Loss of endothelial barrier function requires neutrophil adhesion

The inflammatory response is characterized by cytokine-induced up-regulation of endothelial adhesion molecules followed by polymorphonuclear neutrophil (PMN) adhesion and breakdown of tight junctions between cells. The purpose of this investigation was to determine whether PMN adhesion is an essential element in the alteration of endothelial permeability or whether cytokines alone can produce this change.

Human umbilical vein endothelial cells (HUVECs) were exposed to formylated met-leu-phe-activated PMNs. In a second series of experiments, PMNs were contained in a microporous membrane that allowed passage of secreted cytokines but not cells. Permeability was quantified by using transendothelial electrical resistance (TEER, ohm.cm2,) whereas expressions of two cell adhesion molecules (endothelial leukocyte adhesion molecule-1 [ELAM-1] and intercellular adhesion molecule-1 [ICAM-1]) were measured by flow cytometry (% shift). Cytokine production was monitored with enzyme-linked immunosorbant assays (picograms per milliliter).

Stimulated PMNs secreted comparable amounts of cytokines whether allowed access to HUVECs or trapped in a microporous membrane (interleukin-1 alpha, 5.88 +/- 2.38 versus 3.65 +/- 1.84 pg/ml; tumor necrosis factor-alpha, 10.27 +/- 3.21 versus 6.61 +/- 1.82 pg/ml). Up-regulation of ELAM-1 and ICAM-1 was observed whether PMNs were free or restricted (52.97% +/- 2.14% versus 75.32% +/- 4.19% and 71.66% +/- 7.37% versus 73.66% +/- 4.32%, respectively). TEER was unchanged in controls and when PMNs were membrane restricted. In contrast, TEER decreased precipitously (51% +/- 5.9% of control, p < 0.05) if PMNs were allowed access to HUVECs.

Cytokine secretion by PMNs is independent of endothelial contact and is sufficient to upregulate adhesion molecules. However, PMN adhesion is essential for the loss of endothelial barrier function, which leads to diapedesis of activated PMNs and eventual tissue injury.