Characterization of rat retinal responses to a conventional retinal laser and a novel short pulse duration laser.

Retinal laser photocoagulation is the process whereby laser energy is directed onto the retina and converted to heat within the melanosomes of the retinal pigment epithelium (RPE). Its principle clinical purposes are to reduce macula oedema, inhibit angiogenesis, and create chorioretinal scars. The exact biological mechanisms by which destruction of RPE/photoreceptors leads to a therapeutic resolution of oedema remain to be elucidated. However, it is well documented that laser photocoagulation causes thermal injury to the overlying photoreceptors. The desire to produce a clinically effective and safer retinal laser motivated the development of a novel 3- nanosecond pulse duration laser, Retinal Regeneration Therapy (2RT; Ellex Pty. Ltd., Adelaide), which delivers approximately 0.2% of the energy per pulse compared to a conventional continuous wave photocoagulation laser. Recent laboratory and clinical research has demonstrated that the 2RT laser caused remarkably little destructive injury to the overlying photoreceptor layer, but still effectively ablated RPE cells. The retinal glial response to this laser had not been characterized. Furthermore, previously published research indicated that retinal lasers can stimulate endogenous production of neuronal survival factors, such as heat shock proteins and trophic factors, but the effect of 2RT on retinal trophic factors was unknown. Hence, this thesis was motivated by the notion that 2RT may be capable of inducing an endogenous neuroprotective response in the retina without causing collateral retinal damage. The aims of Part I were: (1) to study the RPE/retinal damage profiles and neuronal effects of a CW laser and the short pulse duration 2RT laser; and (2) to characterize in detail the glial and inflammatory responses to the above retinal lasers. The aims of Part II to were: (1) to examine whether multiple preconditioning pathways are activated by CW and 2RT lasers; (2) to examine whether preconditioning with either laser is neuroprotective in a rat model of retinal ganglion cell degeneration caused by calibrated optic nerve crush. Pigmented Dark Agouti rats were used and treated with either a conventional, thermal, continuous wave (CW; 532nm, 100ms pulse duration) or a short-pulse (2RT; 532nm, Q-switched, 3ns pulse) laser. Settings were at visible threshold for the CW laser (12.7J/cm2/pulse) and at supra- and subvisible threshold for the 2RT laser (“High”, 2RT-H, 163mJ/cm2/pulse; “Low”, 2RT-L, 109mJ/cm2/pulse). At various time points after lasering, rats were killed and analysed for histology, immunohistochemistry, RT-PCR and Western immunoblotting. In Part II, groups of rats were killed at 6 hours, 1 day and 1 week following the laser treatment. Samples were taken for immunohistochemistry, RT-PCR and Western immunoblotting. Rats were randomly assigned to one of three treatment groups: sham, CW or 2RT. For the CW and 2RT groups, 20 laser spots were applied to the mid-central retina of the right eye, while left eyes remained untreated. At 1 day or 7 days after lasering, rats in all three groups received ON crush in the right eye. The left ON remained intact. Rats were allowed to recover and killed 1 week or 2 weeks after optic nerve crush for quantification of RGCs in wholemounts. The results of chapters 2 & 3, showed both lasers caused focal loss of RPE cells with no destruction of Bruch’s Membrane; RPE cells were present at lesion sites again within 7 days of treatments. There were no obvious effects to horizontal, amacrine or ganglion cells, as defined by immunolabeling, but an activation of PKCα within bipolar cells was noted. There was little discernible damage to any cells other than the RPE with the 2RT-L treatment. The CW laser caused outer retinal lesions that were associated with photoreceptor death, astrocyte and Müller cell activation, and infiltration of macrophages and neutrophils. Furthermore, inflammatory cytokines, heat shock proteins, endogenous trophic factors, and matrix metalloproteinases were induced. In comparison, all of these changes were drastically attenuated when the 2RT laser was used, particularly at the sub-threshold setting. In chapter 4, the results showed that both the CW and 2RT lasers induced local glial cell activation and induced localized upregulations of a number of well-documented (CNTF, FGF-2 Hsp27, pAKT) or putative (cFOS, ATF-3, IL-6) RGC survival factors. However, neither laser caused sustained increases in other factors associated with neuronal preconditioning, such as BDNF, Hsp70, IGF-1, bcl-2, and nitric oxide synthase. As regards neuroprotection, analysis of the data revealed that ON crush resulted in the loss of approximately 70% of Brn3a-labelled RGCs after 1 week. In conclusion the CW laser photocoagulation caused death of RPE cells with associated damage to the outer retina but negligible impact on the inner retina. The 2RT laser, at the lower setting, was able to selectively kill RPE cells without causing collateral damage to photoreceptors. It was demonstrated that the CW laser produced astrocyte and Müller cell activation, and infiltration of macrophages and neutrophils. Furthermore, inflammatory cytokines, heat shock proteins, endogenous trophic factors, and matrix metalloproteinases were induced. Similar changes were observed when the 2RT laser was used, but the effects were less striking. Furthermore, both CW and 2RT lasers activated multiple preconditioning pathways but the stimulation of these pathways was insufficient to augment RGC survival after optic nerve crush.Thesis (M.Phil.) -- University of Adelaide, School of Medicine, 201

Cervical Carcinoma Manifesting as Progressive Bilateral Visual Loss

We report a patient with bilateral choroidal metastasis from disseminated cervical squamous cell carcinoma. A 52-year-old woman presented with progressive bilateral visual loss due to choroidal masses in both eyes. The fundus examination revealed posterior serous retinal detachment in both eyes associated with creamy choroidal lesions. A thorough systemic work-up revealed choroidal metastasis from a squamous cell carcinoma of the cervix. This case highlights the importance of a thorough systemic evaluation in patients with choroidal tumours