Effect of H4R Antagonist N-(2-Aminoethyl)-5-Chloro-1H-Indole-2-Carboxamide (Compound A) in a Mouse Model of Allergic Asthma.

Context: Allergic asthma is a multifactorial airway disease characterised by chronic lung inflammation and airway remodelling. The histamine H4 receptor involved in the chemotaxis of leukocytes and mast cells to the site of inflammation is suggested to be a potential drug target for allergy and asthma. In this study we examined the effect of Compound A, N-(2-Aminoethyl)-5-chloro-1H-indol-2-carboxamide a H4 receptor antagonist in allergic asthma mice model. Objective: To investigate the anti-asthmatic effect of compound A in in vivo, airway inflammation in ovalbumin (OVA) induced allergic asthma mouse model was used. Methodology: Allergic asthma was induced in Balb/c mice using ovalbumin. BAL fluid was examined for the level of IgE, IL-4, IL-5, IL-13 and IL-17 using ELISA. Furthermore, infiltration of leucocytes by histopathology and effect of compound A on signalling molecules were examined in lung tissue. Results: In mice pre-treatment with compound A (10 mg/kg, 20 mg/kg, 30 mg/kg) at different concentrations markedly reduced the levels of IgE, Th2 cytokine IL-4, IL-5, IL-13 and Th17 cytokine IL-17 in BAL fluid. Histopathological examination of lung tissue showed that compound A was able to reduce the level of inflammatory infiltrates. Furthermore, lung tissue from Compound A treated group shown to down-regulate the levels of signalling molecules such as ERK1/2, Akt, SAPK/JNK and NF-κB compared to OVA treated group. Discussion and conclusion: Taken together our data demonstrates that compound A has shown to block the H4R-mediated allergic inflammation in this allergic asthma mice model and may be used as a molecule to study the function of H4R. Abbreviations: Compound A, N-(2-Aminoethyl)-5-chloro-1H-indol-2-carboxamide; JNJ7777120, 1-[(5-chloro-1H-indol-2-yl)carbonyl]-4-methylpiperazine; H4R: Histamine 4 Receptor; AHR: Airway hyper responsiveness.

Silencing of H4R inhibits the production of IL-1ß through SAPK/JNK signaling in human mast cells.

CONTEXT: Mast cell (MC) activation through H4R releases various inflammatory mediators which are associated with allergic asthma. OBJECTIVES: To investigate the siRNA-mediated gene silencing effect of H4R on human mast cells (HMCs) functions and the activation of stress-activated protein kinases (SAPK)/jun amino-terminal kinases (JNK) signaling pathways for the release of ineterleukin-1ß (IL-1ß) in HMCs. MATERIALS AND METHODS: H4R expression was analyzed by RT-PCR and western blotting in human mast cell line-1 (HMC-1) cells and H4RsiRNA transfected cells. The effect of H4RsiRNA and H4R-antagonist on H4R mediated MC functions such as intracellular Ca2+ release, degranulation, IL-6 and IL-1ß release, and the activation SAPK/JNK signaling pathways were studied. HMC-1 cells were stimulated with 10 µM of histamine (His) and 4-methylhistamine (4-MH) and pretreated individually with H4R-antagonist JNJ7777120 (JNJ), histamine H1 receptor (H1R)-antagonist mepyramine, and signaling molecule inhibitors SP600125 (SP) and Bay117082. RESULTS: We found that the HMC-1 cells expressed H4R and H4RsiRNA treatment down regulated the H4R expression in HMC-1 cells. Both His and 4-MH induced the intracellular Ca2+ release and degranulation whereas; H4R siRNA and JNJ inhibited the effect. Furthermore, the activation of H4R caused the phosphorylation of SAPK/JNK pathways. H4R gene silencing and pretreatment with SP and JNJ decreased His and 4-MH induced phosphorylation of SAPK/JNK. We found that the activation of H4R caused the release of IL-1ß (124.22 pg/ml) and IL-6 (122.50 pg/ml) on HMC-1 cells. Whereas, SAPK/JNK inhibitor (68.36 pg/ml) inhibited the H4R mediated IL-1ß release. CONCLUSIONS: Taken together, the silencing of H4R inhibited the H4R mediated MC functions and SAPK/JNK phosphorylation. Furthermore, the H4R activation utilized SAPK/JNK signaling pathway for IL-1ß release in HMC-1 cells.

Local and Systemic Profiles of Inflammatory Cytokines in Carrageenan-induced Paw Inflammation in Rats.

OBJECTIVE: Carrageenan (CA)-induced edema has been described as highly reproducible model of acute inflammation. However, little is known about the cytokines attributed to the CA-induced inflammation. In this study, we aimed to investigate the local and systemic expression profiles of various inflammatory cytokines following the subplantar injection of CA in rats. METHODOLOGY: Acute inflammation was induced in male Wistar rats by subplantar injection of CA. Serum and paw tissue were examined for the level of 19 specific inflammatory cytokines using antibody array. Further, the CA-elicited level of key inflammatory cytokines, cytokine-induced neutrophil chemoattractant (CINC)-2, CINC-3, interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α, were quantified by enzyme-linked immunosorbent assay (ELISA). RESULTS: Edema was peaked 3 h postinjection of CA in hind paw. Among 19 specific cytokines profiled using antibody array, CA significantly (p < 0.05) elicited the levels of CINC-2, CINC-3, IL-1ß, IL-6, ß-NGF, TNF-α, and VEGF in paw tissue and that of CINC-2 and CINC-3 in serum. Consistently, levels of CINC-2, CINC-3, IL-1ß, IL-6, and TNF-α in tissue and CINC-2 and CINC-3 in serum were upregulated in CA-treated rats when compared to control, quantified by ELISA. CONCLUSIONS: This study corroborates the distinct pattern of inflammatory cytokines involved during CA-induced acute inflammation. Furthermore, data provide new evidence on elevated expression of rat CXC chemokines: CINC-2 and CINC-3 at the site of inflammation as well as their significant reflection in the circulation, thereby suggesting their frontline role in CA-induced acute inflammation.

Vitex trifolia L. modulates inflammatory mediators via down-regulation of the NF-κB signaling pathway in carrageenan-induced acute inflammation in experimental rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Vitex trifolia L. (V. trifolia L.), commonly known as the three-leaved chaste tree, is extensively employed in traditional Chinese medicine (TCM) to treat various conditions associated with inflammation. AIM OF THE STUDY: The present study aimed to delineate the molecular mechanisms responsible for the anti-inflammatory effect of V. trifolia L. in carrageenan (CA)-induced acute inflammation in experimental rats. MATERIALS AND METHODS: CA-induced rat paw edema model was adopted to investigate the anti-inflammatory effect of methanolic extract from leaves of V. trifolia L. (VTME) in vivo. Leukocyte infiltration into the site of inflammation was determined by histopathological analysis. Further, the effect of VTME on CA-induced local and systemic levels of specific cytokines was quantified by enzyme-linked immunosorbent assay (ELISA). Moreover, its impact on the nuclear translocation of nuclear factor Kappa B (NF-κB) was analyzed by employing the western blotting technique. RESULTS: VTME at the doses of 100 mg/kg and 200 mg/kg significantly inhibited the paw edema induced by CA (p < 0.05) and effectively reduced the inflammatory leukocyte infiltration. Further, VTME markedly inhibited the CA-induced levels of Interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α in tissue, and that of cytokine-induced neutrophil chemoattractant (CINC)-2/C-X-C motif chemokine (CXCL)3 and CINC-3/CXCL2 in tissue as well as in serum. On the other hand, VTME significantly upregulated the tissue concentration of anti-inflammatory cytokine IL-10. Moreover, VTME significantly attenuated the CA-induced IκBα degradation and nuclear translocation of NF-κB p65. CONCLUSIONS: Our results demonstrate the potent anti-inflammatory effect of V. trifolia L. in vivo, providing insight into its molecular mechanism, which is mediated through down-regulation of NF-κB signal transduction.

The Role of Histamine and Histamine Receptors in Mast Cell-Mediated Allergy and Inflammation: The Hunt for New Therapeutic Targets.

Histamine and its receptors (H1R-H4R) play a crucial and significant role in the development of various allergic diseases. Mast cells are multifunctional bone marrow-derived tissue-dwelling cells that are the major producer of histamine in the body. H1R are expressed in many cells, including mast cells, and are involved in Type 1 hypersensitivity reactions. H2R are involved in Th1 lymphocyte cytokine production. H3R are mainly involved in blood-brain barrier function. H4R are highly expressed on mast cells where their stimulation exacerbates histamine and cytokine generation. Both H1R and H4R have important roles in the progression and modulation of histamine-mediated allergic diseases. Antihistamines that target H1R alone are not entirely effective in the treatment of acute pruritus, atopic dermatitis, allergic asthma, and other allergic diseases. However, antagonists that target H4R have shown promising effects in preclinical and clinical studies in the treatment of several allergic diseases. In the present review, we examine the accumulating evidence suggesting novel therapeutic approaches that explore both H1R and H4R as therapeutic targets for histamine-mediated allergic diseases.

A bioinformatics search for selective histamine h4 receptor antagonists through structure-based virtual screening strategies.

The prevalence of allergic disease is increasing dramatically in the developed world. Studies of allergic diseases have clearly demonstrated that histamine plays an important role in the pathogenesis of the early-phase allergic response. Histamine effects are mediated by H1, H2, H3, and H4 receptors. The presence of the histamine H4 receptors on leukocytes and mast cells suggests that the new histamine receptor H4 plays an important role in the modulation of the immune system. Thus, histamine H4 receptor is an attractive target for anti-allergic therapy. In our present study, we have generated a histamine H4 receptor model using I-TASSER based on human B2-adrenergic G-protein-coupled receptor. Structurally similar compounds of the three known antagonists JNJ777120, thioperamide, and Vuf6002 were retrieved from PubChem, and database was prepared. Virtual screening of those databases was performed, and six compounds with high docking score were identified. Also the binding mode revealed that all the six compounds had interaction with Asp94 of the receptor. Our results serve as a starting point in the development of novel lead compounds in anti-allergic therapy.