Stretching the Limits in Infection Mechanobiology: Investigating the Spatiotemporal Dynamics of Bacterial Dissemination in Mechanically Stimulated Cell Monolayers

• Applicants: Effie Bastounis, Ph.D.; Professorin Dr. Iordania Constantinou
• Subject Area: Biophysics ; Cell Biology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 490839690

The rapid rise of multidrug-resistant pathogens poses a global health threat. To combat infections, we must understand how host cells interact with bacterial pathogens in environments rich in biochemical and mechanical cues. In the small intestine, the primary site of infection for many pathogens, epithelial cells form a single-cell barrier that prevents bacterial spread. This barrier experiences constant mechanical forces, such as peristalsis-driven stretching and luminal flow. These forces regulate key cell and barrier functions, and their disruption, particularly during inflammation, correlates with increased infection susceptibility. However, how (extra)cellular mechanics regulate intracellular bacterial spread remains poorly understood. This project aims to investigate how cyclic stretching influences epithelial cell biomechanics in health and during intracellular infection with Listeria monocytogenes (L.m.), a food-borne pathogen that infects intestinal epithelial cells before disseminating systemically. We will develop StretchView 2.0, a high-throughput, multi-directional cell-stretching system compatible with automated video-microscopy, to systematically study the interplay between cell mechanics and infection dynamics. Our preliminary data show that cyclic stretching reduces epithelial motility and enhances E-cadherin localization at junctions, likely strengthening intercellular forces and barrier integrity. To quantify these effects, we will apply force inference methods based on cell shape analysis and assess cytoskeletal dynamics and transcriptomic changes to unravel the crosstalk between cell mechanics and underlying biochemistry in response to stretch. Building on this foundation, we will examine how epithelial stretching impacts L.m. spread. We hypothesize that increased intercellular forces will restrict bacterial cell-to-cell spread, limiting dissemination. At later infection stages, we will investigate stretching’s impact on collective infected cell extrusion (CICE), a process that results from the mechanical battle between infected and uninfected cells driven by ERK activation waves. We hypothesize that stretching will modulate ERK wave propagation, influencing the mechanical competition leading to CICE, which we will test using traction force microscopy and FRET imaging on infected cell monolayers residing in the StretchView 2.0 platform. Finally, we will integrate perfusion on the StretchView 2.0 platform to assess its impact on cytokine distribution, ERK wave frequency, and infected cell clearance. This will reveal whether addition of flow enhances host defense by promoting CICE or facilitates bacterial spread. Beyond enhancing our understanding of how mechanical forces shape host-pathogen interactions, our findings could pave the way for therapies that harness mechanical cues to reinforce host defense against infection.

• DFG Programme: Research Grants