Porcine Alveolar Epithelial Cells in Primary Culture: Morphological, Bioelectrical and Immunocytochemical Characterization

Abstract

The purpose of this study was to establish a primary culture of porcine lung epithelial cells as an alternative to the currently existing cell cultures from other species, such as e.g., rat or human. Primary porcine lung epithelial cells were isolated, cultivated and analyzed at distinct time points after isolation. The main part of the work focused on the morphology of the cells and the detection of alveolar epithelial cell markers by using electron microscopy, immunofluorescence microscopy and immunoblotting. Regarding a later use for in vitro pulmonary drug absorption studies the barrier properties of the cell monolayer were evaluated by monitoring bioelectrical parameters and by marker transport. Epithelial cells isolated from porcine lung grew to confluent monolayers with typical intercellular junctions within a few days. Maximum transepithelial resistance of about 2,000 Omega cm2 was achieved and demonstrated the formation of a tight epithelial barrier. Permeability data of sodium fluorescein recommended a minimal transepithelial resistance of 600 Omega cm2 for transport studies. The cell population changed from a heterogeneous morphology and marker distribution (caveolin-1, pro-SP-C, surface sugars) towards a monolayer consisting of two cell types resembling type I and type II pneumocytes. The porcine alveolar epithelial primary cell culture holds promise for drug transport studies, because it shares major hallmarks of the mammalian alveolar epithelium and it is easily available and scaled up for drug screening.