Abstract

DO.09.08

Müller stem cells – a putative tool to regenerate the retina

G. Astrid Limb

Ocular Biology and Therapeutics, UCL Institute of Ophthalmology, London, United Kingdom

Background and aim

Müller glia has been shown to form a stem cell niche for the neural regeneration of the injured retina in adult zebrafish, newborn chick, and rat. Our studies have identified a population of Müller glia with stem cell characteristics in the adult human eye. We examined the distribution of Müller stem cells in the human retina, and investigated the neurogenic ability of the cells in vitro and in vivo when transplanted into rodent models of retina degeneration. 
Methods
Human Müller stem cells were examined by immunohistochemical studies of the retina and by EGF induced proliferation of the cells in retinal explant cultures. We established Müller stem cell lines and examined the effect of growth and differentiation factors (FGF, DAPT, extracellular matrix) on the neural differentiation in vitro. We investigated the ability of differentiated cells to respond to neural excitatory stimuli in vitro, and to integrate and restore retinal function when transplanted into a NMDA model of ganglion cell depletion. 
Results
Bundles of Müller glia expressing nestin and the neural stem cell markers SOX2, PAX6, NOTCH1 and CHX10, were found in the most anterior, non laminated human neuroretina. In response to EGF Müller stem cells re-enter the cell cycle and express the proliferating antigen Ki67 in situ. Culture of isolated cells with growth and differentiation factors induced their differentiation into retinal neurons, as judged by their expression of photoreceptor (NRL, recoverin, rhodopsin) and ganglion cell (BRN3b, ISL1, HUD) markers. Cells expressing ganglion cell markers responded to nicotinic stimulation (characteristic of RGC) in vitro with an increase in cytosolic calcium levels. When transplanted into the retina of RGC depleted rats, differentiated cells expressing RGC phenotype migrated into the RGC layer, acquired neural morphology, and induced a partial restoration of the negative scotopic threshold response of the ERG.
Conclusions
The data suggest that Müller stem cells from the adult human retina have the ability to differentiate into retinal neurons. Müller stem cell derived neurons with ganglion cell phenotype have the potential to be used in cell based therapies to treat degenerative diseases affecting ganglion cells, such as glaucoma.

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