Mucus hypersecretion and persistent airway inflammation are common features of various airway diseases, such as asthma, microbial infections, chronic obstructive pulmonary disease, and cystic fibrosis. One key question is: Does the associated airway inflammation in these diseases affect mucus production? If so, what is the underlying mechanism? It appears that increased mucus secretion results from increased mucin gene expression and is also frequently accompanied by an increased number of mucous cells (goblet cell hyperplasia/metaplasia) in the airway epithelium. Many studies on mucin gene expression have been directed toward Th2 cytokines such as IL-4, IL-9, and IL-13, because of their known pathophysiological role in allergic airway diseases such as asthma. However, the effect of these cytokines has not been definitely linked to their direct interaction with airway epithelial cells. In order to assess the role of cytokine in the regulation of airway mucous cell differentiation, we initiated a direct treatment of well-differentiated human primary tracheobronchial epithelial cells with a panel of cytokines (interleukin-1a, 1B, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 15, 16, 17, 18, 23 and TNFa). To our surprise, only IL-6 and IL-17, not the Th2 type, were able to stimulate mucin gene (MUC5AC.MUC5B) expression in a time and dose-dependent fashion. Since IL-6 and IL-17 are found in airway lumen associated with chronic inflammation, a further study of the role of these cytokines in the regulation of airway mucous cell differentiation is needed. In this application, we hypothesize that IL-17-enhanced mucin gene expression is through an IL-6 autocrine/paracrine loop-dependent pathway via a transcriptional mechanism. To test this hypothesis, three Specific Aims are proposed.
Aim 1 is to test the hypothesis that IL-17 stimulates IL-6 secretion and the secreted IL-6 is partly responsible for IL-17- dependent stimulation of MUC gene expression in airway epithelium. Using various inhibitors, DN and CA expression clones, siRNA silencer approach as well as the use of lentiviral delivery systems, IL-6-dependent signaling transductions related to IL-17 mediated mucin gene expression will be elucidated.
Aim 2 is to test the hypothesis that cis-acting elements at the 5'-flanking region of MUC5B gene are responsible for both IL-17-mediated IL-6-dependent and -independent mucin gene expression. Using the promoter-reporter gene expression approach and genomic foot printing, the nature of cis-/trans-activation will be elucidated. These studies will further resolve the molecular mechanism underlined these diverse pathways. Lastly, Aim 3 is to develop an in vivo model that the function of IL-17 in mediating MUC gene expression can be specifically carried out in vivo. Using a neutrophil depleted protocol and triple transgenic mice approach to generate an in vivo environment that the effects of IL-17 directly on airway epithelial cell differentiation can be assessed. These in vitro and in vivo approaches will provide the molecular basis and signaling pathways involved in the regulation of airway MUC gene expression. These studies may also provide the basic information for therapeutic application for the treatment of aberrant mucus production associated with various airway diseases.
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