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MRGPRX2 and Immediate Drug Hypersensitivity: Insights From Cultured Human Mast Cells

Elst J1, Sabato V1,2, Faber MA1, Bridts CH1, Mertens C1, Van Houdt M1, Van Gasse AL1,3, Hagendorens MM1,3, Van Tendeloo V4, Maurer M5, Campillo-Davo D4, Timmermans J-P6, Pintelon I6, Ebo DG1,2

1Department of Immunology, Allergology, Rheumatology and the Infla-Med Centre of Excellence, Faculty of Medicine and Health Sciences, University of Antwerp, Antwerp, Belgium and Immunology, Allergology, Rheumatology, Antwerp University Hospital, Antwerp, Belgium
2Department of Immunology, AZ Jan Palfijn Hospital Gent, Ghent, Belgium
3Department of Pediatrics, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp, Belgium
4Laboratory of Experimental Haematology, Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Science, University of Antwerp, Antwerp, Belgium
5Dermatological Allergology, Allergie-Centrum-Charité, Department of Dermatology and Allergy, Charité - Universitätsmedizin Berlin, Berlin, Germany
6Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium

J Investig Allergol Clin Immunol 2021; Vol 31(6) : 489-499
doi: 10.18176/jiaci.0557

Background: Mast cell (MC) degranulation via activation of the Mas-related G protein–coupled receptor X2 (MRGPRX2) plays a key role in immediate drug hypersensitivity (IDH). However, data in humans are limited to observations in specific cell lines.
Objective: To study the usefulness of silencing MRGPRX2 in human MCs with the aim of further unveiling the MRGPRX2 pathway in IDH.
Methods: MCs were cultured from CD34+ progenitor cells obtained from peripheral blood (PBCMCs) and incubated with substance P (as a positive control), rocuronium, moxifloxacin, morphine, or amoxicillin. Immunophenotyping of the cells included flow cytometry and microscopy analyses of the expression of CD117, CD203c, and MRGPRX2. Intracellular calcium was measured using Fluo-4. Degranulation was analyzed by quantifying CD63 expression. For MRGPRX2 silencing, MCs were electroporated with Dicer small interference RNAs.
Results: Incubation of MCs with substance P, morphine, and moxifloxacin increased intracellular calcium levels and triggered MC degranulation, which, for the drugs, is almost completely abolished by selective MRGPRX2 silencing. Despite an increase in intracellular calcium in MRGPRX2+ cells, incubation with nontoxic concentrations of rocuronium does not result in degranulation of PBCMCs. Amoxicillin has no effect on PBCMCs.
Conclusion: The use of MRGPRX2 silencing in human MCs can provide important insights into the role of MRGPRX2 in the pathogenesis of IDH. As induction of calcium signals does not necessarily translate into a secretory response, measurement of the degranulation reaction seems more meaningful in the context of drug testing.

Key words: Amoxicillin. Calcium, CD63, Flow cytometry, Mast cells, Morphine, Moxifloxacin, MRGPRX2, Rocuronium