Immunomodulatory Effect of Adipose Stem Cell-Derived Extra-Cellular Vesicles on Cytokine Expression and Regulatory T Cells in Patients with Asthma

Abstract

Although mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) are as effective as MSCs in the suppression of allergic airway inflammation, few studies have evaluated the immunomodulatory capacity of MSC-derived EVs in patients with asthma. Thus, we assessed the effects of adipose stem cell (ASC)-derived EVs on cytokine expression and regulatory T cells (Tregs) in peripheral blood mononuclear cells (PBMCs) of asthmatic patients. PBMCs (1 × 10⁶ cells/mL) were isolated from asthmatic patient and healthy controls and co-cultured with 1 μg/mL of ASC-derived EVs. Th (T helper) 1-, Th2-, and Treg-related cytokine expression, fluorescence-activated cell sorting analysis of CD4⁺CD25⁺FOXP3⁺ T cells, and co-stimulatory molecules were analyzed before and after ASC-derived EV treatment. The expression levels of IL-4 and costimulatory molecules such as CD83 and CD86 were significantly higher in PBMCs of asthmatic patients than in control PBMCs. However, ASC-derived EV treatment significantly decreased the levels of interleukin (IL)-4 and co-stimulatory molecules such as CD83 and CD86 in the phytohemagglutinin (PHA)-stimulated PBMC of asthmatic patients. Furthermore, ASC-derived EVs remarkably increased the transforming growth factor-β (TGF-β) levels and expression of Tregs in the PBMC of asthmatic patients. ASC-derived EVs induce Treg expansion and have immunomodulatory effects by downregulating IL-4 and upregulating TGF-β in PBMCs of asthmatic patients.

Keywords: T lymphocytes; asthma; extracellular vesicles; immunosuppression therapy; mesenchymal stem cells; regulatory.

Figure 1

Characterization of adipose stem cell (ASC)-derived extracellular vesicles (EVs). (A,B) Transmission electron microscopy demonstrated that ASC-derived EVs have lipid bilayers and spherical shape (original magnification ×50,000 and ×80,000, respectively). (C) The average size of the EVs measured by dynamic light scattering was 130.3 ± 65.7 nm in diameter. The majority of particles isolated from the ultracentrifugation-concentrated EV pellets were in a size range of approximately 100–200 nm.

Figure 2

Effects of adipose stem cell (ASC)-derived extracellular vesicles (EVs) on cytokine levels. IL-4 level was significantly higher in the CON and PHA group of asthmatic patients than healthy controls. ASC-derived EVs markedly decreased the levels of IL-4 but increased levels of TGF-β in the PHA group of asthmatic patients. IFN-γ levels were significantly higher in the PHA group of asthmatic patients than healthy controls. ASC-derived EVs significantly elevated the IFN-γ levels in the CON group of asthmatic patients and IL-17A levels in the CON group of healthy controls and asthmatic patients. The horizontal lines represent the means. CON, control; IFN, interferon; IL, interleukin; PHA, phytohemagglutinin; TGF, transforming growth factor. * p < 0.05; ** p < 0.01.

Figure 3

Effects of adipose stem cell (ASC)-derived extracellular vesicles (EVs) on CD4⁺CD25⁺FOXP3⁺ T cells. Although there were no significant differences in the percentage of CD4⁺CD25⁺FOXP3⁺ T cells among the groups, ASC-derived EVs significantly increased the relative expression of CD4⁺CD25⁺FOXP3⁺ T cells in the CON and PHA group of asthmatic patients. The horizontal lines represent the means. CON, control; PHA, phytohemagglutinin, Rel, relative. * p < 0.05; ** p < 0.001.

Figure 4

Effects of adipose stem cell (ASC)-derived extracellular vesicles (EVs) on CD80, CD83, and CD86. The percentages of CD83 and CD86 were significantly higher in the CON group of asthmatic patients than healthy controls. ASC-derived EVs significantly decreased the percentage of CD83, and CD86 in the PHA-stimulated PBMC of asthmatic patients. The horizontal lines represent the means. CON, control; PHA, phytohemagglutinin. * p < 0.05.