Photoreceptor Cell Death, Proliferation and Formation of Hybrid Rod/S-Cone Photoreceptors in the Degenerating STK38L Mutant Retina

Abstract

A homozygous mutation in STK38L in dogs impairs the late phase of photoreceptor development, and is followed by photoreceptor cell death (TUNEL) and proliferation (PCNA, PHH3) events that occur independently in different cells between 7–14 weeks of age. During this period, the outer nuclear layer (ONL) cell number is unchanged. The dividing cells are of photoreceptor origin, have rod opsin labeling, and do not label with markers specific for macrophages/microglia (CD18) or Müller cells (glutamine synthetase, PAX6). Nestin labeling is absent from the ONL although it labels the peripheral retina and ciliary marginal zone equally in normals and mutants. Cell proliferation is associated with increased cyclin A1 and LATS1 mRNA expression, but CRX protein expression is unchanged. Coincident with photoreceptor proliferation is a change in the photoreceptor population. Prior to cell death the photoreceptor mosaic is composed of L/M- and S-cones, and rods. After proliferation, both cone types remain, but the majority of rods are now hybrid photoreceptors that express rod opsin and, to a lesser extent, cone S-opsin, and lack NR2E3 expression. The hybrid photoreceptors renew their outer segments diffusely, a characteristic of cones. The results indicate the capacity for terminally differentiated, albeit mutant, photoreceptors to divide with mutations in this novel retinal degeneration gene.

Overlap of abnormal photoreceptor development and progressive degeneration in Leber congenital amaurosis caused byNPHP5mutation

“…Rod photoreceptor origin of these dividing cells has been established by showing that the dividing cells are surrounded by a rim of delocalized rod-opsin immunolabeling, and by excluding any contribution of Mü ller glia, microglia or nestin positive stem cells to the dividing cell population. This process was first identified in the STKL38 (erd) mutant retina, and now extended to RPGR (xlpra2) and PDE6B (rcd1) models (27,28). In NPHP5 mutants, there is a burst of photoreceptor mitosis that coincides with the peak of cell death established by TUNEL labeling.…”

“…This is a feature of other early onset photoreceptor degenerations (27,28). To this end, we used the mitosis-specific marker PHH3 to identify dividing cells in the ONL, and found background labeling in control (< 3.9 6 1 cells/10 6 mm 2 ), but very high levels in the 6 week mutant retina (51.4 6 7 cells/10 6 mm 2 ).…”

“…Immunolabeling was performed by standard IHC methods previously described (27,43). Slides were incubated overnight with one primary antibody, and subsequently incubated with a fluorochrome-labeled secondary antibody (Alexa Fluor 568 or 488, Invitrogen, Carlsbad, CA).…”

Cell Cycle Proteins and Retinal Degeneration: Evidences of New Potential Therapeutic Targets

“…In this case, the neurons duplicate the DNA, but there is no data indicating that they actually can divide or whether DNA synthesis is an attempt to repair the DNA. In the retina, no strong evidence of cell division was documented during the course of retinal degeneration with the exception of the STK38L dog (Berta et al 2011), but this mutation affects the function of a gene involved in the regulation of the cell cycle, and in this case we probably face a development problem. Beside cell cycle regulation, cell cycle proteins may have other targets.…”

“…PCNA (proliferation cell nuclear antigen) which favors DNA polymerase action and coordinates the action of other proteins (review (Moldovan et al 2007)) was also documented to be present in the dopaminergic neurons of rodent models of Parkinson's disease (Hoglinger et al 2007). Interestingly, in dogs affected by mutations in STK38L, a kinase controlling the cell cycle, many photoreceptor cells express cell cycle markers during the early stage of the disease, when the animals are aged from 7 to 14 weeks, then the photoreceptor number decreases dramatically (Berta et al 2011). Indeed, PCNA and phospho-H3 are present in the ONL and not in the microglia (CD18) of the STK38L mutant dogs.…”