Variable linkage with platelet functions of genetic mutations in ORAI1, STIM1 or FERMT3 in patients with severe immune deficiencies

M. Nagy1, T. G. Mastenbroek1, N. J. A. Mattheij1, S. de Witt1, K. J. Clemetson2, J. Kirschner3, A. S. Schulz4, J. M. E. M. Cosemans1, B. Zieger3, J. W. M. Heemskerk1 (1Maastricht, The Netherlands, 2Bern, Switzerland, 3Freiburg, Germany, 4Ulm, Germany)

Platelets - Physiology and Disorders of platelet number and function
Date: 17.02.2017,
Time: 17:15 - 18:15

Objective: In patients with severe immune-deficiency and mutations in stromal interaction molecule 1 (STIM1), calcium release-activated calcium channel protein 1 (ORAI1) or integrin-regulating protein kindlin-3 (FERMT3), platelet functions and hemostatic activity can be impaired. Here, we aimed to assess the overall consequences of homozygous or heterozygous mutations in these genes on overall platelet functionality in whole-blood thrombus formation.

Methods: Platelet function was analyzed for nine patients, including parents, with genetic mutations in ORAI1, STIM1 or FERMT3. Intracellular calcium release was measured by calibrated fluorometry. Whole blood thrombus formation was measured at arterial- or venous-shear conditions using a multiparameter microspot assay.

Results: In platelets from 4 out of 6 patients with ORAI1 or STIM1 mutations, store-operated Ca2+ entry (SOCE) was completely or incompletely diminished in comparison to the signal of healthy control platelets. The SOCE was greatly improved in one patient after bone marrow transplantation. All patients showed deficiencies in parameters of thrombus formation on collagen microspot due to reduced platelet activation and/or low platelet count. Parameters of thrombus formation on microspots with von Willebrand factor (VWF)/fibrinogen or VWF/rhodocytin were more closely related to the deficiency in SOCE in comparison to those on collagen microspot. However, the extent of the differences was variable within families. For all surfaces, patients platelets with a partial reduction in SOCE showed a marked decrease in phosphatidylserine exposure in comparison to P-selectin expression and integrin activation, both at low and high shear rate. Heat mapped parameters of thrombus formation on all microspots were most severely reduced with blood from a case patient with FERMT3 mutation, and partly diminished with the blood from both parents. A prediction model was built to relate changes in genotype, platelet count and platelet function in thrombus formation.

Conclusion: Within families, mutations in ORAI1, STIM1 or FERMT3 variably link to multiple parameters of altered thrombus formation due to the combination of relatively low platelet counts and impaired platelet deposition and activation.