Human RPE Stem Cell-Derived RPE Preserves Photoreceptors in the Royal College of Surgeons Rat: Method for Quantifying the Area of Photoreceptor Sparing.

Abstract

Numerous preclinical studies have shown that transplantation of stem cell-derived retinal pigment epithelial cell (RPE) preserves photoreceptor cell anatomy in the dystrophic Royal College of Surgeons (RCS) rat. How rescue is spatially distributed over the eye, relative to the transplantation site, is less clear. To understand spatial variations in transplant efficacy, we have developed a method to measure the spatial distribution of rescued photoreceptor cells. Human RPE Stem Cell-derived RPE (RPESC-RPE) cells were subretinally injected into RCS rat eyes. After tissue recovery and orientating the globe, a series of retinal sections were cut through the injected area. Sections were stained with DAPI (4',6-diamidino-2-phenylindole) and a number of photoreceptor nuclei were counted across the nasal-temporal and superior-inferior axes. These data were used to construct 2D maps of the area of photoreceptor cell saving. Photoreceptor cell preservation was detected in the injected temporal hemisphere and occupied areas greater than 4 mm(2) centered near the injection sites. Rescue was directed toward the central retina and superior and inferior poles, with maximal number of rescued photoreceptor cells proximal to the injection sites. RPESC-RPE transplantation preserves RCS photoreceptor cells. The photoreceptor cell contour maps readily convey the extent of rescue across the eye. The consistent alignment and quantification of results using this method allow the application of other downstream statistical analyses and comparisons to better understand transplantation therapy in the eye.