PS1482 EFFECT OF THROMBOPOIETIN RECEPTOR AGONISTS (TPO‐RAS) ON ENDOTHELIAL CELLS AND NEUTROPHIL EXTRACELLULAR TRAP (NET) FORMATION IN IMMUNE THROMBOCYTOPENIA (ITP)
Authors: Garabet L. , Henriksson C.E. , Ghanima W. , Bussel J. , Mowinckel M.‐C. , Brodin E. , Fernández‐Pérez M.P. , Martínez C. , González‐Conejero R. , Lozano M.L. , Sandset P.M. 
Published: June 2019

Background:

ITP patients have an increased risk of thrombosis, and TPO‐RA‐treatment may further increase this risk. Studies have shown that procoagulant microvesicles (MVs), soluble P‐selectin and plasminogen activator inhibitor‐1 (PAI‐1) are increased in ITP patients and may contribute to the increased thrombotic risk. PAI‐1 and P‐selectin are secreted by the endothelial cells (ECs) suggesting EC involvement. NETs are extracellular DNA fibers comprising histones and neutrophil proteins released to kill pathogens; however, NETs also contribute to inflammation and promote thrombosis. The roles of ECs and NETs in ITP have not been well explored. We hypothesize that MVs interact with ECs, leading to increased expression of adhesive molecules and neutrophil recruitment/adhesion; and that platelet P‐selectin, through binding to PSGL‐1, promotes NET formation that may contribute to the hypercoagulable state.

Aims:

To study the effect of TPO‐RA‐treatment on ECs and NET formation and their roles int he dysregulated coagulation in ITP.

Methods:

We measured intercellular adhesion molecule‐1 (ICAM‐1), vascular adhesion molecular‐1 (VCAM‐1) and soluble thrombomodulin in EDTA‐plasma of 21 ITP patients; and E‐selectin in the serum of 18 ITP patients before, 2 and 6 weeks after the initiation of TPO‐RAs and in matched controls using Quantikine ELISA. We also assayed citrullinated histone H3/DNA(CitH3‐DNA) in EDTA‐plasma (the method by E. Lefrançais‐Looney laboratory) as a measure of NET formation in 15 ITP patients before, 2 and 6 weeks after the initiation of TPO‐RAs and matched controls using ELISA (the method by E. Lefrançais‐Looney Laboratory). Friedman test (non‐parametric)/one‐way ANOVA (parametric) with Dunn's multiple comparisons were used to compare measurements in ITP patients before and after initiation of TPO‐RA‐treatment. Kruskal‐Wallis test was used to compare measurements between ITP‐patients and controls.

Results:

ITP patients pre‐treatment with TPO‐RAs vs controls: ICAM‐1, thrombomodulin, and NETs were higher in ITP patients pre‐treatment (T1) than controls (p = 0.02, p = 0.002, p = 0.003). No differences were found for VCAM‐1 or E‐selectin. (Fig.1)

TPO‐RA‐treated ITP patients (2 and 6 weeks) vs controls: ICAM‐1, thrombomodulin and NETs were higher in ITP patients than controls at 2 weeks (T2) (p = 0.04, p = 0.0005, p = 0.004) and 6 weeks (T3) (p = 0.01, p = 0.0005, p = 0.003) after TPO‐RA‐treatment. No differences were found for VCAM‐1 or E‐selectin. (Fig.1)

TPO‐RA‐treated ITP patients (after 2 and 6 weeks) vs. pre‐treatment: None of the measured parameters showed significant changes after TPO‐RA‐treatment.

Summary/Conclusion:

Elevated ICAM‐1, thrombomodulin, and NETs in ITP patients suggest that EC activation/dysfunction, increased neutrophils adhesion and NET formation may contribute to the hypercoagulable state in ITP patients. However, treatment with TPO‐RAs did not lead to a further increase in EC activation/dysfunction or NET formation.

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