Dorsal-Ventral Differences in Retinal Structure in the Pigmented Royal College of Surgeons Model of Retinal Degeneration.

Retinitis pigmentosa is a family of inherited retinal degenerations associated with gradual loss of photoreceptors, that ultimately leads to irreversible vision loss. The Royal College of Surgeon's (RCS) rat carries a recessive mutation affecting mer proto-oncogene tyrosine kinase (merTK), that models autosomal recessive disease. The aim of this study was to understand the glial, microglial, and photoreceptor changes that occur in different retinal locations with advancing disease. Pigmented RCS rats (RCS-p+/LAV) and age-matched isogenic control rdy (RCS-rdy +p+/LAV) rats aged postnatal day 18 to 6 months were evaluated for in vivo retinal structure and function using optical coherence tomography and electroretinography. Retinal tissues were assessed using high resolution immunohistochemistry to evaluate changes in photoreceptors, glia and microglia in the dorsal, and ventral retina. Photoreceptor dysfunction and death occurred from 1 month of age. There was a striking difference in loss of photoreceptors between the dorsal and ventral retina, with a greater number of photoreceptors surviving in the dorsal retina, despite being adjacent a layer of photoreceptor debris within the subretinal space. Loss of photoreceptors in the ventral retina was associated with fragmentation of the outer limiting membrane, extension of glial processes into the subretinal space that was accompanied by possible adhesion and migration of mononuclear phagocytes in the subretinal space. Overall, these findings highlight that breakdown of the outer limiting membrane could play an important role in exacerbating photoreceptor loss in the ventral retina. Our results also highlight the value of using the RCS rat to model sectorial retinitis pigmentosa, a disease known to predominantly effect the inferior retina.

Retinal and choroidal TGF-beta in the tree shrew model of myopia: isoform expression, activation and effects on function.

A visually evoked signalling cascade, which begins in the retina, transverses the choroid, and mediates scleral remodelling, is considered to control eye growth. The ubiquitous cytokine TGF-beta has been associated with alterations in ocular growth, where alterations in scleral TGF-beta isoforms mediate the scleral remodelling that results in myopia. However, while the TGF-beta isoforms have been implicated in the scleral change during myopia development, it is unclear whether alterations in retinal and choroidal isoforms constitute part of the retinoscleral cascade. This study characterised the retinal and choroidal TGF-beta isoform profiles and TGF-beta2 activation during different stages of myopia development, as induced by form deprivation, in a mammalian model of eye growth. Using quantitative real-time PCR, the mRNA for all three mammalian isoforms of TGF-beta was detected in tree shrew retina and choroid. Distinct tissue-specific isoform profiles were observed for the retina (TGF-beta1:TGF-beta2:TGF-beta3=20:2085:1) and choroid (TGF-beta1:TGF-beta2:TGF-beta3=16:23:1), which remained constant over the development period under investigation. The active and latent pools of retinal TGF-beta2 were quantified using ELISA with the majority (>94%) of total TGF-beta2 found in the latent form. Unlike previous scleral data showing early and continuous decreases in TGF-beta isoform expression during myopia development, the levels of the three isoforms remained within normal ranges for retinal (TGF-beta1, -14 to +14%; TGF-beta2, -2 to +20%; TGF-beta3, -10 to +26%) and choroidal (TGF-beta1, -19 to +21%; TGF-beta2, -26 to +8%; TGF-beta3, -11 to +28%) tissues during myopia development (induction times of 3h, 7h, 11h, 24h, and 5 days). A 40% decrease in retinal TGF-beta2 activation was observed after 5 days of myopia development, however, there was no functional correlate of altered TGF-beta2 activity, as assessed by the retinal ERG response. Overall, these data highlight the specific nature of TGF-beta isoform expression, which reflects the differences in tissue structure and function. While TGF-beta isoforms are involved in scleral regulation during myopia development in mammals, they do not have a primary role in the retinal and choroidal signals. Thus, the regulation of eye growth via the retinoscleral cascade involves more than one factor, which is likely to be tissue-specific in nature.

Expression of the ETS transcription factor ELF3 in the retinal pigment epithelium.

PURPOSE: The ETS family of transcription factors regulate several critical cellular functions. They have also been implicated in invertebrate ocular development. This work was undertaken to determine whether epithelium-specific ETS transcription factors are expressed in the retinal pigment epithelium and to investigate the possible role of these factors in retinal diseases such as age-related macular degeneration. METHODS: The expression of the epithelial ETS transcription factors ELF5, ESE3, and ELF3 was assessed by RT-PCR in the human RPE cell lines D407 and hTERT-RPE1. The full-length coding sequence of rat Elf3 was isolated with 3' rapid amplification of cDNA ends (RACE) and degenerative primers, and its expression was determined in various rat tissues, by RT-PCR and real-time PCR. A polyclonal ELF3 antibody produced from a C-terminal peptide was used to observe the distribution of the transcription factor within the retina. To assess the possible ELF3 regulation of the TIMP3 promoter, transient transfection assays were performed. Promoter activity was determined with a firefly luciferase reporter gene construct. RESULTS: The epithelium-specific ETS transcription factor ELF3 was expressed in the D407 and hTERT-RPE1 cell lines. Neither ESE3 nor ELF5 was detected in the RPE. The cloning of rat Elf3 produced two splice variants, designated Elf3a (1786 bp) and Elf3b (1855 bp). The larger form, Elf3b, contained a 69-bp insert in the coding sequence, which showed high homology to a similar insert previously identified in murine Elf3. Both splice variants were expressed in rat lung, kidney, liver, and retina, but were absent in heart tissue. Real-time PCR analysis showed the retina to contain high levels of Elf3, which was subsequently localized to the RPE. Elf3 upregulated the TIMP3 promoter, with Elf3a and -3b inducing an approximate sixfold increase in activity. CONCLUSIONS: The ELF3 transcription factor is highly expressed in the RPE and can regulate important ocular genes, such as TIMP3, in vitro. The specific expression of ELF3 in the RPE may reflect an important role for this transcription factor in retinal function. Furthermore, its regulation of TIMP3 may have implications for degenerative retinal diseases, such as age-related macular degeneration.