Soluble P-Selectin Levels in Patients with Sickle Cell Disease Reflect Platelets' Activation Rather Than Endothelial Dysfunction

Background: Vascular occlusion is a major cause of the morbidity associated with sickle-cell disease (SCD). The tendency of sickle red blood cells to adhere to the vascular endothelium is believed to be a major contributor and possibly primary cause of the vaso-occlusive process. Therefore mediators of adhesion have become a potential new target for pharmacological therapy to combat the complications of SCD. One of the molecules involved in this process is P-Selectin or CD62P, a cell adherence molecule that is rapidly and chronically expressed on the surface of endothelial cells and platelets when activated. P-Selectin is found in the plasma of normal individuals at ng/mL concentrations. Circulating soluble P-Selectin (sP-Selectin) appears to be slightly smaller than native P-Selectin. An alternatively spliced mRNA encoding a form of human P-Selectin lacking the transmembrane anchoring domain has been reported for both megakaryocytes and endothelial cells, and evidence suggests that the majority of circulating sP-Selectin arises in this manner. We and others have reported elevated sP-Selectin levels in SCD even in steady phase. Recently, a humanized monoclonal antibody that binds to P-Selectin and blocks its interaction with P-selectin glycoprotein ligand 1 (PSGL-1) has been administered to patients with SCD, resulting in an amelioration of painful VOC. In this context we aimed to explore if sP-Selectin levels could be used to choose among the SCD patients those who might benefit from the new therapy.

Patients and Methods: Eighty adult Caucasian patients with HbS/βthal at steady phase [40 patients under hydroxyurea (HU+) treatment and 40 patients without hydroxyurea (HU-) treatment] were included in this study, while 20 apparently healthy individuals of similar age and gender served as controls. Along with sP-Selectin levels, measured with the same method as reported previously (Human sP-Selectin/CD62P Immunoassay, R&D Systems, Minneapolis, MN, USA), other parameters of hemolysis, inflammation, endothelial dysfunction, iron accumulation and clinical features of the disease were evaluated. Results are expressed as median values ± SEM.

Results: We found that sP-Selectin levels were elevated in 45/80 (56%) patients with HbS/βthal compared to controls (108.2±6.3 vs. 69.3±4.1ng/mL, respectively, p<0.001), independently of patients' βthal genotype and correlated strongly with PLT count (r=0.760, p<0.001). Regarding HU treatment, sP-Selectin levels did not differ between (HU+) and (HU-) patients (112.5±9.8 vs. 100.3±7.4ng/mL, respectively, p>0.07). No significance correlation was found between sP-Selectin levels and markers of: hemolysis (RPI: r=0.191, p>0.100); LDH: r=0.103, p>0.360 and bilirubin: r=0.171, p>0.130); inflammation (hs-CRP: r= 0.002, p>0.842); endothelial dysfunction (vWF:antigen: r=0.141, p>0.210 and ADAMTS-13: r=0.089, p>0.507). Regarding iron accumulation no correlation was found between sP-Selectin and ferritin levels (r=0.090, p>0.438), while a weak negative correlation was found with hepcidin-25 levels (r=-0.283, p=0.018). Furthermore, no correlation was found between sP-Selectin levels and history of clinical complications such as VOC (p>0.795), acute chest syndrome, venous and arterial thrombosis and mean pulmonary artery pressure values, (p>0.402).

Conclusion: In this study, we proceeded with an external validation procedure of sP-Selectin determination in patients with HbS/βthal (keeping the same methodology in a different cohort of patients), and we found elevated levels of sP-Selectin with the evidence of PLTs' secretion origin, as no correlation found with other markers of endothelial dysfunction and inflammation. Interestingly, we failed to find a significant link of sP-Selectin levels with other markers of disease severity and/or clinical features of SCD. Thus, we consider that the use sP-Selectin as a biomarker of assessment and treatment of endothelial dysfunction in patients with SCD is of almost negligible importance.