Endothelial cell source dictates the expression and release of fibrinolytic markers in a proinflammatory environment

Endothelial cells (ECs) provide a surface for molecular interactions, secreting various factors that govern hemostasis. Inflammatory cytokines can perturb the vascular microenvironment, potentially causing endothelial dysfunction and dysregulation of hemostasis.

To examine the fibrinolytic balance of ECs from different vascular beds and their response to proinflammatory stimuli.

Primary human umbilical vein ECs (HUVECs), human coronary artery ECs (HCAECs), and immortalized EA.hy926 cells were cultured under venous (2.5 dyne/cm²) and arterial (12 dyne/cm²) shear stress. ECs were stimulated with thrombin, interleukin 6, or tumor necrosis factor (TNF)-α for 24 hours. The expression of coagulation and fibrinolytic proteins was quantified by quantitative polymerase chain reaction and secreted proteins via ELISA or activity assay. Plasma clot lysis on ECs ± 300 pM tissue-type plasminogen activator (tPA) was monitored at 405 nm.

Basal secretion of C-reactive protein, urokinase plasminogen activator (uPA), and plasminogen activator inhibitor-1 (PAI-1) was higher in HCAECs than in HUVECs, but tPA was similar in both. TNF-α stimulation of HCAECs increased secretion of tPA, uPA, and PAI-1. Levels of tPA/PAI-1 and uPA/PAI-1 were higher in media, as was free-active PAI-1. In contrast, stimulation of HUVECs did not significantly alter gene/protein levels. HCAECs and HUVECs delayed clot lysis relative to no-cell controls by 11 ± 8 minutes (P < .01) and 8 ± 6 minutes (P < .05) respectively, but were normalized by neutralizing PAI-1. TNF-α stimulation of HCAECs prolonged clot lysis in a PAI-1-dependent manner.

HCAECs respond more potently to a proinflammatory environment than HUVECs, altering expression of fibrinolytic proteins and promoting a hypofibrinolytic response. These data highlight HCAECs as a model of coronary vasculature with potential for screening novel antithrombotic strategies.