Journal Club 18.01.12.

Transient receptor potential vanilloid 4–expressing macrophages and keratinocytes contribute differentially to allergic and nonallergic chronic itch

TRP4-expressing macrophages and keratinocytes contribute differentially to allergic and nonallergic chronic itch

Jialie Luo, PhD,a* Jing Feng, PhD,a* Guang Yu, PhD,a,b Pu Yang, PhD,a Madison R. Mack, BA,a Junhui Du, PhD,c Weihua Yu, PhD,d Aihua Qian, PhD,e Yujin Zhang, PhD,f Shenbin Liu, PhD,a Shijin Yin, PhD,g Amy Xu, BS,a Jizhong Cheng, PhD,h Qingyun Liu, PhD,i Roger G. O’Neil, PhD,j Yang Xia, PhD,f Liang Ma, PhD,k
Susan M. Carlton, PhD,c Brian S. Kim, MD,a,k Kenneth Renner, PhD,l Qin Liu, PhD,a and Hongzhen Hu, PhDa
St Louis, Mo; Nanjing, Chongqing, Shanghai, and Wuhan, China; Galveston and Houston, Tex; and Vermillion, SD

Background: Chronic itch is a highly debilitating symptom that underlies many medical disorders with no universally effective treatments. Although unique neuronal signaling cascades in the sensory ganglia and spinal cord have been shown to critically promote the pathogenesis of chronic itch, the role of skin- associated cells remains poorly understood.
Objective: We sought to examine the cutaneous mechanisms underlying transient receptor potential vanilloid 4 (TRPV4)–mediated allergic and nonallergic chronic itch. Methods: Expression of TRPV4 in chronic itch and healthy control skin preparations was examined by using real-time RT-PCR. Trpv4eGFP mice were used to study the expression and function of TRPV4 in the skin by means of immunofluorescence staining, flow cytometry, calcium imaging, and patch-clamp recordings. Genetic and pharmacologic approaches were used to examine the role and underlying mechanisms of TRPV4 in mouse models of dry skin–associated chronic itch and spontaneous scratching associated with squaric acid dibutylester–induced allergic contact dermatitis.
Results: TRPV4 is selectively expressed by dermal macrophages and epidermal keratinocytes in mice. Lineage-specific deletion of TRPV4 in macrophages and keratinocytes reduces allergic and nonallergic chronic itch in mice, respectively. Importantly, TRPV4 expression is significantly increased in skin biopsy specimens from patients with chronic idiopathic pruritus in comparison with skin from healthy control subjects. Moreover, TRPV4-dependent chronic itch requires 5-hydroxytryptamine (5-HT) signaling secondary to activation of distinct 5-HT receptors in both patients with allergic and those with nonallergic chronic itch conditions.
Conclusion: Our study reveals previously unrecognized mechanisms by which TRPV4-expressing epithelial and immune cells in the skin critically and dynamically mediate chronic itch and unravels novel targets for therapeutics in the setting of chronic itch. (J Allergy Clin Immunol 2017;nnn:nnn-nnn.)
Key words: Transient receptor potential vanilloid 4, chronic itch, macrophage, keratinocyte

Journal Club 18.01.12. Read More »

2018.01.05

UVB radiation generates sunburn pain and affects skin by activating epidermal TRPV4 ion channels and triggering endothelin-1 signaling.

Moore C1, Cevikbas F, Pasolli HA, Chen Y, Kong W, Kempkes C, Parekh P, Lee SH, Kontchou NA, Yeh I, Jokerst NM, Fuchs E, Steinhoff M, Liedtke WB.

Abstract

At our body surface, the epidermis absorbs UV radiation. UV overexposure leads to sunburn with tissue injury and pain. To understand how, we focus on TRPV4, a nonselective cation channel highly expressed in epithelial skin cells and known to function in sensory transduction, a property shared with other transient receptor potential channels. We show that following UVB exposure mice with induced Trpv4 deletions, specifically in keratinocytes, are less sensitive to noxious thermal and mechanical stimuli than control animals. Exploring the mechanism, we find that epidermal TRPV4 orchestrates UVB-evoked skin tissue damage and increased expression of the proalgesic/algogenic mediator endothelin-1. In culture, UVB causes a direct, TRPV4-dependent Ca(2+) response in keratinocytes. In mice, topical treatment with a TRPV4-selective inhibitor decreases UVB-evoked pain behavior, epidermal tissue damage, and endothelin-1 expression. In humans, sunburn enhances epidermal expression of TRPV4 and endothelin-1, underscoring the potential of keratinocyte-derived TRPV4 as a therapeutic target for UVB-induced sunburn, in particular pain.

1월 UVB radiation generates sunburn pain and affects skin by activating epidermal TRPV4 ion channels and triggering endothelin-1 signaling

supplimentary

2018.01.05 Read More »

2017.12.29

Sensory TRP channels contribute differentially to skin inflammation and persistent itch.

Feng J1, Yang P1, Mack MR2, Dryn D1,3, Luo J1, Gong X1,4, Liu S1, Oetjen LK2, Zholos AV3, Mei Z4, Yin S4, Kim BS2, Hu H5.

Abstract

Although both persistent itch and inflammation are commonly associated with allergic contact dermatitis (ACD), it is not known if they are mediated by shared or distinct signaling pathways. Here we show that both TRPA1 and TRPV1 channels are required for generating spontaneous scratching in a mouse model of ACD induced by squaric acid dibutylester (SADBE), a small molecule hapten, through directly promoting the excitability of pruriceptors. TRPV1 but not TRPA1 channels protect the skin inflammation, as genetic ablation of TRPV1 function or pharmacological ablation of TRPV1-positive sensory nerves promotes cutaneous inflammation in the SADBE-induced ACD. Our results demonstrate that persistent itch and inflammation are mediated by distinct cellular and molecular mechanisms in a mouse model of ACD. Identification of distinct roles of TRPA1 and TRPV1 in regulating itch and inflammation may provide new insights into the pathophysiology and treatment of chronic itch and inflammation in ACD patients.

Sensory TRP channels contribute differentially to skin inflammation and persistent itch.

Supplementary information

2017.12.29 Read More »

Journal Club 17.12.22.

TRPC3 Is Dispensable for β-Alanine Triggered Acute Itch
Peter Dong1, Changxiong Guo2, Shengxiang Huang2,3, Minghong Ma1, Qin Liu2 & Wenqin Luo1

TRPC3 Is Dispensable for β-Alanine Triggered Acute Itch

The detection of pruritic (itchy) stimuli is mediated by a variety of receptors and channels expressed
by primary sensory neurons. The G protein-coupled receptor (GPCR) MRGPRD is selectively expressed by a subset of mouse non-peptidergic nociceptors and functions as the molecular receptor for the itch-inducing chemical β-alanine. However, the channels responsible for generating electrical signals downstream of MRGPRD remain unclear. Here, we found that a member of the canonical TRP channel family, TRPC3, is highly expressed in MRGPRD+ non-peptidergic nociceptors, raising the possibility of whether TRPC3 functions as a downstream channel in the MRGPRD signaling pathway. We tested TrpC3 null mice for β-alanine induced itch, and found that these mice exhibit normal responses to β-alanine. At the cellular level, calcium in ux triggered by β-alanine is also unchanged in cultured DRG neurons from TrpC3 null mice compared to wild type. Together, our results demonstrate that mouse TrpC3 is dispensable for β-alanine-induced acute itch.

Journal Club 17.12.22. Read More »

2017.12.15

Transient Receptor Potential Vanilloid 4 Ion Channel Functions as a Pruriceptor in Epidermal Keratinocytes to Evoke Histaminergic Itch*

Abstract

TRPV4 ion channels function in epidermal keratinocytes and in innervating sensory neurons; however, the contribution of the channel in either cell to neurosensory function remains to be elucidated. We recently reported TRPV4 as a critical component of the keratinocyte machinery that responds to ultraviolet B (UVB) and functions critically to convert the keratinocyte into a pain-generator cell after excess UVB exposure. One key mechanism in keratinocytes was increased expression and secretion of endothelin-1, which is also a known pruritogen. Here we address the question of whether TRPV4 in skin keratinocytes functions in itch, as a particular form of “forefront” signaling in non-neural cells. Our results support this novel concept based on attenuated scratching behavior in response to histaminergic (histamine, compound 48/80, endothelin-1), not non-histaminergic (chloroquine) pruritogens in Trpv4 keratinocyte-specific and inducible knock-out mice. We demonstrate that keratinocytes rely on TRPV4 for calcium influx in response to histaminergic pruritogens. TRPV4 activation in keratinocytes evokes phosphorylation of mitogen-activated protein kinase, ERK, for histaminergic pruritogens. This finding is relevant because we observed robust anti-pruritic effects with topical applications of selective inhibitors for TRPV4 and also for MEK, the kinase upstream of ERK, suggesting that calcium influx via TRPV4 in keratinocytes leads to ERK-phosphorylation, which in turn rapidly converts the keratinocyte into an organismal itch-generator cell. In support of this concept we found that scratching behavior, evoked by direct intradermal activation of TRPV4, was critically dependent on TRPV4 expression in keratinocytes. Thus, TRPV4 functions as a pruriceptor-TRP in skin keratinocytes in histaminergic itch, a novel basic concept with translational-medical relevance.

Transient Receptor Potential Vanilloid 4 Ion Channel Functions as a Pruriceptor in Epidermal Keratinocytes to Evoke Histaminergic Itch

2017.12.15 Read More »

2017.12.08

Targeting human Mas-related G protein-coupled receptor X1 to inhibit persistent pain.

Li Z1, Tseng PY1, Tiwari V2, Xu Q1, He SQ2, Wang Y1, Zheng Q1, Han L1, Wu Z3,4, Blobaum AL5, Cui Y6, Tiwari V2, Sun S1, Cheng Y1, Huang-Lionnet JH2, Geng Y1, Xiao B6, Peng J3,4,7, Hopkins C5, Raja SN2, Guan Y8, Dong X9,10.

Abstract

Human Mas-related G protein-coupled receptor X1 (MRGPRX1) is a promising target for pain inhibition, mainly because of its restricted expression in nociceptors within the peripheral nervous system. However, constrained by species differences across Mrgprs, drug candidates that activate MRGPRX1 do not activate rodent receptors, leaving no responsive animal model to test the effect on pain in vivo. Here, we generated a transgenic mouse line in which we replaced mouse Mrgprs with human MrgprX1 This humanized mouse allowed us to characterize an agonist [bovine adrenal medulla 8-22 (BAM8-22)] and a positive allosteric modulator (PAM), ML382, of MRGPRX1. Cellular studies suggested that ML382 enhances the ability of BAM8-22 to inhibit high-voltage-activated Ca2+ channels and attenuate spinal nociceptive transmission. Importantly, both BAM8-22 and ML382 effectively attenuated evoked, persistent, and spontaneous pain without causing obvious side effects. Notably, ML382 by itself attenuated both evoked pain hypersensitivity and spontaneous pain in MrgprX1 mice after nerve injury without acquiring coadministration of an exogenous agonist. Our findings suggest that humanized MrgprX1 mice provide a promising preclinical model and that activating MRGPRX1 is an effective way to treat persistent pain.

KEYWORDS: DRG neurons; GPCR; MrgprX1; pain; positive allosteric modulator

Targeting human Mas-related G protein coupled receptor X1 to inhibit persistent pain.

2017.12.08 Read More »

IL-31RA Knockout Mice Exhibit Elevated Responsiveness to OSMR

IL-31RA Kockout Mice Exhibit Elevated Responsiveness to OSMR

Abstract

IL-31 signals through the heterodimeric receptor IL-31RA and oncostatin M receptor (OSMR), and has been linked with the development of atopic dermatitis, a Th2 cytokine-associated disease in humans. However, recent studies of IL-31RA knockout (KO) mice have suggested that IL-31 signaling may be required to negatively regulate Th2 type responses rather than exacerbate them. Because those studies were performed on genetically modified mice, we examined whether neutralizing IL-31 with a specific mAb would give similar results to IL-31RA KO mice in two Th2 cytokine-associated immune models. We report no difference in lymphocyte Th2-type cytokine production after Ag immunization between IL-31RA KO mice, mice treated with the IL-31 mAb, or control animals. Second, we tested whether the absence of the IL-31RA subunit in IL-31RA KO mice may allow for increased pairing of the OSMR subunit with another cytokine receptor, gp130, resulting in overrepresentation of the heterodimeric receptor for OSM and increased responsiveness to OSM protein. We found that intranasal OSM challenge of IL-31RA KO mice resulted in increased IL-6 and vascular endothelial growth factor production in the lung compared with wild-type littermate control animals. Moreover, PBS-challenged IL-31RA KO mice already had increased levels of vascular endothelial growth factor, which were further increased by OSM challenge. These data imply that IL-31RA-deficient mice produce increased levels of OSM-inducible cytokines during airway sensitization and challenge, which may be the driving force behind the apparent exacerbation of Th2-type inflammatory responses previously observed in these mice.

IL-31RA Knockout Mice Exhibit Elevated Responsiveness to OSMR Read More »

Journal Club 17.11.24

 

 

Involvement of thromboxane A2 in interleukin-31-induced itch-associated response in mice

저자 Andoh, Tsugunobu ⁎
Li, Sikai
Uta, Daisuke
자료유형 Article
초록 •Interleukinin-31 (IL-31) induced scratching, an itch-associated response, in mice.•IL-31-induced scratching was inhibited by TP thromboxane (TX) receptor antagonist.•IL-31 receptors were expressed mainly in epidermal keratinocytes.•IL-31 induced TXA2 production in the skin and primary cultures of keratinocytes.•The results suggest that IL-31 elicits scratching via epidermal TXA2 production.
Background Atopic dermatitis is a chronic and severe pruritic skin disease. Interlukin-31 (IL-31) has been recently demonstrated to be one of the key pruritogens in atopic dermatitis. However, the mechanisms underlying IL-31-induced itching remains unclear. In our previous study, we have shown that thromboxane (TX) A2 is involved in itch-associated responses in mice with atopy-like skin diseases.
Methods IL-31 was given intradermally into the rostral back of ICR mice and the hind-paw scratching to the injection site were counted. Expression of TX synthase and IL-31 receptors were analyzed using immunohistochemical staining or RT-PCR in mouse skin or primary cultures of mouse keratinocytes. The concentration of TXB2, a metabolite of TXA2, in the skin and the culture medium of primary cultures of mouse keratinocytes was measured using enzyme immunoassay. The concentration of intracellular Ca2+ ions in mouse keratinocytes was measured using the calcium imaging method.
Results An intradermal injection of IL-31 elicited scratching, an itch-related response, in mice. The scratching was inhibited by TP TXA2 receptor antagonist DCHCH. The distribution of TX synthase and IL-31RA receptor was mainly epidermal keratinocytes in the skin. The primary cultures of keratinocytes expressed the mRNAs of TX synthase and IL-31 receptors. IL-31 increased the concentration of TXB2, which was inhibited by TX synthase inhibitor sodium ozagrel and EGTA, in the skin and the culture medium of primary cultures of keratinocytes. IL-31 increased the concentration of intracellular Ca2+ ions in mouse keratinocytes.
Conclusion It is suggested that IL-31 elicits itch-associated responses through TXA2 produced from keratinocytes.
Affiliation Department of Applied Pharmacology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan
Publisher Elsevier Urban & Partner Sp. z o.o.
DOI 10.1016/j.pharep.2017.10.001
Accession Number S1734114017303808
Copyright © 2017 Published by Elsevier Sp. z o.o. on behalf of Institute of Pharmacology, Polish Academy of Sciences.

 

Involvement of thromboxane A2 in interleukin-31-induced itch-associated response in mice

 

 

 

 

Journal Club 17.11.24 Read More »

Journal Club 2017. 11. 03.

Sensory Neurons Co-opt Classical Immune Signaling Pathways to Mediate Chronic Itch

Sensory Neurons Co-opt Classical Immune Signaling Pathways to Mediate Chronic Itch

Landon K. Oetjen,1,2 Madison R. Mack,1,2 Jing Feng,1,3 Timothy M. Whelan,1,2 Haixia Niu,1,2 Changxiong J. Guo,1,3
Sisi Chen,4 Anna M. Trier,1,2 Amy Z. Xu,1,2 Shivani V. Tripathi,1,2 Jialie Luo,1,3 Xiaofei Gao,1,3 Lihua Yang,5
Samantha L. Hamilton,5 Peter L. Wang,6 Jonathan R. Brestoff,6 M. Laurin Council,2 Richard Brasington,7
Andra ́ s Schaffer,2,6 Frank Brombacher,8 Chyi-Song Hsieh,6,7 Robert W. Gereau IV,3 Mark J. Miller,5 Zhou-Feng Chen,1,3 Hongzhen Hu,1,3 Steve Davidson,4 Qin Liu,1,3 and Brian S. Kim1,2,3,6,9,*
1Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO 63110, USA
2Division of Dermatology, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA
3Department of Anesthesiology, Washington University School of Medicine, St. Louis, MO 63110, USA
4Department of Anesthesiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA
5Division of Infectious Diseases, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA 6Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110, USA
7Division of Rheumatology, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA 8International Centre for Genetic Engineering and Biotechnology and Institute of Infectious Disease and Molecular Medicine, Division of Immunology, University of Cape Town, Cape Town 7700, South Africa
9Lead Contact
*Correspondence: briankim@wustl.edu
http://dx.doi.org/10.1016/j.cell.2017.08.006

Mammals have evolved neurophysiologic reflexes, such as coughing and scratching, to expel invading pathogens and noxious environmental stimuli. It is well established that these responses are also asso- ciated with chronic inflammatory diseases, including asthma and atopic dermatitis. However, the mecha- nisms by which inflammatory pathways promote sensations such as itch remain poorly understood. Here, we show that type 2 cytokines directly activate sensory neurons in both mice and humans. Further, we demonstrate that chronic itch is dependent on neuronal IL-4Ra and JAK1 signaling. We also observe that patients with recalcitrant chronic itch that failed other immunosuppressive therapies mark- edly improve when treated with JAK inhibitors. Thus, signaling mechanisms previously ascribed to the immune system may represent novel therapeutic targets within the nervous system. Collectively, this study reveals an evolutionarily conserved para- digm in which the sensory nervous system employs classical immune signaling pathways to influence mammalian behavior.

Journal Club 2017. 11. 03. Read More »

Journal Club 2017.10.27

Contribution of TRPC3 to store-operated calcium entry and inflammatory transductions in primary nociceptors.

Alkhani H, Ase AR, Grant R, O’Donnell D, Groschner K, Séguéla P1

Abstract

BACKGROUND:

Prolonged intracellular calcium elevation contributes to sensitization of nociceptors and chronic pain in inflammatory conditions. The underlying molecular mechanisms remain unknown but store-operated calcium entry (SOCE) components participate in calcium homeostasis, potentially playing a significant role in chronic pain pathologies. Most G protein-coupled receptors activated by inflammatory mediators trigger calcium-dependent signaling pathways and stimulate SOCE in primary afferents. The aim of the present study was to investigate the role of TRPC3, a calcium-permeable non-selective cation channel coupled to phospholipase C and highly expressed in DRG, as a link between activation of pro-inflammatory metabotropic receptors and SOCE in nociceptive pathways.

RESULTS:

Using in situ hybridization, we determined that TRPC3 and TRPC1 constitute the major TRPC subunits expressed in adult rat DRG. TRPC3 was found localized exclusively in small and medium diameter sensory neurons. Heterologous overexpression of TRPC3channel subunits in cultured primary DRG neurons evoked a significant increase of Gd3+-sensitive SOCE following thapsigargin-induced calcium store depletion. Conversely, using the same calcium add-back protocol, knockdown of endogenous TRPC3 with shRNA-mediated interference or pharmacological inhibition with the selective TRPC3 antagonist Pyr10 induced a substantial decrease of SOCE, indicating a significant role of TRPC3 in SOCE in DRG nociceptors. Activation of P2Y2 purinoceptors or PAR2 protease receptors triggered a strong increase in intracellular calcium in conditions of TRPC3 overexpression. Additionally, knockdown of native TRPC3 or its selective pharmacological blockade suppressed UTP- or PAR2 agonist-evoked calcium responses as well as sensitization of DRG neurons. These data show a robust link between activation of pro-inflammatory receptors and calcium homeostasis through TRPC3-containing channels operating both in receptor- and store-operated mode.

CONCLUSIONS:

Our findings highlight a major contribution of TRPC3 to neuronal calcium homeostasis in somatosensory pathways based on the unique ability of these cation channels to engage in both SOCE and receptor-operated calcium influx. This is the first evidence for TRPC3 as a SOCE component in DRG neurons. The flexible role of TRPC3 in calcium signaling as well as its functional coupling to pro-inflammatory metabotropic receptors involved in peripheral sensitization makes it a potential target for therapeutic strategies in chronic pain conditions.

Contribution of TRPC3 to SOCE and inflammatory transductions in primary nociceptors

Journal Club 2017.10.27 Read More »

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