Neutrophil-Derived Exosomes: A New Mechanism Contributing to Airway Smooth Muscle Remodeling.

ABSTRACT

Neutrophils infiltrate the airways of patients with asthma of all severities, yet their role in the pathogenesis of asthma and their contribution to airway remodeling is largely unknown. We hypothesized that neutrophils modulate airway smooth muscle (ASM) proliferation in asthma by releasing bioactive exosomes. These newly discovered nano-sized vesicles have the capacity to modulate immune responses, cell migration, cell differentiation, and other aspects of cell-to-cell communication. The aim of the study is to determine whether bioactive exosomes are released by neutrophils, and, if so, characterize their proteomic profile and evaluate their capacity to modulate ASM cell proliferation. Exosomes were isolated from equine neutrophil supernatants by differential centrifugation and filtration methods, followed by size-exclusion chromatography. Nanovesicles were characterized using electron microscopy, particle size determination, and proteomic analyses. Exosomes were cocultured with ASM cells and analyzed for exosome internalization by confocal microscopy. ASM proliferation was measured using an impedance-based system. Neutrophils release exosomes that have characteristic size, morphology, and exosomal markers. We identified 271 proteins in exosomes from both LPS and unstimulated neutrophils, and 16 proteins that were differentially expressed, which carried proteins associated with immune response and positive regulation of cell communication. Furthermore, neutrophil-derived exosomes were rapidly internalized by ASM cells and altered their proliferative properties. Upon stimulation of LPS, neutrophil-derived exosomes can enhance the proliferation of ASM cells and could therefore play an important role in the progression of asthma and promoting airway remodeling in severe and corticosteroid-insensitive patients with asthma.