Despite advances in the symptomatic treatment of emphysema, the prevalence of this devastating lung disease is increasing worldwide and there remains no cure save for lung transplantation. We propose that novel tissue engineering approaches utilizing embryonic stem cells (ESCs), adult mesenchymal stem cells (MSCs), or induced pluripotent stem cells (iPS) cultured in three-dimensional culture matrices can regenerate functional lung tissue for replacement of emphysematous lung tissue. We have found that MSCs impregnated into de-cellularized mouse lungs re-cellularize both airways and parenchymal lungs regions and acquire phenotypic markers of type 2 alveolar epithelial cells in the parenchymal lung regions. These findings raise the possibility that embryonic or adult stem cells cultured in de-cellularized lungs can be utilized to engineer new lung tissue ex vivo and that this might be a therapeutic strategy for regenerating new alveoli in emphysematous lungs. Further, lung cells are subjected to cyclic stretch and other forces during normal breathing. We have found that cyclic stretch of cultured MSCs specifically induces expression of lung epithelial proteins while decreasing expression of markers of other cell types such as fibroblasts or muscle cells. These preliminary studies raise the possibility that specific manipulation of the culture environment by applying mechanical forces reminiscent of those found in normal lung can further promote development of functional lung tissue from embryonic or adult stem cells. The influences of 3-dimensional culture and of cyclic stretch, in combination with a specific airways growth medium, on development of lung cells from MSCs, ESCs, and iPS will be explored and defined in this proposal. These studies will provide a firm scientific basis for lung tissue engineering utilizing embryonic and adult stem cells as a possible therapeutic approach for emphysema and other lung diseases. PUBLIC HEALTH RELEVANCE: Chronic obstructive pulmonary disease (COPD), the fourth most common cause of death worldwide, remains a devastating fatal lung disease that is one of the only major diseases increasing in prevalence and for which there is no cure save lung transplantation. A new cell-based therapy approach is suggested by our recent work demonstrating that adult mesenchymal stem cells cultured in three-dimensional scaffolds may form viable lung tissue for potential use in regenerating new lung in emphysema. The focus of the current proposal is to further explore whether 3 dimensional scaffolds as well as mechanical stretch may be used to direct generation of functional 3- dimensional lung tissue from mesenchymal stem cells as well as from embryonic and induced pluripotent stem cells. These studies will provide a firm scientific basis for future translational and clinical trials of stem cells for treatment of emphysema.