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Dmitri Serjanov, Galina Bachay, Dale D Hunter, William J Brunken; Laminin β2 Chain Regulates Retinal Progenitor Cell Behavior Via Dystroglycan. Invest. Ophthalmol. Vis. Sci. 2017;58(8):112.
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© ARVO (1962-2015); The Authors (2016-present)
Laminin, a heterotrimeric molecule, is an indispensable organizational component of the extracellular matrix; mutations in laminin genes lead to defective CNS and ocular development in mice and humans. This study investigates the role of laminin β2 chain in regulation of the retinal progenitor cell (RPC) polarity and subsequent proliferation and cell fate adoption. The role of the laminin receptor dystroglycan (DG) was also investigated.
WT and laminin β2-/- retinas were used. IHC was performed using centrosomal and mitotic markers along with DG and integrin β1 (intβ1). RPC mitotic spindles were visualized via 3D-reconstruction of the dividing RPCs and spindle angles were calculated for each nucleus. DG and integrin β1 expression patterns were assayed and quantified digitally. Organotypic cultures were prepared from P0 retinas. Addition of recombinant b2-containing laminin to the retinal surface was used to rescue Lamb2 deletion ex vivo. Reciprocally, addition of DG function-blocking antibodies was used to block DG-mediated signaling in WT cultures.
Laminin β2 chain deletion results in decreased proliferation and premature RPC pool depletion by P5 (down by 70%; p≤0.01), resulting in underproduction of bipolar cells (down by 38%; p≤0.002) and Müller glia (down by 39%; p≤0.003). This is accompanied by a significant shift of the RPC mitotic spindle pole orientation towards asymmetric (neurogenic) cell divisions at P0 and P3 (p≤0.01). Additionally, down-regulation of DG and intβ1 at the retinal vitreal surface was observed. Addition of exogenous β2-containing laminin trimers to the retinal surface ex vivo restored the normal receptor distribution, spindle orientation, and increased RPC proliferation. Addition of dystroglycan-blocking antibodies to WT organotypic cultures phenocopied laminin β2 deletion, resulting in decreased proliferation and asymmetric mitotic spindle orientation, while intβ1-blocking antibodies had no effect.
These data demonstrate that b2-containing laminins at the ILM regulate critical developmental patterning by regulating matrix receptor expression and RPC behavior such as cytokinesis dynamics and proliferation. DG in particular appears to play an important role in laminin β2-mediated ECM-to-RPC signaling. To our knowledge, this is the first report of dystroglycan signaling affecting mitotic spindle orientation.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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