrifuged at 1,400 rpm for 5 min to
slowly and carefully in patients with SCI. 7 remove the enzyme solution. The remaining cells were
In view of these findings, the local application of MSC- washed in Dulbecco’s phosphate-buffered saline (DPBS;
EVs may be an effective anti-inflammatory approach for PanEco, Russia), centrifuged again to remove residual

Volume 4 Issue 4 (2025) 78 doi: 10.36922/AN025110022