Grant: Discovery Grants Program - Individual

In the research program grant, I propose new technologies for the affinity isolation of extracellular vesicles (EVs), the detection and counting of EVs, and comprehensive analysis of EV biomarkers. EVs are small membrane-enclosed structures that are secreted into the extracellular milieu by many cell types. These particles include small (exosomes, 30-150 nm), and large (microvesicles, 100 - 1000 nm; apoptotic bodies, 50 - 5000 nm) vesicles. These vesicles are secreted from cells and released in different fluids of the body. EVs contain a lipid bilayer that protects the cargo, which can consist of nucleic acids, proteins, lipids and metabolites, from degradation. EVs are now central to biological sciences because they seem to constitute a new system of cell-cell communication. Since the cargo of EVs reflects the precise state of a cell, there has been significant interest in utilizing EVs in biomarker analysis. EVs are a plentiful source of biomarkers of traumatic brain injury (TBI) and posttraumatic stress disorder (PTSD) among other conditions. Recently, the recognition that EVs derived from brain cells that carry proteins and nucleic acid from their mother cells and cross the blood brain barrier can be detected in peripheral blood and saliva, offers a new opportunity to evaluate molecular changes in neurons and glia during PTSD and after TBI. Over the next 5 years, my research team and I plan to provide two novel technologies for the production of specific affinity probes for EVs, named nucleic acid aptamers. The resulting aptamers will have switchable binding and high selectivity to exosomes and microvesicles originating from brain cells. Two EV isolation technologies (magnetic bead and column - based) will be also developed and published, for the benefit of the EV research community. We expect to achieve high recovery of isolated EVs from human samples for downstream applications such as miRNA and protein assays. We will develop and share at least two technologies based on capillary electrophoresis and mass spectrometry for comprehensive analysis of proteins and microRNAs of EVs. The data obtained from these approaches will be integrated into public databases. In particular, we will develop a breakthrough diagnostic based on aptamer-assisted isolation of brain EVs and analysis of TBI/PTSD biomarkers to discriminate injury states. In the long-term, I plan to perform large-scale research on selection of aptamers, their structures, functions and related binding partners. I want to study EVs using high-throughput approaches in proteomics and metabolomics, supported by miRNA data and bioinformatic analysis. In general, I will address fundamental issues of EV origins and final destinations, molecular diversity and biological functions. The development of bioassays for brain cell-derived EVs will bring new analytical tools for researchers investigating fundamental processes of TBI, PTSD and degenerative brain disorders.