Fusion of cultured dog kidney (MDCK) cells: I. Technique, fate of plasma membranes and of cell nuclei

Abstract

Summary The evaluation of the intracellular signal train and its regulatory function in controlling transepithelial transport with electrophysiological methods often requires intracellular measurements with microelectrodes. However, multiple impalements in epithelial cells are hampered by the small size of the cells. In an attempt to avoid these problems we fused cells of an established cell line, Madin Darby canine kidney cells, originally derived from dog kidney, to “giant” cells by applying a modified polyethylene glycol method. During trypsin-induced detachment from the ground of the petri dish, individual cells grown in a monolayer incorporate volume and mainly lose basolateral plasma membrane by extrusion. By isovolumetric cell-to-cell fusion, spherical “giant” cells are formed within 2 hr. During this process a major part of the individual cell plasma membranes is internalized. Over three weeks following cell plasma membrane fusion degradation of single cell nuclei and cell nuclear fusion occurs. We conclude that this experimental approach opens the possibility to investigate ion transport of epithelia in culture by somatic cell genetic techniques.