RESUMO
BACKGROUND: The quality of the endothelial cell layer is a major criterion for the approval of organ-cultured human donor-corneas for transplantation. We wanted to compare the predictive capacities of initial endothelial density and endothelium cell morphology for the approval of donor corneas for transplantation and for the clinical outcome after transplantation. METHODS: The endothelial density and endothelium morphology in organ culture were examined by semiautomatic assessment of 1031 donor corneas. We performed a statistical analysis for correlations of donor-data and cultivation parameters regarding their predictive capacities for the final approval of donor corneas for transplantation and the clinical outcome of 202 transplanted patients. RESULTS: Corneal endothelium cell density proved to be the only parameter with a certain predictive capacity with regard to the final decision, whether donor corneas are suitable for transplantation - however, the correlation was low (area under the curve [AUC] = 0.655). Endothelial cell morphology lacked any predictive power (AUC = 0.597). The clinical outcome regarding visual acuity seemed to be largely independent from both corneal endothelial cell density and morphology. Sub-analyses on transplanted patients stratified for their diagnoses vindicated these findings. CONCLUSIONS: Higher endothelial density (above a cut-off level of 2000 cells/mm2), as well as better endothelial morphology do not seem to be critical for transplant-corneal functionality in organ culture and up to 2 years after transplantation. Comparable long-term studies on graft survival are recommended to determine, whether the present endothelial density cut-off levels might be too stringent.
Assuntos
Transplante de Córnea , Endotélio Corneano , Humanos , Córnea , Técnicas de Cultura de Órgãos , Doadores de Tecidos , Contagem de CélulasRESUMO
BACKGROUND: Cataract surgery induces corneal endothelial cell loss (ECL). This study investigates the relationship between bottle height (BH) and ECL induced due to irrigation and aspiration (I/A) in cataract surgery and quantifies protective effects of intraoperatively used ophthalmic viscoelastic substances. METHODS: Intermittent I/A without phacoemulsification was performed in porcine eyes for 10 min with varying BHs of 100 cm (BH100), 125 cm (BH125), 150 cm (BH150) or no treatment (control, no I/A). Additionally, in one group a dispersive ophthalmic viscoelastic substance was injected into the anterior eye chamber before treatment with I/A at a BH of 150 cm (BH150 + V). After exposure of the corneal endothelium to I/A, the corneas were prepared to split corneal buttons on day 0 and cultivated for 15 days. Endothelial cell density (ECD) was analyzed blinded on days 1, 8 and 15. RESULTS: Relative ECL significantly correlated with irrigation BH (control (n = 13): -9.69 ± 6.03% (average ± standard deviation); BH100 (n = 12): -9.69 ± 4.81%-p = 1.000; BH125 (n = 14): -19.44 ± 7.30% - p < 0.001; BH150 (n = 13): -21.99 ± 6.70%-p < 0.001). I/A-induced ECL was significantly decreased by the injection of ophthalmic viscoelastic, as BH150 + V (n = 14; -10.92 ± 4.09%-p = 1.000) showed a cell loss comparable to the control group. CONCLUSIONS: ECL is altered by I/A BH and reduced when viscoelastic substances are used.
Assuntos
Extração de Catarata , Catarata , Facoemulsificação , Animais , Suínos , Células Endoteliais , Endotélio Corneano , Contagem de CélulasRESUMO
There are many disease patterns that are treated jointly by neurologists and ophthalmologists, for which optical coherence tomography (OCT) is of important differential diagnostic significance. In this context neurologists are mainly confronted by two patient collectives: patients with an acute ischemic event, who present with an acute but painless monocular visual deterioration (for central retinal artery occlusion) or with a monocular visual field defect (for arterial branch occlusion or anterior ischemic optic neuropathy). The second collective is patients without ophthalmological symptoms but with conspicuous optic nerve findings (papilledema or optic disc drusen). In this overview article both patient collectives are considered separately. In addition, the most important OCT findings for optic neuritis are presented. Before the disease patterns are described in detail, the normal OCT findings and the diagnostic possibilities of OCT are explained.
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Neurologia , Neurite Óptica , Papiledema , Humanos , Neurite Óptica/diagnóstico por imagem , Papiledema/diagnóstico , Tomografia de Coerência Óptica/métodosRESUMO
In the field of ophthalmology, central retinal artery occlusion is still one of the unsolved pathologies, as there is no widely accepted evidence-based therapeutic concept. Meta-analysis suggests that intravenous fibrinolysis might help to improve the outcome if performed within the first 4.5 hours after symptom onset. However, this short window of opportunity is often missed due to delays during diagnostic testing (e.g. time elapsed before performance of an ophthalmological examination) or patient-processing (referring the patient to a specialized medical facility). This article presents a diagnostic tool in the form of a simple questionnaire, comprising a medical history and examinations, which could help to minimize the time required to initiate the appropriate treatment.
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Oclusão da Artéria Retiniana , Fundo de Olho , Humanos , Oclusão da Artéria Retiniana/diagnóstico , Oclusão da Artéria Retiniana/terapiaRESUMO
The retina is a complex neurological tissue and is extremely sensitive to an insufficient supply of oxygen. Hypoxia plays a major role in several retinal diseases, and often results in the loss of cells that are essential for vision. Cyclosporine A (CsA) is a widely used immunosuppressive drug. Furthermore, treatment with CsA has neuroprotective effects in several neurologic disorders. No data are currently available on the tolerated concentration of CsA when applied to the retina. To reveal the most effective dose, retinal explants from rat eyes were exposed to different CsA concentrations (1-9 µg/mL). Immunohistochemistry with brain-specific homeobox/POU domain protein 3a (Brn3a) and TUNEL staining was performed to determine the percentage of total and apoptotic retinal ganglion cells (RGCs), as well as the responses of micro- and macroglial cells. Furthermore, optical coherence tomography (OCT) scans were performed to measure the changes in retinal thickness, and recordings with multielectrode array (MEA) were performed to evaluate spontaneous RGC spiking. To examine the neuroprotective effects, retinas were subjected to a hypoxic insult by placing them in a nitrogen-streamed hypoxic chamber prior to CsA treatment. In the biocompatibility tests, the different CsA concentrations had no negative effect on RGCs and microglia. Neuroprotective effects after a hypoxic insult on RGCs was demonstrated at a concentration of 9 µg/mL CsA. CsA counteracted the hypoxia-induced loss of RGCs, reduced the percentage of TUNEL+ RGCs, and prevented a decrease in retinal thickness. Taken together, the results of this study suggest that CsA can effectively protect RGCs from hypoxia, and the administered concentrations were well tolerated. Further in vivo studies are needed to determine whether local CsA treatment may be a suitable option for hypoxic retinal diseases.
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Apoptose/efeitos dos fármacos , Hipóxia Celular/efeitos dos fármacos , Ciclosporina/farmacologia , Fármacos Neuroprotetores/farmacologia , Retina/efeitos dos fármacos , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Glaucoma/tratamento farmacológico , Técnicas de Cultura de Órgãos , Ratos , Retina/fisiologia , Doenças Retinianas/tratamento farmacológico , Células Ganglionares da Retina/metabolismo , Oclusão da Veia Retiniana/tratamento farmacológico , Tomografia de Coerência ÓpticaRESUMO
BACKGROUND: Primary open-angle glaucoma (POAG) is a progressive neurodegenerative disease which leads to irreversible blindness. An elevated intraocular pressure (IOP) is considered to be the main risk factor for the disease progression. It is known that retinal blood flow is altered in POAG eyes. Tafluprost, a prostaglandin analogue which lowers the IOP, has shown to also improve the retinal blood flow in animals. METHODS: The current study therefore evaluated the retinal vessel density in the peripapillary and macular region of POAG patients with normal IOP treated with topical Tafluprost (n = 20) compared to surgically treated patients with normal IOP (n = 22) using optical coherence tomography angiography (OCT-A). The retinal flow density was obtained after binarisation and evaluated in five sectors. RESULTS: There was a significantly higher peripapillary flow density in all sectors in Tafluprost treated eyes when compared to post-surgery eyes. The flow density in the inferior sector of the superficial plexus in the macular region was also significantly higher in the Tafluprost group. CONCLUSIONS: These results indicate that Tafluprost not only lowers IOP, but may also enhance retinal blood flow in POAG patients with a normal IOP.
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Glaucoma de Ângulo Aberto , Doenças Neurodegenerativas , Disco Óptico , Angiografia , Animais , Glaucoma de Ângulo Aberto/tratamento farmacológico , Humanos , Pressão Intraocular , Prostaglandinas F , Vasos Retinianos/diagnóstico por imagem , Tomografia de Coerência ÓpticaRESUMO
BACKGROUND: Hypoxia contributes to retinal damage in several retinal diseases, including central retinal artery occlusion, with detrimental consequences like painless, monocular loss of vision. Currently, the treatment options are severely limited due to the short therapy window, as the neuronal cells, especially the retinal ganglion cells (RGCs), are irreversibly damaged within the first few hours. Hypothermia might be a possible treatment option or at least might increase the therapy window. METHODS: To investigate the neuroprotective effect of hypothermia after retinal hypoxia, an easy-to-use ex vivo retinal hypoxia organ culture model developed in our laboratory was used that reliably induced retinal damage on a structural, molecular and functional level. The neuroprotective effect of hypothermia after retinal hypoxia was analysed using optical coherence tomography scans, histological stainings, quantitative real-time polymerase chain reaction, western blotting and microelectrode array recordings. RESULTS: Two different hypothermic temperatures (30°C and 20°C) were evaluated, both exhibited strong neuroprotective effects. Most importantly, hypothermia increased RGC survival after retinal hypoxia. Furthermore, hypothermia counteracted the hypoxia-induced RGC death, reduced macroglia activation, attenuated retinal thinning and protected from loss of spontaneous RGC activity. CONCLUSIONS: These results indicate that already a mild reduction in temperature protects the RGCs against damage and could function as a promising therapeutic option for hypoxic diseases.
Assuntos
Hipotermia Induzida , Hipóxia/patologia , Retina/patologia , Células Ganglionares da Retina/citologia , Animais , Apoptose , Western Blotting , Sobrevivência Celular/fisiologia , Citoproteção , Regulação da Expressão Gênica/fisiologia , Proteína Glial Fibrilar Ácida/genética , Imuno-Histoquímica , Microeletrodos , Técnicas de Cultura de Órgãos , RNA Mensageiro/genética , Ratos , Reação em Cadeia da Polimerase em Tempo Real , Retina/metabolismo , Células Ganglionares da Retina/metabolismo , Tomografia de Coerência ÓpticaRESUMO
To determine the corneal endothelial cell density human donor corneas are usually placed in hypotonic balanced salt solution for unstained cell counting. Experimental studies often stain the corneal endothelium to highlight cells and cell borders and simplify counting. However, staining normally terminates the experiments. Up to date, there is no publication comparing endothelial cell counts before and after staining. This study compared the counting results of unstained (hypotonic balanced salt solution) and stained (trypan blue & alizarin red S) assessment of the corneal endothelial cell density. No evidence for significantly differing cell counts was found between unstained and stained analysis of the corneal endothelial cell density. Therefore, we consider unstained counting non-inferior to stained counting.
Assuntos
Antraquinonas/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Corantes/farmacologia , Endotélio Corneano/citologia , Azul Tripano/farmacologia , Contagem de Células/métodos , HumanosRESUMO
Silicone oil endotamponades need to be injected and removed in a reasonable time and under moderate pressure conditions. However, due to ever-decreasing sizes of incisions and trocars, injection and removal of highly viscous silicone oils is very time-consuming. To address resulting problems like longer treatment times or hypotony, thixotropic silicone oils were developed. These oils are characterized by a diminished viscosity under constant mechanical stress; whilst there is pressure or vacuum acting on it, the oils will become more fluid and, therefore, much easier to be applied. Once the force is being removed from the oil, it will automatically return to its initial viscosity after a short time.
Assuntos
Tamponamento Interno , Descolamento Retiniano , Óleos de Silicone , Humanos , Descolamento Retiniano/terapia , Viscosidade , VitrectomiaRESUMO
Human corneas usually are not available for research, as they are used for transplantation only. At the same time, scientific studies on cultured human endothelial cells can produce misleading results due to inevitable dedifferentiation. Therefore, an organ-culture model of porcine corneas-displaying endothelial cell death rates comparable to those of cultured human corneas-would be very desirable. Fresh pig eyes were prepared under sterile conditions to obtain corneoscleral buttons, corneal buttons and so called "split corneal buttons" (new preparation method) and cultivated for 15 days. Morphology of the endothelial cell layer was observed by light microscopy on day 1, 8 and 15. On day 15 staining with trypan blue and alizarin red S was performed. Photographs were evaluated in a randomized, blinded manner. Here, the morphology of the corneal endothelium and the number of endothelial cells per mm2 were analyzed. After 15 days of cultivation the endothelial cell layer was maintained only in corneal buttons and split corneal buttons. Alizarin red S stained areas and the existence of polymorphisms like rosette figures and reformation figures were significantly less frequent in split corneal buttons than in corneal buttons. Loss of endothelial cells was significantly greater in corneal buttons [575 ± 25/250 cells/mm2 (median ± 25%/75%-quantile); 14.8%] than in split corneal buttons [417 ± 138/179 cells/mm2 (median ± 25%/75%-quantile); 10.2%]. The new preparation method of split corneal buttons allows the cultivation of porcine corneas for 2 weeks with cell death rates comparable to those of the corresponding human tissue in cornea banks without the need to add de-swelling additives to the media. This is therefore a simple and highly reliable method model to be applied in intervention studies on corneal endothelial cells in their natural compound.
Assuntos
Endotélio Corneano/citologia , Técnicas de Cultura de Órgãos/métodos , Animais , Antraquinonas/análise , Contagem de Células , Morte Celular , Endotélio Corneano/ultraestrutura , Coloração e Rotulagem/métodos , Suínos , Azul Tripano/análiseRESUMO
Optical coherence tomography (OCT) dramatically changed the way of diagnostic assessment in retinal diseases during the last years. Using this technique in-vivo in-depth analysis of the retina and its layers is possible. Since animal research is changing by intrinsic and extrinsic pressure to animal-(in-vivo)-free methods, we adapted OCT-measurements to organotypic cultures. An easy to use protocol was generated to assess standardized OCT assessments in organotypic culture. First, two custom-made devices need to be made to change any commercially available OCT for examinations in humans into a device allowing ex-vivo analyses of organotypic culture. The modification is feasible within seconds. After OCT measurement of the ex-vivo tissues, quantitative evaluation of the retinas were performed via ImageJ software. OCT pictures of ex-vivo retinas were obtained for time periods of seven days and the thickness of retinal tissue was evaluated. The reproducibility of the pictures and measurements was very high (SD < 15%). In conclusion, an easy to use protocol for the investigation of different effects on retinal cultures with commercially available OCT devices was successfully established.
Assuntos
Modelos Animais de Doenças , Retina/diagnóstico por imagem , Técnicas de Cultura de Tecidos , Tomografia de Coerência Óptica/métodos , Animais , Ratos , Reprodutibilidade dos Testes , Retina/ultraestrutura , Doenças Retinianas/diagnóstico por imagemRESUMO
The standard of care to treat small- and medium-sized macular holes (<400 µm diameter) consists of a conventional transconjunctival sutureless pars plana vitrectomy followed by ILM peeling and endotamponade, mainly with gas or in some cases with silicone oil, resulting in closure rates of over 90% and good functional results. Large (>400 µm diameter), chronic and persistent macular holes remain a surgical challenge since closure rates and functional results decrease with larger macular hole diameters. Various modifications of the conventional surgical technique were introduced to improve anatomic and functional success in refractory cases not suitable for conventional macular hole surgery. These techniques comprise the positioning of tissue at the top of the hole to improve closure as performed by an inner limiting membrane flap and free flap preparation or the transplantation of autologous retinal tissue, lens capsule or amniotic membrane. For the treatment of very large and persistent macular holes, the induction of a localized retinal detachment at the posterior pole by subretinal injection of balanced salt solution and a subsequent attenuation of the rim of the hole during fluid-air exchange has been suggested as a promising surgical technique. In particular, accurate patient education about the expected surgical outcome in this specific group of patients appears important.
Assuntos
Perfurações Retinianas , Vitrectomia , Humanos , Perfurações Retinianas/cirurgia , Vitrectomia/métodos , Tamponamento Interno/métodos , Tomografia de Coerência Óptica , Acuidade Visual , Retalhos CirúrgicosRESUMO
PURPOSE: Several iatrogenic risk factors during pars plana vitrectomy (PPV) could cause damage to the retina. One mechanism is excitotoxicity. Therefore, neuroprotective irrigation solutions would be desirable. METHODS: Retinal ganglion cells (RGC-5) and retinal whole mounts were incubated in standard irrigation solution (SIS) and Dulbecco's Modified Eagle Medium (DMEM). Cell viability, cell amount, cell survival and caspase 3/7 activity were measured by MTS-Test, crystal-violet staining, Annexin-V/PI flow cytometry and caspase 3/7 activity assay, respectively. The morphology and the function of retinal whole mounts were analysed by Live/Dead(TM) staining and by the b-wave and a-wave of the electroretinogram (ERG). RESULTS: Under excitotoxic conditions (10 mM and 12 mM glutamate) RGC-5 cells incubated in SIS showed a statistically significant reduction in cell viability, cell amount, cell survival and caspase 3/7 activity compared to DMEM. Furthermore, the incubation of retinal whole mounts in DMEM resulted in a significant decrease of cell death under excitotoxic (250 µM glutamate) and standard conditions compared to SIS. ERG b-wave recordings revealed good functional preservation of retinal whole mounts in DMEM, but loss in SIS. CONCLUSION: DMEM seems to support retinal cells very well and to be strongly protective against excitotoxicity. Therefore, DMEM may be considered as possible neuroprotective irrigation solution for PPV.
Assuntos
Meios de Cultura/farmacologia , Fármacos Neuroprotetores/farmacologia , Soluções Oftálmicas/farmacologia , Retina/efeitos dos fármacos , Células Ganglionares da Retina/efeitos dos fármacos , Vitrectomia , Animais , Anexina A5/metabolismo , Caspase 3/metabolismo , Caspase 7/metabolismo , Bovinos , Contagem de Células , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Eletrorretinografia , Ensaio de Imunoadsorção Enzimática , Agonistas de Aminoácidos Excitatórios/toxicidade , Citometria de Fluxo , Ácido Glutâmico/toxicidade , Ratos , Retina/citologia , Retina/enzimologia , Células Ganglionares da Retina/citologia , Células Ganglionares da Retina/enzimologia , Irrigação TerapêuticaRESUMO
The structure of the vitreous body, its interaction with the retinal surface and tractive alterations of the vitreoretinal interface may play a role in the pathogenesis and the development of age-related macular degeneration (AMD). From clinical trials it can be concluded that posterior vitreous detachment (PVD) or vitreous removal may protect against the development of neovascular AMD. Vitreomacular adhesions may promote neovascular AMD and may have an impact on the efficacy and frequency of intravitreal vascular endothelial growth factor (VEGF) inhibition. Therefore, vitreomacular surgery may be considered as a treatment option in selected cases. Peeling of epimacular membranes and the internal limiting membrane (ILM) may contribute to stabilizing visual acuity and reducing the treatment burden of current intravitreal pharmacotherapy. At present, surgical interventions in AMD are mainly performed in cases of submacular hemorrhage involving the fovea. The treatment is not standardized and consists of liquefaction of the blood using recombinant tissue plasminogen activator (rTPA) and its pneumatic displacement, potentially combined with VEGF inhibition. Other submacular surgical strategies, such as choroidal neovascularization (CNV) extraction, macular translocation or transplantation of retinal pigment epithelium (RPE) are currently limited to selected cases as a salvage treatment; however, the development of these submacular surgical interventions has formed the basis for new approaches in the treatment of dry and neovascular AMD including submacular or intravitreal gene and stem cell therapy.
RESUMO
Age-related changes in vitreous molecular and anatomic morphology begin early in life and involve two major processes: vitreous liquefaction and weakening of vitreo-retinal adhesion. An imbalance in these two processes results in anomalous posterior vitreous detachment (PVD), which comprises, among other conditions, vitreo-macular adhesion (VMA) and traction (VMT). VMA is more common in patients with neovascular age-related macular degeneration (nAMD) than age-matched control patients, with the site of posterior vitreous adherence to the inner retina correlating with location of neovascular complexes. The pernicious effects of an attached posterior vitreous on age-related macular degeneration (AMD) progression involve mechanical forces, enhanced fluid influx and inflammation in and between the retinal layers, hypoxia leading to an accumulation of vascular endothelial growth factor (VEGF) and other stimulatory cytokines, and probably an infiltration of hyalocytes. It has been shown that vitrectomy not only mitigates progression to end-stage AMD, but existing choroidal neovascularization regresses after surgery. Thus, surgical PVD induction during vitrectomy or by pharmacologic vitreolysis may be considered in non-responders to anti-VEGF treatment with concomitant VMA.
Assuntos
Degeneração Macular , Doenças Retinianas , Descolamento do Vítreo , Humanos , Corpo Vítreo/cirurgia , Fator A de Crescimento do Endotélio Vascular , Descolamento do Vítreo/complicações , Degeneração Macular/complicações , Doenças Retinianas/complicaçõesRESUMO
BACKGROUND: Intravitreal anti-vascular endothelial growth factor (VEGF) is the standard treatment for exudative age-related macular degeneration (AMD). The constitution of the vitreomacular interface varies greatly in cases of attached (with or without traction) or detached vitreous body, which can impact the effectiveness of the anti-VEGF treatment. OBJECTIVE: Based on the current literature this article displays the current state of the science on whether the constitution of the vitreous body has an effect on the anti-VEGF treatment. MATERIAL AND METHODS: The published data extracted from current trials and post hoc analyses concerning this topic are presented and put into the clinical context. RESULTS: The presence of a vitreomacular adhesion reduces the efficacy of anti-VEGF treatment of exudative AMD. Posterior vitreous body detachment represents a positive prognostic factor concerning the efficacy of anti-VEGF treatment but not necessarily the prognosis for visual acuity. CONCLUSION: Patients with attached vitreous body need a more intensive treatment monitoring compared to patients with detached vitreous body. Therefore, in eyes with initial posterior vitreous body detachment receiving a treat and extend regimen, the interval between anti-VEGF injections can be extended to 4 instead of 2 weeks without endangering the success of treatment.
Assuntos
Degeneração Macular , Doenças Retinianas , Descolamento do Vítreo , Humanos , Corpo Vítreo , Fator A de Crescimento do Endotélio Vascular/uso terapêutico , Degeneração Macular/tratamento farmacológicoRESUMO
Purpose: To investigate the sensitivity and specificity of central retinal artery occlusion (CRAO)-Detection Score in diagnosing CRAO via questionnaire and without fundoscopy. Methods: This prospective study enrolled 176 emergency patients suffering from acute visual loss, of whom 38 were suffering from CRAO. Before conducting any examination, we administered our questionnaire containing six questions, followed by a thorough ophthalmologic examination to make the diagnosis. Statistical analysis involved a LASSO penalised multivariate logistic regression model. Results: Our receiver operating characteristic (ROC) analysis based on a LASSO penalised multivariate logistic regression model showed an area under the curve (AUC) of 0.9 - three out of six questions were selected by LASSO. Interestingly, the unweighted ROC analysis of only two questions (Short CRAO-Detection Score) yielded similar results with an AUC of 0.88. The short CRAO-Detection Score of 2 yielded 14% (4/28) false positive patients. Conclusion: This prospective study demonstrates that a high percentage of CRAO patients are detectable with a questionnaire. The CRAO-Detection Score might be used to triage patients suffering acute visual loss, which is important as intravenous fibrinolysis seem to be time-dependent to be effective.
RESUMO
BACKGROUND: During vitreoretinal surgery, vital dyes are used to visualize anatomical structures. Substances with a density higher than water are added to facilitate sedimentation and staining. BBG with 4% PEG (ILM Blue) and BBG with TB and 4% PEG (MBB Dual) are two new dyes. This study evaluates biocompatibility of the new dye solutions, using cell cultures and electrophysiological evaluation. METHODS: To determine cytotoxicity of ILM Blue and MBB Dual for 30, 60, 120 and 320 seconds, monolayer cultures of retinal ganglion cells (RGC5) were used. For functionality testing, bovine retinas were isolated and superfused with an oxygen-saturated nutrient solution, and the electroretinogram (ERG) was recorded. The two dye solutions were applied epiretinally for 30, 60 and 120 seconds. ERG recovery was monitored. RESULTS: After staining with ILM Blue, no statistical significant reduction of a- or b-wave amplitudes at the end of the wash-out was recorded. For MBB Dual, only a significant reduction of the a-wave amplitudes after 30 seconds of application at the end of the wash-out was noticed, while no statistically different changes for a- and b-wave amplitudes up to 120 seconds were noted. During the MTT assay, we noted no significant difference in cell viability after 30, 60, 120 and 320 seconds of staining with ILM blue, MBB Dual or 4% PEG in comparison to the control group (DMEM, Triton X-100 0.9% as positive control) after formazan extraction. CONCLUSIONS: ILM Blue and MBB Dual seem to be safe for clinical use for a staining period of up to 120 seconds, probably even up to 320 seconds.
Assuntos
Materiais Biocompatíveis/toxicidade , Corantes/toxicidade , Retina/efeitos dos fármacos , Células Ganglionares da Retina/efeitos dos fármacos , Corantes de Rosanilina/toxicidade , Azul Tripano/toxicidade , Animais , Bovinos , Técnicas de Cultura de Células , Sobrevivência Celular/efeitos dos fármacos , Combinação de Medicamentos , Eletrorretinografia/efeitos dos fármacos , Teste de Materiais , Técnicas de Cultura de Órgãos , Perfusão , Coloração e Rotulagem , Fatores de Tempo , Cirurgia VitreorretinianaRESUMO
BACKGROUND: During pars plana vitrectomy, the retina is exposed to several iatrogenic risk factors, including excitotoxicity. A taurine-containing irrigation solution for pars plana vitrectomy (PURI PROTECT) has been developed and is claimed to have neuroprotective properties. METHODS: Retinal ganglion cells (RGC-5) and retinal whole mounts were incubated in standard irrigation solution (SIS) and SIS supplemented with 3 mM taurine (SIS-taurine). Excitotoxicity was induced by the addition of 8, 10, and 12 mM or 250 µM glutamate. Cell viability and cell survival were assessed by the MTT test and Annexin-V/propidium iodide flow cytometry. Whole mounts were stained with the Live/Dead staining assay. Pars plana vitrectomy with SIS or SIS-taurine was performed in rabbits. Animals were followed-up by electroretinography. RESULTS: RGC-5 incubated in SIS-taurine showed a 4.3-fold (P < 0.0005) better overall cell viability and an up to 8.5-fold (P < 0.05) increased cell survival under excitotoxic conditions compared with that incubated in SIS. Whole mounts incubated in SIS-taurine showed a 1.7-fold (P < 0.0005) and 1.6-fold (P < 0.0005) better cell survival under excitotoxic and nonexcitotoxic conditions, respectively. In the immediate postoperative period, b-wave amplitudes were significantly better in animals operated with SIS-taurine compared with control (P < 0.01). CONCLUSION: A taurine-containing irrigation solution may protect retinal ganglion cells against excitotoxicity.