Mycoplasma pneumoniae activation of airway epithelium

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DESCRIPTION (provided by applicant): Mycoplasma pneumoniae is a major cause of lung disease in humans, and the initial interaction with airway epithelial cells is likely to contribute to subsequent disease development. It is clear that immune responses against mycoplasma are a major component of disease with the recruitment and development of immune-mediated inflammatory responses in the submucosa of the respiratory tract. Thus, the factors produced by airway epithelial cells are likely to influence the pathogenesis of these immune-mediated lesions; however, the full complement of cytokines/chemokines produced by airway epithelial cells is unknown. Furthermore, the mechanisms and mycoplasma components involved in the activation of these cells is uncharacterized. If understood, this information could facilitate the development of new approaches to intervene in mycoplasma and other pulmonary diseases. The long-term objective of this project is to determine the molecular mechanisms central to the generation of immune responses in mycoplasma respiratory disease. This project proposes to test the hypothesis that M. pneumoniae stimulate human respiratory bronchoepithelial cells to produce chemokines or other cytokines that are likely to contribute to the inflammatory lesions. Furthermore, we propose that mycoplasma adhesion protein promote the interaction between mycoplasma lipoproteins and toll-like receptors (TLR) of respiratory epithelial cells, which in turn stimulate the production these factors by epithelial cells. Specifically, we proposed to address the following questions: 1) Do M. pneumoniae membrane components activate or damage human bronchoepithelial cells and is this dependent on adhesion molecules? 2) Does M. pneumoniae stimulate human bronchoepithelial cells via toll-like receptors (TLR)? The experimental designs are: 1) Viable mycoplasma, mycoplasma membranes and lipoproteins will stimulate a human bronchoepithelial cell line. Cytokine/chemokine mRNA and protein levels will be determined. We will compare the activity of an adherent parental mycoplasma strain with a nonadherent strain. 2) The expression of TLR by bronchoepithelial cells will be monitored before and after mycoplasma stimulation. Cell lines lacking TLR will be transfected with plasmids expressing each of the human TLR to determine their roles in mycoplasma (cell, membranes and lipoproteins) stimulation of cells. The role of the TLR on human bronchoepithelial cells will be determined using blocking antibodies or transfection with inactive forms of TLR. These studies will provide the foundation for future studies to further elucidate the molecular events involved in these responses, and examine the role of epithelium-derived chemokines/cytokines in disease pathogenesis. Furthermore, it is likely that these events contribute to the exacerbation of asthma and other respiratory diseases. Thus, modulation of each of these events may lead to novel therapies against mycoplasma and other chronic respiratory diseases. The bacterium, Mycoplasma pneumoniae, is responsible for "walking pneumonia" which comprises at least 30% of the pneumonia cases in the United States and associated with severe asthma. The proposed research will study the mechanism through which this bacterium causes lung disease and more severe asthma. Ultimately, we expect these studies will lead to more effective treatments of diseases due to this bacterium by means of promoting favorable, rather than harmful, responses within the lungs of patients.