Effects of divalent cations and various vasoactive and haemostatically active agents on the integrity of monolayers of cultured human endothelial cells

Injury to human endothelial cells in primary culture was evaluated by a ⁵¹Cr-release assay, phase contrast microscopy and the trypan blue exclusion test. Over a period of 24 hours incubation with either normal human serum or standard culture medium, the normal integrity of the confluent endothelium was maintained and total ⁵¹Cr-release showed a slow, linear increase. If Tris-buffered saline was used as incubation medium considerable contraction and detachment of cells and accelerated ⁵¹Cr-release were observed. This evidence of injury was markedly reduced by the addition of small amounts of Ca⁺⁺ ions, while Mg⁺⁺ had no effect. Among 4 haemostatically active agents investigated (ADP, thrombin, endotoxin and sodium polyanethol sulfonate) only thrombin induced morphological evidence of injury accompanied by accelerated release of ⁵¹Cr. These effects of thrombin were inhibited by hirudin. Five vasoactive agents (adrenaline, noradrenaline, histamine, bradykinin and serotonin) had no appreciable effect on endothelial morphology and did not accelerate ⁵¹Cr-release.