Outcome and risk of ocular complications of managing children with chronic anterior uveitis with topical rimexolone 1.

PURPOSE: To investigate the efficacy and safety of 1% rimexolone ophthalmic suspension in children with chronic anterior uveitis under real-life conditions in a tertiary center. METHODS: This is a retrospective longitudinal study. Medical records were analyzed at baseline, 1, 3, 6 and 12 months before and after switching to rimexolone for best-corrected visual acuity (BCVA), oral steroid use, number of flares, IOP and anti-glaucoma management. RESULTS: Twenty-four patients (41 eyes) diagnosed with either anterior uveitis (n = 25, 60.0%) or panuveitis (n = 16, 40%) were enrolled. The mean age was 10.5 years (4-16 years). The number of patients requiring oral prednisolone reduced from 8 patients (32.0%) at baseline to 3 patients (20.0%) at 12 months (P < 0.001). Following baseline, the median number of uveitis flares reduced from 2.0 (inter-quartile range (IQR) 1.0-2.75) to 1.0 (IQR 0.0-1.0) compared to the 12 months before baseline (P < 0.001). The mean IOP reduced from baseline (22.0 ± 7.3 mmHg) to 1 month (18.8 ± 8.7 mmHg, P = 0.01) and remained stable up to 12 months (15.9 ± 5.0 mmHg, P < 0.001). Average BCVA, dose of oral prednisolone and anti-glaucoma treatments did not change compared to the baseline. The development for IOP ≥ 30 mmHg was associated with a known corticosteroid response [odds ratio (OR) 6.8, P = 0.003] and a dose > 7.5 mg/day oral prednisolone (OR 4.4, P = 0.033). CONCLUSIONS: Rimexolone 1% ophthalmic suspension is an effective and safe topical steroid for pediatric anterior uveitis.

Expression of Toll-like receptors in human retinal and choroidal vascular endothelial cells.

Toll-like receptors (TLRs) are a family of proteins that initiate the innate immune response in reaction to invading microbes. Studies confirm the expression of TLRs in a variety of ocular tissues and cells, and it has also been suggested that selected TLRs may be associated with geographic atrophy and neovascularisation in age-related macular degeneration, diabetic retinopathy and other vascular and inflammatory diseases of the ocular posterior segment. However, TLR expression and localisation in the retinal and choroidal vasculature has not been defined. A better understanding of differential TLR expression in the choroid and retina, particularly in endothelial cells would improve our knowledge of vascular and inflammatory diseases in the posterior segment of the eye. In this study the gene (mRNA) expression of TLRs 1-10 was investigated using RT-PCR and comparative qPCR and the protein expression and localisation of selected TLRs (3, 4, 6 and 9) were examined using western blotting, flow cytometry and immunofluorescent staining. PCR showed gene expression of TLR1-6 and 9 in human choroidal endothelial cells (hCEC) and TLR2-6, 9 and 10 in human retinal endothelial cells (hREC). Western blotting detected TLR3, 4 and 9 proteins in both hCEC and hREC with higher levels in hCEC, whilst TLR6 protein was not detectable in either endothelial cell type. Flow cytometry detected all four TLRs (3, 4, 6 and 9) on the cell surface and intracellularly, TLR6 expression was detectable but low. The expression and localisation of TLR3, 4 and 9 were confirmed by immunofluorescent staining in endothelial cells and whole tissue sections and their functionality tested by expression of IL-6 (ELISA) in response to stimulation with specific TLR ligands. This study has, for the first time, identified the differential expression and localisation of TLRs in intraocular endothelial cells. This profiling will help inform our understanding of different retinal and choroidal vascular diseases, as well as the development of future treatments for intraocular vascular diseases.

A glycine site-specific NMDA receptor antagonist protects retina ganglion cells from ischemic injury by modulating apoptotic cascades.

Glutamate neurotoxicity is one of the causative factors leading to neural degeneration including retina. Inhibition of NMDA receptors has been shown neuroprotective effects. However, specifically inhibition of glycine subunit in NMDA receptors and its effects on retina neural protection has not been tested. In this study, using a glycine site-specific NMDA receptor antagonist, we investigated its neuroprotective effects on rat retinal ganglion cells (RGCs) from a transient ischemic injury and its possible underlying mechanisms. Following an ischemia/reperfusion injury the structural damages of rat retinas were assessed by an immunofluorescence method and the apoptosis of retinal neural cells was evaluated by using a terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) method. The survived RGCs were labeled by retrograde manner and counted on whole-mounted retinas. In the presence of glycine site-specific NMDA receptor antagonist, the thickness of retina was sustained, especially in the inner nuclear layers compared with mock controls. While a significantly higher numbers of TUNEL-positive apoptotic cells and fewer of RGCs were observed in the retina without the glycine antagonist, indicating its strong protective roles. Some apoptotic factors such as Bax, Bcl-2, CAMK II, COX1, COX4, Caspase-3, and GRIN1 gene have been tested from retinal samples with or without the glycine antagonist. A significantly lower of expressions of Bax, CAMK II, COX1, COX4, Caspase-3, and GRIN1 have been shown in the retinas with the antagonist. Bcl-2/Bax ratio was significantly higher with the antagonist, suggested that the glycine site-specific NMDA receptor antagonist protecting RGC death might through inhibition of apoptotic signaling.

Suitability of endogenous reference genes for gene expression studies with human intraocular endothelial cells.

BACKGROUND: The use of quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) has become widely applied as a method to measure transcript abundance. In order to be reflective of biological processes during health and disease this method is dependent on normalisation of data against stable endogenous controls. However, these genes can vary in their stability in different cell types. The importance of reference gene validation for a particular cell type is now well recognised and is an important step in any gene expression study. RESULTS: Cultured primary human choroidal and retinal endothelial cells were treated with the immunostimulant polyinosinic: polycytidylic acid or untreated. qRT-PCR was used to quantify the expression levels of 10 commonly used endogenous control genes, TBP, HPRT1, GAPDH, GUSB, PPIA, RPLP0, B2M, 18S rRNA, PGK1 and ACTB. Three different mathematical algorithms, GeNorm, NormFinder, and BestKeeper were used to analyse gene stability to give the most representative validation. In choroidal endothelial cells the most stable genes were ranked as HPRT1 and GUSB by GeNorm and NormFinder and HPRT1 and PPIA by BestKeeper. In retinal endothelial cells the most stable genes ranked were TBP and PGK1 by GeNorm and NormFinder and HPRT1 by BestKeeper. The least stable gene for both cell types was 18S with all 3 algorithms. CONCLUSIONS: We have identified the most stable endogenous control genes in intraocular endothelial cells. It is suggested future qRT-PCR studies using these cells would benefit from adopting the genes identified in this study as the most appropriate endogenous control genes.

Efficacy of tetrandrine on lowering intraocular pressure in animal model with ocular hypertension.

PURPOSE: To compare the efficacy of topical tetrandrine (an alkaloid isolated from the Chinese medicinal herb Radix Stephania tetrandrae S) and timolol 0.5% ophthalmic solution on lowering the intraocular pressure (IOP) in ocular normotensive and hypertensive rats. METHODS: The experiment was designed as 2 parts. In the first part, normal male Sprague-Dawley rats were divided into 4 groups followed by topical administration once of 0.1%, 0.2%, 0.3% tetrandrine, and 0.5% timolol in the right eye, 0.9% saline was administered once at the opposite eye as control. In the second part, the ocular hypertension model was induced unilaterally in the rats by a diode laser treatment. Twice daily applications of the above drugs were delivered to hypertensive eyes. The control group was administered 0.9% saline. The TonoPen XL tonometer was used to determine the IOP levels. RESULTS: No lowering effect on IOP was detected in the normotensive rats treated with tetrandrine whereas timolol significantly reduced IOP in normotensive eyes. Both tetrandrine and timolol significantly reduced the IOP levels in the hypertensive eyes compared with the levels in the saline-treated groups. All concentrations of tetrandrine used in this study showed significant reduction of IOP in the laser-induced hypertensive eyes. Tetrandrine 0.3% had a similar efficacy as 0.5% timolol in reducing elevated IOP in ocular hypertensive eyes. CONCLUSIONS: This study provides evidence that tetrandrine has a major effect on lowering IOP levels in the ocular hypertension rat model. The functional mechanisms of tetrandrine require further investigation.