Role of SNAREs in Atopic Dermatitis–Related Cytokine Secretion and Skin-Nerve Communication

JianghuiMeng¹²JiafuWang¹JoergBuddenkotte³⁴TimoBuhl⁵MartinSteinhoff²³⁴⁶

The role of soluble N-ethylmaleimide-sensitive factor attachment protein receptors in atopic dermatitis (AD) is unknown. This study identifies the function of soluble N-ethylmaleimide sensitive factor attachment protein receptor in AD-related cytokine secretion and epidermis-nerve communication. Herein, we report that various cytokines were simultaneously upregulated and coreleased in innate immunity–activated primary human keratinocytes. AD-related cytokines thymic stromal lymphopoietin, endothelin-1, and inflammatory tumor necrosis factor-α activated distinct but overlapping sensory neurons. Tumor necrosis factor-α potentiated thymic stromal lymphopoietin–induced Ca²⁺-influx, whereas endothelin-1 caused itch-selective B-type natriuretic peptide release. In primary human keratinocytes, B-type natriuretic peptide upregulated genes promoting dermatological and neuroinflammatory diseases and conditions. VAMP3, SNAP-29, and syntaxin 4 proved important in driving cytokine release from primary human keratinocytes. Depletion of VAMP3 inhibited nearly all the cytokine release including thymic stromal lymphopoietin and endothelin-1. Accordingly, VAMP3 co-occurred with endothelin-1 in the skins of patients with AD. Our study pinpoints the pivotal role of soluble N-ethylmaleimide sensitive factor attachment protein receptors in mediating cytokine secretion related to AD. VAMP3 is identified as a suitable target for developing broad-spectrum anticytokine therapeutics for controlling itch and atopic skin inflammation.

Abstract

Atopic dermatitis (AD) is a chronic inflammatory skin disease affecting 15% to 20% of the general population in developed countries.¹ It is characterized by recurrent eczematous legions and intense itch. Because the itch sensation induces scratching behavior, which exacerbates the skin inflammation and disturbs the quality of life of affected individuals, chronic itch is a challenging clinical problem in the treatment of AD.

Itch can be induced by various chemical mediators. Among them, much attention has been paid to IL-31 as an AD-associated itch mediator since the discovery of the pruritogenic action of IL-31 in mice.² IL-31 is mainly produced by CD4⁺ T cells and transmits the signals via a heterodimeric receptor composed of IL-31 receptor A and oncostatin M receptor (OSMR), both of which are expressed in various cell types including dorsal root ganglion (DRG) neurons.³ A recent clinical study has demonstrated that blockade of IL-31 signals by a specific antibody for IL-31 receptor A alleviates pruritus in patients with AD.⁴ However, the neuronal mechanism underlying IL-31–induced itch sensation is poorly understood.

Activation of Mast-Cell-Expressed Mas-Related G-Protein-Coupled Receptors Drives Non-histaminergic Itch

Abstract

Classical itch studies have focused on immunoglobulin E (IgE)-mediated mast cell activation and histamine release. Recently, members of the Mas-related G-protein-coupledreceptor(Mrgpr)familyhavebeen 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 itchsensory 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.

BAM8-22 and its receptor MRGPRX1 may attribute to cholestatic pruritus

Babina Sanjel,^1,2Han-Joo Maeng,^1,2 and  Won-Sik Shim^1,2

Abstract

Pruritus is an unexpected symptom observed in cholestasis and its mechanism is still unclear. Here, we show that bovine adrenal medulla (BAM) 8–22, an endogenous itch-inducing peptide, could be involved in cholestatic pruritus. It was found that bile duct ligation (BDL) mice, an obstructive cholestasis model, showed increased spontaneous scratching behaviour. Importantly, the mRNA level of proenkephalin, a precursor polypeptide of BAM8-22, was significantly increased in the skin of BDL mice. Furthermore, the mRNA level of Mrgprx1, which encodes a receptor for BAM8-22, was significantly increased in the dorsal root ganglia (DRG) of BDL mice. This was further confirmed by elevation of intracellular calcium levels upon BAM8-22 treatment in primarily-cultured DRG neurons. In addition, BDL mice showed augmented scratching behaviour by BAM8-22, indicating enhanced activity of MRGPRX1. Moreover, the skin homogenate of BDL mice induced elevation of intracellular calcium levels through MRGPRX1. Finally, among the various bile acids, chenodeoxycholic acid significantly increased proenkephalin transcription in a human keratinocyte cell line (HaCaT). In conclusion, cholestatic pruritus could be attributed in part to enhanced action of both BAM8-22 in the skin and its receptor MRGPRX1 in sensory neurons.

Subject terms: Sensory processing, Molecular neuroscience

Peripheral sensitisation and loss of descending inhibition is a
hallmark of chronic pruritus

Neurophysiological mechanisms leading to chronicity of pruritus are not yet fully understood and it is not known whether these mechanisms diverge between different underlying diseases of chronic pruritus. This study aimed to detect such mechanisms in chronic pruritus of various origins. One-hundred and twenty patients with chronic pruritus of inflammatory origin (atopic dermatitis), neuropathic origin (brachioradial pruritus) and chronic prurigo of nodular type, the latter as a model for chronic scratching, as well as 40 matched healthy controls participated in this study. Stimulation with cowhage induced a more intensive itch sensation compared to stimulation with other substances in all patient groups but not in healthy controls, arguing for sensitisation of cutaneous mechano- and heat-sensitive C-fibers in chronic pruritus. All patient groups showed a decreased intraepidermal nerve fibre density compared to controls. A decreased condition pain modulation effect was observed in all patient groups compared to controls, suggesting a reduced descending inhibitory system in chronic pruritus. In sum, chronic pruritus of different etiology showed a mixed peripheral and central pattern of neuronal alterations, which might contribute to the chronicity of pruritus with no differences between pruritus entities. Our
findings may contribute to the development of future treatment strategies targeting these pathomechanisms.

Key words: Itch, quantitative sensory testing, conditioned pain modulation,
intraepidermal nerve fibre density, pain

MRGPRX4 is a G protein-coupled receptor activated by bile acids that may contribute to cholestatic pruritus.

Meixiong J, Vasavda C, Snyder SH, Dong X.

Abstract

Patients suffering from cholestasis, the slowing or stoppage of bile flow, commonly report experiencing an intense, chronic itch. Numerous pruritogens are up-regulated in cholestatic patient sera, including bile acids (BAs). Acute injection of BAs results in itch in both mice and humans, and BA-modulating therapy is effective in controlling patient itch. Here, we present evidence that human sensory neuron-expressed Mas-related G protein-coupled receptor X4 (MRGPRX4), an orphan member of the Mrgpr family of GPCRs, is a BA receptor. Using Ca²⁺ imaging, we determined that pathophysiologically relevant levels of numerous BAs activated MRGPRX4. No mouse Mrgpr orthologs were activated by BAs. To assess the in vivo relevance of BA activation of MRGPRX4, we generated a humanized mouse with targeted expression of MRGPRX4 in itch-encoding sensory neurons. BAs activated MRGPRX4⁺ sensory neurons at higher levels compared with WT neurons. Compared with control animals, MRGPRX4⁺ mice scratched more upon acute injection of BAs and in a model of cholestatic itch. Overall, these data suggest that targeting MRGPRX4 is a promising strategy for alleviating cholestatic itch.

TLR3 in Chronic Human Itch: A Keratinocyte-Associated Mechanism of Peripheral Itch Sensitization

Short title: Skin TLR3 and chronic human itch.

Keywords: pruritus, toll-like receptor, endothelin, nerve, keratinocyte

Quorum sensing between bacterial species on the skin protects against epidermal injury in atopic dermatitis.

Pruritus in ordinary scabies: IL-31 from macrophages induced by overexpression of TSLP and periostin.

The TRPV4 Agonist GSK1016790A Regulates the Membrane Expression of TRPV4 Channels

The TRPV4 Agonist GSK1016790A Regulates the Membrane Expression of TRPV4 Channels

Sara Baratchi1*, Peter Keov1,2,3, William G. Darby1, Austin Lai1, Khashayar Khoshmanesh4, Peter Thurgood4, Parisa Vahidi1, Karin Ejendal5 and Peter McIntyre1
1 School of Health and Biomedical Sciences, RMIT University, Melbourne, VIC, Australia, 2 Molecular Pharmacology Division, Victor Chang Cardiac Research Institute, Darlinghurst, NSW, Australia, 3 St Vincent’s Clinical School, University of New South Wales, Darlinghurst, NSW, Australia, 4 School of Engineering, RMIT University, Melbourne, VIC, Australia, 5 Weldon School of Biomedical Engineering, Purdue University, West Lafayette, IN, United States

TRPV4 is a non-selective cation channel that tunes the function of different tissues including the vascular endothelium, lung, chondrocytes, and neurons. GSK1016790A is the selective and potent agonist of TRPV4 and a pharmacological tool that is used to study the TRPV4 physiological function in vitro and in vivo. It remains unknown how the sensitivity of TRPV4 to this agonist is regulated. The spatial and temporal dynamics of receptors are the major determinants of cellular responses to stimuli. Membrane translocation has been shown to control the response of several members of the transient receptor potential (TRP) family of ion channels to different stimuli. Here, we show that TRPV4 stimulation with GSK1016790A caused an increase in [Ca2+]i that is stable for a few minutes. Single molecule analysis of TRPV4 channels showed that the density of TRPV4 at the plasma membrane is controlled through two modes of membrane trafficking, complete, and partial vesicular fusion. Further, we show that the density of TRPV4 at the plasma membrane decreased within 20min, as they translocate to the recycling endosomes and that the surface density is dependent on the release of calcium from the intracellular stores and is controlled via a PI3K, PKC, and RhoA signaling pathway.
Keywords: TRPV4, membrane trafficking, endothelial cells, GSK1016790A, calcium