The influence of X-box Binding Protein 1 and the Unfolded Protein Response on Müller glia differentiation into retinal neurons

Abstract

Purpose : Retinal injury or pathologies often result in the loss of retinal cells and subsequent vision impairment. Endogenous repair of the mammalian retina does not occur to any meaningful extent. After retinal insult, Müller glia can de-differentiate into multipotent progenitor cells. We explore factors that influence Müller glia to act as a stem cell-like population able to re-differentiate into neuronal cell types and potentially repair the retina.

Methods : We isolate Müller glia from adult wild type mice and inducible, cre-activated conditional knockout mice that lack X-box binding protein-1 (XBP1) in Müller glia. In vitro, we measure endoplasmic reticulum (ER) stress and activation of the Unfolded Protein Response (UPR) as Müller glia de-differentiate to a progenitor-like state and re-differentiate into neuronal subtypes. Protein levels for ER stress and cell-specific markers are determined with Western blotting and immunocytochemical staining. After multiple passages, we induce differentiation of Müller glia into neuron-like cells that express neuronal markers and cell-specific markers. Minor alterations in culture conditions allow us to induce Müller glia into photoreceptor-like cells or retinal ganglion-like cells.

Results : We find induction of Müller glia differentiation leads to increased levels of ER stress and activation of all three pathways of the UPR. Conditional deletion of XBP1 in Müller glia amplifies activation of the UPR. Further, we find more rapid and complete differentiation of Müller glia into neuronal cell types in XBP1-negative cells compared to wild type cells. XBP1 deficiency decreases stemness and the proliferative ability of Müller glia while simultaneously increasing the expression of neuronal markers, number of neurites, and neurite length.

Conclusions : We conclude that the activation of the UPR is a crucial parameter in promoting Müller glia differentiation. Specifically, XBP1 acts to decrease ER stress during Müller glia differentiation and preserves stemness and proliferative ability. Therefore, manipulating ER stress pathways (e.g. by overexpressing or down-regulating XBP1) at the appropriate time window after injury or disease might allow the Müller glia to revert to a progenitor-like state and proliferate to act as an endogenous repair system in the mammalian retina.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.