Scanning ion conductance microscopy reveals how a functional renal epithelial monolayer maintains its integrity

Abstract

To function as a transport barrier a renal tubule epithelial monolayer needs to maintain its integrity when, sudden hypertonic stress causes cell shrinkage, new cells are added, or cells in the monolayer die. However, the mechanism used to achieve this is largely unknown. Scanning ion conductance microscopy (SICM) has been shown to be suitable for imaging the surface of live renal cells with high topographic resolution, and can be used to elucidate how a functional renal epithelial monolayer maintains its integrity. SICM was used for high spatial resolution topographic imaging of Xenopuslaevis renal epithelial A6 cells cultured on membrane filter inserts. The SICM images of A6 cells showed that the epithelial monolayer maintains its integrity under hypertonic stress, and during cell division and death. Sequential SICM topographic images revealed detailed structural changes and their time course for these protective processes, which involve highly cooperative cell movement. Some "balloon-like" structures were observed at susceptible tight junction regions, which were proposed to help cell maintaining the monolayer permeability integrity. SICM is a powerful tool for research on living renal epithelial cells, and has been used to elucidate how a functional epithelial monolayer maintains its integrity. Using this technique we have observed that during hypertonic stress and regeneration, an organized sequence of events protect the loss of integrity of monolayer so that tight junctions and cell-cell contact are maintained and disruption to the function of whole monolayer is prevented.