Small extracellular vesicles derived from human MSCs prevent allergic airway inflammation via immunomodulation on pulmonary macrophages.

Allergic airway inflammation is a major public health disease that affects up to 300 million people in the world. However, its management remains largely unsatisfactory. The dysfunction of pulmonary macrophages contributes greatly to the development of allergic airway inflammation. It has been reported that small extracellular vesicles derived from mesenchymal stromal cells (MSC-sEV) were able to display extensive therapeutic effects in some immune diseases. This study aimed to investigate the effects of MSC-sEV on allergic airway inflammation, and the role of macrophages involved in it. We successfully isolated MSC-sEV by using anion exchange chromatography, which were morphologically intact and positive for the specific EV markers. MSC-sEV significantly reduced infiltration of inflammatory cells and number of epithelial goblet cells in lung tissues of mice with allergic airway inflammation. Levels of inflammatory cells and cytokines in bronchoalveolar lavage fluid were also significantly decreased. Importantly, levels of monocytes-derived alveolar macrophages and M2 macrophages were significantly reduced by MSC-sEV. MSC-sEV were excreted through spleen and liver at 24 h post-administration in mice, and were able to be taken in by macrophages both in vivo and in vitro. In addition, proteomics analysis of MSC-sEV revealed that the indicated three types of MSC-sEV contained different quantities of proteins and shared 312 common proteins, which may be involved in the therapeutic effects of MSC-sEV. In total, our study demonstrated that MSC-sEV isolated by anion exchange chromatography were able to ameliorate Th2-dominant allergic airway inflammation through immunoregulation on pulmonary macrophages, suggesting that MSC-sEV were promising alternative therapy for allergic airway inflammation in the future.

Small extracellular vesicles derived from human mesenchymal stromal cells prevent group 2 innate lymphoid cell-dominant allergic airway inflammation through delivery of miR-146a-5p.

Group 2 innate lymphoid cells (ILC2s) are recently reported to play a more critical role in allergic diseases. We previously identified that mesenchymal stromal cells (MSCs) elicited therapeutic effects on allergic airway inflammation. Small extracellular vesicles (sEV) derived from MSCs possess striking advantages including low immunogenicity and high biosafety, and is extremely promising cell-free therapeutic agents. However, the effects of MSC-sEV on ILC2s are still unclear. Additionally, scalable isolation protocols are required for the mass production of homogenous MSC-sEV especially in clinical application. We previously reported that induced pluripotent stem cells-derived MSCs were the ideal cellular source for the large preparation of MSC-sEV. Here we developed a standardized scalable protocol of anion-exchange chromatography for isolation of MSC-sEV, and investigated the effects of MSC-sEV on ILC2 function from patients with allergic rhinitis and in a mouse ILC2-dominant asthma model. The characterization of MSC-sEV was successfully demonstrated in terms of size, morphology and specific markers. Using flow cytometry and human Cytokine Antibody Array, MSC-sEV but not fibroblasts-sEV (Fb-sEV) were found to significantly inhibit the function of human ILC2s. Similarly, systemic administration of MSC-sEV but not Fb-sEV exhibited an inhibition of ILC2 levels, inflammatory cell infiltration and mucus production in the lung, a reduction in levels of T helper 2 cytokines, and alleviation of airway hyperresponsiveness in a mouse model of asthma. Using RNA sequencing, miR-146a-5p was selected as the candidate to mediate the above effects of MSC-sEV. We next revealed the uptake of ILC2s to MSC-sEV, and that transfer of miR-146a-5p in MSC-sEV to ILC2s in part contributed to the effects of MSC-sEV on ILC2s in vitro and in a mouse model. In conclusion, we demonstrated that MSC-sEV were able to prevent ILC2-dominant allergic airway inflammation at least partially through miR-146a-5p, suggesting that MSC-sEV could be a novel cell-free strategy for the treatment of allergic diseases.