Platelet Activation Is Linked to the Severity of Pulmonary Hypertension in Sickle Cell Disease and Normalizes on Sildenafil Therapy.
Authors: Villagra Jose D. , Machado Roberto F. , Hunter Lori A. , Gladwin Mark T. , Kato Gregory J. 
Published: April 2020

Abstract
Pulmonary hypertension (PHT) is a severe complication of sickle cell disease (SCD) characterized by abnormal vascular tone, vascular proliferation and thrombosis, associated with chronic hemolysis, and nitric oxide (NO) consumption by cell-free hemoglobin. Since platelets are central to the development of large-vessel thrombosis, and increased platelet activation has been described independently in patients with primary PHT and in patients with SCD, we sought to define the degree of platelet activation in SCD patients with secondary PHT. We hypothesized that the degree of platelet activation may be linked to the severity of PHT, hemolytic rate and NO bioavailability; conversely, platelet activation may decrease with therapy directed to PHT by increasing NO-dependent signaling. We obtained blood samples from SCD patients with PHT (n=16) and without PHT (n=17) during steady state (not in pain crisis). PHT was defined by a tricuspid regurgitant jet (TRjet) velocity &gt;= 2.5 m/sec by Doppler echocardiography. Platelet activation was measured by the flow cytometry detection of activated fibrinogen receptors (activated GPIIb/IIIa) and surface expression of P-Selectin. The mean percentage of platelets with activated GPIIb/IIIa was 10.3 ± 1.8%( mean ± SEM) for non-PHT SCD patients vs. 27.2 ± 4.7% for PHT SCD patients (p = 0.001). The percentage of platelets positive for P-Selectin was 2.0 ± 0.8% for non-PHT SCD patients vs. 5.1 ± 2.5% for PHT SCD patients (p = 0.02). The percentage of activated platelets strongly correlated with TR jet velocities (r = 0.63 and p = 0.0006). (Figure 1) In addition, the percentage of activated platelet correlated to surrogate markers of hemolysis LDH and reticulocyte count. This link may not prove causation but it suggests a common pathophysiology. Nitric oxide plays a central role in vascular homeostasis by regulating vascular tone and platelet activation through the generation of cyclic guanosine 5′-monophosphate (cGMP). It is possible that decreased NO bioavailability is responsible for both pulmonary vasculopathy and platelet activation. Our pilot studies indicate sildenafil, which enhances NO responsiveness by promoting the accumulation of its downstream mediator cGMP, may be useful in treating PHT.To verify the effect of this therapy on the degree of platelet activation, we studied a group of 12 SCD patients receiving sildenafil. Platelet activation was determined from blood samples obtained before and after (120–150 minutes) a single dose of 100 mg. of sildenafil. The mean percentage of activated platelets was higher before than after sildenafil; for activated GPIIb/IIIa 31.2 ± 4.7% vs. 17.1 ±3.9% (p = 0.001); for P-Selectin 6.3 ± 1.6% vs. 3.2 ±0.6% (p = 0.11).
Our findings show for the first time the association of platelet activation and pulmonary hypertension in SCD. Furthermore, by demonstrating that platelet activation of SCD patients with PHT is decreased by sildenafil, a drug that potentiates the effects of NO, this work suggests a common pathophysiology. Additional research is needed to define whether platelet activation plays a role in the thrombo-vascular events that characterize PHT.