Kapoor, K. S. | Search bioRxiv

bioRxiv2025

Oncogenic KRAS regulates secretion of extracellular vesicles and surface membrane charge via regulation of phosphatidylserine in pancreatic cancer

Extracellular vesicles (EVs) have the potential to be used as a liquid biopsy for cancer detection and treatment response assessment. Although the potential of EVs as disease-specific biomarkers has been promising, the rapid and specific enrichment of EVs from body fluids in the clinical setting remains challenging. To address this limitation, we developed a Microfluidic Electrophoresis (MEP) device, that allows label-free enrichment of EVs based on charge from the serum of patients with pancreatic cancer (PaCa). The ability of MEP to enrich anionic EVs was validated using cell line-derived EVs and PaCa serum-derived EVs. We observed a positive correlation between the KRAS status, secretion of EVs, and net negative zeta potential ({zeta}-potential) of EVs. Further analyses identified phosphatidylserine (PS) and extraluminal DNA as molecular determinants of the enhanced anionic nature of PaCa-derived EVs. Overall, this work introduces a new microfluidic device to enrich cancer EVs in circulation with potential for further rapid detection of PaCa.

bioengineering↗

bioRxiv2023

Single extracellular vesicle imaging and computational analysis identifiesinherent architectural heterogeneity

Evaluating the heterogeneity of extracellular vesicles (EVs) is crucial for unraveling their complex actions and biodistribution. Here, we identify consistent architectural heterogeneity of EVs using cryogenic transmission electron microscopy (cryo-TEM) which has an inherent ability to image biological samples without harsh labeling methods and while preserving their native conformation. Imaging EVs isolated using different methodologies from distinct sources such as cancer cells, normal cells, and body fluids, we identify a structural atlas of their dominantly consistent shapes. We identify EV architectural attributes by utilizing a segmentation neural network model. In total, 7,576 individual EVs were imaged and quantified by our computational pipeline. Across all 7,576 independent EVs, the average eccentricity was 0.5366, and the average equivalent diameter was 132.43 nm. The architectural heterogeneity was consistent across all sources of EVs, independent of purification techniques, and compromised of single spherical (S. Spherical), rod-like or tubular, and double shapes. This study will serve as a reference foundation for high-resolution EV images and offer insights into their potential biological impact.

bioengineering↗