| Recent survey estimated that about300million people have asthma. Though the symptoms could be alleviated by inhaling immunosuppressants (i.e., glucocorticoids) and β-adrenergic receptor agonists in most patients, while5%to10%of asthmatic patients with chronic asthma are resistant to these conventional therapies, which showed poor effect, even no effect. However, the mechanism of steroid resistance in this kind of airway inflammation is not yet clear. Moreover, many drugs for asthma treatment also have a variety of issues, such as long-term side effects, low intestinal absorbance. Thus, elucidation of steroids resistance in asthmatic airway inflammation and the development of more effective drugs become a top project in the field of respiratory diseases. The present study is designed to investigate the mechanim of steroid resistance and neutrophil infiltration in airway inflammation in an asthmatic mice model, and to determine if the anti-inflammatory effect of Vam3, a newly synthetized compound based on resveratrol, on immune complex-mediated inflammation is through suppression of Syk pathway. Neutrophilic airway inflammation is associated with steroid-resistant asthma, but most animal models use only one allergen, which cannot simulate asthma closely as seen in patients. To determine which inflammatory process is involved in this severe status, female BALB/c mice were repetitively challenged with the pooled extract of dust mite, ragweed and Aspergillus species (DRA) or normal saline as control in10weeks. Dexamethasone or its diluent as placebo control was administered in the last two weeks of exposure. We found that DRA challenge without treatment increased IL-10and TGF-β levels and neutrophil recruitment in bronchial alveolar lavage fluid. Dexamethasone suppressed the release of those2cytokines associated with mast cells recruitment, mucus hypersecretion, but increased, not decreased, neutrophil infiltration and elevated the level of keratinocyte-derived chemokine (mKC), a functional homolog of human interleukin8(IL-8). In in vitro studies using human lung alveolar A549cells, pretreatment with Der p1, an extract of house dust mite Dermatophagoides pteronyssinus, increased the expression of IL-8and activity of NF-κB. However, dexamethasone did not suppress the elevated IL-8level as BAY11-7082, a selective NF-κB inhibitor. The results suggested that increased IL-8(mKC) levels may be involved in steroid-resistant neutrophilic airway inflammation through a NF-kB-dependent pathway.Spleen tyrosine kinase (Syk) plays an indispensable role through preliminary extracellular antigen-induced cross-linking of FcεR I in acute and chronic inflammation. The goal of this part of study was to determine whether Vam3modulates immune complex-mediated inflammation through suppression of Syk pathway. In our study, Vam3could selectively inhibit Syk kinase, suppressed rat peritoneal mast cell and RBL-2H3cells degranulation (ICso=2μM) and Ca2+release in a dose order. To investigate the effects of Vam3on intracellular signaling and mast cell activation, RBL-2H3cells were employed. It was found that binding of DNP IgE to FcεRI on mast cell membranes increased intrinsic tyrosine kinase Syk activity, activated downstream events including phosphorylation of phospholipase Cγ1(PLCyl), Linker for Activation of T cells (LAT), cytosolic phospholipases A2(cPLA2), mobilization of intracellular Ca2+. These findings demonstrated that Vam3attenuated degranulation and Ca2+release through the suppression of multiple step signaling and might be beneficial for the prevention of allergic inflammation. |
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