Laminin γ3 expression and its functional role during human retinal morphogenesis

Abstract

Purpose : Laminins (Lam) are heterotrimeric glycoproteins of the extracellular matrix composed of a α, β and γ chains. In mammals, 11 different laminin chains have been identified, assembling into at least 16 laminins. They play a significant role during retinal morphogenesis and several chains are expressed through development. Lam γ3, the latest of all discovered laminin chains, was found in the interphotoreceptor matrix (IPM) and the outer plexiform layer during rat development. Currently, the distribution and function of Lam γ3 in human retinal development is unknown and thus forms the focus of this study.

Methods : Lam γ3 distribution was examined in mouse, monkey and human adult retina, the developing human eye and in human embryonic stem cell and induced pluripotent stem cell derived retinal organoids by immunohistochemistry (IHC). The Lam γ3 function was examined in vitro by the application of a blocking antibody to retinal organoid cultures on days 43 and 150 during differentiation. IHC and qPCR were used to assess the impact of Lam γ3 blocking on the development of retinal organoids.

Results : Lam γ3 expression was observed in the inner nuclear layer (INL) and ganglion cell layer of the mouse retina, in the IPM, INL and inner plexiform layer (IPL) of the monkey retina and across the entire adult human retina. During early human eye development (6.3^th and 8^th post conceptual week [PCW]), Lam γ3 was found throughout the retina. In contrast, from 10^th – 16^th PCW, expression of Lam γ3 became restricted to the IPM, IPL, inner neuroblastic zone and, later in development (18^th PCW), in the outer nuclear layer. Lam γ3 expression in retinal organoids changed during development, first showing basal immunoreactivity (day 35) followed by expression towards the apical aspect of neural retina, at the site of developing photoreceptor inner segments (day 150 and 200). Blocking Lam γ3 in organoids during the early stages of retinal development resulted in disorganisation of the neural retina. Later in development, blocking of Lam γ3 resulted in the loss of ganglion cells and ribbon synapses. Furthermore, the retinal organoid laminar organisation was disrupted.

Conclusions : Lam γ3 has a unique tempo-spatial expression pattern which varies across different adult species. These data indicate an important role for Lam γ3 in ganglion cell survival and the formation of the photoreceptor synapse during human retinal development.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.