Cinnamaldehyde elicits itch behavior via TRPV1and TRPV4 but not TRPA1

Domocos, Dan PhD^a,b; Follansbee, Taylor PhD^a; Nguyen, Amanda BS^a; Nguyen, Tony BS^a; Carstens, Mirela I. BS^a; Carstens, Earl PhD^a,Author Information Itch: July-September 2020 – Volume 5 – Issue 3 – p e36doi: 10.1097/itx.0000000000000036

Abstract

Introduction: 

Cinnamaldehyde (CA) elicits itch sensation in humans. We investigated if CA elicits scratching behavior in mice and determined the roles for TRPV1, TRPA1, and TRPV4.

Materials and Methods: 

Scratching behavior elicited by intradermal injection of CA was assessed in wildtype (WT) mice and knockout (KO) mice lacking TRPV1, TRPA1, TRPV4, or deficient in mast cells. We also assessed scratching and wet dog shakes elicited by low-threshold mechanical stimulation of skin treated topically with CA or vehicle. Using calcium imaging we tested if CA activates dorsal root ganglion (DRG) neurons of each genotype.

Results: 

Intradermal cheek injection of CA elicited dose-dependent hindlimb scratch bouts, with fewer forelimb wipes and facial groom bouts that were not dose-dependent. CA elicited significantly fewer scratch bouts in TRPV1and TRPV4 KO mice, but not TRPA1KOs, compared with WTs. There were no sex differences across genotypes. The histamine H1 antagonist cetirizine did not affect CA-evoked scratching, which was normal in mast cell deficient mice, indicating lack of histamine involvement. Scores for alloknesis were significantly greater following topical application of CA compared with vehicle. Post-CA alloknesis scores were significantly higher in TRPV4KOs of both sexes and in female TRPV1 and TRPA1KOs, compared with WTs. Low threshold mechanical stimuli also elicited significantly more wet dog shakes in mice treated topically with 20% CA, with significantly fewer in TRPV1, TRPA1, and TRPV4KOs compared with WTs. In calcium imaging studies, CA excited 24% of WT DRG cells, significantly fewer (11.5%) in cells from TRPV4KOs, and none in TRPA1KOs. Responses of cells of all genotypes exhibited significant sensitization to repeated CA stimulation. Sensitization was significantly enhanced by IL-4, which itself excited 16% of WT DRG cells and none from TRPA1KOs.

Discussion: 

The results indicate that TRPA1 is dispensable for CA-evoked scratching, which depends partly on TRPV1 and TRPV4.

Baicalin induces Mrgprb2-dependent pseudo-allergy in mice 

Jue Wanga, Yongjing Zhanga, Delu Chea, Yingnan Zenga, Yuanyuan Wub, Qiaohong Qinc, Nan Wanga,*

a School of Pharmacy, Xi’an Jiaotong University, Xi’an, 710061, China
b Department of Respiratory and Critical Care Medicine, The Second Affiliated Hospital of Xi’an Jiaotong University, Xi’an, 710061, China  c Institute of Basic and Translational Medicine, Xi’an Medical University, Xi’an, 710021, China

Baicalin, a component of traditional Chinese medicine, is one of the main compounds present in Scutellaria baicalensis Georgi. Pseudo-allergy induced by the injection of these medicines is a frequent adverse drug reaction. Therefore, elucidation of the anaphylactoid reaction of baicalin and its underlying mechanisms are important. Mast cells are primary effectors of allergic reactions, including pseudo-allergy. Studies have shown that Mrgprx2 in human mast cells is a specific receptor that is crucial for pseudo-allergic drug reactions, Mrgprb3 is the rat ortholog of human Mrgprx2, which in mice is designated as Mrgprb2. Here, we aimed to investigate baicalin- induced pseudo-allergy and the association of Mrgprb3 and Mrgprb2 with this effect. We examined the aller- genic effect of baicalin on RBL-2H3 cells and Mrgprb3-knockdown RBL-2H3 cells. Mrgprb2-expressing HEK293 cells and Mrgprb2-knockout mice were used to evaluate the role of Mrgprb2 in baicalin-induced allergy. Baicalin was found to dose-dependently induce pseudo-allergy both in vitro and in vivo. RBL-2H3 cells were activated by baicalin, whereas in Mrgprb3-knockout RBL-2H3 cells, baicalin showed a negligible effect on cell activation. Furthermore, baicalin activated the Mrgprb2-expressing HEK293 cells. Our data showed that baicalin did not induce allergy in Mpgprb2-knockout mice. We conclude that baicalin induces pseudo-allergy via Mrgprb2 in mice.

Keywords:Baicalin Pseudo-allergy Mrgprb2 Mrgprb3 Degranulation

Periostin Activation of Integrin Receptors on Sensory Neurons Induces Allergic Itch

Santosh K. Mishra,1,2,3,4,9,* Joshua J. Wheeler,1,2 Saumitra Pitake,1 Huiping Ding,5 Changyu Jiang,7 Tomoki Fukuyama,1 Judy S. Paps,8 Patrick Ralph,1 Jacob Coyne,1 Michelle Parkington,1 Jennifer DeBrecht,1 Lauren C. Ehrhardt-Humbert,1 Glenn P. Cruse,1,2 Wolfgang Ba ̈ umer,1,6 Ru-Rong Ji,7 Mei-Chuan Ko,5 and Thierry Olivry2,8
1Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, USA 2Comparative Medicine Institute, North Carolina State University, Raleigh, NC, USA

3The WM Keck Behavioral Center, North Carolina State University, Raleigh, NC, USA
4Program in Genetics, North Carolina State University, Raleigh, NC, USA
5Department of Physiology and Pharmacology, Wake Forest School of Medicine, Winston-Salem, NC, USA
6Institute of Pharmacology and Toxicology, Department of Veterinary Medicine, Freie Universita ̈ t Berlin, Berlin, Germany 7Duke University, Durham, NC, USA
8Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, USA 9Lead Contact
*Correspondence: skmishra@ncsu.edu

SUMMARY

Chronic allergic itch is a common symptom affecting millions of people and animals, but its pathogenesis is not fully explained. Herein, we show that periostin, abundantly expressed in the skin of patients with atopic dermatitis (AD), induces itch in mice, dogs, and monkeys. We identify the integrin aVb3 expressed on a subset of sensory neurons as the periostin receptor. Using pharmacological and ge- netic approaches, we inhibited the function of neuronal integrin aVb3, which significantly reduces periostin-induced itch in mice. Furthermore, we show that the cytokine TSLP, the application of AD- causing MC903 (calcipotriol), and house dust mites all induce periostin secretion. Finally, we establish that the JAK/STAT pathway is a key regulator of periostin secretion in keratinocytes. Altogether, our results identify a TSLP-periostin reciprocal activation loop that links the skin to the spinal cord via periph- eral sensory neurons, and we characterize the non- canonical functional role of an integrin in itch.

https://doi.org/10.1016/j.celrep.2020.03.036

Transient receptor potential vanilloid 4 (TRPV4) channel as a target of crotamiton and its bimodal effects

Hiroki Kittaka1 & Yu Yamanoi1,2,3 & Makoto Tominaga1,2,4

Received: 21 January 2017 / Revised: 2 May 2017 / Accepted: 12 May 2017 / Published online: 13 June 2017 # Springer-Verlag Berlin Heidelberg 2017

Abstract The sensation of itching can be defined as Ban un- pleasant cutaneous sensation that provokes a desire to scratch.^ The perception of itching is not critical for the main- tenance of life, but persistent itching can be extremely irritat- ing and decreases the quality of life. Crotamiton (N-ethyl-o- crotonotoluidide) has been used as an anti-itch agent for humans for around 70 years. In spite of the long use of crotamiton, its mechanism of action remains unknown. We hypothesized that crotamiton might have effects on transient receptor potential (TRP) channels expressed in the peripheral nervous system and the skin. We first examined the effects of crotamiton on TRP channels by whole-cell patch-clamp re- cordings. We found that crotamiton strongly inhibited TRPV (vanilloid) 4 channels followed by large currents after crotamiton washout. In mice, crotamiton inhibited itch- related behaviors induced by a TRPV4-selective agonist (GSK1016790A). We biophysically investigated the large TRPV4 currents after crotamiton washout. Comparing single-channel open probabilities and current amplitudes of TRPV4, increases in both parameters were found to contribute to the large washout currents of TRPV4. Because the change in current amplitudes suggested pore dilation of TRPV4, we examined this possibility with cation replacement experiments and by measuring changes in reversal potentials. Greater cat- ion influxes and changes in reversal potentials upon crotamiton washout were observed, suggesting that the TRPV4 pore dilated in its uninhibited state. From these re- sults, we identified the molecular target of crotamiton as TRPV4 and demonstrated pore dilation of TRPV4 upon crotamiton washout.

Keywords Crotamiton . TRPV4 . Pore dilation . Itch

Spinal-cord-NLRP1-inflommsome-open-access-contributes-to-dry-skin-induced-chronic-itch-in-mice

Abstract
Background: Dry skin itch is one of the most common skin diseases and elderly people are believed to be particularly prone to it. The inflammasome has been suggested to play an important role in chronic inflammatory disorders including inflammatory skin diseases such as psoriasis. However, little is known about the role of NLRP1 inflammasome in dry skin-induced chronic itch.
Methods: Dry skin-induced chronic itch model was established by acetone-ether-water (AEW) treatment. Spontaneous scratching behavior was recorded by video monitoring. The expression of nucleotide oligomerization domain (NOD)-like receptor protein 1 (NLRP1) inflammasome complexes, transient receptor potential vanilloid type 1 (TRPV1), and the level of inflammatory cytokines were determined by western blot, quantitative real-time PCR, and enzyme-linked immunosorbent assay (ELISA) kits. Nlrp1a knockdown was performed by an adeno-associated virus (AAV) vector containing Nlrp1a-shRNA- eGFP infusion. H.E. staining was used to evaluate skin lesion.
Results: AEW treatment triggers spontaneous scratching and significantly increases the expression of NLRP1, ASC, and caspase-1 and the levels of IL-1β, IL-18, IL-6, and TNF-α in the spinal cord and the skin of mice. Spinal cord Nlrp1a knockdown prevents AEW-induced NLRP1 inflammasome assembly, TRPV1 channel activation, and spontaneous scratching behavior. Capsazepine, a specific antagonist of TRPV1, can also inhibit AEW-induced inflammatory response and scratching behavior. Furthermore, elderly mice and female mice exhibited more significant AEW-induced scratching behavior than young mice and male mice, respectively. Interestingly, AEW-induced increases in the expression of NLRP1 inflammasome complex and the levels of inflammatory cytokines were more remarkable in elderly mice and female mice than in young mice and male mice, respectively.
Conclusions: Spinal cord NLRP1 inflammasome-mediated inflammatory response contributes to dry skin-induced chronic itch by TRPV1 channel, and it is also involved in age and sex differences of chronic itch. Inhibition of NLRP1 inflammasome may offer a new therapy for dry skin itch.
Keywords: NLRP1 inflammasome, TRPV1, Chronic itch, Dry skin, Spinal cord

IL-37 is protective in allergic contact dermatitis through mast cell inhibition 

Weihua Lia, Fengmin Dingb, Yi Zhaia, Wenting Taob, Jing Bic, Hong Fanc, Nina Yind,

Zhigang Wangc,⁎

a Department of Cardiology, Affiliated Liyuan Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430077, China b School of Basic Medical Sciences, Hubei University of Chinese Medicine, Wuhan 430065, China
c Department of Pathogen Biology, School of Basic Medical Sciences, Hubei University of Chinese Medicine, Wuhan 430065, China
d Department of Anatomy, School of Basic Medical Sciences, Hubei University of Chinese Medicine, Wuhan 430065, China

ABSTRACT

Allergic contact dermatitis (ACD), characterized predominantly by erythema, vesiculation, and pruritus, is a T cell-mediated skin inflammatory condition. Among immune cells involved in ACD, mast cells (MCs) play an essential role in its pathogenesis. As an inhibitor of proinflammatory IL-1 family members, interleukin 37 (IL-37) has been shown to ameliorate inflammatory responses in various allergic diseases. In this study, we assessed the immunomodulatory effect of IL-37 on allergic inflammation using a 2,4-dinitrofluorobenzene (DNFB)-induced ACD rat model and isolated rat peritoneal mast cells (RPMCs). Systematic application of IL-37 significantly relieved ear swelling, reduced inflammatory cell infiltration, decreased inflammatory cytokine production (TNF- α, IL-1β, IFN-γ, and IL-13), inhibited MC recruitment, lowered IgE levels, and reduced IL-33 production in the local ear tissues with DNFB challenge. Additionally, RPMCs isolated from ACD rats with IL-37 intervention showed downregulation of IL-6, TNF-α, IL-13, and MCP-1 production following IL-33 stimulation, and reduction of β-hexosaminidase and histamine release under DNP-IgE/HSA treatment. Moreover, IL-37 treatment also significantly restrained NF-κB activation and P38 phosphorylation in ACD RPMCs. SIS3, a specific Smad3 in- hibitor, abolished the suppressive effects of IL-37 on MC-mediated allergic inflammation, suggesting the parti- cipation of Smad3 in the anti-ACD effect of IL-37. These findings indicated that IL-37 protects against IL-33- regulated MC inflammatory responses via inhibition of NF-κB and P38 MAPK activation accompanying the regulation of Smad3 in rats with ACD.

Activation of Different Heterodimers of TLR2 Distinctly Mediates Pain and Itch

Ting-Ting Wang, a,c,dy Xian-Yun Xu, a,b,cy Wei Lin, e Dan-Dan Hu, a,b,c Wu Shi, a,b,c Xin Jia, a,b,c Hui Wang, a Ning-Jing Song, d Yu-Qiu Zhang e and Ling Zhang a,b,c*

a The First Rehabilitation Hospital of Shanghai, Tongji University School of Medicine, Shanghai 200090, China
b Key Laboratory of Spine and Spinal Cord Injury Repair and Regeneration of Ministry of Education, Orthopaedic Department of Tongji

Hospital, School of Medicine, Tongji University, Shanghai, 200065, China

c Department of Anatomy and Histology, Tongji University School of Medicine, Shanghai 200092, China

d Department of Dermatology, Tongren Hospital Shanghai Jiao Tong University School of Medicine Shanghai 200336, China

e State Key Laboratory of Medical Neurobiology and MOE Frontiers Center for Brain Science, Department of Translational Neuroscience, Institutes of Brain Science, Fudan University, Shanghai 200032, China

Abstract—Toll-like receptors (TLRs) have been implicated in pain and itch regulation. TLR2, a TLR family member that detects microbial membrane components, has been implicated in pathologic pain. However, the role of TLR2 in pruritic and nociceptive responses has not been thoroughly investigated. In this study, we found that TLR2 was expressed in mouse dorsal root ganglia (DRG) and trigeminal ganglia (TG) neurons. Itch and pain behaviors, including histamine-dependent and histamine-independent acute itching, acetone/diethyl ether/water and 2,4-dini trofluorobenzene-induced chronic itching and inflammatory pain, were largely attenuated in TLR2 knockout (KO) mice. The TLR2 agonist Pam3CSK4, which targets TLR2/1 heterodimers, evoked pain and itch behavior, whereas lipoteichoic acid (LTA) and zymosan, which recognize TLR2/6 heterodimers, produced only pain response. The TLR2 agonist-induced nociceptive and pruritic behaviors were largely diminished in transient receptor potential vanilloid 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) KO mice. Finally, Pam3Csk4 and zymosan increased the [Ca2+]i in DRG neurons from wild-type mice. However, the enhancement of [Ca2+]i was largely inhibited in the DRG neurons from TRPV1 and TRPA1 KO mice. Our results demonstrate that TLR2 is involved in different itch and pain behaviors through activating TLR1/TLR2 or TLR6/TLR2 heterodimers via TRPV1 and TRPA1 channels. ! 2020 IBRO. Published by Elsevier Ltd. All rights reserved.

Key words: TLR2, pain, itch, TRPV1, TRPA1.

PAR2 mediates itch via TRPV3 signaling in keratinocytes

Jiahui Zhao, Admire Munanairi, Xian-Yu Liu, Jie Zhang, Linghan Hu, Meiqin Hu, Dingfang Bu, Lingling Liu, Zhiqiang Xie, Brian S. Kim, Yong Yang, Zhou-Feng Chen

The Journal of Investigative Dermatology

20 August 2019
10 December 2019 12 January 2020

Please cite this article as: Zhao J, Munanairi A, Liu X-Y, Zhang J, Hu L, Hu M, Bu D, Liu L, Xie Z,
Kim BS, Yang Y, Chen Z-F, PAR2 mediates itch via TRPV3 signaling in keratinocytes, The Journal of Investigative Dermatology (2020), doi: https://doi.org/10.1016/j.jid.2020.01.012.

Abstract

Animal studies have suggested that transient receptor potential (TRP) ion channels and G protein-coupled receptors (GPCRs) play important roles in itch transmission. TRPV3 gain-of-function mutations have been identified in patients with Olmsted syndrome which is associated with severe pruritus. However, the mechanisms causing itch remain poorly understood. Here, we show that keratinocytes lacking TRPV3 impair the function of protease activated receptor 2 (PAR2), resulting in reduced neuronal activation and scratching behavior in response to PAR2 agonists. Moreover, we show that TRPV3 and PAR2 were upregulated in skin biopsies from patients and mice with atopic dermatitis (AD), whereas their inhibition attenuated scratching and inflammatory responses in mouse AD models. Taken together, these results reveal a previously unrecognized link between TRPV3 and PAR2 in keratinocytes to convey itch information and suggest that a blockade of PAR2 or TRPV3 individually or both may serve as a potential approach for antipruritic therapy in AD.

Key Words

TRPV3PAR2itchkeratinocytescalciumatopic dermatitis

Abbreviations

ADatopic dermatitisBAM 8-22bovine adrenal medulla peptide 8-22DRGdorsal root gangliaGPCRG protein coupled receptorMrgprMas-related G-protein coupled receptorPAR2protease-activated receptor 2TRPV3transient receptor potential cation channel V3TSLPthymic stromal lymphopoietin

Polymyxin B and polymyxin E induce anaphylactoid response through T mediation of Mas-related G protein–coupled receptor X2

Yingzhuan Zhan, Nan Ma, Rui Liu, Nan Wang, Tao Zhang, Langchong He* School of Pharmacy, Health Science Center, Xi’an Jiaotong University, Xi’an, Shaanxi Province, 710061, PR China

Keywords: Polymyxin B, Polymixin E, Anaphylactoid reaction, MRGPRX2

ABSTRACT

Polymyxin B (PMB) and polymyxin E (PME) are cyclic, peptide antibiotics which derived from various species of Paenibacillus (Bacillus) polymyxa. They are decapeptide antibiotics with an antimicrobial spectrum that includes Gram-negative bacteria, and reused as therapeutic agents due to the emergence of multidrug-resistant (MDR) Gram-positive bacteria. PMB or PME-induced anaphylactoid reactions in the clinic have been documented. However, the mechanism underlying anaphylactoid reaction induced by polymyxin has not yet been reported. Here, we report that human Mas-related G protein-coupled receptor X2 (MRGPRX2) and its mouse homologue Mas-related G protein-coupled receptor B2 (MrgprB2) are the receptors mediating the anaphylactoid response provoked by PMB and PME. We firstly investigated the anaphylactoid reactions induced by PMB and PME in LAD2 cells in vitro and in vivo, and found that treatment with PMB and PME led to significant release of mast cell granules such as histamine and β-hexosaminidase, secretion of pro-inflammatory cytokines, such as TNF-α and PGD2, and provocation of calcium flux in LAD2 cells. Furthermore, treatment with PMB and PME led to reduced release of β-hexosaminidase in MRGPRX2 knockdown-LAD2 cells, and obvious increased calcium release in MRGPRX2 overexpressing HEK293 cells, which suggested that MRGPRX2 are involved in mast cell activation provoked by PMB or PME. In vivo, MRGPRB2 knockout mice exhibited lower pseudo-allergic reactions than wild type mice. Activation of MrgprB2 also triggers increased capillary permeability and paw swelling. Our results elucidated the role of MRGPRX2 in PMB and PME-induced anaphylactoid response and suggested that MRGPRX2 as a potential therapeutic target to control the anaphylactoid reactions which triggered by PMB or PME.

3’-O-Methylorobol Inhibits the Voltage-Gated Sodium Channel Nav1.7 with Anti-Itch Efficacy in A Histamine-Dependent Itch Mouse Model

Fan Zhang †, Ying Wu †, Shuwen Xue, Shuangyan Wang, Chunlei Zhang * and Zhengyu Cao *

State Key Laboratory of Natural Medicines and Jiangsu Provincial Key Laboratory for TCM Evaluation and Translational Development, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, China; zhangfan20111112@126.com (F.Z.); wingwycpu@126.com (Y.W.); xsw19850856166@163.com (S.X.); shuangyanwcpu@163.com (S.W.)

* Correspondence: zhangchunlei11@sina.com (C.Z.); zycao1999@hotmail.com (Z.C.); Tel.: +86-25-8618-5955 (C.Z.); +86-25-8618-5955 (Z.C.)

† These authors contributed equally to this work.
Received: 15 November 2019; Accepted: 29 November 2019; Published: 1 December 2019

􏰀􏰁􏰂􏰀􏰃 􏰅􏰆􏰇

􏰈􏰉􏰊􏰋􏰌􏰂􏰍

Abstract: An itch is a clinical complication that affects millions of patients. However, few treatment options are available. The voltage-gated sodium channel Nav1.7 is predominantly expressed in peripheral sensory neurons and is responsible for the rising phase of action potentials, thereby mediating nociceptive conduction. A gain-of-function mutation of Nav1.7 results in the hyperexcitability of sensory neurons and causes the inherited paroxysmal itch. Conversely, a monoclonal antibody that selectively inhibits Nav1.7 is able to effectively suppress the histamine-dependent itch in mice. Therefore, Nav1.7 inhibitors may possess the potential to relieve the itch. In the present study, using whole-cell voltage-clamp recordings, we demonstrated that 3’-O-methylorobol inhibited Na+ currents in Nav1.7-CHO cells and tetrodotoxin-sensitive Na+ currents in mouse dorsal root ganglion (DRG) neurons with IC50 (half-maximal inhibitory concentration) values of 3.46 and 6.60 μM, respectively. 3’-O-methylorobol also suppressed the tetrodotoxin-resistant Na+ currents in DRG neurons, though with reduced potency (~43% inhibition at 30 μM). 3’-O-methylorobol (10 μM) affected the Nav1.7 by shifting the half-maximal voltage (V1/2) of activation to a depolarizing direction by ~6.76 mV, and it shifted the V1/2 of inactivation to a hyperpolarizing direction by ~16.79 mV. An analysis of 3’-O-methylorobol activity toward an array of itch targets revealed that 3’-O-methylorobol was without effect on histamine H1 receptor, TRPV1, TRPV3, TRPV4, TRPC4 and TRPM8. The intrathecal administration of 3’-O-methylorobol significantly attenuated compound 48/80-induced histamine-dependent spontaneous scratching bouts and the expression level of c-fos in the nuclei of spinal dorsal horn neurons with a comparable efficacy to that of cyproheptadine. Our data illustrated the therapeutic potential for 3’-O-methylorobol for histamine-dependent itching, and the small molecule inhibition of Nav1.7 may represent a useful strategy to develop novel therapeutics for itching.

Keywords: Nav1.7; 3’-O-methylorobol; gating modifier; itch relief; lead compound