NIH Reporter #5P20GM156674-02

Probing extracellular vesicle transport in hydrogels to advance diagnostics

This award funds research to understand extracellular vesicle transport in hydrogels to improve diagnostic assays by measuring EV concentration gradients and uptake using advanced microscopy and interferometry techniques.

Description

Extracellular vesicles (EVs) are membrane-bound, submicron particles that traffic bioactive cargo such as proteins, lipids, and RNAs between cells and mediate intercellular communication. Since EVs secreted from compromised cells contain disease-specific molecules, EVs hold great promise as biomarkers for noninvasive sensing and diagnostics. Development of these assays has been slowed by limited understanding of the fundamental mechanisms of transport in tissue spaces near EV-secreting cells, which dictate the spatial distribution, interstitial concentration, and clearance rates of these indicators. In the proposed work, we leverage unique experimental platforms to measure the evolution of EV concentration gradients within extracellular matrix-mimetic hydrogels across multiple length and time scales. In Aim 1, we expose hydrogels of variable crosslinking density to suspensions of macrophage-derived EVs and track the EV diffusion at micron and Hz resolution using microfluidic Fabry-Perot interferometry (μFPI). Unlike conventional transport characterization techniques (e.g., Fluorescent Recovery After Photobleaching), the refractive index-based method precisely quantifies EV concentration without fluorescent tagging, enabling EVs secreted from diseased populations to be characterized without modification. In Aim 2, we pattern the 10 – 100 μm scale spatial distribution of nanoparticle-amplified EV secreting macrophages and EV-receiving macrophages in hydrogel scaffolds using advective assembly extrusion and measure EV uptake by confocal microscopy. Combining the diffusivity, permeation, and binding measurements obtained in Aim 1 with the tissue analog in Aim 2 will begin to elucidate EV transport mechanisms and limitations across biologically relevant scales. Junior investigator Bayles will build upon the results obtained in this pilot project through subsequent NIH grant applications (e.g., NIH R01, R35), tool development in the DCMBS Core, and collaborations with other investigators in COBRE.

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