| Background.;Reciprocal interactions between lung mesenchymal and epithelial cells play essential roles in lung organogenesis and homeostasis. Altered lung mesenchymal cell number and function are related to many chronic lung diseases, such as pulmonary fibrosis and emphysema. Although the molecular markers and related animal models that target lung epithelial cells are relatively well studied, molecular markers of lung mesenchymal cells and the genetic tools to target and/or manipulate gene expression in a lung mesenchyme-specific manner are not available. The ability to manipulate gene expression in specific cell type in the lung mesenchyme is important for understanding the molecular mechanisms of the lung mesenchymal biology and the related pulmonary diseases. Lack of knowledge of gene regulatory elements that contribute to lung mesenchymal cell specificity is a major barrier for developing such genetic tools.;Result.;We have characterized a mouse Tbx4 gene enhancer that contains conserved DNA sequences across many vertebrate species with lung or lung-like gas exchange organ. Two transgenic mouse lines were then generated to express rtTA/LacZ under the control of this Tbx4 lung enhancer. By combining the Tbx4-rtTA driven Tet-On inducible Cre recombinase expression mouse lines with a mT-mG dual fluorescence reporter mouse line, the spatial-temporal patterns of Tbx4 lung enhancer-targeted lung mesenchymal cells were defined. The Tbx4 lung enhancer was active only in lung mesenchymal cells, but not in other tissues, from prenatal developmental stage to adult. Pulmonary endothelial cells and vascular smooth muscle cells were only targeted by the Tbx4-rtTA driver line prior to E11.5 and E15.5, respectively, while other subtypes of lung mesenchymal cells including airway smooth muscle cells, fibroblasts, pericytes could be targeted during the entire developmental stage.;Conclusion.;Activation of the Tbx4 lung enhancer is only detected in mesenchymal cells of developing lung. With our newly created Tbx4 lung enhancer-driven Tet-On inducible system, lung mesenchymal cells can be specifically and differentially targeted in vivo at the first time by controlling the doxycycline induction time window. This novel system provides a unique tool to study lung mesenchymal cell lineages and gene functions in lung mesenchymal development, injury repair, and regeneration in mice.;Background.;Although bone marrow mesenchymal stem cells (BMMSCs) have been extensively studied, tissue specific MSCs, including lung resident MSCs, are poorly defined and characterized. Studies have shown that stem cell-mediated injury repair and regeneration is critical to maintain normal lung structure and function. The increasing recognition of the properties of mesenchymal stem cells has led to various pathogenesis-oriented studies. The roles of lung resident MSCs in lung diseases have never been studied.;Emphysema is the third leading cause of death in U.S, the pathogenesis of emphysema has not been fully elucidated. No effective therapy is available to treat this disease. Recent studies in our lab have demonstrated that Smad3 knockout mice can spontaneously develop emphysema, which offers a great tool for us to investigate the pathogenic mechanisms and test novel therapeutic approaches for emphysema. Therefore, we have used this emphysema-mouse model to study the role of lung resident mesenchymal progenitor cells in emphysema pathogenesis.;Result.;We have successfully isolated potential lung mesenchymal stem cells from mouse lung tissue using modified method of bone marrow MSC isolation. These cells are plastic-adherent, capable of self-renew and differentiation, and expressing certain mesenchymal stem cell markers, same features as described for BMMSC. Furthermore, with the aforementioned Tbx4-rtTA/TetO-Cre/mT-mG transgenic mice of line 6 in last chapter, we found that the isolated lung resident MSCs consist of at least two subpopulations with different developmental origins and repair/regeneration capacities. Lung resident MSCs isolated from our newly developed transgenic mice, in which MSCs with embryonic lung mesenchyme origin are marked with membrane GFP protein, while MSCs derived from other organs are labeled with membrane Tomato protein.;The deficiency of lung resident MSCs and their pathogenic roles in a Smad3 knockout emphysema mouse model were studied. Impaired lung MSC-mediated repair/regeneration may contribute to emphysema pathogenesis. The quantitative and qualitative alterations of lung MSCs subsets in Smad3 knockout mice need further investigation. To determine the therapeutic efficacy of lung resident mesenchymal stem cells, exogenous MSCs administration into emphysema mice should be considered. |
