Abstract
BackgroundSevere, steroid-resistant asthma (SSRA) is a serious clinical problem in asthma management. Affected patients have severe clinical symptoms, worsened quality of life, and do not respond to steroid, a mainstay steroid treatment of asthma. Thus, effective therapies are urgently needed. Exosomes derived from mesenchymal stem cell (MSC-Exo) has become attractive candidates for the lung inflammatory diseases through its immunomodulatory effects. In this study, we explored the therapeutic effects of MSC-Exo in SSRA and identified the therapeutic mechanism of MSC-Exo.MethodExosomes from human umbilical cord mesenchymal stem cell (hUCMSC) were isolated and characterized by transmission electron microscopy, nanoparticle tracking analysis and flow cytometry analysis. Effects of MSC-Exo on airway hyper responsiveness (AHR), inflammation, histopathology, and macrophage polarization in SSRA in mice were evaluated. Systematic depletion of macrophages determined the role of macrophages in the therapeutic effect of SSRA in mice. LPS-stimulated RAW 264.7 cell model was constructed to determine the underlying mechanism of MSC-Exo on macrophage polarization. qRT-PCR, Western blotting, immunofluorescence, and flow cytometry were performed to evaluate the expression of M1 or M2 markers. Tandem mass tags (TMT)-labeled quantitative proteomics were applied to explore the central protein during the regulation effect of MSC-Exo on macrophage polarization. Knockdown and overexpression of TRAF1 were used to further clarify the role of the central protein on macrophage polarization.ResultWe successfully isolated and characterized exosomes from hUCMSCs. We verified that the intratracheal administration of MSC-Exo reversed AHR, histopathology changes, and inflammation in SSRA mice. Systematic depletion of macrophages weakened the therapeutic effect of MSC-Exo. We found that MSC-Exo treatment inhibited M1 polarization and promoted M2 polarization in LPS-stimulated RAW 264.7 cells. Subsequently, tumor necrosis factor receptor-associated factor 1 (TRAF1) was determined as the central protein which may be closely related to the regulation of macrophage polarization from TMT-labeled quantitative proteomics analysis. Knockdown and overexpression of TRAF1 demonstrated that the effect of MSC-Exo treatment on macrophage polarization, NF-κB and PI3K/AKT signaling was dependent on TRAF1.ConclusionMSC-Exo can ameliorate SSRA by moderating inflammation, which is achieved by reshaping macrophage polarization via inhibition of TRAF1.
Highlights
Severe, steroid-resistant asthma (SSRA) is a serious clinical problem in asthma management
These results indicated that the exosomes from human umbilical cord mesenchymal stem cell (hUCMSC) were successfully extracted
We found that neutrophil numbers were not reduced by DEX treatment in the OVA/Complete Freud’s adjuvant (CFA) group (OVA/ CFA+DEX) but was decreased in the Mesenchymal stem cell (MSC)-Exo treated group (OVA/CFA+Exosomes derived from mesenchymal stem cell (MSC-Exo)) compared with OVA/CFA group (Fig. 2f)
Summary
Steroid-resistant asthma (SSRA) is a serious clinical problem in asthma management. Immune cells and the disease-associated cytokines they secret are important in the pathogenesis of SSRA These mediators such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) exacerbate airway inflammation and lung tissue damage [3,4,5]. During the acute exacerbation of SSRA, M1 macrophages have been shown to secrete large amounts of inflammatory mediators (include TNF-α, IL1β, IL-6, inducible nitric oxide synthase (iNOS)) [9]. These inflammatory factors are associated with neutrophil-rich infiltration, airway hyper-responsiveness (AHR) and airway remodeling [2]. M1 and M2 phenotypic switch might be a potentially novel therapeutic avenue for the treatment of SSRA
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