Effect of intrabronchial treatment with adipose stem cell derived extracellular vesicles on pulmonary inflammation and miRNA in blood and bronchoalveolar fluid in horses with severe asthma.

Severe equine asthma (SEA) is a common chronic respiratory disease that poses a significant problem to the health and welfare of horses. The main clinical features are due to airway hyperresponsiveness and underlying airway inflammation, mucus accumulation, reversible obstruction, and airway remodelling. These features and the course of the underlying inflammation are similar in humans. Therefore, the horse is an appropriate model for the study of human asthma. The diversity of phenotypes and subphenotypes associated with asthma in both species is poorly understood, limiting the development and use of targeted therapies. Traditionally, treatment of SEA has included reduction of exposure to allergens and treatment with local or systemic anti-inflammatory drugs and bronchodilators, and often antibiotics. These therapeutic strategies only relieve symptoms and do not address airway wall remodelling. Treatment with systemic corticosteroids may also be associated with adverse side effects, while treatment with antibiotics, generally considered as unnecessary, tends to promote antibiotic resistance in bacteria in the community. Treatment with inhaled corticosteroids and bronchodilators, which is not associated with significant side effects, has become a relatively effective treatment for controlling the symptoms of SEA. Regardless of treatment protocol, recurrent asthma attacks occur in 79% of treated patients. Regenerative cell-based therapies are becoming an increasingly important treatment option for a variety of clinical problems. We have recently shown that treatment with mesenchymal stem cells (MSCs) is safe and has the ability to modulate lung inflammation. Recent evidence suggests that miRNAs play a central role in the pathogenesis of asthma. Their influence can be positive and/or negative, local, or systemic. Therefore, it is important to investigate the effects of specific miRNAs or their combinations on specific pathology. Our preliminary studies have identified a number of miRNAs in the bronchoalveolar fluid (BALf) of horses with SEA that are associated with the pathogenesis of asthma, as well as miRNAs for which no association with asthma is currently known. The dynamics of miRNA transfer are greatly enhanced by the transfer of miRNA via extracellular vesicles (EV), which optimizes intercellular communication between endothelial cells, epithelial cells, dendritic cells, and MSCs. The transfer of biological molecules by EV alters the biological activity of recipient cells and may thus influence inflammation in the lung. This study will follow our established research model, previously conducted as a randomized controlled trial. Horses with SEA will be randomly divided into two groups. One group will be treated by intrabronchial administration of autologous adipose tissue-derived MSCs (AD-MSCs) and the other by EVs derived from AD -MSCs. BALf and peripheral venous blood will be collected from all horses before and three weeks after treatment. The clinical course of the disease will be recorded, and mRNA levels of interleukins and their proteins will be measured. miRNA will be analyzed in all samples, including MSC and EV samples before application, using a high-performance sequencing method. The results of this study will be multifaceted. We will attempt to associate the identified miRNAs with specific asthma phenotypes/sub-phenotypes. We will document the relationship between miRNA and inflammatory cytokines before and after treatment in blood and BALf. Based on our previous research, we hypothesise that treatment with EVs will have a beneficial effect on airway inflammation in SEA. EVs may be a more specific and stable form of treatment compared to MSCs. They can also be administered locally in the form of inhalation, which would be the next step in developing specific ""off-the-shelf"" therapies for specific asthma phenotypes.