NONADHESIVE STATIONARY ORGAN-CULTURE OF HUMAN BRONCHIAL-MUCOSA

The supply of fresh bronchial tissue from human donors for in vitro cu lture is limited. Routine fiberoptic bronchoscopy offers a safe and ea sy procedure for obtaining minor biopsies and we wanted to see if the material provided could be used for organ culture by using a simple li quid overlay technique. Bronchial biopsies were cut into fragments 400 -500 mu m and kept immersed in a standard serum-supplemented medium fo r 40 days. An agar base prevented adhesion of the tissue. By light and electron microscopy it was shown that the tissue fragments had a diff erentiated epithelium at their surface throughout the culture period. An outgrowth of epithelial cells on the scaffold of the exposed stroma , covering the surface of the whole fragment, occurred within the firs t 5 days of culture. This epithelium was partly ciliated, 2-4 cell lay ers thick with squamous and cuboidal cells and expressed epithelial ma rkers (cytokeratin and Ber-Ep4). The amount of cilia increased during the first 15 days of culture. The epithelium rested on a neosynthesize d basement membrane as visualized by electron microscopy and immunohis tochemistry with antibodies directed against collagen IV, laminin, and fibronectin. The central stroma consisted of loose connective tissue with fibroblasts. This simple tissue culture model combines maintenanc e and neoformation of bronchial epithelium on top of a living natural substrate, thus enabling direct biological studies on clinical biopsy material under perfectly viable conditions.