mMrgprA3/mMrgprC11/hMrgprX1: potential therapeutic targets for allergic contact dermatitis induced pruritus in mice and human

Fengxian Li , Changming Wang , Danyou Hu , Xinyu Zhang , Ran Shen , Yuan Zhou , Yan Yang , Chan Zhu , Zongxiang Tang , Guang Yu 

•Department of Anesthesiology, Zhujiang Hospital of Southern Medical University, Guangzhou, Guangdong, China •School of Medicine & Holistic Integrative Medicine, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China •Key Laboratory for Chinese Medicine of Prevention and Treatment in Neurological Diseases, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China

Abstract

Background: Although the Mas-related G-protein-coupled receptors (Mrgprs) play essential roles in itch detection, their contribution to allergic contact dermatitis (ACD) associated itch remains unclear.

Objectives: To investigate whether Mrgprs are involved in ACD and whether Mrgprs can be identified as potential therapeutic targets.

Methods: Mrgpr-clusterΔ^-/- mice and human MrgprX1 (hMrgprX1) transgenic mice were applied to evalute the function of Mrgprs in oxazolone-induced ACD.

Results: Utilizing ACD model, we find that Mrgpr-clusterΔ^-/- mice display significantly reduced pruritus. Among 12 Mrgprs deleted in Mrgpr-clusterΔ^-/- mice, the expression of MrgprC11 and MrgprA3 are significantly increased in ACD model, which also innervate the skin and spinal cord at higher-than-normal densities, the proportions of dorsal root ganglia neurons responding to bovine adrenal medulla peptide 8-22 and chloroquine are also remarkably increased in ACD model and result in enhancing itch behavior. To study the function of human Mrgprs in ACD-induced itch, we utilize hMrgprX1 transgenic mice, which rescues the severe itch defect of Mrgpr-clusterΔ^-/- mice in ACD model. Remarkably, pharmacological blockade of hMrgprX1 significantly attenuates ACD itch in hMrgprX1 transgenic mouse.

Conclusions: Our study provides the first evidence that Mrgprs are involved in ACD induced chronic itch, which provides new avenues for itch management in ACD. This article is protected by copyright. All rights reserved.

Keywords: Allergic contact dermatitis; Mrgprs; pruritus; sensory neurons.

MRGPR-mediated activation of local mast cells clears cutaneous bacterial infection and protects against reinfection

Mohammad Arifuzzaman, Yuvon R. Mobley, HAE WOONG CHOI,

PRADEEP BISTCRISTINA A. SALINASZACHARY D. BROWNSWAINE L. CHEN, HERMAN F. STAATS, SOMAN N. ABRAHAM 

Published in Science advances, 2 Jan 2019

Abstract

Mast cells (MCs) are strategically distributed at barrier sites and prestore various immunocyte-recruiting cytokines, making them ideal targets for selective activation to treat peripheral infections. Here, we report that topical treatment with mastoparan, a peptide MC activator (MCA), enhances clearance of Staphylococcus aureus from infected mouse skins and accelerates healing of dermonecrotic lesions. Mastoparan functions by activating connective tissue MCs (CTMCs) via the MRGPRX2 (Mas-related G protein-coupled receptor member X2) receptor. Peripheral CTMC activation, in turn, enhances recruitment of bacteria-clearing neutrophils and wound-healing CD301b⁺ dendritic cells. Consistent with MCs playing a master coordinating role, MC activation also augmented migration of various antigen-presenting dendritic cells to draining lymph nodes, leading to stronger protection against a second infection challenge. MCAs therefore orchestrate both the innate and adaptive immune arms, which could potentially be applied to combat peripheral infections by a broad range of pathogens.

Histamine enhances ATP-induced itching and responsiveness to ATP in keratinocytes

Yoshihiro Inami, Miki Fukushima, Toshiaki Kume, Daisuke Uta

Abstract

Mechanical stimulation of cultured keratinocytes and a living epidermis increases intracellular calcium ion concentrations ([Ca²⁺]_i) in stimulated cells. This action propagates a Ca²⁺ wave to neighboring keratinocytes via ATP/P2Y₂ receptors. Recent behavioral, pharmacological studies revealed that exogenous ATP induces itching via P2X₃ receptors in mice. We previously showed that alloknesis occurs when an external stimulus is applied to the skin with increased epidermal histamine in the absence of spontaneous pruritus. Based on these results, we investigated the effects of histamine at a concentration that does not cause itching on ATP-induced itching. The mean number of scratching events induced by the mixture of ATP and histamine increased by 28% over the sum of that induced by histamine alone or ATP alone. A317491, a P2X₃ receptor antagonist, suppressed the mixture-induced scratching more often than the ATP-induced scratching. Next, we examined the ATP-induced [Ca²⁺]_i change before and after histamine stimulation using normal human epidermal keratinocytes. Some cells did not respond to ATP before histamine stimulation but responded to ATP afterward, the phenomenon suppressed by chlorpheniramine maleate. These findings suggest that histamine enhances ATP-induced itching and that a potential mechanism could involve increased responsiveness to ATP in keratinocytes.

Keywords: ATP, Histamine, Keratinocytes, Itch, Calcium

presentation by Jong hyun

Type I hypersensitivity promotes Aedes aegypti blood feeding

Michael J. Conway, published in Scientific reports

Abstract:

Mosquitoes play a major role in human disease by serving as vectors of pathogenic microorganisms. Mosquitoes inject saliva into host skin during the probing process. Mosquito saliva contains a number of proteins that facilitate blood feeding by preventing hemostasis. Mosquito saliva also contains potent allergens that induce type I hypersensitivity reactions in some individuals. Type I hypersensitivity reactions in skin involve IgE-mediated degranulation of mast cells, which leads to vasodilation and an itch sensation. We hypothesized that hypersensitivity to mosquito saliva influences blood feeding. To test this hypothesis, we recruited human subjects who consented to Aedes aegypti bites. We measured their first sensation of itch, the strength of their itch sensation, the number of times mosquitoes attempted to feed, the number of times mosquitoes probed their skin, feeding time, engorgement status, and wheal diameter. Here we show that hypersensitive subjects had a stronger itch sensation, and that the time to first itch sensation was inversely correlated with wheal diameter; however, mosquitoes tended to probe less and engorge more on these subjects. Follow-up experiments testing the impact of oral antihistamine treatment on mosquito feeding parameters failed to reveal a statistically significant result. Histamine also failed to promote blood feeding on an artificial membrane feeder. This study suggests that mosquito saliva-induced type
I hypersensitivity promotes blood feeding but that this may be independent from histamine or histamine signaling.

TRPC3 Antagonizes Pruritus in A Mouse Contact Dermatitis Model

Katherine Beattie , Haowu Jiang , Mayank Gautam , Mary K MacVittie , Barbara Miller , Minghong Ma , Qin Liu , Wenqin Luo 

Abstract

Contact dermatitis (CD), including allergic and irritant CD, are common dermatological diseases and characterized by an erythematous rash and severe itch. In this study, we investigated the function of TRPC3, a canonical TRP channel highly expressed in type 1 non-peptidergic (NP1) nociceptive primary afferents and other cell types, in a mouse CD model. Though TrpC3 null mice had little deficits in acute somatosensation, they showed significantly increased scratching with CD. In addition, TrpC3 null mice displayed no differences in mechanic and thermal hypersensitivity in an inflammatory pain model, suggesting that this channel preferentially functions to antagonize CD-induced itch. Using dorsal root ganglia (DRG) and pan-immune-specific TrpC3 conditional KO (CKO) mice, we determined that TrpC3 in DRG neurons, but not in immune cells, is required for this phenotype. Furthermore, the number of MRGPRD⁺ NP1 afferents in CD-affected DRGs is significantly reduced in TrpC3 mutant mice. Taken together, our results suggest that TrpC3 plays a critical role in NP1 afferents to cope with CD-induced excitotoxicity, and that degeneration of NP1 fibers may lead to an increased itch of CD. Our study identified a role of TrpC3 and NP1 afferents in CD pathology.

Keywords: Contact dermatitis; DRG; MRGPRD; TRPC3; non-peptidergic nociceptors; pruritus.

Dissecting the precise nature of itch-evoked scratching

Nivanthika K Wimalasena ¹, George Milner ², Ricardo Silva ³, Cliff Vuong ³, Zihe Zhang ¹, Diana M Bautista ⁴, Clifford J Woolf ⁵

Abstract

Itch is a discrete and irritating sensation tightly coupled to a drive to scratch. Acute scratching developed evolutionarily as an adaptive defense against skin irritants, pathogens, or parasites. In contrast, the itch-scratch cycle in chronic itch is harmful, inducing escalating itch and skin damage. Clinically and preclinically, scratching incidence is currently evaluated as a unidimensional motor parameter and believed to reflect itch severity. We propose that scratching, when appreciated as a complex, multidimensional motor behavior, will yield greater insight into the nature of itch and the organization of neural circuits driving repetitive motor patterns. We outline the limitations of standard measurements of scratching in rodent models and present new approaches to observe and quantify itch-evoked scratching. We argue that accurate quantitative measurements of scratching are critical for dissecting the molecular, cellular, and circuit mechanisms underlying itch and for preclinical development of therapeutic interventions for acute and chronic itch disorders.

Keywords: high-speed recording; itch; pruritis; rodent models; scratching behavior.

Statin-boosted cellular uptake of penetratin due to reduced membrane dipole
potential

published in bioRxiv, 2020.08.04

Gyula Batta, Levente Kárpáti, Gabriela Fulaneto Henrique, Szabolcs Tarapcsák, Tamás Kovács, Florina Zákány, István M. Mándity, Peter Nagy

Abstract

Since cell penetrating peptides are promising tools for delivery of cargo into cells, factors limiting or facilitating their cellular uptake are intensely studied. Using labeling with pH-insensitive and pH-sensitive dyes we report that escape of penetratin from acidic endo-lysosomal compartments is retarded compared to its cellular uptake. The membrane dipole potential, known to alter transmembrane transport of charged molecules, is shown to be negatively correlated with the concentration of penetratin in the cytoplasmic compartment. Treatment of cells with therapeutically relevant concentrations of atorvastatin, an inhibitor of HMG-CoA reductase and cholesterol synthesis, significantly increased the release of penetratin from acidic endocytic compartments in two different cell types. This effect of atorvastatin correlated with its ability to decrease the membrane dipole potential. These results highlight the importance of the dipole potential in regulating cellular uptake of cell penetrating peptides and suggest a clinically relevant way of boosting this process.

Identification of the dog orthologue of human MAS-related G protein coupled receptor X2 (MRGPRX2) essential for drug-induced pseudo-allergic reactions

Published in scientifc reports, 30 september 2020

Eri Hamamura-Yasuno, Takuma Iguchi, Kazuyoshi Kumagai, Yoshimi tsuchiya&Kazuhiko Mori

Abstarct

MAS-related G protein coupled receptor-X2 (MRGPRX2), expressed in human mast cells, is associated with drug-induced pseudo-allergic reactions. Dogs are highly susceptible to drug-induced anaphylactoid reactions caused by various drugs; however, the distribution and physiological function of canine MRGPR family genes, including MRGPRX2, remain largely unknown. In the present study, we clarified the distribution of dog MRGPR family genes by real-time quantitative PCR and in situ hybridisation. We also investigated the stimulatory effects of various histamine-releasing agents, including fluoroquinolones, on HEK293 cells transiently transfected with dog MRGPR family genes to identify their physiological function. Dog MRGPRX2 and MRGPRG were distributed in a limited number of tissues, including the skin (from the eyelid, abdomen, and cheek), whereas MRGPRD and MRGPRF were extensively expressed in almost all tissues examined. Histochemical and in situ hybridisation analyses revealed that MRGPRX2 was expressed in dog connective tissue-type mast cells in the skin. Intracellular Ca²⁺ mobilisation assay revealed that HEK293 cells, expressing dog MRGPRX2 or human MRGPRX2, but not dog MRGPRD, MRGPRF, and MRGPRG, responded to histamine-releasing agents. Our results suggest that dog MRGPRX2 is the functional orthologue of human MRGPRX2 and plays an essential role in drug-induced anaphylactoid reactions in dogs.

Presented by Jong hyun Im

The TRPM3 ion channel mediates nociception but not itch evoked by endogenous pruritogenic mediators

Biochemical Pharmacology, ~ 5.0 (IF)

BalázsKelemen, SilviaPinto, NawooKim, ErikaLisztes, MartinHanyicska, AnitaVladár, AttilaOláh, ZsófiaPénzes, BrianShu, JorisVriens, TamásBíró, TiborRohács, ThomasVoets, Balázs IstvánTóth

Abstract

During the molecular transduction of itch, the stimulation of pruriceptors on sensory fibers leads to the activation or sensitization of ion channels, which results in a consequent depolarization of the neurons. These ion channels mostly belong to the transient receptor potential (TRP) channels, which are involved in nociception and thermosensation. In particular, TRPV1 and TRPA1 were described in the transduction of both thermal nociception as well as histaminergic and non-histaminergic itch. The thermosensitive TRPM3 plays an indispensable role in heat nociception together with TRPV1 and TRPA1. However, the role of TRPM3 in the development of pruritus has not been studied yet. Therefore, in this study we aimed at investigating the potential role of TRPM3 in the transduction of pruritus and pain by investigating itch- and nociception-related behavior of Trpm3^+/+ and Trpm3^−/− mice, and by studying the activation of somatosensory neurons isolated from trigeminal ganglia upon application of algogenic and pruritogenic substances. Activators of TRPM3 evoked only nocifensive responses, but not itch in Trpm3^+/+ animals, and these nocifensive responses were abolished in the Trpm3^−/− strain. Histamine and endogenous non-histaminergic pruritogens induced itch in both Trpm3^+/+and Trpm3^−/− mice to a similar extent. Genetic deletion or pharmacological blockade diminished TRPM3 mediated Ca²⁺ responses of sensory neurons, but did not affect responses evoked by pruritogenic substances. Our results demonstrate that, in contrast to other thermosensitive TRP channels, TRPM3 selectively mediates nociception, but not itch sensation, and suggest that TRPM3 is a promising candidate to selectively target pain sensation.

Hierarchical Specification of Pruriceptors by Runt-Domain Transcription Factor Runx1

Lu Qi, Chengcheng Huang, Xiaohua Wu, Yeqi Tao, Jingjing Yan, Tianyong Shi, Cheng Cao, Lu Han, Mengsheng Qiu, Qiufu Ma, Zijing Liu , Yang Liu 

The Journal of Neuroscience, May 31, 2017

Abstract

The somatic sensory neurons in dorsal root ganglia (DRG) detect and transmit a diverse array of sensory modalities, such as pain, itch, cold, warm, touch, and others. Recent genetic and single-cell RNA sequencing studies have revealed a group of DRG neurons that could be particularly relevant for acute and chronic itch information transmission. They express the natriuretic peptide type B (NPPB), as well as a cohort of receptors and neuropeptides that have been implicated in chronic itch manifestation, including the interleukin-31 receptor A (IL-31ra) and its coreceptor oncostatin M receptor (Osmr), the cysteinyl leukotriene receptor 2 (Cysltr2), somatostatin, and neurotensin. However, how these neurons are generated during development remains unclear. Here we report that Runx1 is required to establish all these molecular features of NPPB⁺ neurons. We further show that while early embryonic Runx1 activity is required for the formation of NPPB⁺ cells, at later stages Runx1 switches to a genetic repressor and thus its downregulation becomes a prerequisite for the proper development of these pruriceptors. This mode by Runx1 is analogous to that in controlling another group of pruriceptors that specifically express the chloroquine receptor MrgprA3. Finally, behavioral studies using both sexes of mice revealed marked deficits in processing acute and chronic itch in Runx1 conditional knock-out mice, possibly attributable to impaired development of various pruriceptors.

SIGNIFICANCE STATEMENT 

Our studies reveal a generalized control mode by Runx1 for pruriceptor development and consolidate a hierarchical control mechanism for the formation of sensory neurons transmitting distinct modalities. Among dorsal root ganglion neurons that initially express the neurotrophin receptor TrkA, Runx1 is necessary for the proper development of those neurons that innervate tissues derived from the ectoderm such as skin epidermis and hair follicles. These Runx1-dependent cutaneous sensory neurons are then divided into two groups based on persistent or transient Runx1 expression. The Runx1-persistent group is involved in transmitting mechanical and thermal information, whereas the Runx1-transient group transmits pruriceptive information. Such hierarchical control mechanisms may provide a developmental solution for the formation of sensory circuits that transmit distinct modalities.

Keywords: NPPB; Runx1; chronic itch; development; pruriceptor; transcriptional regulation.