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

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          Abstract

          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.

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          Cellular remodeling in mammalian retina: results from studies of experimental retinal detachment.

          Retinal detachment, the separation of the neural retina from the retinal pigmented epithelium, starts a cascade of events that results in cellular changes throughout the retina. While the degeneration of the light sensitive photoreceptor outer segments is clearly an important event, there are many other cellular changes that have the potential to significantly effect the return of vision after successful reattachment. Using animal models of detachment and reattachment we have identified many cellular changes that result in significant remodeling of the retinal tissue. These changes range from the retraction of axons by rod photoreceptors to the growth of neurites into the subretinal space and vitreous by horizontal and ganglion cells. Some neurite outgrowths, as in the case of rod bipolar cells, appear to be directed towards their normal presynaptic target. Horizontal cells may produce some directed neurites as well as extensive outgrowths that have no apparent target. A subset of reactive ganglion cells all fall into the latter category. Muller cells, the radial glia of the retina, undergo numerous changes ranging from proliferation to a wholesale structural reorganization as they grow into the subretinal space (after detachment) or vitreous after reattachment. In a few cases have we been able to identify molecular changes that correlate with the structural remodeling. Similar changes to those observed in the animal models have now been observed in human tissue samples, leading us to conclude that this research may help us understand the imperfect return of vision occurring after successful reattachment surgery. The mammalian retina clearly has a vast repertoire of cellular responses to injury, understanding these may help us improve upon current therapies or devise new therapies for blinding conditions.
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            Apoptotic photoreceptor degeneration in experimental retinal detachment.

            To investigate the possibility that cell death in retinal detachment may occur by reactivation of apoptotic programmed cell death mechanisms. Unilateral retinal detachments were created in adult cats using 0.25% sodium hyaluronate; detached and control retinas were studied at different intervals. Internucleosomal DNA fragmentation (one of the landmarks of apoptosis) was investigated in tissue sections with the TUNEL technique, which uses terminal transferase to label with biotinylated nucleotides the 3' ends of DNA fragments. Sections also were labeled with propidium iodide, which intensely stains pyknotic nuclei. In addition, one time point was selected for analysis with electron microscopy. TUNEL-positive (T+) and propidium iodide-positive (PI+) cells almost never were observed in retinas from control eyes, but they were abundant at defined time points after retinal detachment, appearing almost exclusively in the photoreceptor layer. Their frequency was particularly high 1 to 3 days after detachment but declined rapidly over the next several weeks. T+ cells were still present 28 days after retinal detachment. Electron microscopy also revealed evidence of apoptotic cells after retinal detachment. Results are consistent with the hypothesis that photoreceptor degeneration after retinal detachment occurs through apoptosis, usually associated with intrinsic, programmed cell death mechanisms. The detection of a rapid wave of photoreceptor degeneration seems to suggest that early therapeutic interventions might be recommended; agents capable of interfering with the apoptotic mechanism could have a role in the prevention of cell losses that represent a critical complication of retinal detachment.
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              The mononuclear phagocyte system revisited.

              The mononuclear phagocyte system (MPS) was defined as a family of cells comprising bone marrow progenitors, blood monocytes, and tissue macrophages. In this review, we briefly consider markers for cells of this lineage in the mouse, especially the F4/80 surface antigen and the receptor for macrophage colony-stimulating factor. The concept of the MPS is challenged by evidence that there is a separate embryonic phagocyte lineage, the blurring of the boundaries between macrophages and other cells types arising from phenotypic plasticity and transdifferentiation, and evidence of local renewal of tissue macrophage populations as opposed to monocyte recruitment. Nevertheless, there is a unity to cells of the MPS suggested by their location, morphology, and shared markers. We discuss the origins of macrophage heterogeneity and argue that macrophages and antigen-representing dendritic cells are closely related and part of the MPS.
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                Author and article information

                Journal
                Mol Vis
                MV
                Molecular Vision
                Molecular Vision
                1090-0535
                2008
                28 November 2008
                : 14
                : 2148-2156
                Affiliations
                [1 ]University of Valladolid, Instituto Universitario de Oftalmobiologia Aplicada (IOBA), Campus Miguel Delibes, Valladolid, Spain
                [2 ]University of Valladolid, Faculty of Medicine, Celullar Biology, Histology and Pharmacology Department, Ramon y Cajal nº 7, Valladolid, Spain
                Author notes
                Correspondence to: Ivan Fernandez-Bueno, University of Valladolid, Instituto Universitario de Oftalmobiologia Aplicada (IOBA), Campus Miguel Delibes s/n Valladolid, Valladolid 47011, Spain; Phone: +34 983 184 753; FAX: +34 983 210 824; email: ifernandezb@ 123456ioba.med.uva.es
                Article
                252 2008MOLVIS0183
                2593001
                19052655
                61a754c0-0a68-42bf-9fd2-d3511a18eb26
                Copyright @ 2008

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 02 June 2008
                : 20 November 2008
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                Vision sciences
                Vision sciences

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