RESUMO
Mast cells have existed long before the development of adaptive immunity, although they have been given different names. Thus, in the marine urochordate Styela plicata, they have been designated as test cells. However, based on their morphological characteristics (including prominent cytoplasmic granules) and mediator content (including heparin, histamine, and neutral proteases), test cells are thought to represent members of the lineage known in vertebrates as mast cells. So this lineage presumably had important functions that preceded the development of antibodies, including IgE. Yet mast cells are best known, in humans, as key sources of mediators responsible for acute allergic reactions, notably including anaphylaxis, a severe and potentially fatal IgE-dependent immediate hypersensitivity reaction to apparently harmless antigens, including many found in foods and medicines. In this review, we briefly describe the origins of tissue mast cells and outline evidence that these cells can have beneficial as well as detrimental functions, both innately and as participants in adaptive immune responses. We also discuss aspects of mast cell heterogeneity and comment on how the plasticity of this lineage may provide insight into its roles in health and disease. Finally, we consider some currently open questions that are yet unresolved.
Assuntos
Suscetibilidade a Doenças , Inflamação/etiologia , Inflamação/metabolismo , Mastócitos/imunologia , Mastócitos/metabolismo , Imunidade Adaptativa , Animais , Biomarcadores , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Imunidade Inata , Inflamação/diagnóstico , Mediadores da Inflamação/metabolismo , Transdução de SinaisRESUMO
Mast cells are rare tissue-resident cells of importance to human allergies. To understand the structural basis of principle mast cell functions, we analyzed the proteome of primary human and mouse mast cells by quantitative mass spectrometry. We identified a mast-cell-specific proteome signature, indicative of a unique lineage, only distantly related to other immune cell types, including innate immune cells. Proteome comparison between human and mouse suggested evolutionary conservation of core mast cell functions. In addition to specific proteases and proteins associated with degranulation and proteoglycan biosynthesis, mast cells expressed proteins potentially involved in interactions with neurons and neurotransmitter metabolism, including cell adhesion molecules, ion channels, and G protein coupled receptors. Toward targeted cell ablation in severe allergic diseases, we used MRGPRX2 for mast cell depletion in human skin biopsies. These proteome analyses suggest a unique role of mast cells in the immune system, probably intertwined with the nervous system.
Assuntos
Mastócitos/citologia , Mastócitos/imunologia , Animais , Biomarcadores/metabolismo , Degranulação Celular , Linhagem da Célula , Células Cultivadas , Tecido Conjuntivo/imunologia , Humanos , Imunoterapia , Mastócitos/metabolismo , Proteínas de Membrana/imunologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/imunologia , Proteínas do Tecido Nervoso/metabolismo , Neuroimunomodulação , Proteoglicanas/biossíntese , Proteoma , Receptores Acoplados a Proteínas G/imunologia , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/imunologia , Receptores de Neuropeptídeos/metabolismo , Pele/imunologiaRESUMO
Classical itch studies have focused on immunoglobulin E (IgE)-mediated mast cell activation and histamine release. Recently, members of the Mas-related G-protein-coupled receptor (Mrgpr) family have been identified as mast cell receptors, but their role in itch is unclear. Here, we report that mast cell activation via Mrgprb2 evoked non-histaminergic itch in mice independently of the IgE-Fc epsilon RI (FcεRI)-histamine axis. Compared with IgE-FcεRI stimulation, Mrgprb2 activation of mast cells was distinct in both released substances (histamine, serotonin, and tryptase) and the pattern of activated itch-sensory neurons. Mrgprb2 deficiency decreased itch in multiple preclinical models of allergic contact dermatitis (ACD), a pruritic inflammatory skin disorder, and both mast cell number and PAMP1-20 concentrations (agonist of the human Mrgprb2 homolog, MRGPRX2) were increased in human ACD skin. These findings suggest that this pathway may represent a therapeutic target for treating ACD and mast-cell-associated itch disorders in which antihistamines are ineffective.
Assuntos
Mastócitos/imunologia , Proteínas do Tecido Nervoso/metabolismo , Prurido/patologia , Receptores Acoplados a Proteínas G/metabolismo , Receptores de IgE/metabolismo , Receptores de Neuropeptídeos/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Feminino , Histamina/metabolismo , Antagonistas dos Receptores Histamínicos/uso terapêutico , Humanos , Imunoglobulina E/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Fragmentos de Peptídeos/metabolismo , Receptores Acoplados a Proteínas G/genética , Serotonina/metabolismo , Pele/metabolismo , Triptases/metabolismo , Adulto JovemRESUMO
Allergic asthma is a chronic inflammatory disease of airways involving complex mechanisms, including MAS-related GPR family member X2 (MRGPRX2) and its orthologue MRGPRB2 on mast cells (MCs). Although miRNAs have been previously shown to related to allergic asthma, the role of miR-212/132 in this process has not been studied. In this study, the predicted pairing of miRNAs and MRGPRX2 (MRGPRB2) mRNAs was carried out by online databases and the function was verify using in vivo and in vitro experiments. Database prediction showed that miR-212/132 interact with MRGPRX2 and MRGPRB2. miR-212/132 mimics alleviated MRGPRB2 mRNA expression as well as pathology changes in lungs and AHR of mice with airway inflammation in vivo. The expression level of MRGPRB2 in the mice lungs after inhaled OVA was also decreased by miR-212/132 mimics. Meanwhile, miR-212/132 inhibited MCs degranulation and cytokines release triggered by C48/80 in vitro. Further, ASAP1 (ARF GTPase-Activating Protein 1) was selected from the junction related pathways using RNAseq and KEGG enrichment. ASAP1 mRNA level was upregulated in airway inflammation and MCs activation and decreased by miR-212/132 mimics. miR-212/132 attenuated OVA-induced airway inflammation by inhibiting MCs activation through MRGPRX2 and ASAP1.
Assuntos
Asma , Mastócitos , MicroRNAs , Animais , Camundongos , Asma/induzido quimicamente , Asma/genética , Asma/metabolismo , Degranulação Celular , Inflamação/induzido quimicamente , Inflamação/metabolismo , Mastócitos/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Ovalbumina/efeitos adversos , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , RNA Mensageiro/metabolismo , HumanosRESUMO
BACKGROUND: Drug-induced anaphylaxis is triggered by the direct stimulation of mast cells (MCs) via Mas-related G protein-coupled receptor X2 (MRGPRX2; mouse ortholog MRGPRB2). However, the precise mechanism that links MRGPRX2/B2 to MC degranulation is poorly understood. Dedicator of cytokinesis 2 (DOCK2) is a Rac activator predominantly expressed in hematopoietic cells. Although DOCK2 regulates migration and activation of leukocytes, its role in MCs remains unknown. OBJECTIVE: We aimed to elucidate whether-and if so, how-DOCK2 is involved in MRGPRX2/B2-mediated MC degranulation and anaphylaxis. METHODS: Induction of drug-induced systemic and cutaneous anaphylaxis was compared between wild-type and DOCK2-deficient mice. In addition, genetic or pharmacologic inactivation of DOCK2 in human and murine MCs was used to reveal its role in MRGPRX2/B2-mediated signal transduction and degranulation. RESULTS: Induction of MC degranulation and anaphylaxis by compound 48/80 and ciprofloxacin was severely attenuated in the absence of DOCK2. Although calcium influx and phosphorylation of several signaling molecules were unaffected, MRGPRB2-mediated Rac activation and phosphorylation of p21-activated kinase 1 (PAK1) were impaired in DOCK2-deficient MCs. Similar results were obtained when mice or MCs were treated with small-molecule inhibitors that bind to the catalytic domain of DOCK2 and inhibit Rac activation. CONCLUSION: DOCK2 regulates MRGPRX2/B2-mediated MC degranulation through Rac activation and PAK1 phosphorylation, thereby indicating that the DOCK2-Rac-PAK1 axis could be a target for preventing drug-induced anaphylaxis.
Assuntos
Anafilaxia , Hipersensibilidade a Drogas , Humanos , Camundongos , Animais , Anafilaxia/induzido quimicamente , Degranulação Celular , Mastócitos/metabolismo , Receptores de Neuropeptídeos/genética , Receptores de Neuropeptídeos/metabolismo , Hipersensibilidade a Drogas/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismoRESUMO
Two potent and selective KRASG12D inhibitors, ERAS-4693 and ERAS-5024, were generated as possible clinical candidates to treat patients harboring G12D mutations in solid tumors. Both molecules exhibited strong anti-tumor activity in the KRASG12D mutant PDAC xenograft mouse models while ERAS-5024 also showed tumor growth inhibition when administered on an intermittent dosing regimen. Acute dose-limiting toxicity consistent with an allergic reaction was observed for both molecules shortly after administration at doses just above those which demonstrated anti-tumor activity, indicative of a narrow therapeutic index. A series of studies were subsequently conducted to identify a common underlying mechanism for the observed toxicity, including CETSA® (Cellular Thermal Shift Assay) as well as several functional off-target screens. Both ERAS-4693 and ERAS-5024 were identified to agonize MRGPRX2 which has been linked to pseudo-allergic reactions. In vivo toxicologic characterization of both molecules included repeat-dose studies in the rat and dog. Dose-limiting toxicities were observed in both species with ERAS-4693 and ERAS-5024 and plasma exposure levels at the maximum tolerated doses were generally below that which caused strong anti-tumor activity, supporting the initial observation of a narrow therapeutic index. Additional overlapping toxicities included a reduction in reticulocytes and clinical pathological changes suggestive of an inflammatory response. Furthermore, increases in plasma histamine were observed in dogs administered ERAS-5024, supporting the hypothesis that MRGPRX2 agonism may be the cause of the pseudo-allergic reaction. This work highlights the importance of balancing both the safety and efficacy of KRASG12D inhibitors as this class of molecules begins to enter clinical development.
Assuntos
Hipersensibilidade , Neoplasias Pancreáticas , Humanos , Camundongos , Ratos , Animais , Cães , Proteínas Proto-Oncogênicas p21(ras)/genética , Neoplasias Pancreáticas/patologia , Mutação , Proteínas do Tecido Nervoso , Receptores de Neuropeptídeos/genética , Receptores Acoplados a Proteínas G/genéticaRESUMO
Hydroquinone (HQ) is one of the most effective drugs to treat hyperpigmentary disorders, but often causes skin irritation in clinic. Mast cell plays an important role in contact dermatitis and triggering pseudo-allergic reactions via MRGPRX2. Whether HQ-induced skin irritant reaction through activating mast cells via MRGPRX2 remains unknown. To investigate the role of mast cells in HQ-induced skin irritant reaction and verify whether MRGPRX2 participated in the HQ effect on mast cells which contributed to the pathogenesis of skin irritant reaction, a mouse model of HQ-induced skin irritation was established to observe the local and systemic inflammation associated with mast cell receptor MrgprB2. Human mast cell LAD2 was used to verify the effect of HQ on mast cells via MRGPRX2 by knocking down with siRNA. As a result, mast cells were involved in the development of HQ-induced irritant reaction, and local inflammation is closely related to mast cell receptor MrgprB2. HQ could activate mast cells via MRGPRX2, causing changes in calcium concentration, degranulation and release of inflammatory cytokines which lead to skin irritant reaction. In conclusion, HQ-induced skin irritant reaction could be skin pseudo-allergic reactions achieved by activating mast cells via MRGPRX2.
Assuntos
Dermatite Atópica , Hipersensibilidade , Animais , Camundongos , Humanos , Mastócitos/patologia , Irritantes/toxicidade , Hidroquinonas/efeitos adversos , Receptores Acoplados a Proteínas G/genética , Inflamação/patologia , Dermatite Atópica/patologia , Degranulação Celular , Proteínas do Tecido Nervoso/genética , Receptores de Neuropeptídeos/genéticaRESUMO
MRGPRX2, a novel Gaq -coupled human mast cell receptor, mediates non-immune adverse reactions without the involvement of antibody priming. Constitutively expressed by human skin mast cells, MRGPRX2 modulates cell degranulation producing pseudoallergies manifesting as itch, inflammation and pain. The term pseudoallergy is defined in relation to adverse drug reactions in general and immune/non-immune-mediated reactions in particular. A list of drugs with MRGPRX2 activity is presented, including a detailed examination of three important and widely used approved therapies: neuromuscular blockers, quinolones and opioids. For the clinician, the significance of MRGPRX2 is considered as an aid in distinguishing and ultimately identifying specific immune and non-immune inflammatory reactions. Anaphylactoid/anaphylactic reactions, neurogenic inflammation and inflammatory diseases with a clear or strongly suspected association with MRGPRX2 activation are examined. Inflammatory diseases include chronic urticaria, rosacea, atopic dermatitis, allergic contact dermatitis, mastocytosis, allergic asthma, ulcerative colitis and rheumatoid arthritis. MRGPRX2- and allergic IgE/FcεRI-mediated reactions may be clinically similar. Importantly, the usual testing procedures do not distinguish the two mechanisms. Currently, identification of MRGPRX2 activation and diagnosis of pseudoallergic reactions is generally viewed as a process of exclusion once other non-immune and immune processes, particularly IgE/FcεRI-mediated degranulation of mast cells, are ruled out. This does not take into account that MRGPRX2 signals via ß-arrestin, which can be utilized to detect MRGPRX2 activation by employing MRGPRX2 transfected cells to assess MRGPRX2 activation via two pathways, the G-protein-independent ß-arrestin pathway and the G-protein-dependent Ca2+ pathway. Testing procedures, interpretations for distinguishing mechanisms, patient diagnosis, agonist identification and drug safety evaluations are addressed.
Assuntos
Anafilaxia , Receptores de IgE , Humanos , Receptores de IgE/metabolismo , Receptores de Neuropeptídeos/metabolismo , Mastócitos/metabolismo , Inflamação , Imunoglobulina E , Proteínas de Ligação ao GTP/metabolismo , beta-Arrestinas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Proteínas do Tecido Nervoso/metabolismoRESUMO
Insights into the pathophysiology of many non-immune-mediated drug reactions referred to as toxicities, sensitivities, intolerances, or pseudoallergies have resulted from research identifying the mastocyte-related G-protein-coupled receptor (GPCR) member X2 (MRGPRX2), a human mast cell receptor mediating adverse reactions without the involvement of antibody priming. Opioid-induced degranulation of mast cells, particularly morphine, provoking release of histamine and other preformed mediators and causing hemodynamic and cutaneous changes seen as flushing, headache and wheal and flare reactions in the skin, is an example of results of MRGPRX2 activation. Opioids including morphine, codeine, dextromethorphan and metazocine as well as endogenous prodynorphin opioid peptides activate MRGPRX2 at concentrations causing mast cell degranulation. Unlike the canonical opioid receptors, MRGPRX2 shows stereochemical recognition preference for dextro rather than levo opioid enantiomers. Opioid analgesic drugs (OADs) display a range of histamine-releasing potencies from the strong releaser morphine to doubtful releasers like hydromorphone and the non-releaser fentanyl. Whether there is a correlation between histamine release by individual OADs, MRGPRX2 activation, and presence or absence of adverse cutaneous effects is not known. To investigate the question, ongoing research with recently pursued methodologies and strategies employing basophil and mast cell tests resulting from MRGPRX2 insights should help to elucidate whether or not an opioid's histamine-releasing potency, and its property of provoking an adverse reaction, are each a reflection of its activation of MRGPRX2.
Assuntos
Analgésicos Opioides , Hipersensibilidade , Humanos , Analgésicos Opioides/toxicidade , Histamina/farmacologia , Receptores de Neuropeptídeos , Receptores Acoplados a Proteínas G , Derivados da Morfina/farmacologia , Mastócitos , Degranulação Celular , Proteínas do Tecido NervosoRESUMO
Vinpocetine injection is often used in clinical treatment of acute cardiovascular and cerebrovascular diseases. However, it was reported that vinpocetine injection caused allergic reactions in clinical use; therefore, its safety needs urgent attention. Until now, research on its sensitization is rarely reported. Here, the components contained in three vinpocetine injections were examined. It was found that besides vinpocetine, the synthetic raw material vincamine, the excipients benzyl alcohol and ethyl p-toluenesulfonate, and the impurities A, B, C, and D, which are excipients specified in the European Pharmacopoeia, were also present in them. Then the Mas-related G-protein-coupled receptor X2 (MRGPRX2)-HEK293 cell membrane chromatography was used to investigate the affinity of them with MRGPRX2 and found that vinpocetine, vincamine, and impurities A, B, C, and D bind to MRGPRX2. Afterwards, these compounds were further used to investigate the local sensitization ability in vivo. The results showed that vinpocetine, vincamine, and impurity C could induce swelling of the paw and decrease body temperature in mice, but only impurity C could cause local skin mast cell degranulation and serum histamine release increase. In vitro, the results also indicated that impurity C could increase intracellular [Ca2+ ] in MRGPRX2-HEK293 cells, whereas vinpocetine and vincamine did not. Therefore, the impurity C was the potential anaphylactoid component in vinpocetine injection, which may be one of the reasons for the occurrence of allergic reactions in the clinical use of vinpocetine injection. This work provides evidence on the sensitization of impurity C and also contributes to promoting the clinical safety of vinpocetine injection.
Assuntos
Anafilaxia , Vincamina , Humanos , Animais , Camundongos , Células HEK293 , Anafilaxia/induzido quimicamente , Vincamina/metabolismo , Vincamina/uso terapêutico , Excipientes , Receptores Acoplados a Proteínas G/metabolismo , Membrana Celular/metabolismo , Cromatografia , Mastócitos/metabolismo , Degranulação Celular , Proteínas do Tecido Nervoso/metabolismo , Receptores de Neuropeptídeos/metabolismo , Receptores de Neuropeptídeos/uso terapêuticoRESUMO
BACKGROUND: Chronic prurigo (CPG) is characterized by intensive itch and interactions among nerves, neuropeptides, and mast cells (MCs). The role of some neuropeptides such as cortistatin (CST) and its receptor, Mas-related G protein-coupled receptor X2 (MRGPRX2), in CPG remains poorly investigated. OBJECTIVES: We evaluated first whether CST activates human skin MCs, and second whether CST and MRGPRX2 are expressed in the skin of CPG patients, and by which cells. METHODS: Skin prick tests and microdialysis with CST were performed in 6 and 1 healthy volunteers, respectively. Degranulation of human skin MCs was assessed using ß-hexosaminidase and histamine release assays. Skin samples from 10 patients with CPG and 10 control subjects were stained for CST, MCs, and MRGPRX2 (protein and mRNA) using immunohistochemistry, immunofluorescence, and/or in situ hybridization. Flow cytometry was used to assess CST in human skin MCs. MRGPRX2 levels were measured in serum by ELISA. RESULTS: CST induced concentration-dependent degranulation of human skin MCs in vivo and ex vivo. Skin lesions of CPG patients exhibited markedly higher numbers of CST-expressing cells, CST-expressing MCs, MRGPRX2-expressing cells, and MRGPRX2 mRNA-expressing cells than nonlesional skin. MCs were the main MRGPRX2 mRNA-expressing cells in the lesions of most CPG patients (70%). Stimulation of human skin MCs with anti-IgE led to a release of CST. The number of MRGPRX2-expressing cells correlated with disease severity (r = 0.649, P = .04). MRGPRX2 serum levels in CPG patients correlated with disease severity (r = 0.704, P = .023) and quality-of-life impairment (r = 0.687, P = .028). CONCLUSIONS: CST and MRGPRX2 may contribute to the pathogenesis of CPG and should be evaluated in further studies as potential biomarkers and novel therapeutic targets.
Assuntos
Neuropeptídeos , Prurigo , Degranulação Celular , Humanos , Mastócitos/fisiologia , Proteínas do Tecido Nervoso/genética , RNA Mensageiro , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/genéticaRESUMO
BACKGROUND: P17, a peptide isolated from Tetramorium bicarinatum ant venom, is known to induce an alternative phenotype of human monocyte-derived macrophages via activation of an unknown G protein-coupled receptor (GPCR). OBJECTIVE: We sought to investigate the mechanism of action and the immunomodulatory effects of P17 mediated through MRGPRX2 (Mas-related G protein-coupled receptor X2). METHODS: To identify the GPCR for P17, we screened 314 GPCRs. Upon identification of MRGPRX2, a battery of in silico, in vitro, ex vivo, and in vivo assays along with the receptor mutation studies were performed. In particular, to investigate the immunomodulatory actions, we used ß-hexosaminidase release assay, cytokine releases, quantification of mRNA expression, cell migration and differentiation assays, immunohistochemical labeling, hematoxylin and eosin, and immunofluorescence staining. RESULTS: P17 activated MRGPRX2 in a dose-dependent manner in ß-arrestin recruitment assay. In LAD2 cells, P17 induced calcium and ß-hexosaminidase release. Quercetin- and short hairpin RNA-mediated knockdown of MRGPRX2 reduced P17-evoked ß-hexosaminidase release. In silico and in vitro mutagenesis studies showed that residue Lys8 of P17 formed a cation-π interaction with the Phe172 of MRGPRX2 and [Ala8]P17 lost its activity partially. P17 activated LAD2 cells to recruit THP-1 and human monocytes in Transwell migration assay, whereas MRGPRX2-impaired LAD2 cells cannot. In addition, P17-treated LAD2 cells stimulated differentiation of THP-1 and human monocytes, as indicated by the enhanced expression of macrophage markers cluster of differentiation 11b and TNF-α by quantitative RT-PCR. Immunohistochemical and immunofluorescent staining suggested monocyte recruitment in mice ears injected with P17. CONCLUSIONS: Our data provide novel structural information regarding the interaction of P17 with MRGPRX2 and intracellular pathways for its immunomodulatory action.
Assuntos
Peptídeos/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Animais , Sítios de Ligação , Permeabilidade Capilar/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Quimiotaxia/efeitos dos fármacos , Cricetulus , Citocinas/metabolismo , Edema/imunologia , Edema/metabolismo , Azul Evans/metabolismo , Inativação Gênica , Humanos , Masculino , Mastócitos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Receptores Acoplados a Proteínas G/genéticaRESUMO
Anaphylaxis is a life-threatening or even fatal systemic hypersensitivity reaction. The incidence of anaphylaxis has risen at an alarming rate in the past decades in the majority of countries. Generally, the most common causes of severe or fatal anaphylaxis are medication, foods and Hymenoptera venoms. Anaphylactic reactions are characterized by the activation of mast cells and basophils and the release of mediators. These cells express a variety of receptors that enable them to respond to a wide range of stimulants. Most studies of anaphylaxis focus on IgE-dependent reactions. The mast cell has long been regarded as the main effector cell involved in IgE-mediated anaphylaxis. This paper reviews IgE-independent anaphylaxis, with special emphasis on mast cells, basophils, anaphylactic mediators, risk factors, triggers, and management.
Assuntos
Anafilaxia , Estimulantes do Sistema Nervoso Central , Humanos , Mastócitos , Basófilos , Imunoglobulina ERESUMO
Mast cells (MCs) are tissue-resident immune cells of a hematopoietic origin that play vital roles in innate and adaptive immunity. Human MCs can be isolated and differentiated from various tissue sources, including cord blood, when supplemented with cytokines such as stem cell factor, interleukin 3, and interleukin 6. Our current research study has shown significant differences in the marker expressions of human cord blood-derived mast cells (hCBMCs) based on donor dependency and the type of medium used for culturing and differentiation. These findings are particularly relevant given the challenges of obtaining specialty media influencing MC phenotypic marker expressions. We found that hCBMCs cultured in StemSpanTM-XF medium had a moderate expression of mast/stem cell growth factor receptor Kit (c-KIT) (mRNA and protein), low expressions of FcεRI (mRNA) and TLR2 (mRNA and protein) but had high levels of MRGPRX2 (mRNA and protein) expressions. In contrast, hCBMCs cultured in Stem Line II medium expressed FcεRI and TLR2 (mRNA and protein) with higher c-KIT but had lower MRGPRX2 expressions compared to the hCBMCs cultured in the StemSpanTM-XF medium. These results suggest that it is crucial to consider both donor dependency and the medium when investigating MC functions and that further research is needed to fully understand the impact of these factors on the hCBMC marker expressions.
Assuntos
Sangue Fetal , Mastócitos , Humanos , Mastócitos/metabolismo , Receptor 2 Toll-Like , Células Cultivadas , Diferenciação Celular , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas do Tecido Nervoso , Receptores de Neuropeptídeos , Receptores Acoplados a Proteínas GRESUMO
Tightly controlled inflammation is an indispensable mechanism in the maintenance of cellular and organismal homeostasis in living organisms. However, aberrant inflammation is detrimental and has been suggested as a key contributor to organ injury with different etiologies. Substance P (SP) is a neuropeptide with a robust effect on inflammation. The proinflammatory effects of SP are achieved by activating its functional receptors, namely the neurokinin 1 receptor (NK1R) receptor and mas-related G protein-coupled receptors X member 2 (MRGPRX2) and its murine homolog MRGPRB2. Upon activation, the receptors further signal to several cellular signaling pathways involved in the onset, development, and progression of inflammation. Therefore, excessive SP-NK1R or SP-MRGPRX2/B2 signals have been implicated in the pathogenesis of inflammation-associated organ injury. In this review, we summarize our current knowledge of SP and its receptors and the emerging roles of the SP-NK1R system and the SP-MRGPRX2/B2 system in inflammation and injury in multiple organs resulting from different pathologies. We also briefly discuss the prospect of developing a therapeutic strategy for inflammatory organ injury by disrupting the proinflammatory actions of SP via pharmacological intervention.
Assuntos
Receptores da Neurocinina-1 , Substância P , Camundongos , Animais , Substância P/metabolismo , Receptores da Neurocinina-1/metabolismo , Inflamação/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/metabolismoRESUMO
The signs and symptoms of chronic urticaria (CU) are caused by the activation and degranulation of skin mast cells (MCs). Recent studies have added to our understanding of how and why skin MCs are involved and different in CU. Also, novel and relevant mechanisms of MC activation in CU have been identified and characterized. Finally, the use of MC-targeted and MC mediator-specific treatments has helped to better define the role of the skin environment, the contribution of specific MC mediators, and the relevance of MC crosstalk with other cells in the pathogenesis of CU. Here, we review these recent findings and their impact on our understanding of CU, with a focus on chronic spontaneous urticaria (CSU). Also, we highlight open questions, issues of controversy, and unmet needs, and we suggest what studies should be performed moving forward.
Assuntos
Urticária Crônica , Urticária , Humanos , Urticária/diagnóstico , Mastócitos , Pele/patologiaRESUMO
Morphine derivatives are clinically important anesthetic and sedative drugs, which often show anaphylactic side effects. Mas-related G-protein coupled receptor member X2 (MRGPRX2) triggers mast cell degranulation, which is important process in anaphylactic reactions. MRGPRX2-HEK293 and LAD2 cell membrane chromatographic (CMC) models were used to screen morphine derivatives binding to MRGPRX2. Furthermore, most morphine derivatives significantly enhanced Ca2+ mobilization. More importantly, thebaine was found to effectively promote histamine release. Thebaine induced the increased release of ß-hexosaminidase and high secretion level of cytokines, confirming that thebaine could further trigger anaphylactic reactions and promote subsequent inflammatory reactions. Moreover, the ability of thebaine inducing degranulation and the release of allergenic mediators in mast cells was significantly decreased after MRGPRX2 knockdown, which proved that MRGPRX2 is the key media for thebaine-induced anaphylactic reactions. Significant hind paw swelling and hypothermia in mice after injecting thebaine suggested that thebaine could trigger anaphylactic reactions in vivo.
Assuntos
Anafilaxia , Mastócitos , Proteínas do Tecido Nervoso , Receptores Acoplados a Proteínas G , Receptores de Neuropeptídeos , Tebaína , Anafilaxia/induzido quimicamente , Animais , Degranulação Celular , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/genética , Tebaína/efeitos adversosRESUMO
Mas-related G protein-coupled receptor X2 (MRGPRX2) mediates the itch response in neurons and is involved in atopic dermatitis (AD)-associated inflammation and itch. Potent and MRGPRX2-selective ligands are essential to an understanding of the detailed function of the receptor and to develop new therapeutic agents for its related diseases. (+)-TAN-67 (1), the enantiomer of the δ-opioid receptor (DOR) selective ligand (-)-TAN-67 (1), has been reported to activate MRGPRX2, although (+)-1 also interacts with DOR, which prevents investigators from interrogating the function of MRGPRX2. Here, we have succeeded in developing a novel unnatural morphinan compound (+)-2a by a transformation based on the structure of (+)-1, which removes the DOR binding affinity. (+)-2a activated both human MRGPRX2 and the mouse orthologue Mrgprb2 in in vitro experiments and induced itch-like behaviors in mice to the same extent as (+)-1. The (+)-2a-induced itch response in mice was suppressed by administration of the tripeptide QWF, an MRGPRX2/Mrgprb2 antagonist, or the antipruritic drug nalfurafine. Together, (+)-2a serves as a useful tool to elucidate the itch-related function/action of MRGPRX2 and its mouse orthologue Mrgprb2.
Assuntos
Comportamento Animal/efeitos dos fármacos , Desenvolvimento de Medicamentos , Morfinanos/efeitos adversos , Proteínas do Tecido Nervoso/metabolismo , Prurido/induzido quimicamente , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/metabolismo , Animais , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Humanos , Ligantes , Camundongos , Estrutura Molecular , Morfinanos/síntese química , Morfinanos/química , Proteínas do Tecido Nervoso/antagonistas & inibidores , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores de Neuropeptídeos/antagonistas & inibidores , Receptores Opioides delta , Relação Estrutura-AtividadeRESUMO
Pseudo-allergic reactions frequently occur following clinical drug use and sometimes even cause mortal danger. Mas-related G-protein-coupled receptor member X2 (MRGPRX2) is a novel receptor that mediates pseudo-allergy and is an important target in the treatment of allergies. However, to date, there are no synthetic small-molecule inhibitors that prevent anaphylactoid reactions through this pathway. Our preliminary research suggested that B10-S and mubritinib effectively inhibited LAD2 cells. Therefore, two novel derivatives were synthesized by integrating the active substructures of B10-S and mubritinib, according to the molecular docking results. The antiallergic inhibitory effects of the two compounds were preliminarily evaluated in vitro using ß-hexosaminidase release, histamine release, and intracellular Ca2+ mobilization assays, and their binding sites on MRGPRX2 were analyzed by molecular docking. Both substances inhibited ß-hexosaminidase and histamine release in LAD2 cells and decreased intracellular Ca2+ by inhibiting MRGPRX2 in MRGPRX2-HEK293 cells treated with C48/80 in a dose-dependent manner. The docking results suggested that the molecules could competitively bind to the active site on MRGPRX2 and Glu141, which were combined by C48/80. Our study indicated that the two compounds have potential anti-allergic properties, which may provide evidence that will facilitate the development of synthetic molecules with anti-pseudo-allergic activity for clinical use in the future.
Assuntos
Anafilaxia/tratamento farmacológico , Antialérgicos/farmacologia , Hipersensibilidade/tratamento farmacológico , Proteínas do Tecido Nervoso/metabolismo , Oxazóis/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Neuropeptídeos/metabolismo , Triazóis/farmacologia , Anafilaxia/metabolismo , Antialérgicos/síntese química , Antialérgicos/química , Linhagem Celular , Relação Dose-Resposta a Droga , Células HEK293 , Humanos , Hipersensibilidade/metabolismo , Estrutura Molecular , Oxazóis/síntese química , Oxazóis/química , Relação Estrutura-Atividade , Triazóis/síntese química , Triazóis/químicaRESUMO
WHAT IS KNOWN AND OBJECTIVE: Red man syndrome (RMS) is a non-IgE-mediated anaphylactoid adverse event frequently witnessed after a rapid infusion of vancomycin. This study aims to unravel drugs and associated off-label targets that induce RMS by exploiting FDA Adverse Event Reporting System (FAERS) and Pharmacovigilance/Pharmacogenomics Insilico Pipeline (PHARMIP). METHODS: The case/non-case retrospective observational study was conducted in the FAERS database. Reporting odds ratio (ROR) and proportional reporting ratio (PRR) data mining algorithms were used to evaluate the strength of the signal. The off-label targets of the drugs with potential signals were obtained using online servers by applying a similarity ensemble approach and a reverse pharmacophore database, which was further validated by molecular docking studies. RESULTS AND DISCUSSION: Oritavancin exhibited a strong positive signal (PRR:1185.20 and ROR:1256), which suggests a higher risk for causing RMS. The literature search revealed the involvement of the MRGPRX2 gene in the development of RMS. PHARMIP study unearthed Carbonic anhydrase II (CA2) as the common off-label target among the drugs causing RMS. The results obtained from molecular docking studies reinforced the findings as mentioned earlier, wherein the highest docking score was disinterred for oritavancin (-9.4 for MRGPRX2 and - 8.7 for CA2). WHAT IS NEW AND CONCLUSION: Many antibiotics and other classes of medications have been discovered in the quest for drugs that may induce RMS, although a causal relationship could not be established. The implication of MRGPX2 and CA2 in the initial stages of pathogenesis necessitates the development of inhibitors that could be used as potential therapeutic agents against RMS.