journal club 2015.01.23.

Protein kinase Cdelta mediates histamine-evoked itch and responses in pruriceptors

Molecular Pain 2015, 11:1 doi:10.1186/1744-8069-11-1

Manouela V Valtcheva (valtchevam@wusm.wustl.edu) Steve Davidson (sdavidson@wustl.edu) Chengshui Zhao (zhaoc@anest.wustl.edu) Michael Leitges (michael.leitges@biotek.uio.no) Robert W Gereau IV (gereaur@wustl.edu)

1744-8069-11-1

Abstract Background

Itch-producing compounds stimulate receptors expressed on small diameter fibers that innervate the skin. Many of the currently known pruritogen receptors are Gq-Protein Coupled Receptors (GqPCR), which activate Protein Kinase C (PKC). Specific isoforms of PKC have been previously shown to perform selective functions; however, the roles of PKC isoforms in regulating itch remain unclear. In this study, we investigated the novel PKC isoform PKCδ as an intracellular modulator of itch signaling in response to histamine and the non- histaminergic pruritogens chloroquine and β-alanine.

Results

Behavioral experiments indicate that PKCδ knock-out (KO) mice have a 40% reduction in histamine-induced scratching when compared to their wild type littermates. On the other hand, there were no differences between the two groups in scratching induced by the MRGPR agonists chloroquine or β-alanine. PKCδ was present in small diameter dorsal root ganglion (DRG) neurons. Of PKCδ-expressing neurons, 55% also stained for the non-peptidergic marker IB4, while a smaller percentage (15%) expressed the peptidergic marker CGRP. Twenty-nine percent of PKCδ-expressing neurons also expressed TRPV1. Calcium imaging studies of acutely dissociated DRG neurons from PKCδ-KO mice show a 40% reduction in the total number of neurons responsive to histamine. In contrast, there was no difference in the number of capsaicin-responsive neurons between KO and WT animals. Acute pharmacological inhibition of PKCδ with an isoform-specific peptide inhibitor (δV1-1) also significantly reduced the number of histamine-responsive sensory neurons.

Conclusions

Our findings indicate that PKCδ plays a role in mediating histamine-induced itch, but may be dispensable for chloroquine- and β-alanine-induced itch.

journal club 2015.01.23. Read More »

Journal Club 2015.1.16

Cathepsin S signals via PAR2 and generates a novel tethered ligand receptor agonist
Filename : cathepsin-s-signals-via-par2-and-generates-a-novel-tethered-ligand-receptor-agonist.pdf (789 KB)
Caption :

Cathepsin S signals via PAR2 and generates a novel tethered ligand receptor agonist.

Elmariah SB1, Reddy VB1, Lerner EA1.

Author information

Abstract

Protease-activated receptor-2 is widely expressed in mammalian epithelial, immune and neural tissues. Cleavage of PAR2 by serine proteases leads to self-activation of the receptor by the tethered ligand SLIGRL. The contribution of other classes of proteases to PAR activation has not been studied in detail. Cathepsin S is a widely expressed cysteine protease that is upregulated in inflammatory conditions. It has been suggested thatcathepsin S activates PAR2. However, cathepsin S activation of PAR2 has not been demonstrated directly nor has the potential mechanism of activation been identified. We show that cathepsin S cleaves near the N-terminus of PAR2 to expose a novel tethered ligand, KVDGTS. The hexapeptide KVDGTS generates downstream signaling events specific to PAR2 but is weaker than SLIGRL. Mutation of the cathepsin S cleavage site prevents receptor activation by the protease while KVDGTS retains activity. In conclusion, the range of actions previously ascribed to cysteine cathepsins in general, and cathepsin S in particular, should be expanded to include molecular signaling. Such signaling may link together observations that had been attributed previously to PAR2 or cathepsin S individually. These interactions may contribute to inflammation.

Journal Club 2015.1.16 Read More »

Journal club 2015-01-05

LPS sensitizes TRPV1 via activation of TLR4 in trigeminal sensory neurons.

Diogenes A1, Ferraz CC, Akopian AN, Henry MA, Hargreaves KM.

Abstract

Recent studies have demonstrated that the lipopolysaccharide (LPS) receptor (TLR4) is expressed in TRPV1 containing trigeminal sensory neurons. In this study, we evaluated whether LPS activates trigeminal neurons, and sensitizes TRPV1 responses via TLR4. To test this novel hypothesis, we first demonstrated that LPS binds to receptors in trigeminal neurons using competitive binding. Second, we demonstrated that LPS evoked a concentration-dependent increase in intracellular calcium accumulation (Ca(2+))(i) and inward currents. Third, LPS significantly sensitized TRPV1 to capsaicin measured by (Ca(2+))(i), release of calcitonin gene-related peptide, and inward currents. Importantly, a selective TLR4 antagonist blocked these effects. Analysis of these data, collectively, demonstrates that LPS is capable of directly activating trigeminal neurons, and sensitizing TRPV1 via a TLR4-mediated mechanism. These findings are consistent with the hypothesis that trigeminal neurons are capable of detecting pathogenic bacterial components leading to sensitization of TRPV1, possibly contributing to the inflammatory pain often observed in bacterial infections.

LPS

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Journal Club 2014.12.29

TRPA1 chanels mediate acute neurogenic inflammation and pain produced by bacterial endotoxins
Filename : trpa1-chanels-mediate-acute-neurogenic-inflammation-and-pain-produced-by-bacterial-endotoxins.pdf (2 MB)
Caption :

TRPA1 channels mediate acute neurogenic
inflammation and pain produced by bacterial
endotoxins
Victor Meseguer1
, Yeranddy A. Alpizar2, Enoch Luis1
, Sendoa Tajada3, Bristol Denlinger1
, Otto Fajardo1
,Jan-Albert Manenschijn1
, Carlos Ferna´ndez-Pen˜a1
, Arturo Talavera2,4, Tatiana Kichko5, Bele´n Navia6,
Alicia Sa´nchez2, Rosa Sen˜arı´s6, Peter Reeh5, Marı´a Teresa Pe´rez-Garcı´a3, Jose´ Ramo´n Lo´pez-Lo´pez3,
Thomas Voets2, Carlos Belmonte1
, Karel Talavera1,2,* & Fe´lix Viana1,*

Gram-negative bacterial infections are accompanied by inflammation and somatic or
visceral pain. These symptoms are generally attributed to sensitization of nociceptors
by inflammatory mediators released by immune cells. Nociceptor sensitization during
inflammation occurs through activation of the Toll-like receptor 4 (TLR4) signalling pathway
by lipopolysaccharide (LPS), a toxic by-product of bacterial lysis. Here we show that LPS
exerts fast, membrane delimited, excitatory actions via TRPA1, a transient receptor potential
cation channel that is critical for transducing environmental irritant stimuli into nociceptor
activity. Moreover, we find that pain and acute vascular reactions, including neurogenic
inflammation (CGRP release) caused by LPS are primarily dependent on TRPA1 channel
activation in nociceptive sensory neurons, and develop independently of TLR4 activation. The
identification of TRPA1 as a molecular determinant of direct LPS effects on nociceptors offers
new insights into the pathogenesis of pain and neurovascular responses during bacterial
infections and opens novel avenues for their treatment.

Journal Club 2014.12.29 Read More »

journal club – 2014.12.15.

A natural dye, Niram improves atopic dermatitis through down-regulation of TSLP

Na-Ra Hana, Jin-Young Parkb, Jae-Bum Jangb, Hyun-Ja Jeongc,∗, Hyung-Min Kima,∗∗

a Department of Pharmacology, College of Korean Medicine, Kyung Hee University, 1 Hoegi-dong, Dongdaemun-gu, Seoul 130-701, Republic of Korea
b Regional Innovation Center and Inflammatory Disease Research Center, Hoseo University, 165, Sechul-ri, Baebang-myun, Asan, Chungnam 336-795, Republic of Korea

c Department of Food Technology, Biochip Research Center, and Inflammatory Disease Research Center, Hoseo University, 165, Sechul-ri, Baebang-myun, Asan, Chungnam 336-795,Republic of Korea

1-s2.0-S1382668914002476-main

abstract

Naju Jjok (Polygonum tinctorium Lour.) has been known to treat skin diseases in traditional Korean medicine. A natural textile dye, Niram made from Naju Jjok has traditionally been used to dye clothes. Thymic stromal lymphopoietin (TSLP) plays an important role in the development of atopic dermatitis (AD). Thus, we investigated that Niram might amelio- rate AD through regulation of TSLP. Niram significantly inhibited the levels of TSLP through blockade of caspase-1/receptor-interacting protein 2 pathway in stimulated mast cells. Fur- ther, Niram ameliorated clinical symptoms in AD mouse. Niram significantly inhibited the infiltration of inflammatory cells in lesional skin. The levels of TSLP, caspase-1, IL-4, and IL-6 were inhibited in lesional skin applied topically with Niram. Niram significantly inhibited the serum levels of IgE and histamine in AD mouse. Finally, Niram significantly inhibited the levels of TSLP in polyriboinosinic polyribocytidylic acid-stimulated human keratinocyte HaCaT cells. These results establish Niram as a functional dye embracing the aspects of not only a traditional use but also a pharmacological effect.

© 2014 Elsevier B.V. All rights reserved.

Keywords : Niram, TSLP, Caspase-1, Mast cell, Atopic dermatitis, NC/Nga mice

journal club – 2014.12.15. Read More »

journal club 2014.12.01

Antipruritic mechanisms of topical E6005, a phosphodiesterase 4 inhibitor: Inhibition of responses to proteinase-activated receptor 2 stimulation mediated by increase in intracellular cyclic AMP

Abstract

Background

Phosphodiesterase 4 (PDE4), which catalyses the conversion of cyclic adenosine 3′,5′-monophosphate (cAMP) to 5′-AMP, plays a critical role in the pathogenesis of inflammatory disorders. Pruritus is the main symptom of dermatitides, such as atopic dermatitis, and is very difficult to control. Recent studies have shown that the activation of proteinase-activated receptor 2 (PAR2) is involved in pruritus in dermatoses in humans and rodents.

Objective

To investigate the inhibitory effect of E6005, a topically effective PDE4 inhibitor, on PAR2-associated itching in mice.

Methods

Mice were given an intradermal injection of SLIGRL-NH2 (100 nmol/site), a PAR2 agonist peptide, into the rostral part of the back. E6005 and 8-bromo-cAMP were applied topically and injected intradermally, respectively, to the same site. Scratching bouts were observed as an itch-related behavior, and firing activity of the cutaneous nerve was electrophysiologically recorded. Keratinocytes were isolated from the skin of neonatal mice and cultured for in vitro experiments. The concentrations of cAMP and leukotriene B4 (LTB4) were measured by enzyme immunoassay. The distribution of PDE4 subtypes in the skin was investigated by immunostaining.

Results

Topical E6005 and intradermal 8-bromo-cAMP significantly inhibited SLIGRL-NH2-induced scratching and cutaneous nerve firing. Topical E6005 increased cutaneous cAMP content. Topical E6005 and intradermal 8-bromo-cAMP inhibited cutaneous LTB4 production induced by SLIGRL-NH2, which has been shown to elicit LTB4-mediated scratching. E6005 and 8-bromo-cAMP inhibited SLIGRL-NH2-induced LTB4 production in the cultured murine keratinocytes also. PDE4 subtypes were mainly expressed in keratinocytes and mast cells in the skin.

Conclusions

The results suggest that topical E6005 treatment inhibits PAR2-associated itching. Inhibition of LTB4 production mediated by an increase in cAMP may be partly involved in the antipruritic action of E6005.

Abbreviations

  • cAMP, cyclic adenosine 3′,5′-monophosphate;
  • EIA, enzyme immunoassay;
  • IgG, immunoglobulin G;
  • LTB4, leukotriene B4;
  • PDE, phosphodiesterase;
  • PAR2, proteinase-activated receptor 2;
  • PBS, phosphate-buffered saline

dec presentation

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Journal Club 2014.11.24

Descending Control of Itch Transmission by the Serotonergic System via 5-HT1A-Facilitated GRP-GRPR Signaling

Highlights

  • •Central 5-HT signaling facilitates itch transmission
  • •5-HT1A potentiates GRPR-mediated itch signaling
  • •5-HT1A and GRPR are present in close proximity
  • •Blockade of 5-HT1A function reduces chronic itch

Summary

Central serotonin (5-hydroxytryptophan, 5-HT) modulates somatosensory transduction, but how it achieves sensory modality-specific modulation remains unclear. Here we report that enhancing serotonergic tone via administration of 5-HT potentiates itch sensation, whereas mice lacking 5-HT or serotonergic neurons in the brainstem exhibit markedly reduced scratching behavior. Through pharmacological and behavioral screening, we identified 5-HT1A as a key receptor in facilitating gastrin-releasing peptide (GRP)-dependent scratching behavior. Coactivation of 5-HT1A and GRP receptors (GRPR) greatly potentiates subthreshold, GRP-induced Ca2+ transients, and action potential firing of GRPR+ neurons. Immunostaining, biochemical, and biophysical studies suggest that 5-HT1A and GRPR may function as receptor heteromeric complexes. Furthermore, 5-HT1A blockade significantly attenuates, whereas its activation contributes to, long-lasting itch transmission. Thus, our studies demonstrate that the descending 5-HT system facilitates GRP-GRPR signaling via 5-HT1A to augment itch-specific outputs, and a disruption of crosstalk between 5-HT1A and GRPR may be a useful antipruritic strategy.

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Journal Club 2014.11.17

Neural peptidase endothelin-converting enzyme 1 regulates endothelin 1-induced pruritus

supplementary neural peptidase

Neural peptidase endothelin-converting enzyme 1 regulates endothelin 1-induced pruritus.

Kido-Nakahara M, Buddenkotte J, Kempkes C, Ikoma A, Cevikbas F, Akiyama T, Nunes F, Seeliger S, Hasdemir B, Mess C, Buhl T, Sulk M, Müller FU, Metze D,Bunnett NW, Bhargava A, Carstens E, Furue M, Steinhoff M.

Abstract

In humans, pruritus (itch) is a common but poorly understood symptom in numerous skin and systemic diseases. Endothelin 1 (ET-1) evokes histamine-independent pruritus in mammals through activation of its cognate G protein-coupled receptor endothelin A receptor (ETAR). Here, we have identified neural endothelin-converting enzyme 1 (ECE-1) as a key regulator of ET-1-induced pruritus and neural signaling of itch. We show here that ETAR, ET-1, and ECE-1 are expressed and colocalize in murine dorsal root ganglia (DRG) neurons and human skin nerves. In murine DRG neurons, ET-1 induced internalization of ETAR within ECE-1-containing endosomes. ECE-1 inhibition slowed ETAR recycling yet prolonged ET-1-induced activation of ERK1/2, but not p38. In a murine itch model, ET-1-induced scratching behavior was substantially augmented by pharmacological ECE-1 inhibition and abrogated by treatment with an ERK1/2 inhibitor. Using iontophoresis, we demonstrated that ET-1 is a potent, partially histamine-independent pruritogen in humans. Immunohistochemical evaluation of skin from prurigo nodularis patients confirmed an upregulation of the ET-1/ETAR/ECE-1/ERK1/2 axis in patients with chronic itch. Together, our data identify the neural peptidase ECE-1 as a negative regulator of itch on sensory nerves by directly regulating ET-1-induced pruritus in humans and mice. Furthermore, these results implicate the ET-1/ECE-1/ERK1/2 pathway as a therapeutic target to treat pruritus in humans.

Journal Club 2014.11.17 Read More »

journal club 2014-11-10

The Gardenia jasminoides extract and its constituent, geniposide, elicit anti-allergic effects on atopic dermatitis by inhibiting histamine in vitro and in vivo

Abstract

Ethnopharmacological relevance

Gardenia jasminoides Ellis has been used in traditional medicine for treatment of inflammation, edema, and dermaitis. The aim of this study was to investigate the mechanism by which Gardenia jasminoides extract (GJE) elicits anti-allergic effects in mast cells and in mice with atopic dermatitis (AD).

Materials and methods

We investigated the effects of GJE and its fractions on compound 48/80-induced histamine release from MC/9 cells and Dermatophagoides farinae-exposed NC/Nga mice. The effects of its constituents on histamine release from MC/9 cells were also investigated.

Results

GJE and its ethyl acetate fraction (GJE-EA) inhibited compound 48/80-induced histamine release from MC/9 mast cells. The topical application of GJE or GJE-EA to Dermatophagoides farinae-exposed NC/Nga mice reduced the symptoms of AD, inhibited the infiltration of inflammatory cells, and lowered the serum levels of immunoglobulin E and histamine. Both GJE and GJE-EA reduced the expression of cytokines (interleukin [IL]-4, IL-6, and tumor necrosis factor-alpha) and adhesion molecules (intercellular adhesion molecule-1 and vascular cell adhesion molecule-1) in ear lesions. In addition, the quantitative analysis of GJE and GJE-EA by high-performance liquid chromatography revealed the presence of crocin and geniposide. Geniposide, but not crocin, inhibited the release of histamine from mast cells, which may contribute to the anti-allergic effect of GJE and GJE-EA.

Conclusions

These results suggest that GJE and GJE-EA can suppress mast cell degranulation-induced histamine release, and geniposide may be potential therapeutic candidates for AD

 

gardenia paper

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Journal Club 2014.11.3

 

TLR4 enhances histamine-mediated pruritus by potentiating TRPV1 activity (1)
Filename : tlr4-enhances-histamine-mediated-pruritus-by-potentiating-trpv1-activity-1.pdf (2 MB)
Caption :

TLR4 enhances histamine-mediated pruritus by potentiating TRPV1 activity.

Min H, Lee H, Lim H, Jang YH, Chung SJ, Lee CJ, Lee SJ1.

Author information

Abstract

BACKGROUND:

Recent studies have indicated that Toll-like receptor 4 (TLR4), a pathogen-recognition receptor that triggers inflammatory signals in innate immune cells, is also expressed on sensory neurons, implicating its putative role in sensory signal transmission. However, the possible function of sensory neuron TLR4 has not yet been formally addressed. In this regard, we investigated the role of TLR4 in itch signal transmission.

RESULTS:

TLR4 was expressed on a subpopulation of dorsal root ganglia (DRG) sensory neurons that express TRPV1. In TLR4-knockout mice, histamine-induced itch responses were compromised while TLR4 activation by LPS did not directly elicit an itch response. Histamine-induced intracellular calcium signals and inward currents were comparably reduced in TLR4-deficient sensory neurons. Reduced histamine sensitivity in theTLR4-deficient neurons was accompanied by a decrease in TRPV1 activity. Heterologous expression experiments in HEK293T cells indicated thatTLR4 expression enhanced capsaicin-induced intracellular calcium signals and inward currents.

CONCLUSIONS:

Our data show that TLR4 on sensory neurons enhances histamine-induced itch signal transduction by potentiating TRPV1 activity. The results suggest that TLR4 could be a novel target for the treatment of enhanced itch sensation.

Journal Club 2014.11.3 Read More »

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