Li Zhuoran, Schneikert Jean, Bal Gürkan, Zuberbier Torsten, Babina Magda
Institute of Allergology, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Berlin, Germany.
Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, Immunology and Allergology, Berlin, Germany.
Biomolecules. 2025 Aug 25;15(9):1224. doi: 10.3390/biom15091224.
MRGPRX2 (Mas-related G protein-coupled receptor member X2) is implicated in mast cell (MC)-driven disorders due to its ability to bind diverse ligands, which may be G-protein-biased or balanced, with the latter activating both G-proteins and the β-arrestin pathway. Icatibant, a peptide drug, produces injection-site reactions in most patients and is used experimentally to probe MRGPRX2 function in skin tests. While reported to be G-protein-biased, it is unknown how skin MCs respond to icatibant, although these are the primary target cells during therapy. We therefore compared responses to icatibant with those induced by the balanced agonist substance P (SP) in skin MCs. Degranulation and desensitization were assessed via β-hexosaminidase release, receptor internalization by flow cytometry, and downstream signaling by immunoblotting. Skin MCs degranulated in response to SP and icatibant, relying on Gi proteins and calcium channels; Gq and PI3K (Phosphoinositide 3-kinase) contributed more strongly to exocytosis following icatibant, while JNK (c-Jun n-terminal kinase) was more relevant for SP. Both agonists activated ERK, PI3K/AKT, and (weakly) p38. Surprisingly, and in contrast to the LAD2 (Laboratory of Allergic Diseases 2 mast cell line) MC line, icatibant was at least as potent as SP in eliciting MRGPRX2 internalization and (cross-)desensitization in skin MCs. These findings suggest that icatibant functions differently in primary versus transformed MCs, acting as a fully balanced ligand in the former by triggering not only degranulation but also receptor internalization and desensitization. Therefore, not only the ligand but also the MRGPRX2-expressing cell plays a decisive role in whether a ligand is balanced or biased. These findings are relevant to our understanding of icatibant's clinical effects on edema and itch.
MRGPRX2（Mas相关G蛋白偶联受体X2成员）因其能够结合多种配体而与肥大细胞（MC）驱动的疾病有关，这些配体可能偏向G蛋白或平衡型，后者可同时激活G蛋白和β-抑制蛋白途径。肽类药物艾替班特在大多数患者中会产生注射部位反应，并在皮肤试验中用于实验性探究MRGPRX2的功能。虽然据报道它偏向G蛋白，但尚不清楚皮肤MCs对艾替班特的反应如何，尽管这些细胞是治疗期间的主要靶细胞。因此，我们比较了皮肤MCs对艾替班特与平衡型激动剂P物质（SP）诱导反应的差异。通过β-己糖胺酶释放评估脱颗粒和脱敏情况，通过流式细胞术检测受体内化，通过免疫印迹检测下游信号传导。皮肤MCs对SP和艾替班特均有脱颗粒反应，依赖Gi蛋白和钙通道；艾替班特作用后，Gq和PI3K（磷脂酰肌醇3激酶）对胞吐作用的贡献更强，而JNK（c-Jun氨基末端激酶）对SP更重要。两种激动剂均激活ERK、PI3K/AKT和（微弱地）p38。令人惊讶的是，与LAD2（变态反应疾病实验室2肥大细胞系）MC系不同，艾替班特在诱导皮肤MCs中MRGPRX2内化和（交叉）脱敏方面至少与SP一样有效。这些发现表明，艾替班特在原代MCs与转化MCs中的作用不同，在前者中它通过不仅触发脱颗粒，还触发受体内化和脱敏而作为一种完全平衡的配体发挥作用。因此，不仅配体，而且表达MRGPRX2的细胞在配体是平衡型还是偏向型方面都起着决定性作用。这些发现与我们对艾替班特对水肿和瘙痒的临床作用的理解相关。
