Development of Methods for Longitudinal Sampling of Neuronally Derived Exosomes in Neurodegenerative Disease
We are working to develop novel technologies based on extracellular vesicles (EVs) for “reading out” the RNA and protein profiles of neurons in human and apply them to study neurodegeneration. We are doing this by developing a framework for isolating cell type-specific EVs in human biological fluids using a variety of unbiased computational and experimental techniques. We will then apply this framework to isolating neuron specific EVs from human cerebrospinal fluid (CSF) and blood plasma. In order to isolate cell type-specific EVs from biological fluids, we will develop robust immuno-isolation techniques to capture EV subpopulations based on specific protein markers on their surface. We plan to use human induced pluripotent stem (iPS) cell derived neurons as a relevant model system before applying our isolation techniques to CSF and plasma. Although isolating neuron-derived EVs from plasma is the ultimate goal, it is also the most challenging, as we expect the proportion of neuron-derived EVs to be low. Thus, we are following a three-step approach in which we first develop our technique in human iPS-derived neurons, then in CSF, and then in plasma. After validating our isolation technique, we plan to use ultrasensitive protein measurement and high-throughput RNA sequencing to characterize the contents of neuron-specific extracellular vesicles. We will apply our methods to profile the RNA and protein repertoires of neuron-derived EVs in CSF and plasma samples obtained from Parkinson’s Disease (PD) patients. These methods will allow us to identify new biological mechanisms associated with PD and study the progression of neurodegeneration in humans.
