Newswise \u2014 COLUMBUS, Ohio \u2013 The tiny bubbles that carry signals between cells throughout the body were discovered decades ago, but precisely what they contain and how their cargo affects recipient cells is still largely a mystery.\u00a0<\/p>\n
Though research to date shows that these bubbles, called extracellular vesicles and particles<\/a>, influence human health and disease, they come in so many different sizes and contain such a huge range of contents that it\u2019s difficult to pinpoint their specific functions. Found circulating in biological fluids and embedded in tissues, they are being explored for applications ranging from early disease detection to drug delivery.\u00a0 \u00a0\u00a0\u00a0\u00a0<\/p>\n In a study published recently in Nature Methods<\/a>, scientists at The Ohio State University report on a new approach to immobilize extracellular vesicles in a way that mimics their interactions with tissues, a tricky environment compared to bodily fluids because these particles have a specific characteristic: They\u2019re sticking to a surface.\u00a0<\/p>\n \u201cThe extracellular vesicles present in tissues are very poorly understood in terms of how the particles actually interact with cells in our body,\u201d said senior author Eduardo Re\u00e1tegui<\/a>, associate professor of chemical and biomolecular engineering at Ohio State<\/a>.<\/p>\n The new technique enables label-free immobilization of extracellular vesicles and particles without damaging them, allowing researchers to study them individually or in clumps and observe how they interact with cells.\u00a0<\/p>\n \u201cWhat we want to do is not only understand what these vesicles contain, but also identify their tissue of origin, and how they interact with cells. One way to do that is to analyze them without destroying them,\u201d said Re\u00e1tegui, also a member of the Cancer Biology Program<\/a> in The Ohio State University Comprehensive Cancer Center.<\/p>\n Researchers began by coating a glass surface with a chemical layer, and then used UV light to \u201cetch\u201d a micropattern on that layer, creating tiny spaces with an attractive electrostatic charge to which the proteins on the extracellular vesicles\u2019 outer layers would stick. Computer simulations confirmed that electrostatic attractions dominate the interactions between the tiny spaces and the particles.\u00a0<\/p>\n The researchers found that every extracellular vesicle type used in experiments adhered to the surface only where the light-induced micropattern was produced, without seeping outside those micropatterns.<\/p>\n The team calls the technology Light-Induced Extracellular Vesicle and Particle Adsorption, or LEVA.\u00a0<\/p>\n This analytical advance opens the door to studying these particles and their complex interactions with cells at a whole new level, such as exploring their contents for disease biomarker discovery or loading them with therapeutic molecules and watching cells respond, to name just a few possibilities.<\/p>\n Previously, Re\u00e1tegui and colleagues developed a method of using antibodies to immobilize these extracellular vesicles and particles for analysis, which allowed the team to identify molecules inside specific types of particles that were biomarkers for brain cancer or indicators of immunotherapy response. But the method had a limitation: Only EVs with a specific molecule on their surface that would be recognized by the antibody could be isolated for analysis.\u00a0<\/p>\n With this new technique focused on surface-based extracellular vesicles and particles (EVPs), the UV light degrades regions of a coating to coax particle adsorption to the surface that is dictated by electrostatic interactions rather than any biological signature.\u00a0<\/p>\n \u201cWe started to think about how we can remove that bias in terms of pre-selecting this extracellular vesicle population with antibodies,\u201d he said. \u201cAnd now we can basically immobilize all of them, and have the ability to interrogate them with molecular probes or even cells.\u201d\u00a0<\/p>\n In one application of the study, researchers showed that the new technique can be used to study early stages of inflammation by mimicking the response of immune cells to pathogens such as bacteria or fungi \u2013 but instead of using bacteria such as E. coli, they used the extracellular vesicles produced by the bacteria. They found that the EVs emitted by E. coli induced what is known as a neutrophil swarming \u2013 the coordinated recruitment and migration of neutrophils<\/a> toward sites of infection, along the micropatterns where the bacterial EVs were bound. \u00a0<\/p>\n \u201cThis showed that, first, we can generate matrix-bound EVPs in different contexts for easy analysis, and second, this approach allows exploration of EVP interactions in tissue,\u201d Re\u00e1tegui said.<\/p>\n This work was supported by the Ohio State Center for Cancer Engineering-Curing Cancer Through Research in Engineering Sciences; the National Institutes of Health; the Burroughs Wellcome Fund; and Ohio State postdoctoral scholars programs.\u00a0<\/p>\n Co-authors include Colin Hisey, Xilal Rima, Jacob Doon-Ralls, Chiranth Nagaraj, Sophia Mayone, Kim Truc Nguyen, Sydney Wiggins, Kalpana Deepa Priya Dorayappan, Xin Huang, Karuppaiyah Selvendiran, David Wood, Chunyu Hu, Divya Patel, Andre Palmer and Derek Hansford of Ohio State; Mangesh Hade and Setty Maga\u00f1a of Nationwide Children\u2019s Hospital; and James Higginbotham, Oleg Tutanov, Jeffrey Franklin and Robert Coffey of Vanderbilt University.<\/p>\n #<\/p>\n Contact: Eduardo Re\u00e1tegui, Reategui.8@osu.edu<\/a><\/p>\n