American Journal of Translational Research
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2025
·Vol. 17
·期 3
·2178
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IF 1.6
Yu Zhang,Fang-Mei Liu,Cun-Yu Li,Xue-Jiao Leng,Yun-Feng Zheng,Guo-Ping Peng
Objective: Qingkailing Injection (QKLI) is a traditional Chinese medicine injection mainly used for sedation, heat clearing, and other treatments. However, recent clinical studies have shown a risk of pseudo-allergic reactions. The purpose of this study is to elucidate the underlying mechanism of QKLI-induced mast cell degranulation in Laboratory of Allergic Diseases 2 (LAD2) and to validate QKLI-induced activation of guinea pig IgE-independent allergic responses. Methods: Levels of β-hexosaminidase (β-Hex), histamine (His), and complement pathway-related indicators in guinea pigs and LAD2 cells were assayed using the Enzyme-linked Immunosorbent Assay (ELISA). The release rates of β-Hex and His from LAD2 cells were measured using the o-phthalaldehyde (OPA) fluorimetric method. The antagonist for complement component 3a (C3a) receptors, SB290157 and siRNAs were used to inhibit the C3a pathway and the Mas-related G-protein-coupled receptor X2 (MRGPRX2) pathway. The MRGPRX2 pathway and its downstream proteins were detected by Western Blot (WB). Results: The results show that QKLI significantly increased levels of β-Hex, His, C3a, complement component 5a (C5a), and terminal complement complex C5b-9 (SC5b-9) in guinea pigs, while levels of interleukin 4 (IL-4), interleukin 13 (IL-13), and interleukin 6 (IL-6) were unaffected. The C3a receptor inhibitor SB290157 significantly reduced levels of β-Hex and His. In LAD2 cells, QKLI increased the release rates of β-Hex and His in a time-dependent manner and decreased the phosphorylation of Extracellular Signal-regulated Kinase 1/2 (ERK1/2) proteins downstream of the MRGPRX2 pathway. The effective components of QKL, baicalin (BA) and geniposide (GE), individually enhance the allergic responses of LAD2 cells to some extent. However, the use of QKL is significantly superior to the individual use of its components. Conclusions: We found that QKLI induced pseudoanaphylaxis via an IgE-independent response in guinea pigs and through the MRGPRX2 pathway in human LAD2 cells. Among these, the main ingredients causing pseudoallergic reactions in QKLI were BA and GE. Our research contributes to a better understanding of the mechanisms underlying drug hypersensitivity reactions (DHRs).
研究领域:
药理学
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免疫学
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DOI
PubMed
高影响因子
DYES AND PIGMENTS
·
2025
·
IF 4.2
Xiaoxue Mo,Yuexin Lv,Jiayu Lu,Yajing Hou,Hua Qian,Cheng Wang,Huaizhen He
MAS-related G protein-coupled receptor X2 (MRGPRX2) is a key target receptor and an important drug target for triggering pseudo-allergic reactions. Studying its ligands will help to understand the mechanism of pseudo-allergic reactions and discover anti-allergic drugs. Small-molecule fluorescent probes are widely used to study the localization of G protein-coupled receptors (GPCRs) on the cell membrane, the transport and internalization of GPCRs, and the interaction between receptors and ligands due to their high selectivity, sensitivity and safety. MRGPRX2 is one of GPCRs with an active hydrophobic ligand binding domain, so designing small-molecule fluorescent probes with environmental sensitivity is very important for the study of MRGPRX2 ligands and the visualization of this receptor. In this study, we selected the agonist of MRGPRX2 ( R )-ZINC-3573 as the recognition group and used different fluorophore skeleton structures as the luminescent group to synthesize a series of small-molecule fluorescent probes targeting MRGPRX2. Among them, PoX2-B with BODIPY FL as the fluorescent group had good photostability and environmentally sensitive properties to the hydrophobic ligand binding domain of MRGPRX2, and could be used for living cell imaging. Furthermore, PoX2-B could specifically recognize MRGPRX2, and it was expected to be used to screen effective ligands of MRGPRX2 in the future, so as to play a role in the development of anti-allergy drugs and the pathogenesis of allergic diseases.
研究领域:
药理学
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生物化学
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分子生物学
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DOI
高影响因子
ANALYTICAL BIOCHEMISTRY
·
2021
·
IF 3.365
Yuanyuan Ding,Xiaoqian Li,Qingpeng Gao,Xinyan Dong,Liyun Kong,Shengli Han,Tao Zhang,Langchong He
Mas-related G-protein-coupled receptor X2 (MRGPRX2) has recently been reported to be associated with anaphylaxis. Detection of MRGPRX2 levels in human peripheral blood might serve as a powerful tool for predicting the predisposition of patients to anaphylactic reactions. For rapid measurement of MRGPRX2, we established a paper-based double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) using mouse monoclonal antibody and horseradish peroxidase (HRP)-labelled rabbit polyclonal antibody as capture antibody and detection antibody, respectively. We avoided chemical functionalization of the cellulose paper by introducing bovine serum albumin (BSA) to provide COOH and NH 2 groups for covalent immobilization of the capture antibody. Through amide condensation , a two-layer immobilization strategy was applied with BSA-BSA and BSA-capture antibody networks as the first and second layers, respectively. This strategy improved the quantity, activity and stability of the immobilized antibody . We then established a paper-based ELISA to detect MRGPRX2 in human peripheral blood. Our method is less laborious, easier to implement, and more cost-effective than conventional ELISA, while offering similar sensitivity, specificity, and accuracy. Therefore, it could serve as an innovative clinical point-of-care diagnostic tool, especially in areas that lack advanced clinical equipment.
研究领域:
医学
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免疫学
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生物化学
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高影响因子
Toxicology Research
·
2021
·Vol. 10
·期 3
·467-475
·
IF 3.524
Gao Jiapan,Che Delu,Du Xueshan,Zheng Yi,Jing Huiling,Wang Nan
Imidazolidinyl urea (IU) is used as an antimicrobial preservative in cosmetic and pharmaceutical products. IU induces allergic contact dermatitis, however, the mechanism has not yet been elucidated. Mas-related G protein-coupled receptor-X2 (MRGPRX2) triggers drug-induced pseudo-allergic reactions. The aims of this study were to determine whether IU activated mast cells through MRGPRX2 to further trigger contact dermatitis. Wild-type (WT) and KitW-sh/HNihrJaeBsmJNju (MUT) mice were treated with IU to observe its effects on local inflammation and mast cells degranulation in vivo. Laboratory of allergic disease 2 cells were used to detect calcium mobilization and release of inflammatory mediators in vitro. WT mice showed a severe local inflammatory response and contact dermatitis, whereas only slight inflammatory infiltration was observed in MUT mice. Thus, MRGPRX2 mediated the IU-induced activation of mast cells. However, histamine, a typical allergen, was not involved in this process. Tryptase expressed by mast cells was the major non-histaminergic inflammatory mediator of contact dermatitis. IU induced anaphylactic reaction via MRGPRX2 and further triggering non-histaminergic contact dermatitis, which explained why antihistamines are clinically ineffective against some chronic dermatitis.
研究领域:
免疫学
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皮肤科
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药理学
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DOI
PubMed
高影响因子
INTERNATIONAL IMMUNOPHARMACOLOGY
·
2023
·
IF 5.6
Yuanyuan Ding,Tianyou Ma,Yonghui Zhang,Chenrui Zhao,Chao Wang,Zhao Wang
Background Allergic contact dermatitis (ACD) is one of the most common dermatoses, which has high disease burden and quality of life impairment. Anti-histamine is not effective in a part of the ACD patients. Thus, the discovery of novel antipruritic therapy is of highly demand. Objective In this study, we investigated the anti-pruritic effects of rosmarinic acid (RA) and explored the underlying mechanism. Method SPF Balb/c mice were randomly divided into control group, ACD model group, RA group (1.0 mg/kg) and loratadine (LORA) group (1.5 mg/kg). Back epidermal thickness was recorded. H&E staining was used for pathological observation. Mast cell degranulation was assessed by toluidine blue staining. ELISA assay was employed to detect cytokines levels. Cortistatin-14 (CST-14) and Mas-related G protein-coupled receptor X2 (MRGPRX2) expression was detetcted by RT-PCR and western blot. Molecular docking assay was used to predict the affinity of RA and MRGPRX2. Surface plasmon resonance (SPR) assay was used to verify structure affinity of RA and MRGPRX2. Results RA treatment significantly decreased epidermal keratinization and inflammatory cell infiltration in ACD mouse model. Administration of RA significantly reduced secretion of histamine, IL-13, and mRNA expression of CST-14. Furthermore, RA treatment increased mRNA expression of MRGPRX2. In addition, Molecular docking results predict that RA has a good affinity with MRGPRX2. RA displayed a structure affinity ( K D = 8.89 × 10 -4 ) with MRGPRX2 by SPR. RA inhibited CST-14 and Compound 48/80 (C48/80)-induced mast cell activation via MRGPRX2-PLCγ1-PKC-NF-κB signaling pathway. Conclusion RA exhibits anti-pruritic and anti-inflammatory effects in ACD mice by inhibiting MRGPRX2-PLCγ1-PKC-NF-κB signaling pathway. RA might emerge as a potential drug for the treatment of pruritus and skin inflammation in the setting of ACD.
研究领域:
皮肤科
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免疫学
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药理学
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产品关联
DOI
PubMed
高影响因子
BIOCHEMICAL PHARMACOLOGY
·
2020
·
IF 4.96
Nan Wang,Jue Wang,Yongjing Zhang,Yingnan Zeng,Shiling Hu,Haoyun Bai,Yajing Hou,Cheng Wang,Huaizhen He,Langchong He
Background Allergic asthma is a common inflammatory lung disease associated with complex pathogenesis. Mast cell (MC) is one of the key drivers of allergic asthma, Mas-related G protein-coupled receptor X2 (MRGPRX2) on the MC could mediate MC activation and trigger a pseudo-allergic reaction. Imperatorin (IMP), the main active compound of Radix Angelicae Dahuricae , has been reported to exert various pharmacological effects. In this study, we focused on the therapeutical mechanism of IMP on MRGPRX2-induced pseudo-allergy and allergic asthma. Methods We examined the effect of IMP on MRGPRX2 related mast cell activation in mouse peritoneal MC (MPMC), Human Laboratory of Allergic Disease 2 MCs (LAD2 cells) and Mrgprx2 -expressing HEK293 cells. Molecular docking and Surface plasmon resonance (SPR) were taken to reveal the binding character between IMP and MRPGRX2. MRGPRX2 downstream proteins were also detected by western blotting . IgE-independent responses was evaluated by using passive cutaneous anaphylaxis (PCA) and active systemic anaphylaxis (ASA) models. The therapeutic effect of IMP on asthma was evaluated by a lung inflammation mouse model which was induced by ovalbumin (OVA). Results IMP was found to reduce substance P (SP) induced calcium flux and suppressed degranulation of MC. SP can promote the phosphorylation of ERK and CamKII, which regulates the synthesis of inflammatory factors such as MIP-2 and TNF-α in MC. In vivo assays revealed that IMP can mitigate SP-induced mouse PCA and ASA. IMP could also mitigate lung inflammation in an OVA induced mice model by inhibiting MC activation in the lung tissue. Furthermore, IMP binds well to MRGPRX2 protein. The binding constant ( K D ) is 4.48 ± 0.49 × 10 −7 M. The data suggeste that IMP is a novel inhibitor of MRGPRX2 to treat allergic asthma.
研究领域:
药理学
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免疫学
·
过敏
·
呼吸系统疾病
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DOI
PubMed
高影响因子
MICROCHEMICAL JOURNAL
·
2025
·
IF 5.1
Nannan Ren,Ping Wang,Chao Wang,Liyun Kong,Jiangang Long
Background Mas-related G-protein-coupled receptor X2 (MRGPRX2) is critical for mast cell degranulation and pseudo-allergic reactions. Detecting MRGPRX2 in human peripheral blood serum is essential for studying mast cell-mediated disorders like chronic spontaneous urticaria (CSU), asthma, and atopic dermatitis. While enzyme-linked immunosorbent assay (ELISA), known for sensitivity and specificity, is limited by the instability of enzymes and antibodies, hindering cost-effectiveness and robustness. Metal-organic frameworks (MOFs) with mild synthesis conditions, excellent biocompatibility, and peroxidase-like catalytic activity were selected as alternatives to conventional enzymes, thereby offering a promising approach to enhance the stability and efficiency of ELISA. Results In this study, Cu-MOF, which exhibiting excellent biocompatibility and catalytic performance, was selected as an alternative to traditional enzymes. The in-situ synthesized anti-MRGPRX2@Cu-MOF retained its antigen recognition capability and enzyme-like catalytic activity across a broad pH range (3–10) and at elevated temperatures (45°C-85°C). The established ELISA method demonstrated a detection limit of 1.660 ng/mL for MRGPRX2. The method exhibited satisfactory accuracy, with recovery rates ranging from 97.95% to 108.68% for intra-assay assessment and from 95.18% to 110.66% for inter-assay assessment. The precision was also acceptable, showing intra- and inter-assay coefficients of variation below 13.94% and 14.41%, respectively. The concentration of MRGPRX2 antigen in human serum was successfully detected using the established ELISA method based on anti-MRGPRX2@Cu-MOF. This method is characterized by its low cost, high stability, and excellent recovery rates and precision. Significance The established ELISA method based on the nanozyme-labeled antibody demonstrated favorable recovery rates and precision for detecting the CSU biomarker MRGPRX2. This method offers several advantages such as cost-effectiveness and the capability for visual result interpretation, making it a promising candidate for future applications in portable detection devices and clinical diagnostics.
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高影响因子
BIOMEDICINE & PHARMACOTHERAPY
·
2022
·
IF 7.419
Cheng Wang,Yajing Hou,Shuai Ge,Jiayu Lu,Xiangjun Wang,Yuexin Lv,Nan Wang,Huaizhen He
Anaphylaxis is a severe systemic allergic reaction that exhibits multiple clinical symptoms. The Mas-related G protein-coupled receptor X2 (MRGPRX2) is recognized as a key cell receptor mediating allergic diseases and drug-induced anaphylactoid reactions. Thus, it has been a promising target for preventing and treating these reactions. Based on the potential activity of imperatorin and active structural feature of MRGPRX2, we first demonstrated that the synthetic imperatorin derivatives (IDs) could significantly inhibit MRGPRX2 agonist-induced degranulation and cytokine release in LAD2 cells, as well as alleviate local and systemic anaphylaxis in mice. The IC 50 value of the most promising compound is an order of magnitude lower than that of imperatorin. IDs were further identified to display anti-pseudo-allergic activity by binding MRGPRX2 with the tertiary nitrogen substructures, just liking the reported MRGPRX2-ligand. These results would propose evidence for discovery of agents for treating MCs-dependent allergic disorders.
研究领域:
免疫学
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药理学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
TALANTA
·
2025
·
IF 6.1
Yuanyuan Ding,Ping Wang,Na Wang,Liyun Kong,Min Li,Jing Wang,Tao Zhang
The immunoassay methods, stand as the gold standard in biomedicine, are widely used in diagnostics. However, they often encounter inherent limitations in practical applications, such as the poor stability of enzymes and antibodies, and the laborious process of preparing enzyme-antibody conjugates through covalent bonding. Mas-related G-protein-coupled receptor X2 (MRGPRX2) has been identified as a blood-based diagnostic biomarker for chronic spontaneous urticaria (CSU). In this study, a bifunctional enzyme-labeled antibody (anti-MRGPRX2&HRP@ZIF-8) was successfully constructed by integrating horseradish peroxidase (HRP) into ZIF-8 during its synthesis, followed by adsorption of anti-MRGPRX2 onto the surface of HRP@ZIF-8. The resulting anti-MRGPRX2&HRP@ZIF-8 complex not only preserved its antigen-capturing capability but also exhibited improved stability. The established chemiluminescence immunoassay (CLIA) method based on anti-MRGPRX2&HRP@ZIF-8 demonstrated enhanced stability and achieved a lower detection limit (1.035 ng/mL) compared to conventional enzyme-linked immunosorbent assay (ELISA) method. Furthermore, MRGPRX2 was successfully detected in serum samples by the established method, demonstrating its suitability for the diagnosis of CSU in real clinical samples. Consequently, this study presented a promising approach for simplifying the preparation of enzyme-antibody conjugates and highlighted the potential of metal-organic framework (MOF)-based composites in biomedical applications.
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PubMed
高影响因子
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