The initiation and maintenance of chronic interstitial pulmonary inflammation is often due to dynamic interactions between an inciting agents, and structural cells of the lung. Clinically, the therapeutic modalities that are available to treat these diseases are limited, which likely reflects an insufficient understanding of the pathophysiologic mechanisms that mediate-these disorders. Independent of the various etiologies, many chronic interstitial lung diseases appear to possess a number of similar pathologic responses, including an initial elicitation of various leukocyte populations to the lung, subsequent fibroblast activation and proliferation, and deposition of extracellular matrix. Understanding the cellular and molecular mechanisms which are responsible for fibroblast activation during the initiation, maintenance, and resolution of interstitial lung inflammation are the broad, long-term objectives of this application. The working hypothesis of this proposal is the following: an exacerbation of fibroblast activation and tissue fibrosis during the evolution of chronic. interstitial lung inflammation is dependent upon the expression of a specific disease phenotype characterized by the predominance of Th2-type cytokines. This hypothesis will be addressed by focusing on fibrotic mechanisms which occur during the development of a Thl (minimally fibrotic) versus a Th2 type (fibrotic) immune response in the lung. Specific areas of cytokine biology that w:Ill be assessed include: l)- the contribution of Th2-like cytokines, especially interleukin-4, interleukin-5, and interleukin-10, to fibroblast activation leading to pulmonary fibrosis; 2) the role of Thl like cytokines, especially interleukin-12 and gamma interferon, in altering fibroblast activation during pulmonary fibrosis; 3) a mechanistic analysis of the contribution of cytokines to interstitial inflammation via adenovirus/cytokine cDNA transfection studies; 4) the contribution of activated fibroblasts isolated from either Th1 or Th2 lung lesions to progressive lung inflammation; and 5) the expression and regulation of Th2-like cytokine profiles in human patients and correlations with interstitial lung fibrosis. Animal models of chronic interstitial lung inflammation will be utilized to assess the contribution of Th1 like and Th2 like cytokines during pulmonary fibroblast activation and lung collagen deposition. A number of techniques will be employed in this application, including the use of Northern blot, reverse transcription-polymerase chain reaction (RT-PCR), mRNA stability, and in situ hybridization analyses for studying the regulation of gene expression; the use of immunohistochemistry for antigen localization and ELISAs for the quantitation of cytokines and endogenous mediators of cytokine activity; gene transfection strategies using adenovirus containing either a murine IL-4, IL-5, IL-10, gammaIFN, or IL-12 cDNA cassette and bioassays for assessing functional activities of specific mediators.
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