Abstract
Purpose: :
Changes in RPE morphology occur during and after retinal degeneration. We hypothesized that similar shared changes in RPE morphology occur in remodeling of divergent retinal degenerations. To test this we compared the morphology of RPE cells from c57BL/6J and rd10 mouse eyes with normal and age related macular degeneration (AMD) human donor eyes.
Methods: :
Mouse eyes were fixed for 10 min in formalin, retina removed, RPE flatmounted, and stained with anti-ZO-1 and propidium iodide. Fundus photos of human donor eyes were taken and eyes fixed for 4 h in formalin and the choroid/RPE layer taken. The latter was stained with AF635-phalloidin and propidium iodide. Confocal microscopy was used to image the flatmounts. Photoshop CS2 or Autopano Pro v2 was used to photomerge the images. Analysis of cell shape, size, number of neighbors etc., was performed using Cell Profiler.
Results: :
Between normal mouse and human, morphology of the RPE sheet is similar with a regular hexagonal array of cells of uniform size. Towards the far periphery, this regular array turns into a soft network. At the macula, human RPE has a higher density of cells than the mid periphery, whereas in mice the RPE sheet has nearly the same density in posterior and mid periphery. Many changes occur in the RPE sheet with disease progression. Affected mice and humans show great disruptions in the central region of the RPE sheet, lesser disruptions in mid-periphery, and almost normal morphology at the far periphery. These disruptions include large multinucleate cells, holes in the RPE sheet, and rosette structures where RPE cells have grown to fill in space where cells have died. In disease, we found bigger cell size, increased variability in cell size, disrupted cell forms, and increased variability in number of neighboring cells. Some changes occurring in disease are unique to either the rd10 model or AMD donors. These include a region of compressed cells in mid-periphery that develops in rd10 mice early in disease progression. Disease patterns in human eyes vary greatly among individuals, but in mice the pattern is very consistent. There are RPE changes in humans associated with drusen that we do not detect in our mouse model.
Conclusions: :
Although mouse rd10 does not show all features of AMD in humans, there are many morphology changes that are similar and likely use the same molecular mechanisms, e.g., filling in of space that was occupied by a cell that has died results in the formation of a rosette. rd10 provides insights into early changes that occur in the RPE concurrently with retinal degeneration that cannot be captured in humans.
Keywords: retinal pigment epithelium • comparative anatomy • age-related macular degeneration