Müller and macrophage-like cell interactions in an organotypic culture of porcine neuroretina

Purpose: To analyze the in vitro Müller cell modifications in an organotypic culture of porcine neuroretina in response to the addition of a blood-derived mononuclear fraction (MNF; monocytes and lymphocytes) as a source of macrophages. Methods: Control and MNF-stimulated neuroretinal explants were examined at 3, 6, and 9 days of culture. Specimens were processed for epoxy-resin embedding and cryosectioning. Light and immunofluorescence microscopy were performed, using toluidine blue staining and antibodies against glial fibrillary acidic protein (GFAP), as a reactive gliosis marker, and cellular retinaldehyde-binding protein (CRALBP), as a Müller cell marker. Results: Compared to controls, explants cocultured with MNF displayed increased cellular disorganization and larger tissue invasion of the subretinal space at 9 days of culture. Immunostaining of the MNF-treated explants revealed evidence of more reactive gliosis and greater number of GFAP-immunoreactive Müller cells that had increased width and processes extending into the subretinal space and forming a multilayer tissue. Astrocytes also responded to the MNF addition, producing extensions that invaded the neuroretinal outer layers. Conclusions: Addition of MNF stimulates modifications of Müller cells, producing a wider intraretinal reactive gliosis and tissue proliferation at the subretinal space (outer layers of the retina). These findings emphasize the role of macrophage-like cells in the production of changes in retinal structure observed after retinal detachment in humans.