Investigating the Role of Exosomes in the Development of Neuromuscular Junctions

McKnight, Adam

Student Work

Investigating the Role of Exosomes in the Development of Neuromuscular Junctions

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The development of the neuromuscular junction involves communication between neuronal and muscle cells as they differentiate. Recently, exosomes have become a popular focus of research on cellular communication and have been found to be involved in the formation of neuromuscular junctions. The goal of this study is to isolate exosomes from a coculture of neuronal and muscle cells, characterize them, and investigate their application towards neuromuscular development alongside a pharmaceutical used to inhibit their biogenesis. This goal was achieved by utilizing an ultracentrifugation protocol to isolate exosomes from a coculture of PC12 and WKO-3M22 cells and characterizing their size using Dynamic Light Scattering. Cultures were then treated with exosomes and the neutral sphingomyelinase inhibitor GW4869 and studied for the degree of neuronal differentiation and neuromuscular junction formation. Sonication and filtration were found to improve sample homogeneity and reduce exosome aggregation following isolation, although further optimizations are necessary. The addition of exosomes or the inhibitor alone did not substantially affect coculture development; however, when added together there was an increase in both the number of neurites per cell, and the development of neuromuscular junctions when compared to control. This observation may be due to altering the ratios of sphingolipids such as ceramide and sphingomyelin, which change during neuronal differentiation. Blocking the conversion of sphingomyelin to ceramide with GW4869 while adding an enriched source of sphingolipids in the form of exosomes may be responsible for enhanced differentiation, presenting an opportunity for future work.

This report represents the work of one or more WPI undergraduate students submitted to the faculty as evidence of completion of a degree requirement. WPI routinely publishes these reports on its website without editorial or peer review.

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