Isolation and Characterization of Glial Cell Lines from Xenopus Neuroepithelium and Retinal Pigment Epithelium

Abstract

We have isolated several immortal cell lines from Xenopus neuroepithelium and retinal pigment epithelium. These cell lines were initially isolated from primary cultures by serial passaging of proliferating cells, followed by subcloning with limiting dilution techniques. Several morphologically distinct cell lines have been isolated using these procedures. On the basis of immunocytochemical characterization using specific antibodies, we have established that three of these cell lines, the XR1, XRpe1, and XRpe2 cell lines, are glial-like in nature. These cell lines were extensively labeled by antibodies against glial fibrillary acidic protein and vimentin, markers used to identify glial cells. Mono-layers of these cell lines served as useful substrates for axon outgrowth from developing retinal ganglion cells. In addition, analysis of cell-free substrates, prepared by treatment of cell line monolayers with Triton X- 100, revealed that the XR1, XRpe1, and XRpe2 cell lines produce an extracellular matrix (ECM) with potent neurite outgrowth-promoting activity. In contrast, other established retinal and nonretinal Xenopus cell lines were relatively ineffective and did not support axon outgrowth. We propose that neurite outgrowth-promoting activity produced by these cell lines is associated with their ECM and may be glial cell specific. In addition, to further characterize these cell lines, we have recently imaged live cells, using the atomic force microscope (AFM). The use of AFM on living, cultured cells provides a new, high-resolution method for examining dynamic cytoskeletal and morphological events.