PB1285 - Glycosylation and functional defects of platelets from XMEN disease patients

Tuesday, June 25, 2024

13:45 – 14:45 ICT

Room: Exhibition Hall

Presenting Author(s)

NB

Nora Butta

Researcher
Grupo de Coagulopatías y Trastornos de la Hemostasia, IdiPaz, Madrid, Spain
Madrid, Madrid, Spain

Co-Author(s)

Ld

Lucía del Pino Molina

Researcher
Clinical Immunology Unit, Hospital Universitario La Paz – IdiPAZ. Center for Biomedical Network Research on Rare Diseases (CIBERER U767)
Madrid, Madrid, Spain
EM

Elena Monzón Manzano

Researcher
Department of Haematology, Hospital Universitario La Paz, Madrid, Spain. IdiPAZ, Madrid, Spain
Madrid, Madrid, Spain
Elena G.Arias Salgado

Researcher
Grupo de Coagulopatías y Trastornos de la Hemostasia, IdiPaz, Madrid, Spain
Madrid, Madrid, Spain
EL

Eduardo López Granados

MD, PhD
Clinical Immunology Unit, Hospital Universitario La Paz – IdiPAZ,. Center for Biomedical Network Research on Rare Diseases (CIBERER U767)
Madrid, Madrid, Spain
PA

Paula Acuña Butta

researcher
Department of Haematology, Hospital Universitario La Paz-IdiPaz
Madrid, Madrid, Spain
CG

Carla Gianelli

MD, PhD
Clinical Immunology Unit, Hospital Universitario La Paz – IdiPAZ. Center for Biomedical Network Res
Madrid, Madrid, Spain
Víctor Jiménez-Yuste

Head of Hematology Service
Hospital Universitario La Paz-IdiPaz
Madrid, Madrid, Spain
María Teresa Alvarez-Román, MD, PhD

Head of the Haemostasis Unit
Department of Hematology, University Hospital La Paz, Autonoma University of Madrid, Madrid, Spain
Madrid, Madrid, Spain

Background: XMEN disease is caused by loss of function mutations in MAGT1, a plasma membrane transporter of magnesium that is also a subunit of the oligosaccharyl-transferase complex involved in the N-glycosylation of proteins in the endoplasmic reticulum.

Aims: To characterize glycome and function of platelets from XMEN patients

Methods: Blood samples from 3 brothers and from one unrelated patient genetically diagnosed with XMEN and from healthy donors were collected in citrate.
Intracellular Ca2+ fluxes were determined by flow cytometry in washed platelets loaded with 1μM Fura Red and stained with mAb anti-CD61 PE. Basal intracellular Ca2+ levels were measured for 60 s, followed by stimulation with 100 µmol/L TRAP. Finally platelets were stimulated with 2 μg/mL ionomycin to measure maximum Ca2+ signaling.
Platelet activation markers and receptors were determined in diluted platelet rich plasma. Platelets were incubated with 100 µmol/L TRAP and either FITC-PAC1, or FITC-anti-P-selectin mAb or FITC-anti-CD63 mAb; and with PE-anti-CD41 mAb to determine fibrinogen receptors. All samples were analysed by flow cytometry.
Lectin binding studies were performed in washed platelets labeled with FITC- lectins listed in Table 1 and analysed by flow cytometry.
Informed consent was obtained and the study was approved by Ethics Committee of Hospital Universitario La Paz.

Results: MAGT1 mutation caused a reduction in Ca2+ fluxes induced by TRAP (Fig. 1A). Accordingly, activation of fibrinogen receptor, and P-selectin and CD63 exposure induced by TRAP was reduced in XMEN patients (Fig. 1B).Diminution in TRAP-induced activation was not due to a reduction in the content of fibrinogen receptors on platelets from XMEN patients.
Regarding glycoside residues on platelet surface, an increase was observed in binding of concavalin and a reduction in RCA, Dolichos biflorus and Wheat germ Agglutinin (Fig.1C).

Conclusion(s): These results suggest that reduced functionality of MAGT1 impaired platelet function and induced changes in platelet’s glycome.

Work supported by Cátedra Roche-UAM: 063203.