1Molecular Therapy, Institute of Ophthalmology, London, United Kingdom
2Developmental Biology Unit, Institute of Child Health, London, United Kingdom
3Moorfields Eye Hospital, London, United Kingdom
Commerical Relationships: A. MacNeil, None; R.A. Pearson, None; R.E. MacLaren, None; P.K. Buch, None; A.J. Smith, None; J.C. Sowden, None; R.R. Ali, None.
Support: Medical Research Council UK, Royal Blind Asylum and School, The National Institute for the War Blinded, The Trustees of Moorfields Eye Hospital.
Purpose:The identification of stem cell populations with retinal potential is of considerable interest because of their hypothesized use as donor cells to replace the photoreceptors lost in disease. The porcine eye provides an excellent model, since its morphology is similar to the human eye and the anatomical regions can be accurately dissected. We sought to determine the potential for harvesting neural stem cells from different regions of porcine eye, particularly those most readily accessible during routine eye surgery. We wished to evaluate the phenotypic properties cells from each region under proliferation or differentiation conditions.
Methods:Porcine samples from the anterior neural retina, pars plana, ciliary body and iris were dissected and dissociated. Cells were re-suspended to allow the formation of clonally derived neurospheres. Cultures from each region were assessed for neurosphere number and size. Immunostaining was used to confirm the presence of retinal stem cell markers and assess the differentiation potential of cells within neurospheres and monolayer cultures. The proportion of proliferating cells in monolayer cultures from each region was determined by pulse labelling with BrDU.
Results:We demonstrate the presence of stem/progenitor cells in the pig ciliary epithelium, pars plana and iris, regions that are readily accessible during routine eye surgery. In the presence of growth factors, these stem/progenitor cells form neurospheres that contain cells expressing the retinal stem/progenitor markers Pax6 and Sox2. Using an adherent monolayer culture system, these cells could be readily expanded to increase their number more than a million-fold and maintain expression of Sox2 in a high proportion of cells. When grown on the substrate laminin, in the presence of serum, spheres and monolayer-derived cells differentiated into neurons, although retinal cell type specific markers were not detected.
Conclusions:These results suggest that cells derived from the iris, pars plana and ciliary body have stem/progenitor properties and neurogenic potential, thereby providing potential novel sources of donor cells for transplantation studies.
Keywords: iris retina ciliary body
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