Bawazir Maram, Sutradhar Sangita, Roy Saptarshi, Ali Hydar
Department of Basic and Translational Sciences, School of Dental Medicine, University of Pennsylvania, Philadelphia, Pa; Department of Oral Diagnostic Sciences, Faculty of Dentistry, King Abdulaziz University, Jeddah, Saudi Arabia.
Department of Basic and Translational Sciences, School of Dental Medicine, University of Pennsylvania, Philadelphia, Pa.
J Allergy Clin Immunol. 2025 Mar;155(3):974-987.e1. doi: 10.1016/j.jaci.2024.11.021. Epub 2024 Nov 22.
In addition to FcεRI, a subtype of human mast cells (MCs) expresses Mas-related G protein-coupled receptor X2 (MRGPRX2; mouse counterpart MrgprB2). Although MrgprB2 contributes to IgE-mediated passive systemic anaphylaxis (PSA) in vivo, an MRGPRX2 inhibitor, compound 9 (C9), does not block MrgprB2- or IgE-mediated MC degranulation in vitro.
Our aim was to generate mice expressing human MRGPRX2 to study receptor function in vitro and PSA in vivo.
The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-mediated gene editing approach was utilized to replace endogenous MrgprB2 with human MRGPRX2 in mice (MRGPRX2-KI mice). MRGPRX2 expression in the skin, gingiva, trachea, and colon were evaluated by using an anti-human MRGPRX2 antibody. Peritoneal MCs (PMCs) cultured from wild-type, MRGPRX2-KI, and MrgprB2 mice were used to study agonists-induced degranulation. The effects of selective MRGPRX2 inhibitors (C9 and compound 9-6 [C9-6]) on substance P- or IgE-mediated MC degranulation in vitro and IgE-mediated PSA in vivo were tested.
MRGPRX2-expressing MCs were present in tissues of MRGPRX2-KI mice. Most of the agonists tested induced greater degranulation at lower concentrations in PMCs from MRGPRX2-KI mice than in cells from wild-type mice. Furthermore, C9 and C9-6 inhibited degranulation in MRGPRX2-KI PMCs in response to substance P. In contrast, they had no effect on IgE-mediated degranulation in vitro but did inhibit PSA in MRGPRX2-KI mice in vivo.
MRGPRX2-KI mice provide a readily available source of primary MCs for signaling studies. Furthermore, transactivation of MRGPRX2 contributes to IgE-mediated PSA, suggesting that MRGPRX2-KI mice could be utilized as a preclinical model for testing novel therapeutics targeting MRGPRX2 and its cross talk with FcεRI.
除了FcεRI外，人类肥大细胞（MC）的一种亚型表达与Mas相关的G蛋白偶联受体X2（MRGPRX2；小鼠对应物为MrgprB2）。尽管MrgprB2在体内促成IgE介导的被动全身过敏反应（PSA），但一种MRGPRX2抑制剂化合物9（C9）在体外并不阻断MrgprB2或IgE介导的MC脱颗粒。
我们的目的是培育表达人类MRGPRX2的小鼠，以研究该受体的体外功能及体内PSA。
利用成簇规律间隔短回文重复序列（CRISPR）/Cas9介导的基因编辑方法，在小鼠体内用人类MRGPRX2替代内源性MrgprB2（MRGPRX2基因敲入小鼠）。使用抗人类MRGPRX2抗体评估皮肤、牙龈、气管和结肠中MRGPRX2的表达。从野生型、MRGPRX2基因敲入和MrgprB2小鼠培养的腹膜MC（PMC）用于研究激动剂诱导的脱颗粒。测试了选择性MRGPRX2抑制剂（C9和化合物9-6 [C9-6]）对体外P物质或IgE介导的MC脱颗粒以及体内IgE介导的PSA的影响。
MRGPRX2基因敲入小鼠的组织中存在表达MRGPRX2的MC。与野生型小鼠细胞相比，大多数测试的激动剂在较低浓度下诱导MRGPRX2基因敲入小鼠的PMC脱颗粒程度更高。此外，C9和C9-6抑制MRGPRX2基因敲入小鼠的PMC对P物质的脱颗粒反应。相反，它们对体外IgE介导的脱颗粒无影响，但在体内确实抑制了MRGPRX2基因敲入小鼠的PSA。
MRGPRX2基因敲入小鼠为信号研究提供了现成的原代MC来源。此外，MRGPRX2的反式激活促成IgE介导的PSA，这表明MRGPRX2基因敲入小鼠可作为一种临床前模型，用于测试靶向MRGPRX2及其与FcεRI相互作用的新型疗法。
