Journal Article

Functional MRGPRX2 expression on peripheral blood-derived human mast cells increases at low seeding density and is suppressed by interleukin-9 and fetal bovine serum

Persoons, Rune, Dupont, Lieven, Goossens, Janne, Schrijvers, Rik, Dilissen, Ellen, Cremer, Jonathan, Roosens, Willem, Jonckheere, Anne-Charlotte, Breynaert, Christine, Bullens, Dominique M. A., Vandenberghe, Peter, Ieven, Toon

2024

Overview

Primary human mast cells (MC) obtained through culturing of blood-derived MC progenitors are the preferred model for the ex vivo study of MRGPRX2- vs. IgE-mediated MC activation. In order to assess the impact of culture conditions on functional MRGPRX2 expression, we cultured CD34 + -enriched PBMC from peripheral whole blood (PB) and buffy coat (BC) samples in MethoCult medium containing stem cell factor (SCF) and interleukin (IL)-3, modified through variations in seeding density and adding or withholding IL-6, IL-9 and fetal bovine serum (FBS). Functional expression of MRGPRX2 was assessed after 4 weeks via flow cytometry. We found similar proportions of CD34 + MC-committed progenitors in BC and PB. Higher seeding densities (≥ 1x10 5 cells/mL) and exposure to IL-9 and FBS suppressed functional MRGPRX2 expression at 4 weeks, while leaving MC yield largely unaffected. IL-6 had no impact on MRGPRX2 expression. MRGPRX2-expressing MC upregulated CD63 upon stimulation with polyclonal anti-IgE, substance P and compound 48/80 at 4 weeks. Ketotifen and dasatinib but not cromolyn sodium inhibited both IgE- and MRGPRX2-dependent pathways. Our results confirm the feasibility of functional MC activation studies on PB-derived MC after a short 4-week culture and highlight the impact of culture conditions on functional MRGPRX2 expression.

• Publisher: Frontiers Media SA,Frontiers Media S.A
• Subject: Animals / Anticoagulants / blood / Blood & organ donations / Buffy coat / Cattle / CD34 antigen / CD63 antigen / Cell culture / Cells, Cultured / Compound 48/80 / culture / Cytokines / Fetal calf serum / Flow cytometry / Hemopoiesis / human / Humans / Immunoglobulin E / Immunology / Interleukin 6 / Interleukin 9 / Interleukin-9 - metabolism / Irritable bowel syndrome / mast cell / Mast cells / Mast Cells - immunology / Mast Cells - metabolism / MRGPRX2 / Nerve Tissue Proteins - metabolism / Peripheral blood mononuclear cells / Receptors, G-Protein-Coupled - genetics / Receptors, G-Protein-Coupled - metabolism / Receptors, Neuropeptide - genetics / Receptors, Neuropeptide - metabolism / Stem cell factor / Stem cells / Substance P