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Yamit Cohen, Pablo Blinder, Maria Idelson, Benjamin reubinoff, shalev Itzkovitz, Ruth Ashery-Padan; Pax6 role in the regulation of retinal pigmented epithelium maturation. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6055.
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© ARVO (1962-2015); The Authors (2016-present)
Haploinsufficiency of Pax6 was long shown to cause aniridia and was reported to involve neovascularization of the cornea. However, Pax6 involvement in vascularization remained elusive and lack of imaging techniques for the choroid vasculature limited further inquiry regarding the effect on this tissue. This research aims to determine the molecular mechanism in which Pax6 regulates retinal pigmented epithelium (RPE) maturation and choroid development using somatic mutagenesis in mouse models and RPE cells derived from human embryonic stem cells (hES-RPE).
To this purpose, conditional mutations are induced in mice and gain of function analysis is conducted using sub-retinal injections followed by electroporation. Quantitative expression levels are measured using single molecule FISH and the choroidal phenotype is analyzed using a novel perfusion technique. Further investigation of the molecular mechanism involved in this regulation is performed using hES-RPE cells and includes chip-seq, RNA-seq and knockdown using viral infections.
Pax6 deletion in the specified RPE resulted in a phenotype of aniridia and among the changes in gene expression we observed an up-regulation in Sox9 expression level, a key transcription factor in organogenesis which is related to late stages of RPE differentiation. Pax6 and Sox9 expression patterns were determined in course of RPE differentiation in wild type mice and mice with Pax6 specific ablation from the RPE. Quantitative expression analysis illustrated opposite correlation and gain of function analysis confirmed this observation since miss-expression of Pax6 resulted in inhibition of Sox9 expression.
This study is the first to document Pax6 role in timing RPE differentiation through its regulatory relations with Sox9. Future efforts are aimed to elucidate the molecular mechanism of this regulation and examine a possible effect on the choroid vasculature.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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