Delu Che

Typical antimicrobials induce mast cell degranulation and anaphylactoid reactions via MRGPRX2 and its murine homologue MRGPRB2

Mast cells are unique immune cells that function as sentinels in host defence reactions, including immediate hypersensitivity responses and allergic responses. The mast cell‐specific receptor named MAS‐related G protein‐coupled receptor X2 (MRGPRX2) triggers mast‐cell degranulation, a key process in anaphylactoid reactions. It is widely observed that antimicrobials can induce pseudo‐allergic reactions (i.e. IgE‐independent mechanism) with symptoms ranging from skin inflammation to life‐threatening systemic anaphylaxis. However, their direct involvement and the mechanisms underlying anaphylactoid reactions caused by antimicrobials have not been demonstrated. Structurally different antimicrobials were screened by Ca2+ imaging using MRGPRX2 overexpressing HEK293 cells. MRGPRX2 related anaphylactoid reactions induced by these components were investigated by body temperature drop and mast cell degranulation assays. We showed that MRGPRX2 is involved in allergic‐like reactions to three types of antimicrobials in a dose‐dependent manner. However, mast cells lacking the receptor show reduced degranulation. Furthermore, mice without MAS‐related G protein‐coupled receptor B2 (the orthologous gene of MRGPRX2) exhibited reduced substance‐induced inflammation. Interestingly, β‐lactam and antiviral nucleoside analogues did not induce anaphylactic reactions, which were also observed in vitro. These results should alarm many clinicians that such drugs might induce anaphylactoid reactions and provide guidance on safe dosage of these drugs.

Interactions between histamine H1 receptor and its antagonists by using cell membrane chromatography method.

A high histamine H1 receptor (H1R) expression cell membrane chromatography (CMC) method was developed to investigate the affinity of ligands for H1R.

Saikosaponin A inhibits compound 48/80-induced pseudo-allergy via the Mrgprx2 pathway in vitro and in vivo

Graphical abstract Figure. No caption available. ABSTRACT Pseudo‐allergic reactions‐adverse, non‐immunologic, anaphylaxis‐like sudden onset reactions mediated through an IgE‐independent pathway—are activated by various basic compounds and occur at least as frequently as IgE‐mediated reactions to drugs. A large family of G protein coupled receptors (Mas‐related genes; Mrgprs) is closely related to pseudo‐allergies. However, few therapies can directly target pseudo‐allergies and related Mrgprs. Saikosaponin A (SSA) is effective in the treatment of passive cutaneous anaphylaxis (PCA), adjuvant arthritis, and delayed hypersensitiveness. In this study, we investigated the anti‐pseudo‐allergy effect of SSA and its underlying mechanism. We examined the effect of SSA on both IgE‐independent and IgE‐dependent responses using PCA and active systemic anaphylaxis models, as well as in vitro‐cultured mast cells. We also evaluated whether the anti‐allergy effect is related to Mrgprs by using in vitro Mrgprx2‐expressing HEK293 cells. SSA dose dependently suppressed compound 48/80 (C48/80)‐induced PCA and mast cell degranulation in mice. When SSA and C48/80 were administered together through the vein, C48/80‐induced systemic anaphylaxis did not occur, and C48/80‐induced shock ratio decreased dose‐dependently upon SSA treatment. However, SSA did not affect IgE‐dependent allergy. When administered topically 24 h before antigen challenge, Evans blue leakage and paw swelling were induced in the SSA‐treated group and the vehicle group. Our in vitro studies revealed that SSA reduced C48/80‐induced calcium flux and suppressed degranulation in LAD2 cells. SSA could also dose‐dependently inhibit C48/80‐induced Mrgprx2‐expressing HEK293 cell activation. As a conclusion, SSA could inhibits IgE‐independent allergy, but not IgE‐dependent allergy, and this effect involves the Mrgprx2 pathway. This study provided a new sight on pseudo‐allergy and its therapy.

Use of the relative release index for histamine in LAD2 cells to evaluate the potential anaphylactoid effects of drugs

Anaphylactoid reactions are common clinical acute adverse drug reactions that can exacerbate a patient’s condition and produce effects that may become life-threatening. Therefore, it is important to establish a novel method to evaluate drugs for anaphylactoid reactions. In this study, we developed a sensitive and rapid method to detect histamine release from LAD2 cells using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and constructed a relative release index based on various release curve parameters, including allergen release time and sudden change rate, to evaluate the potential and strength of allergen-induced anaphylactoid reactions. This LAD2 release model was used to evaluate anaphylactoid reactions induced by ciprofloxacin, norfloxacin, lomefloxacin, moxifloxacin, and baicalin. The results positively correlated with those obtained with an Evans blue ear test and negatively correlated with the Ca2+ influx EC50. In summary, the current study established a novel in vitro method to analyze the properties of histamine release from LAD2 cells and characterize the sensitization and strength of sensitization of drugs or components that may induce anaphylactoid reactions.

MRGPRX2 is essential for sinomenine hydrochloride induced anaphylactoid reactions

Graphical abstract Figure. No Caption available. ABSTRACT Mast cells are unique immunocytes that function as sentinel cells in host defense reactions such as immediate hypersensitivity responses and anaphylactic responses. The mast cell specific receptor MRGPRX2 (Mas‐related G protein‐coupled receptor X2) triggers mast cell degranulation—a key process in anaphylactic reactions. We sought to better understand anaphylactic reaction induced by sinomenine hydrochloride (SH). MRGPRX2‐related pseudo‐allergic reactions induced by SH were investigated using the hindpaw swelling and extravasation assay in vivo and mast cell degranulation assays in vitro. MrgprB2 knockout mice exhibit a reduced SH‐induced inflammation effect. Furthermore, MRGPRX2 (the orthologous gene of MrgprB2) related human mast cells are activated by SH in a dose‐dependent manner; however, MRGPRX2 knockdown mast cells showed reduced degranulation. The results showed a kind of mechanism that SH‐induced anaphylactoid reactions were mediated by MRGPRX2 via activating PLC molecular signaling pathways to provoke mast cells Ca2+ mobilization and degranulation.

The anti-anaphylactoid effects of hydroxysafflor yellow A on the suppression of mast cell Ca2+ influx and degranulation

BACKGROUND Anaphylaxis is a type of potentially fatal hypersensitivity reaction resulting from the activation of mast cell mediators, especially histamine and lipid mediators. Non-IgE-mediated anaphylaxis can occur because of the direct activation of mast cells. Hydroxysafflor yellow A (HSYA) is the main chemical component of safflower (Carthamus tinctorius) and has been reported to have pharmacological activities. However, the anti-anaphylactoid effect of HSYA has not yet been investigated. PURPOSE The aims of this study were to evaluate the anti-anaphylactoid activity of HSYA in vivo and to investigate the underlying mechanism in vitro. METHODS The anti-anaphylactoid activity of HSYA was evaluated in a mouse model of hindpaw extravasation. Calcium imaging was used to assess intracellular Ca2+ mobilization. The levels of cytokines and chemokines released by stimulated mast cells were measured using enzyme immunoassay kits. Western blotting was used to explore the related molecular signaling pathways. RESULTS HSYA markedly inhibited mast cell degranulation by suppressing the activation of intracellular Ca2+ mobilization and preventing the release of cytokines and chemokines from mast cells in a dose-dependent manner via the PKC-PLCγ-IP3R signaling pathway. CONCLUSION In summary, HSYA has anti-anaphylactoid pharmacological activity, which makes it a potential candidate for the development of a novel agent to suppress drug-induced anaphylactoid reactions.

Simultaneous identification of three pseudoallergic components in Danshen injection by using high-expression Mas-related G protein coupled receptor X2 cell membrane chromatography coupled online to HPLC-ESI-MS/MS

Adverse drug reactions of Danshen injection mainly manifested as pseudoallergic reactions. In the present study, salvianolic acid A and a pair of geometric isomers (isosalvianolic acid C and salvianolic acid C) were identified as pseudoallergic components in Danshen injection by a high-expression Mas-related G protein coupled receptor X2 cell membrane chromatography coupled online with high-performance liquid chromatography with electrospray ionization tandem mass spectrometry. Their pseudoallergic activities were evaluated by in vitro assay, which were consistent with the retention times on the cell membrane chromatography column. Salvianolic acid C, the most outstanding compound, was further found to induce pseudoallergic reaction through Mas-related G protein coupled receptor X2. All the results above indicated that the system developed in this study is an effective method for simultaneously analyzing pseudoallergic components, even those with similar structures and the microcomponents in complex samples (salvianolic acid C in Danshen injection).

Relationship between MRGPRX2 and pethidine hydrochloride- or fentanyl citrate-induced LAD2 cell degranulation

Pethidine hydrochloride (PH) and fentanyl citrate (FC) are opioid receptor agonists commonly used to treat pain clinically. PH and FC have been reported to have a high potential for pseudoallergic effects, but the underlying mechanism has not been well studied. MRGPRX2 is a novel atypical opioid receptor that is mainly expressed in human mast cells and considered to mediate drug‐induced pseudoallergic reactions. This study aimed to investigate the allergy effect of these two opioid receptor agonists and the possible association of MRGPRX2 with this response.

Cisatracurium induces mast cell activation and pseudo-allergic reactions via MRGPRX2

Background Pseudo‐allergic reactions occur when patients receive muscle relaxants during perioperative anesthesia. These reactions may result in a serious threat to the patients life, particularly to a childs life. Cisatracurium, a relatively new NMBA, has resulted in bronchospasms and cardiovascular collapse. However, the mechanisms underlying the anaphylactoid reactions caused by cisatracurium have not been fully elucidated. Methods In the present study, the MRGPRX2‐related pseudo‐allergic reactions induced by cisatracurium were investigated using hindpaw swelling and extravasation assays in vivo and mast cell degranulation assays. Results Cisatracurium caused anaphylactoid reactions in wild‐type mice. However, cisatracurium did not induce a similar phenomenon in KitW‐sh/W‐sh mice. Furthermore, mast cell‐related G protein‐coupled receptor B2‐knockout mice did not display an inflammatory response upon treatment with cisatracurium. Cisatracurium induced LAD2 cell degranulation, leading to the dose‐dependent release of &bgr;‐hexosaminidase, histamine and TNF‐&agr;. However, cisatracurium only induced the release of low levels of these mediator LAD2 cells transfected with MRGPRX2 siRNA. Cisatracurium also stimulated intracellular Ca2+ influx in MRGPRX2‐HEK293 cells compared with that in NC‐HKE293 cells. Interestingly, cytokine release was not observed in LAD2 cells even with high dose of cisatracurium. Conclusions Cisatracurium activated MRGPRX2 and triggered mast cell degranulation, leading to anaphylactoid reactions. Therefore, strategies targeting MRGPRX2 might potentially block cisatracurium‐induced pseudo‐allergic reactions. HighlightsCisatracurium induces mast cell degranulation in a dose‐dependent manner.Cisatracurium induces a local inflammatory reaction via MRGPRB2.Cisatracurium induces pre‐synthesis inflammatory modulator release.MRGPRX2 mediates cisatracurium‐induced mast cell activation.

Harpagoside-induced anaphylactic reaction in an IgE-independent manner both in vitro and in vivo

Abstract Background: Harpagoside (HAR) is an active component of Scrophularia ningpoensis (SN), which has anti-inflammatory and anti-immune effects. SN is used widely in China to treat various diseases. Recently, SN has been used as a traditional Chinese medicine injection and used clinically. However, allergic responses to these injections are frequently reported. Aim: We examined whether the main component of SN, HAR, is associated with the allergic reaction to SN. Methods: This study assessed the effects of HAR in mice and mast cell activation to characterize its anaphylactic effects and underlying mechanisms. Mice hindpaw swelling, serum allergy factor detection, enzyme-linked immunosorbent assays, and degranulation assays were performed to measure allergic mediators both in vivo and in vitro. Results: The present study indicated that HAR induced paw swelling, interleukin-6, inositol triphosphate, tumor necrosis factor-α, and histamine increases in mice. Our in vitro data also showed that HAR induced β-hexosaminidase, inositol triphosphate, and interleukin-6 release, leading to mast cell degranulation. In contrast, neither C48/80 nor HAR induced local anaphylaxis in STOCK KitW-sh/HNihrJaeBsmJNju mice. Conclusions: HAR is a potential sensitization compound in SN, and these results provide information for the safe clinical use of SN.