Antagonism of the prostaglandin D(2 )receptor CRTH2 attenuates asthma pathology in mouse eosinophilic airway inflammation

BACKGROUND: Mast cell-derived prostaglandin D(2 )(PGD(2)), may contribute to eosinophilic inflammation and mucus production in allergic asthma. Chemoattractant receptor homologous molecule expressed on TH(2 )cells (CRTH2), a high affinity receptor for prostaglandin D(2), mediates trafficking of TH(2)-cells, mast cells, and eosinophils to inflammatory sites, and has recently attracted interest as target for treatment of allergic airway diseases. The present study involving mice explores the specificity of CRTH2 antagonism of TM30089, which is structurally closely related to the dual TP/CRTH2 antagonist ramatroban, and compares the ability of ramatroban and TM30089 to inhibit asthma-like pathology. METHODS: Affinity for and antagonistic potency of TM30089 on many mouse receptors including thromboxane A(2 )receptor mTP, CRTH2 receptor, and selected anaphylatoxin and chemokines receptors were determined in recombinant expression systems in vitro. In vivo effects of TM30089 and ramatroban on tissue eosinophilia and mucus cell histopathology were examined in a mouse asthma model. RESULTS: TM30089, displayed high selectivity for and antagonistic potency on mouse CRTH2 but lacked affinity to TP and many other receptors including the related anaphylatoxin C3a and C5a receptors, selected chemokine receptors and the cyclooxygenase isoforms 1 and 2 which are all recognized players in allergic diseases. Furthermore, TM30089 and ramatroban, the latter used as a reference herein, similarly inhibited asthma pathology in vivo by reducing peribronchial eosinophilia and mucus cell hyperplasia. CONCLUSION: This is the first report to demonstrate anti-allergic efficacy in vivo of a highly selective small molecule CRTH2 antagonist. Our data suggest that CRTH2 antagonism alone is effective in mouse allergic airway inflammation even to the extent that this mechanism can explain the efficacy of ramatroban

Sensory mechanisms in the human upper airway - Role of TRP ion channels

The nasal mucosa is richly innervated with sensory nerves having the ability to detect changes in the environment and mediate symptoms such as secretion and itch. Some TRP ion channels such as TRPV1 and TRPA1 are abundantly expressed in sensory nerves and respond to a wide variety of stimuli, including irritants, inflammatory mediators and temperatures. Another interesting TRP ion channel is TRPV4, which is present in the airway epithelium and capable of sensing warm temperatures and osmotic changes. TRP ion channels may have important roles in airway physiology and pathology. We studied the responses to TRP activation in the upper airway of healthy subjects and patients with allergic rhinitis. Furthermore, we examined the effect of TRPV1 antagonism in allergic rhinitis. We showed that activation of TRPV1 and TRPA1 lead to secretion of MUC5B from submucosal glands, and that TRPV1 activation results in an enhanced rhinorrhea and itch response in patients with allergic rhinitis during pollen season. However, our initial studies were unable to demonstrate any symptomatic relief by a TRPV1 antagonist in allergic rhinitis. Analyses of mRNA of different TRP ion channels indicated TRPV1 expression in nasal epithelium, but we were unable to detect any functional response in ciliated epithelial cells. TRPV4 presented the highest mRNA expression of tested TRP ion channels, and further functional studies implied that TRPV4 may be involved in ciliary beat frequency regulation in primary human epithelial cells. Our studies indicate that TRPV1 expressing sensory neurons and possibly TRPV1 itself are involved in sensory hyperresponsiveness during allergic inflammation, and have a role in itch and mucus secretion. In addition, we suggest that TRPV1 and TRPV4 may contribute to mucociliary defence due to the proposed role of TRPV4 in epithelial ciliary regulation and of TRPV1 in secretion. Further studies are warranted to clarify the roles of TRPV1, TRPA1 and TRPV4 in human airway physiology and in airway disease

Scrotal Calcinosis Treated with Carbon Dioxide Laser : Two Cases and a Short Literature Review

Scrotal calcinosis (SC) is a rare and benign condition characterized by multiple, calcified, usually asymptomatic, nodules in the scrotal skin (1). SC often appears in adolescence or early adulthood and may cause embarrassment (1, 2). The standard therapeutic approach is surgical excision of the affected skin (3, 4). We report here 2 cases of SC treated successfully with carbon dioxide (CO2) laser, enabling precise removal of the cysts, while sparing the surrounding tissue. To our knowledge, there are no previous reports in the literature describing treatment of SC with CO2 laser

TRPV1-mediated itch in seasonal allergic rhinitis.

Background: Patients with allergic rhinitis may be abnormally sensitive to stimulation of the ion channel transient receptor potential vanilloid-1 (TRPV1). Aim of the study: To examine effects of various TRP ion channel activators on sensory symptoms in allergic rhinitis prior to and during seasonal allergen exposure. Methods: Nasal challenges were carried out with the TRPV1-activators capsaicin, anandamide and olvanil. Moreover, challenges were performed with mustard oil (allylisothiocyanate) and cinnamaldehyde as well as menthol, activators of TRPA1 and TRPM8, respectively. Nasal symptoms were monitored after each challenge and compared with symptoms reported following corresponding sham challenges. Symptoms recorded after challenge prior to pollen season were also compared with challenge-induced symptoms during pollen season. Results: The TRPV1, TRPA1 and TRPM8-activators produced sensory symptoms dominated by pain and smart. During seasonal allergen exposure, but not prior to season, TRPV1-activators also induced itch. Furthermore, the seasonal challenge to the TRPV1-activator olvanil was associated with rhinorrhoea. Conclusion: Patients with allergic rhinitis feature an increased itch response to TRPV1 stimulation at seasonal allergen exposure. We suggest that this reflects part of the hyperresponsiveness that characterizes on-going allergic rhinitis. Intervention with the TRPV1-signalling pathway may offer potential treatments of this condition

TRPV1 and TRPA1 stimulation induces MUC5B secretion in the human nasal airway in vivo.

Aim: Nasal transient receptor potential vanilloid 1 (TRPV1) stimulation with capsaicin produces serous and mucinous secretion in the human nasal airway. The primary aim of this study was to examine topical effects of various TRP ion channel agonists on symptoms and secretion of specific mucins: mucin 5 subtype AC (MUC5AC) and B (MUC5B). Methods: Healthy individuals were subjected to nasal challenges with TRPV1 agonists (capsaicin, olvanil and anandamide), TRP ankyrin 1 (TRPA1) agonists (cinnamaldehyde and mustard oil) and a TRP melastatin 8 (TRPM8) agonist (menthol). Symptoms were monitored, and nasal lavages were analysed for MUC5AC and MUC5B, i.e. specific mucins associated with airway diseases. In separate groups of healthy subjects, nasal biopsies and brush samples were analysed for TRPV1 and MUC5B, using immunohistochemistry and RT-qPCR. Finally, calcium responses and ciliary beat frequency were measured on isolated ciliated epithelial cells. Results: All TRP agonists induced nasal pain or smart. Capsaicin, olvanil and mustard oil also produced rhinorrhea. Lavage fluids obtained after challenge with capsaicin and mustard oil indicated increased levels of MUC5B, whereas MUC5AC was unaffected. MUC5B and TRPV1 immunoreactivities were primarily localized to submucosal glands and peptidergic nerve fibres, respectively. Although trpv1 transcripts were detected in nasal brush samples, functional responses to capsaicin could not be induced in isolated ciliated epithelial cells. Conclusion: Agonists of TRPV1 and TRPA1 induced MUC5B release in the human nasal airways in vivo. These findings may be of relevance with regard to the regulation of mucin production under physiological and pathophysiological conditions

Effect of Mucosal TRPV1 Inhibition in Allergic Rhinitis.

Transient receptor potential vanilloid-1 (TRPV1) has been implicated as a mediator of itch in allergic rhinitis. To address this possibility, we synthesized a TRPV1 blocker (SB-705498) for nasal administration in patients with seasonal allergic rhinitis. The pharmacological activity of SB-705498 was confirmed on human TRPV1-expressing HEK293 cells, using fluorometric calcium imaging, and in patients with allergic rhinitis subjected to nasal capsaicin challenges. The effect of SB-705498 was studied in patients with seasonal allergic rhinitis subjected to daily allergen challenges for seven days, using a double-blind, placebo-controlled, randomized and cross-over design. SB-705498 was delivered by nasal lavage 10 min. before each allergen challenge. Primary end-point was total nasal symptom score on days 5 to 7. Nasal peak inspiratory flow and eosinophil cationic protein content in nasal lavages were also monitored. Daily topical applications of SB-705498 at a concentration that inhibited capsaicin-induced nasal symptoms had no effect on total symptom score, nasal peak inspiratory flow and eosinophil cationic protein levels in allergen-challenged patients with seasonal allergic rhinitis. The individual symptom nasal itch or sneezes was also not affected. These findings may indicate that TRPV1 is not a key mediator of the symptoms in allergic rhinitis. However, additional studies, using drug formulations with a prolonged duration of action, should be conducted before TRPV1 is ruled out as a drug target in allergic rhinitis

TRPV4-mediated calcium influx and ciliary activity in human native airway epithelial cells.

The transient receptor potential, vanilloid 4 (TRPV4), is a calcium permeable ion channel expressed in airway epithelial cells. Based on studies of cell lines and animals, TRPV4 has been suggested to play a role in the regulation of ciliary beat frequency (CBF). Whether the same is true for human ciliated epithelial cells is not known. Therefore, the aim was to examine the expression and function of TRPV4 in human native nasal epithelial cells. Expression of TRPV4 mRNA in nasal epithelial cells and in the cell lines BEAS2B and 16HBE was confirmed by quantitative real-time PCR. A marked apical TRPV4 immunoreactivity was observed in nasal epithelial cells using immunocytochemistry. Responses to pharmacological modulation of TRPV4 were assessed with calcium imaging and CBF measurements. The TRPV4 agonist GSK1016790A produced concentration-dependent calcium responses in TRPV4-expressing HEK293, BEAS2B and 16HBE cells, and the TRPV4 antagonist HC067047 caused a rightward shift of the GSK1016790A concentration-response curves. Nasal epithelial cells responded to the TRPV4 agonist GSK1016790A with increased intracellular calcium signals and increased CBF, followed by cessation of ciliary beating and cell death. These effects were prevented or inhibited by the TRPV4 antagonist HC067047, the TRP channel blocker ruthenium red or removal of extracellular calcium. We conclude that TRPV4 is expressed in human primary nasal epithelial cells and modulates epithelial calcium levels and CBF. Thus, TRPV4 may participate in mucociliary clearance and airway protection. However, exaggerated activation of TRPV4 may result in epithelial cell death. This article is protected by copyright. All rights reserved

TRPA1 mediates spinal antinociception induced by acetaminophen and the cannabinoid Delta(9)-tetrahydrocannabiorcol

TRPA1 is a unique sensor of noxious stimuli and, hence, a potential drug target for analgesics. Here we show that the antinociceptive effects of spinal and systemic administration of acetaminophen (paracetamol) are lost in Trpa1(-/-) mice. The electrophilic metabolites N-acetyl-p-benzoquinoneimine and p-benzoquinone, but not acetaminophen itself, activate mouse and human TRPA1. These metabolites also activate native TRPA1 and, as a consequence, reduce voltage-gated calcium and sodium currents in primary sensory neurons. The N-acetyl-p-benzoquinoneimine metabolite l-cysteinyl-S-acetaminophen was detected in the mouse spinal cord after systemic acetaminophen administration. In the hot-plate test, intrathecal administration of N-acetyl-p-benzoquinoneimine, p-benzoquinone and the electrophilic TRPA1 activator cinnamaldehyde produced antinociception that was lost in Trpa1(-/-) mice. Intrathecal injection of a non-electrophilic cannabinoid, Delta(9)-tetrahydrocannabiorcol, also produced TRPA1-dependent antinociception in this test. Our study provides a molecular mechanism for the antinociceptive effect of acetaminophen and discloses spinal TRPA1 activation as a potential pharmacological strategy to alleviate pain