The retinal pigment epithelium undergoes massive apoptosis during early differentiation and pigmentation of the optic cup

Purpose: The aim of our work was to study apoptosis during the development of the retinal pigment epithelium (RPE) in mice between embryonic day (E) 10.5 and E12.5 and to examine a possible link between apoptosis and pigmentation. Methods: We collected mouse embryos at E10.5, E11.5, and E12.5 and labeled apoptotic cells in 5-µm paraffin sections, using the terminal deoxynucleotidyl transferase dUTP nick end labeling technique. We counted the total number of cells and the number of apoptotic cells in the early developing RPE and calculated the percentage of apoptosis at each stage. Results: In the C57BL/6J mouse, 17% of the RPE cells were apoptotic at E10.5 compared to 0.9% at E12.5. At E11.5, three-quarters of the RPE cells began to pigment, and apoptotic cells were located mostly in the nonpigmented part. In contrast, in the BALB/c mouse (tyrosinase-deficient) and pJ mouse (carrying mutations in the p gene) hypopigmented strains, the RPE contained significantly fewer apoptotic cells (7.5% and 10.1%, respectively) at E10.5 than controls. Subsequently at E11.5 and E12.5, the two hypopigmented strains displayed different apoptotic patterns; the BALB/c RPE had a similar percentage of apoptotic cells to controls (1.5% and 1.1%, respectively, for BALB/c versus 3.0% and 0.9%, respectively, for C57BL/6J), whereas the pJ RPE contained significantly more apoptosis (7.5% and 3.5%, respectively). Overall we observed differences in the evolution of the relative total number of RPE cells between the three strains. Conclusions: Apoptosis is a main event during the first stages of normal RPE development, indicating an essential role during RPE differentiation. Moreover, the early apoptotic pattern and possibly the whole early development of the RPE is different between hypopigmented and pigmented strains, as well as between BALB/c and pJ mice. This suggests the existence of regulatory and developmental differences with a more complex origin than just differing pigmentation levels.