The Effect of Stromal Components on the Phenotype of Normal Human Bronchial Epithelial Cells in 3D Culture.

摘要

Classical studies in embryology demonstrated that stroma is necessary for the proper specification and differentiation of epithelial tissues. Recently, it was shown that the stroma is involved in the homeostatic maintenance of adult tissues, and under pathologic conditions, promotes the development and progression of diseases such as cancer. Hence, pulmonary diseases such as asthma, fibrosis and cancer can be understood in the context of altered communications between the epithelial and stromal tissue compartments. Bronchi are the conducting airways of the lung.;Bronchi trap and eliminate inhaled particles through the coordinated actions of mucus secretion and the beating of cilia. However, inhaled toxicants and carcinogens are linked to several broncho-pulmonary pathologies, including asthma and lung cancer, which is the single deadliest cancer in the United States; since most lung cancers are attributed to tobacco smoke, it is also one of the most avoidable of cancers.;This thesis describes the construction of an in vitro three-dimensional (3D) model of the human bronchus specifically designed for the study of epithelial-stromal interactions that regulate bronchial homeostasis and pathogenesis. This model mimics the morphology and function of the bronchial mucosa and consists of a type-I collagen matrix, either normal human fetal lung fibroblasts (IMR-90) or primary human adult lung cancer-associated fibroblasts (LuCAFs), and a surface epithelium of normal human bronchial epithelial cells (HBECs). When cultured at an air-liquid interface (ALI), the epithelium generated a well-differentiated pseudostratified bronchial epithelium that contained basal, ciliated, and non-ciliated (secretory) epithelial cells. IMR-90 and LuCAFs differentially altered the phenotype of HBECs in distinct ways. While IMR-90 permitted HBECs to form a typical respiratory surface epithelium, LuCAFs promoted the invasion of HBECs into collagen gel forming both epithelial nodules and cysts. This suggests that LuCAFs may alter the HBEC phenotype by modifying biomechanical signals conveyed through the extracellular matrix (ECM). Furthermore, LuCAFs secreted soluble factors that induced HBECs to express genes associated with immune responses, apoptosis, mitosis, cell survival, differentiation and cancer. In conclusion, we have created a 3D model of the human bronchial mucosa that allows investigators to study epithelial-mesenchymal interactions as determinants of tissue architecture as well as the effects of inhaled toxicants.