Effects of nitric oxide (NO) on platelets in neonates

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Nitric oxide (NO) is known to play a crucial role in primary hemostasis due to its platelet-inhibitory properties. To investigate the effects of NO on neonatal platelets, umbilical cord blood from 10 healthy term neonates was obtained immediately after birth. Blood samples from both the mothers and non-pregnant female blood donors (n = 10) were analyzed in parallel as controls. Citrated platelet rich plasma was incubated with the NO-donor SIN-1 (10 μM), an active metabolite of molsidomine, prior to activation with 10 μM adenosine-diphosphate (ADP) or 0.05 U/ml human α-thrombin (both ED₅₀), or with agonists or buffer only. Platelet activation was analyzed in a two-color flow cytometry assay using phycoerythrine (PE) or fluoresceine-isothiocyanate (FITC) conjugated monoclonal antibodies directed against glycoprotein-(GP)-Ib (CD42b) and activated fibrinogen receptor GPIIb-IIIa (PAC-1), respectively. No significant differences for PAC-1 binding to resting platelets (baseline) was observed between neonate, mother or control, indicating absence of pregnancy- or delivery-induced activation of circulating platelets (see figure). No significant differences of platelet response to ADP or thrombin were found between controls and mothers, indicating normal platelet response of the mother. Compared to both the mother and control, neonatal GPIIb-IIIa activation was significantly (P < 0.05) depressed, ie newborn platelets revealed hyporeactivity to ADP and thrombin. NO significantly (P = 0.001) inhibited GPIIb-IIIa activation in all groups, however, percentual downregulation of GPIIb-IIIa-exposure was not significantly different in newborns, mothers, or controls (ca. 70 ± 5%). This demonstrates depressed response of neonatal platelets to agonists, but normal reaction to platelet inhibitory compounds such as NO. Since newborn platelets are starting from a lower level of activation, NO nearly diminished platelet activation. This might be of clinical relevance, since hemorrhagic complications remain a serious problem in critically ill term neonates, or even more in premature newborns. Several studies showed reduced platelet function in both healthy volunteers and patients with adult respiratory distress syndrome (ARDS) during inhalation of gaseous NO. NO-inhalation is well established in the pediatric ICU for treatment of infant respiratory distress syndrome (IRDS) or persistent pulmonary hypertension. The presented data indicate that inhalation of NO might be an additional risk-factor for bleeding complications in critically ill neonates.

Figure (abstract P028)

Effects of SIN-1 on GPIIb-IIIa activation in platelets from cord blood (neonate), mother and female control, activated with ADP or thrombin