Recent studies have indicated the phenotype of the hematopoietic stem cell may show dramatic cell cycle related plasticity as relates to long-term engraftment, progenitor numbers, and probably differentiation profile. These data have led to the concept that at the primitive marrow hematopoietic stem cell level, there is a continuum rather than a hierarchy with a continuously shifting phenotype. Other data have shown that marrow cells and purified marrow stem cells when infused into mice under varying conditions may give rise to nonhematopoietic cells in liver, skin, lung, brain, heart, and GI tract. In several instances, both quantitatively and functionally, significant transdifferentiation has been attained and shown to impact on specific disease manifestations, i.e., in the liver and heart. These data form the basis for the current proposal in which we will be exploring whether or not the transdifferentiation of marrow to lung cells is a phenotype which also shifts with phase of cell cycle. We will study whole marrow cells or purified marrow stem cells stimulated to transit cell cycle by IL-3, IL-6, IL-11, and Steel factor and at different points in cell cycle we will evaluated their ability to form lung cells. Host animals will be subjected to different lung injuries including intertracheal bleomycin and irradiation. Marrow cells will be infused at varying times after injuries. We will utilize B6129SF1/J, Rosa26, and C57BL/6 mice for these studies and track donor cells determining lacZ expression. In separate experiments, we will establish Rosa 26 donor marrow chimerism in lethally irradiated mice, allow for a period of stability, and then evaluate the ability of those chimeric cells subjected to varying stem cell mobilizations to produce lung cells in damaged lung. We plan to evaluate clonality as a measure of the heritogeneity of potential marrow to lung cells and an evaluation of the surface phenotype of stem cells at particular lung differentiation hot spots. We will evaluate adhesion proteins, cytokine receptors, and other epitopes of interest. These studies utilize MoFIo high-speed cell sorting, standard engraftment techniques, phage display biopanning, and immunohistochemical staining. We will also evaluate specific lungs from mice with specific injuries for possible homing peptides using phage display. This proposal hopes to define populations of cells either with cytokine stimulation at particular points in cell cycle or selected from whole marrow populations which will have a phenotype of producing lung cells and which might provide an approach to get quantitatively and functionally significant lung cell production, such that a pre-clinical model might be established. This could then lead to therapies of various lung disorders such as acute respiratory distress syndrome.
