Abstract
Purpose :
Individuals with Oculocutaneous albinism have low ocular pigmentation and decreased best-corrected visual acuity due to defects in fovea development. Since fovea development starts prenatally and continues to develop after birth, it provides the possibility that childhood intervention to improve pigmentation in eye tissues might help rescue vision defects. In adults, benefits may be limited to symptoms associated with glare and photosensitivity, but children may realize more dramatic effects if fovea development can even be partially restored. The purpose of our study is to develop disease in a dish model for Oculocutaneous albinism type 2 (OCA2) and identify drugs that can improve pigmentation.
Methods :
Fibroblast cells from three unrelated genotyped OCA2 individuals were reprogrammed to induced pluripotent stem cell (OCA2-iPSC) lines using Cyto-tune II™. iPS cells were characterized using immunostaining of pluripotency markers, embryoid body formation assay and in vivo teratoma formation assay. The OCA2-iPS cells were differentiated to retinal pigment epithelium (OCA2-RPE) cells using a developmentally guided differentiation protocol and characterized at the molecular and functional level by immunostaining, trans-epithelial resistance (TER) measurements, phagocytosis assay, and electron microscopy analysis.
Results :
The OCA2-iPS cells expressed OCT4, SOX2, TRA-1-81 and TRA-1-60, formed embryoid bodies that can be differentiated to multiple cell types and when implanted in vivo resulted in teratomas. OCA2-RPE were cultured on semi-permeable membranes for 8 weeks to obtain a functionally mature and polarized monolayer tissue that were similar to RPE derived from unaffected control individual and to human fetal (hf) RPE (Control-RPE) for expression and localization of cell-cell junctional markers ZO1, β-CATENNIN, and cell surface proteins NaK+ ATPase and SCRIBBLE. Functionally OCA2-RPE displayed TER and photoreceptor phagocytosis comparable to control RPE. Using electron microscopy we observed significantly reduced and degenerating melanosomes in OCA2-RPE as compared to Control-RPE.
Conclusions :
We show that a fully-functional RPE can be derived from OCA2 patient fibroblast cells that faithfully recapitulate the patient phenotype of pigmentation defects in a cell culture system. The OCA2 patient derived RPE cells will be further used for pigmentation rescue studies.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.