Newborn platelets: lower levels of protease-activated receptors cause hypoaggregability to thrombin.

Newborn platelets show in vitro hypoaggregability to thrombin. Sensitivity of platelets to such a potent agonist is crucial for a functional clot formation. Nevertheless, newborns have an excellent hemostasis. We wanted to investigate the reason for this impairment by comparatively analysing levels of receptors known to be involved in thrombin signaling in newborn and adult platelets. Platelets of adult and cord blood were isolated, washed, and lysed. Resulting protein samples were separated by SDS-PAGE and blotted on nitrocellulose membranes. Receptors were visualized using immunodetection and evaluated densitometrically. Thrombin receptor activating peptide induced platelet aggregation was measured in citrated whole blood on a Multiplate analyzer. Statistical analysis was performed using SPSS 16.0. Significantly lower levels of protease-activated receptors (PAR1, PAR4) and higher levels of glycoprotein Ibα (GPIbα) were found in newborn platelets as compared to adult platelets. Platelet aggregation was lower in newborn samples than in adult controls and values correlated with the corresponding PAR levels. Our results suggest that lower levels of protease-activated receptors contribute to the poor thrombin induced aggregation observed with newborn platelets, which can not be compensated by higher levels of GPIbα.

High microparticle concentration in cord plasma.

UNLABELLED: We investigated if differences in the microparticle concentration and activity between newborn cord plasma and adult plasma exist. METHODS: To enumerate and characterize microparticles (MP) FACS and ELISA were used.The effect of microparticles derived tissue factor (TF) on thrombin generation was measured indirectly by CAT (calibrated automated thrombography). RESULTS: The flow cytometric measurements revealed an increased microparticle concentration in newborn cord compared with adult plasma. By the use of ELISA a significantly increased procoagulant activity of microparticles was found in newborn cord plasma as compared to adult plasma. Initiation of thrombin generation by adding phospholipids alone resulted in a significant lower prolongation of the lag time, time to peak in cord plasma, while the decrease of endogenous thrombin potential (ETP) and peak was comparable between newborns and adults. CONCLUSION: Our results show a higher impact of microparticles on the haemostatic system of newborns than on that of adults. The three methods suggest a somewhat increased microparticle activity in newborn cord plasma, but argue against strong platelet activation during birth.

Comparative evaluation of PAR1, GPIb-IX-V, and integrin αIIbß3 levels in cord and adult platelets.

BACKGROUND: Newborn platelets show hyposensitivity in vitro to many important agonists. However, sensitivity of platelets to these agonists is crucial for a functional clot formation. Nevertheless newborns have an excellent hemostasis. Hence, we examined levels of PAR1 thrombin receptor, GPIb-IX-V (CD42b), and Integrin αIIbß3 in newborn and adult platelets using Western blot analysis. Materials, methods: Platelets of adult and cord blood were isolated, washed, and lysed. Protein samples were separated by SDS-PAGE and blotted on nitrocellulose membranes. Receptors were visualized using immunodetection and evaluated densitometrically. Statistical analysis was performed using SPSS 16.0. RESULTS: We found significantly lower levels of PAR1-receptors and higher levels of CD42b in newborn platelets as compared to adult platelets. Levels of Integrin αIIbß3 in newborn platelets were comparable to adult platelets. CONCLUSION: A lower content of PAR1-receptors explains very well the hyposensitivity of cord platelets to thrombin. Higher levels of CD42b may additionally support the effect of larger more adhesive multimeric vWF in newborn plasma.