Vertebrate retinal development follows a highly stereotyped pattern, in which the retinal progenitor cells (RPCs) give rise to all retinal types in a conserved temporal sequence. Ensuring the proper control over RPC cell cycle exit and re-entry is, therefore, crucially important for the generation of properly functioning retina. In this study, we demonstrate that laminins, indispensible ECM components, at the retinal surface, regulate the mechanisms determining whether RPCs generate proliferative or post-mitotic progeny. In vivo deletion of laminin β2 in mice resulted in disturbing the RPC cell cycle dynamics, and premature cell cycle exit. Specifically, the RPC S-phase is shortened, with increased numbers of cells present in its late stages. This is followed by an accelerated G2-phase, leading to faster M-phase entry. Finally, the M-phase is extended, with RPCs dwelling longer in prophase. Addition of exogenous β2-containing laminins to laminin β2-deficient retinal explants restored the appropriate RPC cell cycle dynamics, as well as S and M-phase progression, leading to proper cell cycle re-entry. Moreover, we show that disruption of dystroglycan, a laminin receptor, phenocopies the laminin β2 deletion cell cycle phenotype. Together, our findings suggest that dystroglycan-mediated ECM signaling plays a critical role in regulating the RPC cell cycle dynamics, and the ensuing cell fate decisions.

Purpose:Birdshot Retinochoroiditis (BRC) is an uncommon but distinct form of bilateral posterior uveitis. It is generally of indolent onset, making early natural history difficult to study. Our report seeks to expand knowledge on the natural history of the onset of BRC.Observations:Our patient presented with clinical features that were consistent with unilateral BRC, despite it being defined as a bilateral condition. Over the course of one year he developed retinal vasculitis, vitritis and fundus features of BRC in the second eye.Conclusions and Importance:Although BRC is a bilateral disease, our case demonstrates that the onset may sometimes be sequential instead of simultaneous. Unilateral disease that is characteristic of BRC should be monitored for second-eye involvement with multi-modal imaging including fundus photography, angiography, perimetry, electroretinography, and optical coherence tomography of the macula with emphasis on the choroidal thickness.

Proper arteriovenous morphogenesis is crucial for maintaining normal tissue perfusion. However, our understanding of how arterial morphogenesis is regulated in the CNS is incomplete. In this study, we asked whether vascular basement membrane (BM) laminins, specifically the γ3-containing isoforms, regulate retinal arterial morphogenesis. We provide evidence that Laminin-γ3 is deposited at both arterial and venous BMs during arteriogenesis. Vascular BM Laminin-γ3 bound dystroglycan (DG), a laminin receptor preferentially expressed by arterial endothelial cells (ECs) during arteriogenesis. Blockade of laminin-DG binding in vitro led to decreased Delta-like ligand (DLL)-4 expression in ECs. Moreover, genetic deletion of the Laminin-γ3- and EC-specific deletion of DG led to similar defects in retinal arteriogenesis, including reduced Dll4 expression, hyperbranching and reduced smooth muscle coverage. These results implicate a newly identified Laminin-γ3-DG signaling cascade that regulates arterial Dll4/Notch signaling to specify and stabilize retinal arteries.-Biswas, S., Watters, J., Bachay, G., Varshney, S., Hunter, D. D., Hu, H., Brunken, W. J. Laminin-dystroglycan signaling regulates retinal arteriogenesis.

Retinal vascular diseases are among the leading causes of acquired blindness. In recent years, retinal microglia have been shown to influence vascular branching density and endothelial cell proliferation. However, how microglial recruitment and activation are regulated during development remains unclear. We hypothesized that microglial recruitment, activation, and down-stream signaling are modulated by components of the mural basement membrane. We used a reverse genetic approach to disrupt laminin expression in the vascular basement membrane and demonstrate that microglia respond to the mural basement membrane in an isoform-specific manner. Microglial density is significantly increased in the Laminin gamma 3 null (Lamc3(-/-)) retinal superficial vascular plexus and consequently the vascular branching density is increased. Microglia also respond to astrocyte-derived matrices and become hyperactivated in the Lamc3(-/-) retina or when tested in vitro with cell-derived matrix. Pharmacological activation of microglia in the wild-type retina produced an Lamc3(-/-)-like vascular phenotype, whereas pharmacological blocking of microglial activation in the Lamc3(-/-) retina rescued the wild-type vascular phenotype. On the molecular level, microglial transforming growth factor-beta 1 expression is down regulated in the Lamc3(-/-) retina, and SMAD signaling decreased in endothelial cells with a consequent increase in endothelial proliferation. The reverse effects were seen in the Lamb2(-/-) retina. Together, our results demonstrate a novel mechanism by which Laminins modulate vascular branching and endothelial cell proliferation during retinal angiogenesis.

The retina expresses several laminins in the outer plexiform layer (OPL), where they may provide an extracellular scaffold for synapse stabilization. Mice with a targeted deletion of the laminin β2 gene (Lamb2) exhibit retinal disruptions: photoreceptor synapses in the OPL are disorganized and the retinal physiological response is attenuated. We hypothesize that laminins are required for proper trans‐synaptic alignment. To test this, we compared the distribution, expression, association and modification of several pre‐ and post‐synaptic elements in wild‐type and Lamb2‐null retinae. A potential laminin receptor, integrin α3, is at the presynaptic side of the wild‐type OPL. Another potential laminin receptor, dystroglycan, is at the post‐synaptic side of the wild‐type OPL. Integrin α3 and dystroglycan can be co‐immunoprecipitated with the laminin β2 chain, demonstrating that they may bind laminins. In the absence of the laminin β2 chain, the expression of many pre‐synaptic components (bassoon, kinesin, among others) is relatively undisturbed although their spatial organization and anchoring to the membrane is disrupted. In contrast, in the Lamb2‐null, β‐dystroglycan (β‐DG) expression is altered, co‐localization of β‐DG with dystrophin and the glutamate receptor mGluR6 is disrupted, and the post‐synaptic bipolar cell components mGluR6 and GPR179 become dissociated, suggesting that laminins mediate scaffolding of post‐synaptic components. In addition, although pikachurin remains associated with β‐DG, pikachurin is no longer closely associated with mGluR6 or α‐DG in the Lamb2‐null. These data suggest that laminins act as links among pre‐ and post‐synaptic laminin receptors and α‐DG and pikachurin in the synaptic space to maintain proper trans‐synaptic alignment.

Vertebrate retinal development follows a pattern during which retinal progenitor cells (RPCs) give rise to all retinal cell types in a highly conserved temporal sequence. RPC proliferation and cell cycle exit are tightly coordinated to ensure proper and timely production of each of the retinal cell types. Extracellular matrix (ECM) plays an important role in eye development, influencing RPC proliferation and differentiation. In this study, we demonstrate that laminins, key ECM components, in the inner limiting membrane, control mitotic spindle orientation by providing environmental cues to the RPCs. In vivo deletion of laminin β2 in mice of both sexes results in a loss RPC basal processes and contact with the ECM, leading to a shift of the mitotic spindle pole orientation toward asymmetric cell divisions. This leads to decreased proliferation and premature RPC pool depletion, resulting in overproduction of rod photoreceptors at the expense of bipolar cells and Müller glia. Moreover, we show that deletion of laminin β2 leads to disruption and mislocalization of its receptors: dystroglycan and β1-integrin. Addition of exogenous β2-containing laminins to laminin β2-deficient retinal explants stabilizes the RPC basal processes and directs their mitotic spindle orientation toward symmetric divisions, leading to increased RPC proliferation, as well as restores proper receptor localization at the retinal surface. Finally, functional blocking of dystroglycan in wild-type retinal explants phenocopies laminin β2 ablation. Our data suggest that dystroglycan-mediated signaling between RPCs and the ECM is of key importance in controlling critical developmental events during retinogenesis.SIGNIFICANCE STATEMENT The mechanisms governing retinogenesis are subject to both intrinsic and extrinsic signaling cues. Although the role of intrinsic signaling has been the subject of many studies, our understanding of the role of the microenvironment in retinal development remains unclear. Using a combination of in vivo and ex vivo approaches, we demonstrate that laminins, key extracellular matrix components, provide signaling cues that control retinal progenitor cell attachment to the basement membrane, mitotic axis, proliferation, and fate adoption. Moreover, we identify, for the first time, dystroglycan as the receptor responsible for directing retinal progenitor cell mitotic spindle orientation. Our data suggest a mechanism where dystroglycan-mediated signaling between the cell and the extracellular matrix controls the proliferative potential of progenitors in the developing CNS.

While genetic networks and other intrinsic mechanisms regulate much of retinal development, interactions with the extracellular environment shape these networks and modify their output. The present review has focused on the role of one family of extracellular matrix molecules and their signaling pathways in retinal development. In addition to their effects on the developing retina, laminins play a role in maintaining Müller cell polarity and compartmentalization, thereby contributing to retinal homeostasis. This article which is intended for the clinical audience, reviews the fundamentals of retinal development, extracellular matrix organization and the role of laminins in retinal development. The role of laminin in cortical development is also briefly discussed.