Quantifying Adhesion of Inflammatory Cells to the Endothelium In Vitro.

Abstract

We present a simple and quantitative assay system that accurately models human endothelium by use of primary human umbilical vein endothelial cells (HUVECs) in cell culture plates coated with gelatin, a matrix that mimics basal lamina, the matrix that is tightly associated with the vascular endothelium and is critical for its proper function. We describe using this system to quantitatively measure adhesion of the inflammatory cells - monocytic THP-1 cell line to the HUVEC monolayer. The THP-1 cells are fluorescently labeled which allows to quantify the number of the fluorescent THP-1 cells adhering to the endothelium under a microscope and the level of florescence - a quantitative measure of the number of adhering fluorescent THP-1 cells using a fluorescent plate reader. After optimization, we were able to detect increased adhesion of the THP-1 cells to the endothelium in response to the inflammatory cytokine TNFα in a dose-dependent manner like what has been observed in vivo.