A Live Cell Imaging Microfluidic Model for Studying Extravasation of Bloodborne Bacterial Pathogens

Abstract

Bacteria that migrate (extravasate) out of the bloodstream during vascular dissemination can cause secondary infections in many tissues and organs, including the brain, heart, liver, joints, and bone with clinically serious and sometimes fatal outcomes. The mechanisms by which bacteria extravasate through endothelial barriers in the face of blood flow-induced shear stress are poorly understood, in part because individual bacteria are rarely observed traversing endothelia in vivo, and in vitro model systems inadequately mimic the vascular environment. To enable the study of bacterial extravasation mechanisms, we developed a transmembrane microfluidics device mimicking human blood vessels. Fast, quantitative, three-dimensional live cell imaging in this system permitted single-cell resolution measurement of the Lyme disease bacterium Borrelia burgdorferi transmigrating through monolayers of primary human endothelial cells under physiological shear stress. This cost-effective, flexible method was 10,000 times more sensitive than conventional plate reader-based methods for measuring transendothelial migration. Validation studies confirmed that B. burgdorferi transmigrate actively and strikingly do so at similar rates under static and physiological flow conditions. This method has significant potential for future studies of B. burgdorferi extravasation mechanisms, as well as the transendothelial migration mechanisms of other disseminating bloodborne pathogens.