• The MRGPR family is a type of GPCR that is responsible for specifically sensing itch, pain, and inflammation. MRGPRX2 is a receptor expressed on mast cells found in barrier tissues such as the skin, airways, and gastrointestinal tract. It can detect a variety of secretagogues, including polycations and peptides, that trigger the mast cell degranulation response, leading to pseudo-allergic reactions. While itching can prompt scratching behaviors that serve a protective defense function for the body, excessive activation of the receptor can result in the development of various diseases, including allergies and inflammation. Therefore, by blocking the activation of MRGPRX2 and mast cell degranulation, MRGPRX2 antagonists hold potential for the broad treatment of mast cell-mediated diseases.
• The human MRGPRX2 protein was detectable only in mast cells of heterozygous and homozygous humanized mice.
• After the induction by substance P (SP), allergic reactions can be observed in both heterozygous and homozygous mice. SP produced an increase in extensively degranulated cells.
• B-hMRGPRX2 mice strain was obtained by mating a mouse with a loxP-flanked STOP cassette that prevents transcription of a CAG promoter-driven human MRGPRX2, all inserted into the Gt(ROSA)26Sor locus, with a Mrgprb2 Cre-mediated mouse. B-hMRGPRX2 mice express robust human MRGPRX2 following Cre-mediated recombination.

Gene targeting strategy for B-hMRGPRX2 mice. B-hMRGPRX2 mice strain was obtained by mating a mouse with a loxP-flanked STOP cassette that prevents transcription of a CAG promoter-driven human MRGPRX2, all inserted into the Gt(ROSA)26Sor locus, with a Mrgprb2 Cre-mediated mouse. B-hMRGPRX2 mice express robust human MRGPRX2 following Cre-mediated recombination. This strain is congenic on the C57BL/6JNifdc genetic background.

MRGPRX2 expression analysis in homozygous B-hMRGPRX2 mice by flow cytometry. Peritoneal lavage fluid cells were collected from wild-type C57BL/6JNifdc mice (+/+), heterozygous B-hMRGPRX2 mice (H/+) and homozygous B-hMRGPRX2 mice (H/H), and analyzed by flow cytometry with anti-human MRGPRX2 antibody(BioLegend, 359004). Human MRGPRX2 was detectable on peritoneal lavage mast cells in heterozygous and homozygous humanized mice.

Substance P induced Mast cell degranulation. Mice received substance P (MCE; HY-P0201) injection on the left ear, PBS injection on the right ear. After 2 min later, an intravenous injection of Evans blue (0.5%, 200 μL) was given to mice 20 min after Evans blue injection (A). Ears of mice were collected, the amount of dye was determined after extraction with 700 μl of Formamide (B). The intensity of absorbance (100 μL) was measured at 620 nm (C). Two-way ANOVA with Sidak's test *P < 0.05, **P < 0.01, n=6. After the induction by substance P, allergic reactions can be observed in both heterozygous and homozygous mice.

Substance P induced Mast cell degranulation.  Ears were collected for Swiss-Giemsa stain Quantification of MC degranulation in the ears 20 min after injection; MC degranulation determined by classification in 3 categories: not degranulated (None), moderately degranulated (Mod), and extensively degranulated (Ext) (A). SP produced an increase in extensively degranulated cells (B, C, D)