Objectives: Platelet concentrates (PCs) have recently been used successfully in a variety of clinical applications aiming to regenerate hard and soft tissues. However, other studies have reported conflicting results, reinforcing the need for a better characterization of PCs at the molecular level. We have previously demonstrated that growth factor (GF) release from PCs, namely VEGF, TGF-beta, PDGF and bFGF is regulated by calcium and thrombin concentrations and that PC supernatants strongly promote endothelial cell (HUVEC) division. Results from neutralization experiments suggest that other mediators released by PCs seem to play regulatory roles on HUVEC division. We have therefore determined the concentrations of IGF-1, ANG-2, TGF-alpha, EGF and IL-1-beta released from PCs and studied the effects of calcium and thrombin on HUVEC division. Methods: Whole blood (WB) and PCs were prepared from five volunteers and treated with various concentrations of calcium and thrombin. Sandwich-ELISA experiments specific for IGF-1, ANG-2, TGF-alpha, EGF and IL-1-beta were performed in triplicate on PC and WB supernatants harvested at various times (0 to 6 days) after addition of calcium and thrombin. These supernatants were added to HUVEC cultures in order to determine their mitogenic potential in vitro. Results: GF concentration varies significantly from one individual to another. Kinetic experiments reveal a strong induction of EGF and ANG-2 release upon addition of high calcium and thrombin concentrations. PC supernatants, treated with high calcium and thrombin concentrations promote strong HUVEC division, when incubated for short periods of time. However, when PCs are incubated for longer periods, supernatants from PCs treated with lower calcium and thrombin conditions appear to be more mitogenic. Conclusions: PCs contain significantly more GFs than WB and promote stronger HUVEC division. Definitive conclusions on optimal concentrations of calcium and thrombin needed in clinical applications will be assessed in vivo.

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