LPS exacerbates TRPV4-mediated itch through the intracellular TLR4-PI3K signalling

Yanping Hao1,2,3 | Liyan Wu1,2 | Yuhui Wang4 | Dongmei Shan1,2 | Yifei Liu1 | Jing Feng1,2 | Yi Chang3 | Ting Wang1,2,5,6

J Cell Mol Med  2024 Jul;28(13):e18509. doi: 10.1111/jcmm.18509.

• ¹Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China.
• ²University of Chinese Academy of Sciences, Beijing, China.
• ³Yangpu Hospital, School of Medicine, Tongji University, Shanghai, China.
• ⁴Department of Anesthesiology, Plastic Surgery Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
• ⁵Yunnan Key Laboratory of Gastrodia and Fungi Symbiotic Biology, Zhaotong University, Zhaotong, Yunnan, China.
• ⁶Yunnan Engineering Research Center of Green Planting and Processing of Gastrodia, Zhaotong University, Zhaotong, Yunnan, China.

PMID: 38957035 PMCID: PMC11220342 DOI: 10.1111/jcmm.18509

Abstract

Pruritus is often accompanied with bacterial infections, but the underlying mechanism is not fully understood. Although previous studies revealed that lipopolysaccharides (LPS) could directly activate TRPV4 channel and TRPV4 is involved in the generation of both acute itch and chronic itch, whether and how LPS affects TRPV4-mediated itch sensation remains unclear. Here, we showed that LPS-mediated TRPV4 sensitization exacerbated GSK101-induced scratching behaviour in mice. Moreover, this effect was compromised in TLR4-knockout mice, suggesting LPS acted through a TLR4-dependent mechanism. Mechanistically, LPS enhanced GSK101-evoked calcium influx in mouse ear skin cells and HEK293T cells transfected with TRPV4. Further, LPS sensitized TRPV4 channel through the intracellular TLR4-PI3K-AKT signalling. In summary, our study found a modulatory role of LPS in TRPV4 function and highlighted the TLR4-TRPV4 interaction in itch signal amplification.

Keywords: LPS; PI3K; TLR4; TRPV4; itch sensitization.

Neuronal BST2: A Pruritic Mediator alongside Protease-Activated Receptor 2 in the IL-27eDriven Itch Pathway

Yanqing Li1, Weiwei Chen1, Xingyun Zhu1, Huiyuan Mei1, Martin Steinhoff2,3,4,5,6,7,
Joerg Buddenkotte2,3,4, Jinhai Wang1, Wenhao Zhang1, Zhenghui Li8, Xiaolong Dai1, Chunxu Shan9, Jiafu Wang9,10 and Jianghui Meng9,10

1School of Life Sciences, Henan University, Henan, China; 2Department of Dermatology and Venereology, Hamad Medical Corporation, Doha, Qatar; 3Translational Research Institute, Academic Health System, Hamad Medical Corporation, Doha, Qatar; 4Dermatology Institute, Academic Health System, Hamad Medical Corporation, Doha, Qatar; 5Department of Dermatology, Weill Cornell Medicine-Qatar, Doha, Qatar; 6College of Medicine, Qatar University, Doha, Qatar; 7Israel Englander Department of Dermatology, Weill Cornell Medicine, New York, New York, USA; 8Department of Neurosurgery, The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, China; and 9School of Biotechnology, Faculty of Science and Health, Dublin City University, Dublin, Ireland 10These authors contributed equally as senior authors.\

Correspondence: Jianghui Meng, School of Biotechnology, Faculty of Science and Health, Dublin City University, Glasnevin Avenue, Dublin 9, Ireland. E-mail: Jianghui.meng@dcu.ie

Abbreviations: AD, atopic dermatitis; HC, healthy control; LAD, lesional atopic dermatitis; mTGN, murine trigeminal ganglionic neuron; PAR2, pro- tease-activated receptor 2; phKC, primary human keratinocyte; STAT, signal transducer and activator of transcription; Th, T helper

Received 29 October 2023; revised 11 January 2024; accepted 27 January 2024; accepted manuscript published online XXX; corrected proof published online XXX

Chronic itch is a common and complex symptom often associated with skin diseases such as atopic dermatitis (AD). Although IL-27 is linked to AD, its role and clinical significance in itch remain undefined. We sought to investigate IL-27 function in itch using tissue-specific transgenic mice, various itch models, behavior scoring, RNA sequencing, and cytokine/kinase array. Our findings show that IL-27 receptors were overexpressed in human AD skin. Intradermal IL-27 injection failed to directly induce itch in mice but upregulated skin protease- activated receptor 2 (PAR2) transcripts, a key factor in itch and AD. IL-27 activated human keratinocytes, increasing PAR2 transcription and activity. Coinjection of SLIGRL (PAR2 agonist) and IL-27 in mice heightened PAR2-mediated itch. In addition, IL-27 boosted BST2 transcription in sensory neurons and keratinocytes. BST2 was upregulated in AD skin, and its injection in mice induced itch-like response. BST2 colocalized with sensory nerve branches in AD skin from both human and murine models. Sensory neurons released BST2, and mice with sensory neuronespecific BST2 knockout displayed reduced itch responses. Overall, this study provides evidence that skin IL-27/PAR2 and neuronal IL-27/BST2 axes are implicated in cutaneous inflammation and pruritus. The discovery of neuronal BST2 in pruritus shed light on BST2 in the itch cascade.

Keywords: Atopic dermatitis, BST2, IL-27, itch, PAR2
Journal of Investigative Dermatology (2024) -, -e-; doi:10.1016/j.jid.2024.01.025

20-HETE mediated TRPV1 activation drives allokinesis via MrgprA3+ neurons in chronic dermatitis

Guang Yu1,2*, Pei Liu1*, Xiaobao Huang3*, Mingxin Qi2, Xue Li2, Weimeng Feng1, Erxin Shang1, Yuan Zhou2, Changming Wang2, Yan Yang2, Chan Zhu2, Fang Wang3, Zongxiang Tang2, Jinao Duan1

1. Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, Jiangsu Key Laboratory for High Technology Research of TCM Formulae, Nanjing University of Chinese Medicine, Nanjing, China.
2. Key Laboratory for Chinese Medicine of Prevention and Treatment in Neurological Diseases, School of Medicine, Nanjing University of Chinese Medicine, Nanjing, China.
3. Department of Dermatology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

* Guang Yu, Pei Liu and Xiaobao Huang are co-first authors.

Corresponding author: Guang Yu, E-mail: yuguang@njucm.edu.cn; Zongxiang Tang, E-mail: tangzxlab@njucm.edu.cn; Jinao Duan, E-mail: dja@njucm.edu.cn (J.D.).

© The author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.

Received: 2023.04.12; Accepted: 2024.01.26; Published: 2024.02.04

Abstract
Rationale: Noxious stimuli are often perceived as itchy in patients with chronic dermatitis (CD); however, itch and pain mechanisms of CD are not known.

Methods: TRPV1 involvement in CD was analyzed using a SADBE induced CD-like mouse model, and several loss- and gain-of-function mouse models. T rigeminal TRPV1 channel and Mrgpr A3+ neuron functions were analyzed by calcium imaging and whole-cell patch-clamp recordings. Lesional CD-like skin from mice were analyzed by unbiased metabolomic analysis. 20-HETE availability in human and mouse skin were determined by LC/MS and ELISA. And finally, HET0016, a selective 20-HETE synthase inhibitor, was used to evaluate if blocking skin TRPV1 activation alleviates CD-associated chronic itch or pain.

Results: While normally a pain inducing chemical, capsaicin induced both itch and pain in mice with CD condition. DREADD silencing of MrgprA3+ primary sensory neurons in these mice selectively decreased capsaicin induced scratching, but not pain-related wiping behavior. In the mice with CD condition, MrgprA3+ neurons showed elevated ERK phosphorylation. Further experiments showed that MrgprA3+ neurons from MrgprA3;Braf mice, which have constitutively active BRAF in MrgprA3+ neurons, were significantly more excitable and responded more strongly to capsaicin. Importantly, capsaicin induced both itch and pain in MrgprA3;Braf mice in an MrgprA3+ neuron dependent manner. Finally, the arachidonic acid metabolite 20-HETE, which can activate TRPV1, was significantly elevated in the lesional skin of mice and patients with CD. Treatment with the selective 20-HETE synthase inhibitor HET0016 alleviated itch in mice with CD condition.

Conclusion: Our results demonstrate that 20-HETE activates TRPV1 channels on sensitized MrgprA3+ neurons, and induces allokinesis in lesional CD skin. Blockade of 20-HETE synthesis or silencing of TRPV1-MrgprA3+ neuron signaling offers promising therapeutic strategies for alleviating CD-associated chronic itch.

Keywords: 20-HETE, TRPV1, allokinesis, MrgprA3+ neurons, chronic dermatitis

IL-31–generating network in atopic dermatitis comprising macrophages, basophils, thymic stromal lymphopoietin, and periostin

Takashi Hashimoto, MD, PhD,a Hiroo Yokozeki, MD, PhD,b Hajime Karasuyama, MD, PhD,c and Takahiro Satoh, MD, PhDa Tokorozawa and Tokyo, Japan

From a the Department of Dermatology, National Defense Medical College, Tokorozawa, and b the Department of Dermatology, Graduate School of Medical and Dental Sciences, and c the Inflammation, Infection and Immunity Laboratory, Advanced Research Institute, Tokyo Medical and Dental University (TMDU), Tokyo. This study was partially supported by Japan Society for the Promotion of Science (JSPS) KAKENHI Grant-in-Aid for Scientific Research (C) (grant numbers 17K16328, 19K08743, and 22K08395 to T.H. and 19K08805 and 22K08444 to T.S.). Disclosure of potential conflict of interest: The authors declare that they have no relevant conflicts of interest. Received for publication July 18, 2022; revised October 22, 2022; accepted for publication November 11, 2022. Available online November 19, 2022. Corresponding author: Takashi Hashimoto, MD, PhD, Department of Dermatology, National Defense Medical College, 3-2, Namiki, Tokorozawa, Saitama 359-8513, Japan. E-mail: hashderm@ndmc.ac.jp. The CrossMark symbol notifies online readers when updates have been made to the article such as errata or minor corrections 0091-6749/$36.00 2022 American Academy of Allergy, Asthma & Immunology https://doi.org/10.1016/j.jaci.2022.11.009

Background: IL-31 is a type 2 cytokine involved in the itch sensation in atopic dermatitis (AD). The cellular origins of IL-31 are generally considered to be TH2 cells. Macrophages have also been implicated as cellular sources of IL-31. Objective: We sought to determine the expression of IL-31 by macrophages and to elucidate the productive mechanisms and contributions to itch in AD skin lesions. Methods: Expression of IL-31 by macrophages, expressions of thymic stromal lymphopoietin (TSLP) and periostin, and presence of infiltrating basophils in human AD lesions were examined through immunofluorescent staining, and correlations were assessed. Furthermore, mechanisms of inducing IL-31– expressing macrophages were analyzed in an MC903-induced murine model for AD in vivo and in mouse peritoneal macrophages ex vivo.

Results: A significant population of IL-311 cells in human AD lesions was that of CD681 cells expressing CD163, an M2 macrophage marker. The number of IL-311/CD681 cells correlated with epidermal TSLP, dermal periostin, and the number of dermal-infiltrating basophils. In the MC903-induced murine AD model, significant scratching behaviors with enhanced expressions of TSLP and periostin were observed, accompanied by massive infiltration of basophils and IL-311/ MOMA-21/Arg-11 cells. Blockade of IL-31 signaling with anti– IL-31RA antibody or direct depletion of macrophages by clodronate resulted in attenuation of scratching behaviors. Toeffectively reduce lesional IL-311 macrophages and itch, basophil depletion was essential in combination with TSLP- and periostin-signal blocking. Murine peritoneal macrophages produced IL-31 when stimulated with TSLP, periostin, and basophils.

Conclusions: A network comprising IL-31–expressing macrophages, TSLP, periostin, and basophils plays a significant role in AD itch. (J Allergy Clin Immunol 2023;151:737-46.)

Key words: Atopic dermatitis, basophil, IL-31, itch, macrophage, periostin, thymic stromal lymphopoietin

Discovery of a Small Molecule Activator of Slack (Kcnt1) Potassium Channels That Significantly Reduces Scratching in Mouse Models of Histamine-Independent and Chronic Itch

Annika Balzulat, W. Felix Zhu, Cathrin Flauaus, Victor Hernandez-Olmos, Jan Heering, Sunesh Sethumadhavan, Mariam Dubiel, Annika Frank, Amelie Menge,
Maureen Hebchen, Katharina Metzner, Ruirui Lu, Robert Lukowski, Peter Ruth,
Stefan Knapp, Susanne Müller, Dieter Steinhilber, Inga Hänelt, Holger Stark, Ewgenij Proschak,* and Achim Schmidtko*

ABSTRACT
Various disorders are accompanied by histamine-independent itching, which is often resistant to the currently available therapies. Here, it is reported that the pharmacological activation of Slack (Kcnt1, KNa1.1), a potassium channel highly expressed in itch-sensitive sensory neurons, has therapeutic potential for the treatment of itching. Based on the Slack-activating antipsychotic drug, loxapine, a series of new derivatives with improved pharmacodynamic and pharmacokinetic profiles is designed that enables to validate Slack as a pharmacological target in vivo. One of these new Slack activators, compound 6, exhibits negligible dopamine D2 and D3 receptor binding, unlike loxapine. Notably, compound 6 displays potent on-target antipruritic activity in multiple mouse models of acute histamine-independent and chronic itch without motor side effects. These properties make compound 6 a lead molecule for the development of new antipruritic therapies targeting Slack.

Borneol exerts its antipruritic effects by inhibiting TRPA1 and activating TRPM8

Miao Luoa,1, Jinfeng Hea,1, Liang Yina, Ping Zhanb, Zhongqiu Zhaoc, Hui Xionga,d,**, Zhinan Meia,e,*

a School of Pharmaceutical Sciences, South-Central Minzu University, Wuhan, 430074, China
b Dermatology Hospital of Jiangxi Province, Nanchang, 330000, China
c Barnes-Jewish Hospital, St. Louis, MO, 63110, USA
d Ethnopharmacology Level 3 Laboratory of National Administration of Traditional Chinese Medicine, South-Central Minzu University, Wuhan, 430074, China e College of Plant Science and Technology, Huazhong Agricultural University, Wuhan, 430070, China

Ethnopharmacological relevance: Borneol is a long-established traditional Chinese medicine that has been found to be effective in treating pain and itchy skin. However, whether borneol has a therapeutic effect on chronic itch and its related mechanisms remain unclear.
Aim of the study: To investigate the antipruritic effect of borneol and its molecular mechanism.

Materials and methods: DrugBAN framework and molecular docking were applied to predict the targets of borneol, and the calcium imaging or patch-clamp recording analysis were used to detect the effects of borneol on TRPA1, TRPM8 or TRPV3 channels in HEK293T cells. In addition, various mouse models of acute itch and chronic itch were established to evaluate the antipruritic effects of borneol on C57BL/6J mice. Then, the borneol- induced pruritic relief was further investigated in Trpa1− /− , Trpm8− /− , or Trpa1− /− /Trpm8− /− mice. The effects of borneol on the activation of TRPM8 and the inhibition of TRPA1 were also measured in dorsal root ganglia neurons of wild-type (WT), Trpm8− /− and Trpv1− /− mice. Lastly, a randomized, double-blind study of adult patients was conducted to evaluate the clinical antipruritic effect of borneol.

Results: TRPA1, TRPV3 and TRPM8 are the potential targets of borneol according to the results of DrugBAN algorithm and molecular docking. Calcium imaging and patch-clamp recording analysis demonstrated that borneol activates TRPM8 channel-induced cell excitability and inhibits TRPA1 channel-mediated cell excitability in transfected HEK293T cells. Animal behavior analysis showed that borneol can significantly reduce acute and chronic itch behavior in C57BL/6J mice, but this effect was eliminated in Trpa1− /− , Trpm8− /− mice, or at least in Trpa1− /− /Trpm8− /− mice. Borneol elicits TRPM8 channel induced [Ca2+]i responses but inhibits AITC or SADBE- induced activation of TRPA1 channels in dorsal root ganglia neurons of WT and Trpv1− /− mice, respectively. Furthermore, the clinical results indicated that borneol could reduce itching symptoms in patients and its efficacy is similar to that of menthol.

Conclusion: Borneol has therapeutic effects on multiple pruritus models in mice and patients with chronic itch, and the mechanism may be through inhibiting TRPA1 and activating TRPM8.

TRPV3-ANO1 interaction positively regulates wound healing in keratinocytes

Yu Yamanoi1,2,3, Jing Lei1,2, Yasunori Takayama4, Shigekuni Hosogi5, Yoshinori Marunaka6,7 &Makoto Tominaga1,2

Communications Biology volume 6, Article number: 88 (2023) Cite this article

Abstract

Transient receptor potential vanilloid 3 (TRPV3) belongs to the TRP ion channel super family and functions as a nonselective cation channel that is highly permeable to calcium. This channel is strongly expressed in skin keratinocytes and is involved in warmth sensation, itch, wound healing and secretion of several cytokines. Previous studies showed that anoctamin1 (ANO1), a calcium-activated chloride channel, was activated by calcium influx through TRPV1, TRPV4 or TRPA1 and that these channel interactions were important for TRP channel-mediated physiological functions. We found that ANO1 was expressed by normal human epidermal keratinocytes (NHEKs). We observed that ANO1 mediated currents upon TRPV3 activation of NHEKs and mouse skin keratinocytes. Using an in vitro wound-healing assay, we observed that either a TRPV3 blocker, an ANO1 blocker or low chloride medium inhibited cell migration and proliferation through p38 phosphorylation, leading to cell cycle arrest. These results indicated that chloride influx through ANO1 activity enhanced wound healing by keratinocytes.