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Bacterial pathogens can cause systemic infections by disseminating from the initial focus of infection to distant organs through the blood vasculature. To colonize tissues from the bloodstream, many bacteria need to adhere to endothelial cells (ECs) that line the vessels’ inner lumen, invade them and then spread from cell to cell. ECs are highly mechanosensitive and respond to mechanical cues, including the stiffness of the matrix on which they adhere, by dynamically re-arranging their cytoskeleton, cell-cell adhesions, and adhesions to the extracellular matrix. We hypothesize that bacterial infection of ECs might also be affected by the mechanics of the host ECs, the latter being influenced by the stiffness of the matrix on which ECs reside. To test this hypothesis, we developed a novel multi-well format assay that allows assessing in a highly quantitative manner the differential effect of matrix stiffness on EC bacterial infection. We demonstrate that, using our assay, we can quantify infection efficiency through flow cytometry and microscopy, and also measure the effects of infection on EC mechanics through traction force microscopy. Our method allows for the analysis of the effect of tissue-relevant mechanics on EC infection, which is a critical step towards understanding the biomechanical interactions between ECs, their extracellular matrix and pathogenic bacteria.

athens journal of sciences

A Highly Quantitative Multi-Well Format Assay for Studying the Effect of Extracellular Matrix Mechanics on the Bacterial Infection of Endothelial Cells