Preoperative tranilast as adjunctive therapy to primary pterygium surgery with a 1-year follow-up.

PURPOSE: To determine the efficacy of tranilast as an adjunctive therapy in conjunctival autograft. METHODS: Twenty-nine patients were randomly allocated to the Tranilast Group (n=15) or the Control Group (n=14). The Tranilast Group received a subconjunctival injection of 0.5% tranilast 30 days prior to surgery. Conjunctival autograft was performed in both groups using fibrin sealant and 0.02% subconjunctival mitomycin C at the end of the surgery. After the resection of the pterygium, immunohistochemistry was performed with 100 cells to identify epithelial cells positive for transforming growth factor-ß (TGF-ß). Subjective symptoms were evaluated using a 5-point scale, and the recurrence rate was assessed. RESULTS: Both groups showed improvements in their symptoms and similar clinical results. Compared with the Control Group, the Tranilast Group failed to show a decreased recurrence rate (p=0.59). However, the number of epithelial cells expressing TGF-ß was lower in the Tranilast Group (5 cells; 95% CI: 2.56-13.15; Control Group, 16 cells, 95% CI: 11.53-24.76; p=0.01). Minimal but reversible complications, including glaucoma secondary to corticosteroids and granuloma, occurred during the study. CONCLUSION: Tranilast was effective in decreasing the number of pterygium epithelial cells expressing TGF-ß.

Preoperative tranilast as adjunctive therapy to primary pterygium surgery with a 1-year follow-up / Terapia auxiliar com o tranilast pré-operatório na cirurgia do pterígio primário com um ano de seguimento

Purpose: To determine the efficacy of tranilast as an adjunctive therapy in conjunctival autograft. Methods: Twenty-nine patients were randomly allocated to the Tranilast Group (n=15) or the Control Group (n=14). The Tranilast Group received a subconjunctival injection of 0.5% tranilast 30 days prior to surgery. Conjunctival autograft was performed in both groups using fibrin sealant and 0.02% subconjunctival mitomycin C at the end of the surgery. After the resection of the pterygium, immunohistochemistry was performed with 100 cells to identify epithelial cells positive for transforming growth factor-β (TGF-β). Subjective symptoms were evaluated using a 5-point scale, and the recurrence rate was assessed. Results: Both groups showed improvements in their symptoms and similar clinical results. Compared with the Control Group, the Tranilast Group failed to show a decreased recurrence rate (p=0.59). However, the number of epithelial cells expressing TGF-β was lower in the Tranilast Group (5 cells; 95% CI: 2.56-13.15; Control Group, 16 cells, 95% CI: 11.53-24.76; p=0.01). Minimal but reversible complications, including glaucoma secondary to corticosteroids and granuloma, occurred during the study. Conclusion: Tranilast was effective in decreasing the number of pterygium epithelial cells expressing TGF-β. .

Objetivo: Determinar a eficácia do tranilast, como terapia auxiliar no transplante autólogo de conjuntiva. Métodos: Vinte e nove pacientes foram randomizados em dois grupos: Grupo Tratado (15) e Grupo Controle (14). Trinta dias antes da cirurgia, o Grupo Tratado recebeu uma injeção subconjuntival de tranilast a 0,5%. O transplante autólogo de conjuntiva foi realizado em ambos os grupos, usando-se a cola de fibrina e a mitomicina 0,02% subconjuntival, ao final da cirurgia. Cada paciente foi examinado por 12 meses de acompanhamento. A imuno-histoquímica foi realizada, mediante um total de 100 células, a fim de que se contassem as células epiteliais positivas, para o fator de crescimento transformador beta (TGF-β), após a cirurgia do pterígio. Os sintomas subjetivos foram avaliados usando-se uma escala de cinco pontos, e a taxa de recorrência foi avaliada. Resultados: Os 2 grupos apresentaram melhora dos sintomas e com resultados clínicos similares. Quando comparado com o Grupo Controle, o Grupo Tratado falhou em mostrar uma diminuição da taxa de recorrência (p=0,59). Entretanto o número de células epiteliais expressando o TGF-β foi menor no Grupo Tratado (5 células; 95% CI=2,56-13,15; Grupo Controle, 16 células; 95% CI: 11,53-24,76, p=0,01). Complicações mínimas, mas reversíveis, ocorreram durante o estudo, incluindo glaucoma secundário ao uso de corticoide e granuloma. Conclusão: O tranilast foi efetivo em diminuir o número células epiteliais do pterígio expressando o TGF-β. .

Biochemical analysis and decomposition products of indocyanine green in relation to solvents, dye concentrations and laser exposure.

PURPOSE: To investigate pH, ions, osmolarity and precipitation of indocyanine green (ICG), as well as the profile of ICG decomposition products (DPs) after laser exposure and the interaction with quenchers. METHODS: ICG was diluted in water, 5% glucose (GL) or balanced salt solution (BSS) to achieve concentrations of 2.5, 1, 0.25 and 0.1 mg/ml. Osmolarity, pH and precipitation were analyzed immediately and after 24 h. Precipitation analyses were done with a scanning electron microscope. Anion and iodate analyses of ICG and infracyanine green (IfCG) were performed by capillary zone electrophoresis. With regard to DPs, 0.5 mg/ml of ICG was assessed with high-performance liquid chromatography (HPLC) after 810-nm laser irradiation. DP profiles were evaluated with ICG dilution in quenchers (Trolox, histidine and DABCO) in 3 concentrations (0.1, 1 and 10 M). RESULTS: BSS promoted iso-osmotic ICG solutions of 208 mOsm (147-266) compared to GL with 177 mOsm. BSS solutions had a higher physiological pH of 7.2 compared with the GL one of 6.55. ICG precipitated more when diluted with BSS (5.95 mg); in contrast, GL showed less precipitate (3.6 mg). IfCG has no iodine derivates and other ICGs have an average 4.6% of iodate derivates. From HPLC analysis, 5 DPs were observed. The rate of DPs was higher when BSS was used (p < 0.05). Five DPs have been generated with ICG, and they may be altered with the quenchers DABCO, histidine and Trolox. CONCLUSIONS: BSS dilution induces more precipitation and DPs. ICG dilution in any solvent induces DPs. Quencher use reduces the amount of toxic DPs.

Corneal absorption of a new riboflavin-nanostructured system for transepithelial collagen cross-linking.

Corneal collagen cross-linking (CXL) has been described as a promising therapy for keratoconus. According to standard CXL protocol, epithelium should be debrided before treatment to allow penetration of riboflavin into the corneal stroma. However, removal of the epithelium can increase procedure risks. In this study we aim to evaluate stromal penetration of a biocompatible riboflavin-based nanoemulsion system (riboflavin-5-phosphate and riboflavin-base) in rabbit corneas with intact epithelium. Two riboflavin nanoemulsions were developed. Transmittance and absorption coefficient were measured on corneas with intact epithelia after 30, 60, 120, 180, and 240 minutes following exposure to either the nanoemulsions or standard 0.1% or 1% riboflavin-dextran solutions. For the nanoemulsions, the epithelium was removed after measurements to assure that the riboflavin had passed through the hydrophobic epithelium and retained within the stroma. Results were compared to de-epithelialized corneas exposed to 0.1% riboflavin solution and to the same riboflavin nanoemulsions for 30 minutes (standard protocol). Mean transmittance and absorption measured in epithelialized corneas receiving the standard 0.1% riboflavin solution did not reach the levels found on the debrided corneas using the standard technique. Neither increasing the time of exposure nor the concentration of the riboflavin solution from 0.1% to 1% improved riboflavin penetration through the epithelium. When using riboflavin-5-phosphate nanoemulsion for 240 minutes, we found no difference between the mean absorption coefficients to the standard cross-linking protocol (p = 0.54). Riboflavin nanoemulsion was able to penetrate the corneal epithelium, achieving, after 240 minutes, greater stromal concentration when compared to debrided corneas with the standard protocol (p = 0.002). The riboflavin-5-phosphate nanoemulsion diffused better into the stroma than the riboflavin-base nanoemulsion.

Effects of light exposure, pH, osmolarity, and solvent on the retinal pigment epithelial toxicity of vital dyes.

PURPOSE: To investigate the in vitro effect of pH, osmolarity, solvent, and light interaction on currently used and novel dyes to minimize dye-related retinal toxicity. DESIGN: Laboratory investigation. METHODS: Retinal pigment epithelium (RPE) human cells (ARPE-19) were exposed for 10 minutes to different pH solutions (4, 5, 6, 7, 7.5, 8, and 9) and glucose solutions (2.5%, 5.0%, 10%, 20%, 40%, and 50%) with osmolarity from 142 to 2530 mOsm, with and without 0.5 mg/mL trypan blue. R28 cells were also incubated with glucose (150, 310, and 1000 mOsm) and mannitol used as an osmotic control agent in both experiments. Dye-light interaction was assessed by incubating ARPE-19 for 10 minutes with trypan blue, brilliant blue, bromophenol blue, fast green, light green, or indigo carmine (0.05 mg/mL diluted in balanced saline solution) in the presence of high-brightness xenon and mercury vapor light sources. RESULTS: Solutions with nonphysiologic pH, below 7 and above 7.5, proved to be remarkably toxic to RPE cells with or without trypan blue. Also, all glucose solutions were deleterious to RPE (P < .001) even in iso-osmolar range. No harmful effect was found with mannitol solutions. Among the dyes tested, only light green and fast green were toxic to ARPE-19 (P < .001). Light exposure did not increase RPE toxicity either with xenon light or mercury vapor lamp. CONCLUSIONS: Solutions containing glucose as a dye solvent or nonphysiologic pH should be used with care in surgical situations where the RPE is exposed. Light exposure under present assay conditions did not increase the RPE toxicity.

Use of lutein and zeaxanthin alone or combined with Brilliant Blue to identify intraocular structures intraoperatively.

PURPOSE: To determine whether a natural dye solution based on lutein and zeaxanthin alone or combined with Brilliant Blue stains and facilitates peeling of intraocular membranes in human eyes. METHODS: In this study of 60 cadaveric eyes, open-sky vitrectomy including posterior hyaloid detachment was performed. Different lutein and zeaxanthin concentrations (0.01-20%) were tested alone or combined with different Brilliant Blue concentrations (0.0125-0.025%) in the corneal endothelium, corneal epithelium, anterior and posterior capsule, vitreous cavity through the macula including the posterior hyaloid, and internal limiting membrane. The various dye solutions were in contact with the intraocular membranes for <1 minute and then were removed by mechanical aspiration or membrane peeling initiated and completed with intraocular forceps. The specimens were examined by light and electron transmission microscopy. RESULTS: Contact between lutein and zeaxanthin and the retinal, lens, and vitreous surface resulted in orange and greenish staining of the intraocular membranes, which facilitated surgical steps in all eyes. Lutein and zeaxanthin alone was useful for vitreous identification and lutein and zeaxanthin combined with Brilliant Blue had strong affinity for internal limiting membrane and anterior capsule. Light microscopy confirmed internal limiting membrane removal in all eyes tested. No dye solutions remained in the eyes after the membrane removal. CONCLUSION: A natural dye solution based on lutein and zeaxanthin alone or combined with Brilliant Blue efficiently stained the anterior capsule, vitreous, and internal limiting membrane in human cadaveric eyes and may be a useful tool for vitreoretinal or cataract surgery.

Vital dyes and light sources for chromovitrectomy: comparative assessment of osmolarity, pH, and spectrophotometry.

PURPOSE: To investigate the in vitro pH, osmolarity, spectral, and photostability properties of nine vital dyes for vitreoretinal surgery. METHODS: Nine dyes-indocyanine green (ICG), trypan blue (TB), brilliant blue (BriB), bromophenol blue (BroB), Congo red (CR), light green (LG), fast green (FG), indigo carmine (IC) and Evans blue (EB)-diluted in three solvents (saline solution, glucose 5%, and water) were tested for osmolarity and pH. Spectrophotometry was used to determine absorbance properties of 27 solutions. Irradiance emission spectra of seven endoillumination light sources and fiber-optics were compared with dye absorbance curves. RESULTS: Dye osmolarity in saline solution and glucose 5% varied widely (257-385 mOsm) and was lower (0-54 mOsm) when dyes were dissolved in water. Dyes diluted in three solvents showed pH values varying from 2.6 to 9.85. ICG, LG, TB, BroB, CR, and IC demonstrated different absorbances, depending on the solvent. BriB and FG showed similar absorbance curves with different solvents. Spectrophometric analysis showed that all dyes except ICG had remarkable spectral overlap with the light sources. Among endoillumination fiber-optics, overlap was greatest with dual-output illumination with an integrated laser pathway and least with a mercury vapor lamp. CONCLUSIONS: Vital dyes showed variable osmolarity and pH, which also depended on the solvent used. Interaction of light from endoillumination source and vital dye may increase or decrease the risk for toxicity, making appropriate selection of both a desirable way to minimize the risk for phototoxic effects.

Immunofluorescence confocal microscopy of porcine corneas following collagen cross-linking treatment with riboflavin and ultraviolet A.

PURPOSE: To assess ultrastructural stromal modifications in porcine corneas after riboflavin and ultraviolet A (UVA) exposure using immunofluorescence confocal imaging. METHODS: Twenty-five freshly enucleated porcine eyes were enrolled in the study. Five eyes served as control (group I). Twenty eyes had their epithelium removed (groups I, II, IV, and V) and five eyes had their epithelium intact (group III). Groups II and III were cross-linked with riboflavin 0.1% solution (10 mg riboflavin-5-phosphate in 10 mL 20% dextran-T-500) and exposed to UVA (365 nm, 3 mW/cm2) for 30 minutes. Group IV included five eyes soaked with riboflavin without posterior irradiation, and group V included five eyes irradiated, without previous exposure to riboflavin. Ultra-thin sections (8 microm) of the corneas were stained with anti-collagen I and DAPI and their fluorescence was revealed under confocal microscopy. RESULTS: Only the cross-linked corneas (group II) showed a pronounced, highly organized anterior fluorescence zone of 182.5 +/- 22.5 microm. Using DAPI staining, an anterior and concentrated displacement of cell nuclei due to collagen compaction was observed after crosslinking (group II). No structural changes were observed in all other groups. CONCLUSIONS: The cross-linking treatment effect can be directly visualized using confocal fluorescence imaging, allowing for a quantitative analysis. Cross-linked corneas showed a pronounced and limited anterior zone of organized collagen fibers, which was not observed in the other groups. Treatment of the cornea with riboflavin and UVA without previous deepithelialization did not induce any cross-linking effect. Consequently, to facilitate diffusion of riboflavin throughout the corneal stroma, the epithelium should be removed as an important initial step in the treatment.

Light and transmission electronic microscopy evaluation of lyophilized corneas.

PURPOSE: Cornea storage for longer periods is still a challenge for corneal surgeons. The purpose of this study was to find a method to lyophilize corneas for anterior lamellar transplant and to evaluate them by light and transmission electronic microscopy. METHODS: Corneal flaps were created by using a microkeratome. Corneas were lyophilized with a cryoprotectant (2.3 mol sacarousis for 40 minutes) and without a cryoprotectant in a lyophilization machine (Modulyon D). The corneas were rehydrated with distilled water, balanced saline solution (BSS), and phosphate-buffered saline, after which they were evaluated by microscopy. A cornea that did not undergo lyophilization served as a control. RESULTS: Lyophilization without a cryoprotectant did not preserve the corneal structure. This finding was also observed when lyophilizing and rehydrating the corneas with distilled water or phosphate-buffered saline. We found that lyophilizing corneas and rehydrating them with 11 mL of BSS for 30 minutes preserved the general corneal structure, the parallelism of the collagen fibers, the Bowman layer, and the epithelial basement membrane for 15 and 30 days and for as long as 1 year or more. CONCLUSIONS: Lyophilization with sacarousis and rehydration with BSS may be a good method for anterior lamellar transplantation.

Subretinal injection of preservative-free triamcinolone acetonide and supernatant vehicle in rabbits: an electron microscopy study.

BACKGROUND: To evaluate the effect of injections of benzyl alcohol (BA)-free triamcinolone acetonide (TA) solution (MTA-PF) and the supernatant vehicle of TA (STA) containing BA into the subretinal space of rabbit eyes. METHODS: Sixteen rabbits underwent vitrectomy and subretinal injection of 0.02 ml of either 40 mg/ml MTA-PF, 40 mg/ml STA, or balanced salt solution (BSS). The animals were examined 6, 12, and 24 hours and 14 days after the procedure by fundus examination and fluorescein angiography (FA), as well as histological studies by light and transmission electron microscopy. The histological injury was classified in four stages: (1) stage 1, photoreceptor outer segment injury, (2) stage 2, stage 1 + photoreceptor inner segment injury, (3) stage 3, stage 2 + outer nuclear layer damage, and (4) stage 4, stage 3 + retinal pigment epithelium (RPE) damage. RESULTS: FA showed no window defects in areas where MTA-PF, STA, or BSS have been injected. Histological examination revealed that subretinal BSS-injection resulted in stage 1 damage during entire follow-up. Subretinal injection of MTA-PF resulted in damage stage 2 at 24 h and 14 days after surgery. However, at the STA position, stage 3 damage was noted 24 h and 14 days postoperatively. No RPE or choroidal damage was observed. CONCLUSIONS: The histological lesions induced by subretinal STA were more relevant than the damage induced by MTA-PF. The vehicle BA may be involved in these abnormalities. The data indicate that care must be taken when using TA during internal limiting membrane peeling in macular hole surgery, due to the possibility of unintentional subretinal migration and for retinal pharmacotherapy.

Effects of intravitreal triamcinolone acetonide injection with and without preservative.

AIMS: To evaluate the effects of intravitreal injection of preservative-free triamcinolone acetonide (PFTA) and TA containing preservative (KE). METHODS: A retrospective review was conducted of 646 intravitreal 4 mg/0.1 ml steroid injections in 471 eyes. A total of 577 intravitreal injections of PFTA and 69 injections of KE were administered in non-randomised eyes. No supernatant removal from KE was performed. Non-infectious endophthalmitis was defined as pseudohypopyon/hypopyon with or without an inflammatory reaction that regressed after steroid eye drop instillation. Ocular hypertension was defined as more than 23 mm Hg with Goldman applanation tonometry. Patients were followed and examined 1, 7 and 28 days, and 3, 4, 6 and 12 months after injection and annually thereafter. Statistical analysis was performed using Fisher's exact test and Chi(2) test. p Values <0.05 were considered significant. RESULTS: Both groups did not differ in demographics (p>0.05). Follow-up ranged from 6 to 57 months (mean 13, SD 7.5). Ocular hypertension was present in 127 eyes (20%), but both groups did not differ significantly (p = 0.167). Four eyes (3.15%) required trabeculectomy. Non-infectious endophthalmitis developed in 12 eyes (1.9%) and varied significantly in both groups (p = 0.005). One eye developed bacterial endophthalmitis (0.15%). CONCLUSIONS: Non-infectious endophthalmitis was observed significantly more often after KE injections (7.3%) than after PFTA injections (1.2%) (p<0.05). An inflammatory reaction was more clinically relevant in the KE group than in the PFTA group.