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W. J. Brunken, G. Bachay, G. Gnanaguru, S. Varshney, G. Pinzón-Duarte; Interactions with Laminin in ILM Confers Polarity Cues for Müller Cells and Retinal Ganglion Cells. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1300.
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© ARVO (1962-2015); The Authors (2016-present)
Genetic ablation of both β2 and γ3 chains of laminin leads to the disruption of the ILM causing a retinal dysplasia. The development and survival of Müller cells (MCs) and ganglion cells (RGCs) are dependent on the integrity of the ILM. Here, we show that disruptions in laminin expression alters MC and RGC polarity and subcellular organization.
Retinas from wild type (WT), β2-/-, γ3-/- and β2-/-:γ3-/- mice were collected from P0 to 15 and examined by conventional microscopy, immunohistochemistry and electron microscopy. In vitro assays were performed on Müller cell lines and organotypic cultures of early postnatal retina.
MCs, and the neuroprogenitors from which they arise, span the neural retina; at their apical surface, they make tight junctions while on their basal side, they form adhesion complexes. In the laminin β2-/- and β2-/-:γ3-/- animals, the polarity of MCs is lost. Normal apical processes and tight junctions are disrupted as are the regular array of MC endfeet that normally surround cells in the RGC layer. MC sprouting from the apical and basal side is apparent in the β2-/- and β2-/-:γ3-/- retinae. At the basal endfoot, disruptions in adhesion complex were seen including a loss of dystroglycan localization. Exogenous addition of laminins to a MC line, in vitro induces dystroglycan clustering supporting a laminin-mediated scaffolding of these complexes. One consequence of these disruptions is increased proliferation of neural progenitor cells in early postnatal retina (P0-P5) and cells we presume to be MCs in late postnatal retina (P15). RGCs organization is progressively disrupted in the β2-/- and β2-/-:γ3-/- mice. At P15, the optic nerves of laminin nulls are hypoplastic; we are currently charting the time course of RGC death. RGC polarity is disorganized; neurotubulin expression is delocalized and a progressive disorganization of the RGC dendritic tree and axonal trajectories is apparent. In organotypic cultures of WT retina, portions of the retina were stripped of the ILM. In these stripped regions, RGCs polarity is disrupted as it is in the laminin nulls. MCs are also disorganized, in these same regions. Whether the effect on RGCs is mediated by direct contact with the ILM or via interactions with MCs is under investigation.
Adhesion to the laminins in the ILM provides polarity cues for both MCs and RGCs. In the absence of such cues, MCs re-entry the cell cycle and proliferate while RGCs degenerate. These studies have implications for the surgical stripping of the ILM and may be important in proliferative vitreoretinopathy.
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