PECAM-1 (CD31) is required for interactions of platelets with endothelial cells after irradiation.

Sustained adhesion of platelets to endothelial cells (EC) is believed to contribute to thrombosis and vascular occlusions following radiation exposure leading to organ functional impairment and even death. Our objective was to evaluate the role of platelet endothelial cell adhesion molecule (PECAM)-1 in the prothrombotic response of EC after irradiation. Endothelial PECAM-1 expression was determined by cell-enzyme linked immunosorbent assay (ELISA) on human microvascular EC from lung (HMVEC-L) up to 21 days after a 10 Gy irradiation. Platelet- and leukocyte-endothelial cell interactions were assessed using a flow adhesion assay with fluorescently labeled whole blood, and the function of PECAM-1 in these processes was measured by using blocking antibody. PECAM-1 expression was significantly increased on irradiated HMVEC-L and remained elevated at 21 days. Anti-PECAM-1 antibody significantly inhibited adhesion of single platelets and thrombi on irradiated HMVEC-L. This inhibitory effect persisted at day 21. Anti-PECAM-1 also reduced leukocyte adhesion to irradiated HMVEC-L. The up-regulation of endothelial PECAM-1 following radiation exposure is persistent. PECAM-1 plays a key role platelet adhesion/aggregation on irradiated EC. Therefore, strategies targeting this adhesion molecule may prevent the development of radiation pathologies.

Radiation effects on circulating and endothelial cell interactions studied by quantitative real-time videomicroscopy.

PURPOSE: To quantify in vitro the functional consequences of irradiation on the interactions between leukocytes or platelets and endothelial cells (EC) in flowing whole blood using a parallel-plate flow chamber and real-time videomicroscopy. MATERIALS AND METHODS: The parallel-plate flow chamber was calibrated to determine the dynamic parameters of the flow channel. Fluorescent-labelled whole blood was perfused at wall shear rates of 25, 75 and 500 s(-1) over a monolayer of human microvascular EC-lung (HMVEC-L) with or without irradiation at 10 Gy. The adhesion of leukocytes and platelets on EC was quantified by videomicroscopy and image analysis. RESULTS: Calibration of the parallel-plate flow chamber showed that flow in the chamber was laminar and steady and had a parabolic velocity profile, thus simulating physiological flow conditions. Flow assay revealed that rolling, mean rolling velocity and firm adhesion of leukocytes was increased following irradiation of EC. Irradiation also favoured platelet adhesion to EC. CONCLUSIONS: The results of an in vitro flow assay with whole blood showed that under physiological flow conditions, irradiation affected the function of EC; pro-inflammatory and thrombogenic responses were enhanced, which may contribute to in vivo radiation-induced vascular occlusion and fibrosis.

Defect of heparin binding in plasma and recombinant von Willebrand factor with type 2 von Willebrand disease mutations.

The aim of our study was to characterise heparin-binding properties of mutated von Willebrand factor (VWF) in 24 patients plasmas with type 2 von Willebrand disease (VWD). and in 15 recombinant VWF (rVWF) with the corresponding mutations. Binding of mutated rVWF or plasma VWF was compared to that of WT-rVWF or normal pool plasma VWF. Four mutations, at positions C509, V551, R552 and R611 lead to significantly decreased binding to heparin in both plasma and rVWF. Interestingly, whereas these four residues are distant in the primary structure of VWF-A1domain, they are close to each other in its three-dimensional structure. Structural analysis suggested how folding problems and destabilisation due to these mutations could induce reorganisation of surface regions involved in heparin binding. In contrast, no heparin-binding defect was found associated with different type 2 VWF mutants, at positions G561, E596, I662, R543, R545, V553, R578 or L697.