Analysis of monosomy-3 in immunomagnetically isolated circulating melanoma cells in uveal melanoma patients.

Monosomy-3 in primary uveal melanoma (UM) is associated with a high risk of metastasis and mortality. Although circulating melanoma cells (CMC) can be found in most UM patients, only approximately 50% of the patients develop metastases. We utilized a novel immuno-FISH assay to detect chromosome-3 in intact CMC isolated by dual immunomagnetic enrichment. Circulating melanoma cells were detected in 91% of the patients (n = 44) with primary non-metastatic UM, of which 58% were positive for monosomy-3. The monosomy-3 status of CMC corresponded to the monosomy-3 status of the primary tumor in 10 of the 11 patients where this could be tested. Monosomy-3 in the CMC was associated with an advanced tumor stage (P = 0.046) and was detected in all four patients who developed metastasis within the follow-up period of 4 yr. This non-invasive technique may enable the identification of UM patients at risk for metastasis particularly when a primary tumor specimen is unavailable.

CD147 and matrix-metalloproteinase-2 expression in metastatic and non-metastatic uveal melanomas.

BACKGROUND: Extracellular matrix remodelling regulated by matrix-metalloproteinase (MMP) inducer (CD147) is a crucial process during tumor cell invasion and regulation of blood supply. In this study, we evaluated the correlation of CD147 and MMP-2 expression with major prognostic factors for uveal melanoma and the development of metastasis. METHODS: The expression of CD147 and MMP-2 was analyzed in 49 samples of uveal melanomas. Triple immunofluorescence stainings using markers against glial cells (GFAP), endothelial cells (CD34) and macrophages (CD68) were performed to further analyse the exact localisation of CD147 and MMP-2 positivity. In 28 cases clinical metastatic disease were found. The remaining 21 cases showed no signs of metastatic disease for an average follow-up of 10 years. Correlation analysis (Pearson correlation) was performed to analyse the association of CD147 and MMP-2 expression with known prognostic factors, vasculogenic mimicry (VM), the mature vasculature (von Willebrand Factor) and tumor induced angiogenesis (by means of Endoglin expression). RESULTS: CD147 and MMP-2 were expressed in 47 (96.0 %) of the uveal melanomas. CD147 up-regulation was significantly correlated with a higher MMP-2 expression. The overall expression analysis revealed no significant difference in the metastatic (p = 0.777) and non-metastatic subgroup (p = 0.585). No correlation of CD147 expression and any system of blood supply was evident. In the non-metastatic sub-group a significant correlation of clustered CD147 positive cells with largest basal diameter (p = 0.039), height (p = 0.047) and TNM-stage (p = 0.013) was evident. CONCLUSIONS: These data may indicate that CD147 regulates MMP-2 expression in uveal melanoma cells.

Analysis of caveolin-1 and phosphoinositol-3 kinase expression in primary uveal melanomas.

BACKGROUND: To evaluate the regulation of blood supply in primary uveal melanomas through caveolin-1 (Cav-1)/phosphoinositol-3 kinase (PI3K). METHODS: The expression of Cav-1 and PI3K was analysed in 51 paraffin sections of metastatic (n = 30) and non-metastastic uveal melanomas (n = 21). Two trained observers quantified Cav-1 and PI3K immunofluorescensce expression by determining intensity of staining and percentage of positive cells. The expression was correlated with known prognostic factors. Besides angiogenesis by means of endoglin expression, the normal vasculature (von Willebrand Factor expression) was evaluated semi-quantitatively. Vasculogenic mimicry (VM) was analysed by CD31/PAS staining. RESULTS: All examined specimens expressed Cav-1 with a mean of 90.34% Cav-1 positive cells (range, 3.23-100%). Metastatic disease was associated with a higher Cav-1 expression. The correlation of Cav-1 with well-established prognostic factors showed a significant association between Cav-1 expression and largest tumour diameter (P = 0.022), tumour node metastasis classification (P = 0.008) and invasion of optic nerve head (P = 0.048). PI3K was expressed by all uveal melanomas with a mean of 87.28% cells showing PI3K expression. A higher level of PI3K was significantly associated with larger height (P = 0.042) and progressed tumour node metastasis stage (P = 0.016). The percentage of PI3K and Cav-1 positive cells were significantly associated (P = 0.034). For PI3K and Cav-1 expression a non-significant association with VM was shown (P = 0.064 and P = 0.072, respectively). No correlation of PI3K or Cav-1 with angiogenesis or mature vasculature was seen (P > 0.05). CONCLUSIONS: Cav-1 expression may be especially up-regulated in larger uveal melanomas. As it was correlated with PI3K expression and VM in this series of uveal melanoma, Cav-1 might induce the formation of VM via the PI3K-signalling cascade.

A p38 MAPK inhibitor improves outcome after glaucoma filtration surgery.

PURPOSE: The aim of this study is to explore the effects of SB 202190, a highly selective p38 MAPK inhibitor, on bleb survival following glaucoma filtering surgery. MATERIALS AND METHODS: Human Tenon's fibroblasts were treated with SB 202190 (0 to 100 µM) to determine IC50, and cell proliferation and migration. Twenty rabbits were divided into 4 groups (G1-G4): G1 animals received only a trabeculectomy. G2-G4 animals had trabeculectomy plus one of the following subconjunctival adjuvants, given intraoperatively and postoperatively: G2=sham, G3=20 µM SB 202190, and G4=50 µM SB 202190. The blebs were assessed using the Indiana Bleb Appearance Grading Scale. The intraocular pressure (IOP) was expressed as the right to left eye ratio (R/L ratio). For morphometric bleb analysis the JMicrovision software was used. RESULTS: SB 202190 inhibits human Tenon's fibroblasts proliferation and migration in vitro (IC50=17.2 µM). In vivo subconjunctival application of SB 202190 after glaucoma filtration surgery significantly increases bleb height, bleb extension, and bleb survival time compared with the control. In all groups, the IOP ratio correlates with the fibrotic process. G3 shows a significantly reduced IOP ratio at day 14 compared with the control. Analysis of the bleb histology shows that G3 has a significant smaller fibrosis area compared with G1 and G2. Application of the highest dose (50 µM SB 202190) is associated with hyphema in 2 of 5 animals (40%). CONCLUSION: Application of SB 202190 significantly improves bleb characteristics and IOP control after filtering glaucoma surgery in a rabbit model.

Serum cytokines as biomarkers for age-related macular degeneration.

PURPOSE: This study evaluates the potential of serum pro-inflammatory cytokines as AMD biomarkers. METHODS: Serum samples from 30 age-related macular degeneration (AMD) patients and 15 age-matched controls were examined for 16 inflammatory cytokines using multiplex ELISA. Patients were divided into three subgroups (improvement/no change/deterioration during anti-VEGF treatment) by OCT and funduscopy, and correlated to the cytokine levels. RESULTS: Serum concentrations of IL-1α, IL-1ß, IL-4, IL-5, IL-10, IL-13, and IL-17 were significantly higher in AMD patients than in controls. None of the co-variables expressed a significant effect on the tested cytokines. Only IL-1a and IL-17 showed a statistically significant difference between groups (improved, unchanged, deteriorated) as determined by one-way ANOVA. Patients with increased macular thickness during treatment showed significantly lower levels of IL-17 compared to improved cases and to unchanged cases (p = 0.004, 0.03 respectively, Dunnett's T3 post hoc multiple test). TNF-α was significantly higher in improved cases compared to deteriorated cases (p =0.03, Dunnett's T3 post hoc multiple test). IL-17 was a significant predictor for macular oedema using linear regression (ß = -0.888, p <0.05). CONCLUSION: Elevation of IL-1α, IL-1ß, IL-4, IL-5, IL-10, IL-13, and IL-17 in the serum of AMD patients supports the hypothesis of AMD as an inflammatory disease. Patients with high IL-17 and TNF-α serum levels were more likely to have a favourable course under VEGF therapy. These cytokines may be used as easy-to-obtain biomarkers.

Testing the effects of the dye acid violet-17 on retinal function for an intraocular application in vitreo-retinal surgery.

PURPOSE: To facilitate epiretinal or inner limiting membrane peeling, dyes like Indocyanine Green (ICG) as well as Trypan Blue (TB) were used so far. However, toxic effects on the retina were described for both dyes. The aim of our study was to investigate the effects of a novel vital dye Acid violet-17 (AV-17) on retinal histology and function to assess a possible application in vitreo-retinal surgery. METHODS: AV-17 was dissolved in a solvent with heavy water. An electroretinogram was recorded on perfused bovine retina. After reaching stable b-wave amplitudes, AV-17 (0.125-0.5 mg/ml) or the solvent was applied epiretinally for 30-300 seconds. The b-wave amplitudes were recorded before, during, and after treatment. Cultures of bovine retina were incubated for 30 or 300 seconds with the dye or solvent and processed for live/dead staining, immunohistochemistry, and immunoblotting. RESULTS: Reductions of the b-wave amplitudes were observed directly after the exposure to AV-17, which were rapidly and completely reversible within the recovery period for all exposure times at the concentrations of 0.125 and 0.25 mg/ml as opposed to the partial recovery after exposure to 0.5 mg/ml. A high degree of damage in the ganglion cell layer (GCL) and glial reactivity were detected at the concentrations of 0.25 and 0.5 mg/ml but not after exposure to lower concentrations or the solvent. CONCLUSION: Application of AV-17 at a concentration of up to 0.125 mg/ml was well tolerated in terms of retinal function, survival in the GCL, and glial reactivity whereas higher concentrations are not recommended.

Identification of circulating melanoma cells in uveal melanoma patients by dual-marker immunoenrichment.

PURPOSE: Despite successful local tumor control, uveal melanoma (UM) patients may develop lethal metastases. To reliably identify circulating melanoma cells (CMC) in UM patients, we set out to test a new immunomagnetic enrichment assay and screened UM patients for the presence of CMC. We also determined whether we could find CMC in culture; for example, for future drug testing. METHODS: A dual-immunomagnetic enrichment assay using antibodies against two melanoma markers (NKI/C3 and NKI/beteb) was used to determine the presence of UM cells in blood. The sensitivity of the assay was determined by spiking normal blood with 92.1 cells (concentration range, 1-10(4) cells/mL). Isolated cells were characterized by immunocytochemistry directly after immunoenrichment and after a 2-week culture. The presence of CMC was determined in the peripheral blood of 31 patients with UM, and results were compared to clinical prognostic factors at the time of presentation. RESULTS: The CMC were detected in 93.5% (n = 29 of 31) of the patients with primary nonmetastatic UM at a median density of 3.5 cells/10 mL blood (range, 0-10.2 cells), as well as in blood cultures. No significant association was observed between the presence or number of CMC and any clinical prognostic factors. CONCLUSIONS: The improved dual-immunoenrichment assay enabled the detection of intact and viable CMC in the majority of UM patients. We also were able to identify CMC after short-term culturing. Molecular characterization of the CMC rather than the prevalence of these cells is expected to provide relevant information on the individual risk of metastasis.

The retinal biocompatibility of dyes in the ex vivo model of the isolated superfused vertebrate retina.

BACKGROUND: Peeling of the internal limiting membrane or epiretinal membranes is a successful principle in macular surgery to achieve a functional benefit. Different dyes are used to facilitate the identification of intraocular tissues. The aim of our work was to investigate the retinal tolerance to the different dyes and their solvent carriers to provide valuable data for surgeons in handling for an optimal intraoperative use. METHODS: Using the ex vivo model of the isolated superfused vertebrate retina technique, the effects of the dyes were tested on human and bovine retinal function. The retinas were perfused with an oxygen preequilibrated standard solution. The electroretinogram (ERG) was recorded using Ag/AgCl electrodes. After recording stable ERG amplitudes, the dyes brilliant blue G, indocyanine green, trypan blue, patent blue, triamcinolone and their solvent carriers were investigated. RESULTS: Reductions of the ERG amplitudes were found for each tested dye. The effects after application of the dyes were dependent on time and concentration of the applied dyes, which were different for each dye. CONCLUSION: In part, the ERG has shown strong effects already after a short period of dye application. Surgeons who rely on the intraocular use of the dyes should keep in mind our findings, and the use of some dyes should be limited to selected cases. The well-considered use of the dyes by the surgeons could lead to a better functional outcome and avoid a possible harmful outcome of the surgery after mishandling.

Morphological and functional outcome after brilliant blue G-assisted macular hole surgery.

PURPOSE OF THE STUDY: To investigate the outcome of brilliant blue G (BBG)-assisted internal limiting membrane peeling in macular hole surgery after a follow-up of 6 months. PROCEDURES: In this retrospective study 20 eyes have been treated with BBG-assisted macular hole surgery. After a follow-up of 6 months, the functional and anatomical outcomes have been analyzed. RESULTS: The mean preoperative best-corrected visual acuity (BCVA) was 0.7 logMAR units (mean ± SD 0.66 ± 0.27). After 3 months, the mean BCVA increased not significantly to 0.4 (0.54 ± 0.30), but a significant improvement to 0.2 logMAR units (0.28 ± 0.23) could be detected after 6 months compared to baseline (p < 0.01). Primary macular hole closure after a single surgery was found in 17 of 20 eyes. CONCLUSION AND MESSAGE: BBG exhibits sufficient staining qualities and safety profile leading to a significant functional improvement after successful macular hole surgery.

Ranibizumab as adjuvant in the treatment of rubeosis iridis and neovascular glaucoma--results from a prospective interventional case series.

BACKGROUND: To evaluate the capability of adjuvant intraocular ranibizumab (Lucentis) injections in the treatment of rubeosis and intraocular pressure in patients with rubeosis and neovascular glaucoma. METHODS: Ten eyes with rubeosis (R) and ten eyes with neovascular glaucoma (NVG) received Lucentis injections (ranibizumab 0.5 mg/0.05 ml) in this prospective, monocenter, 12-months, interventional case series. The primary efficacy outcome measure was the change of degree of iris rubeosis as documented by iris fluorescein angiography measured after 12 months. Secondary outcomes were intraocular pressure (IOP), best-corrected visual acuity (BCVA, logMAR), numbers of additional interventions or antiglaucoma medications administered after injection, the gonioscopic status of the anterior chamber angle, and central retinal thickness. RESULTS: In the R group, 3.6 injections and in the NVG group 2.3 injections of Lucentis were administered. Additional treatments were photocoagulation (n = 19), cyclodestructive procedures (n = 9), cryopexy (n = 3), and vitrectomy (n = 1). The mean stage of rubeosis was 3.4 ± 0.7 in the R group and 3.6 ± 0.8 in the NVG group at baseline. At month 12, the rubeosis was almost resolved in the R group (0.1 ± 0.3, p < 0.001), and decreased significantly in the NVG group (0.7 ± 1.1, p < 0.001). In the NVG subgroup, mean IOP was 41.4 ± 13.4 mmHg at baseline, which decreased rapidly (18.2 ± 12.3, day-14, p = 0.005) and stabilized during the follow-up (15.6 ± 2.0 mmHg, p < 0.05). BCVA improved significantly in both groups (p < 0.05, at month 12). CONCLUSIONS: Injection of 0.5 mg ranibizumab appears to be beneficial as an adjuvant treatment in neovascular glaucoma and rubeosis due to its anti-angiogenic properties and its ability to prevent establishment or progression of anterior chamber angle obstruction. Conventional therapeutic procedures addressing the retinal ischemia are still required in a stage-wise treatment approach.

Effect of high-intensity irradiation from dental photopolymerization on the isolated and superfused vertebrate retina.

BACKGROUND: Light or electromagnetic radiation may damage the neurosensory retina during irradiation of photopolymerizing resinous materials. Direct and indirect effects of irradiation emitted from polymerisation curing light may represent a severe risk factor for the eyes and the skin of the lamp operators, as well as for the patient's oral mucosa. METHODS: Bovine superfused retinas were used to record their light-evoked electroretinogram (ERG) as ex vivo ERGs. Both the a- and the b-waves were used as indicators for retinal damage on the functional level. The isolated retinas were routinely superfused with a standard nutrient solution under normoglycemic conditions (5 mM D-glucose). The change in the a- and b-wave amplitude and implicit time, caused by low and high intensity irradiation, was calculated and followed over time. RESULTS: From the results, it can be deduced that the irradiation from LED high-power lamps affects severely the normal physiological function of the bovine retina. Irradiations of 1,200 lx irreversibly damaged the physiological response. In part, this may be reversible at lower intensities, but curing without using the appropriate filter will bleach the retinal rhodopsin to a large extent within 20 to 40 s of standard application times. CONCLUSION: Constant exposure to intense ambient irradiation affects phototransduction (a-wave) as well as transretinal signalling. The proper use of the UV- and blue-light filtering device is highly recommended, and may prevent acute and long lasting damage of the neurosensory retina.

The novel use of decorin in prevention of the development of proliferative vitreoretinopathy (PVR).

BACKGROUND: The cytokine transforming growth factor-ß (TGF-ß) is a pivotal contributor to tissue fibrosis and a key cytokine in the pathogenesis of cellular transdifferentiation, epithelial-mesenchymal transition (EMT), and cell adhesion. This study evaluates the effect of decorin, a naturally occurring TGF-ß inhibitor, in an experimental rabbit model for proliferative vitreoretinopathy (PVR). METHODS: Traumatic PVR was induced in 50 rabbits divided into ten groups (n = 5). One group (GI) reveals a control with no treatment after trauma. Groups (GII-GIV) consisted of subgroups receiving phacovitrectomy at three different time points; (a) at the time of trauma, (b) 1 week following trauma, and (c) 2 weeks following trauma. GIII and GIV received 100 µg or 200 µg decorin, respectively. PVR severity was scored from 0 to 4. The amount of fibrosis was quantified using JMicroVision© software. RESULTS: The control group GI developed severe PVR with tractional retinal detachment (TRD); (PVR score ≥2) in four rabbits out of five. Vitrectomy had a positive effect (p < 0.05) on PVR development when preformed immediately, however the developed fibrosis was high. The best results were obtained when surgery was used in conjunction with decorin that reduced both the PVR score and fibrosis development significantly (p < 0.05). Depending on dosage and time of vitrectomy, PVR could be completely avoided (PVR score = 0) in 16 rabbits out of 30. TRD was prevented in 13 rabbits out of 15 in GIII to 14 rabbits out of 15 in GIV. In decorin-treated eyes, vitrectomy outcome was best when preformed at 1 week after trauma. There were no drug-related toxic effects evident on clinical and histopathological examination. CONCLUSIONS: In conclusion, in this rabbit model of PVR, adjuvant decorin application during vitrectomy effectively reduces fibrosis and TRD development. In conjunction with no obvious histopathological toxicity signs, decorin represents a promising substance to inhibit PVR reactions.

Ca(v)2.3 Ca2+ channel interacts with the G1-subunit of V-ATPase.

BACKGROUND: Calcium channels are essential in coupling action potential to signal transduction in cells. There are several types of calcium channels, which can be pharmacologically classified as L-, N-, P/Q-, R- and T-type. But molecular basis of R-type channels is less clearly understood compared the other channel types. Therefore the current study aims at understanding the molecular functions of R-type calcium channels by identifying interaction partners of the channel. METHODS: In order to do so, a yeast two hybrid (Y2H) screen, with carboxy terminus of α1 subunit of the channel, as the bait, was performed. G1 subunit of v-ATPase was identified as a putative interaction partner of human Ca(v)2.3 by using the Y2H screening. The interaction was confirmed by immunoprecipitation. To study the functional importance of the interaction, bafilomycin A(1), a potent and specific inhibitor of v-ATPase was used in patch-clamp recordings in Ca(v)2.3 stably-transfected HEK-293 cells (2C6) as well as in electroretinography of the isolated bovine retina expressing R-type Ca(2+) channels. RESULTS: G1 subunit of v-ATPase interacts with C-terminal tail of Ca(v)2.3 and bafilomycin A(1) reduces Ca(v)2.3 mediated calcium currents. Additionally peak I(Ca) is inhibited in retinal signal transduction when recorded as ERG b-wave. CONCLUSIONS: The results suggest that v-ATPase interacts physically and also functionally with Ca(v)2.3. This is the first demonstration of association of Ca(v)2.3 C-terminus with a protein complex which is involved in transmembrane signalling.

Expression of c-Kit and its ligand SCF in primary uveal melanoma.

PURPOSE: To evaluate the concurrent rate of expression of c-Kit and its ligand stem cell factor (SCF) in uveal melanoma in respect to the further clinical course and the correlation of these markers. METHODS: Paraffin sections from 35 primary uveal melanomas were evaluated immunohistochemically for the expression of c-Kit and SCF. Sixteen cases developed systemic metastasis during the follow-up (median: 5 years after diagnosis). The patients who did not develop metastasis (n = 19) had a mean follow-up of 10.6 (9-13) years. Radiation was performed in 6 patients. RESULTS: c-Kit and SCF were expressed in all patients who did or did not develop metastasis in the further clinical course. A mean SCF expression of 77.2% (range 52.7%-97.5%) of tumors that did not develop systemic metastasis and 30.1% (range 2.9%-61.5%) of tumors with systemic metastasis were evident. Uveal melanomas revealing an increased SCF expression were found to develop no metastasis more frequently (p<0.0001; hazard ratio (HR) = 0.963, 95% confidence interval (CI) 0.945-0.981). A mean c-Kit expression of 58.01% (range 5.9%-97.9%) in the group who did not develop metastasis and 48.9% (range 5.0%-95.9%) in the group with systemic metastasis were observed. c-Kit expression was not associated with increased rates of metastasis formation (p = 0.7329; HR = 0.997, 95% CI 0.982-1.013). The correlation between SCF and c-Kit is weak (0.39; 95% CI 0.06-0.63). CONCLUSIONS: c-Kit expression was not found to be associated with metastasis formation. A high SCF expression of primary choroidal melanomas was significantly associated with a lower incidence of systemic metastasis, which indicated SCF as a benign prognostic factor in the further clinical course.

The effect of adjuvant dimethylenastron, a mitotic Kinesin Eg5 inhibitor, in experimental glaucoma filtration surgery.

BACKGROUND: The aim of the study was to analyze the effect of Dimethylenaston, a mitotic kinesin 5 (Eg5) inhibitor, in an experimental setting of glaucoma filtration surgery. METHODS: On 37 chinchilla rabbits (ChBBCH), glaucoma filtration surgery similar to clinical practice, was performed. The animals received either no adjuvant, one unilateral subconjunctival injection of Dimethylenastron (1.0 µmol, 3.0 µmol), or the vehicle alone at baseline and in two further groups additionally at days 3 and 7 thereafter (1.0 µmol, 3.0 µmol). The evaluation of antifibrotic efficacy was performed by clinical response, histological examination, and immunohistochemical staining for smooth muscle actin (SMA) and CD 31. The animals were sacrificed on day 14, and the eyes processed for histology. RESULTS: The vehicle was well tolerated. Except for two cases of transient fibrinous reaction after the injection of 3.0 µmol Dimethylenastron, no adverse effects, such as inflammation or blurring of the optical media, were observed. A bleb scarring occurred in the group that received surgery only, adjuvant DMSO, or Dimethylenastron 3.0 µmol. Dimethylenastron (1.0 µmol) induced a milder scarring compared with the control group but the length of bleb survival was not significantly prolonged (p = 0.053, Kaplan-Meier log rank test). In all groups, the intraocular pressure correlated with the fibrotic process and reached normal levels within 14 days after surgery. Those groups injected with 1.0 µmol Dimethylenastron revealed a significantly reduced ratio of intraocular pressure and a milder, but not sufficiently reduced, subconjunctival fibrotic reaction according to the histological and immunohistochemical analysis. CONCLUSIONS: The subconjunctival administration of Dimethylenastron 1.0 µmol induced a milder conjunctival scarring. The applied concentrations of Dimethylenastron did not improve the surgical outcome of glaucoma filtration treatments in rabbits sufficiently.

Effects of pegaptanib sodium on retinal function in isolated perfused vertebrate retina.

PURPOSE: To evaluate the short-term toxic effects of pegaptanib sodium on retinal function. At present, intraocular anti-vascular endothelial growth factor (VEGF) therapy is the primary choice of treatment for neovascular maculopathy. The isoform VEGF(165) is specifically inhibited by pegaptanib sodium. Therefore, since VEGF(165) has neuroprotective effects against apoptosis of neuronal cells, blockage of VEGF(165) by pegaptanib could induce retinal dysfunction. In the present study, we used an electrophysiological technique for testing retinal toxicity in order to evaluate the short-term toxic effects of pegaptanib sodium on retinal function in a model of isolated perfused vertebrate retina. METHODS: Isolated bovine retinas were perfused with an oxygenated, pre-incubated nutrient solution. Electroretinograms (ERGs) were recorded as trans-retinal potentials using Ag/AgCl electrodes. Pegaptanib sodium (0.006, 0.06, or 0.2 mg/ml) and solvent carrier were added to the nutrient solution for 45 min. ERGs were monitored before, during, and after exposure. RESULTS: No significant reductions of b-wave (p = 0.357, p = 0.31, and p = 0.11, respectively) or a-wave (p = 0.189, p = 0.46, and p = 0.23, respectively) amplitudes were detected during application of pegaptanib (0.006, 0.06, or 0.2 mg/ml). The solvent carrier alone had no effect on ERG b- or a-waves (p = 0.98 and p = 0.42, respectively). During washout, ERG amplitudes of all test series remained unchanged. CONCLUSION: Results suggest that both pegaptanib sodium and its solvent carrier have good safety profiles. Intraocular application of 0.3 mg pegaptanib sodium induced no significant changes in ERGs in our ex vivo model and, thus, appears to be safe.

The safety profile of alkylphosphocholines in the model of the isolated perfused vertebrate retina.

BACKGROUND: Alkylphosphocholines (APCs) are synthetic phospholipid derivatives, and have been demonstrated to inhibit ocular cell proliferation in vitro and in vivo. Currently, they are applied clinically for their antitumoral and antiparasitic properties, but have not yet been implemented for clinical use in proliferative ophthalmic disorders. The purpose of this study was to assess the safety of APC in the ex vivo model of the isolated perfused vertebrate retina. METHODS: Bovine retina preparations were perfused with an oxygen pre-equilibrated standard solution. The electroretinogram (ERG) was recorded using Ag/AgCl-electrodes. After recording stable b-wave amplitudes, an APC was applied at the following concentrations to the nutrient solution: 0.25 microM, 2.5 microM and 25 microM. To investigate the effects of APC on photoreceptor function, a test series at the same concentrations was performed to evaluate the effects of APC on the a-wave amplitude. Aspartate at a concentration of 1 mM was added to the nutrient solution to obtain stable a-wave amplitudes. Thereafter, APC was applied at the same concentrations to the nutrient solution. The recovery of the ERG amplitudes was followed up for 75 minutes. RESULTS: No reduction of the a- and b-wave amplitude was found at the end of the exposure time with APC added in each test series. No differences were found between the ERG amplitudes before and after application of APC at the end of the washout. CONCLUSIONS: In the ex vivo model of the isolated perfused vertebrate retina, APC has proved to be a safe compound in the concentrations applied. Thus, APCs should further be considered as promising candidates for future clinical applications in ophthalmology.

Intravitreal bevacizumab treatment of macular edema in central retinal vein occlusion: one-year results.

PURPOSE: To report on the anatomic and visual acuity response after intravitreal bevacizumab injection in patients with macular edema due to non-ischemic central retinal vein occlusion (CRVO). METHODS: In a retrospective study, 21 consecutive patients (21 eyes) with non-ischemic CRVO underwent, on average, 3.7 intravitreal bevacizumab injections (1.25 mg). Ophthalmic examination included best corrected visual acuity (BCVA) and central retinal thickness (CRT) at baseline and follow-up visits. Fluorescein angiography was performed at baseline and at follow-up visits if needed. Primary outcomes were change of BCVA and CRT. RESULTS: The follow-up period for all of the included patients was 12 months. The mean BCVA was unchanged at the 12-month examination (baseline: 20/160; 12 months: 20/160) (P = 0.771). The mean CRT decreased from 780 microm (standard deviation [SD] +/- 324 microm) at the baseline to a mean of 462 microm (SD +/- 248 microm) at 12 months (P < 0.05). CONCLUSION: Intravitreal bevacizumab resulted in a significant decrease in CRT without significant improvement of visual acuity in patients with non-ischemic CRVO after a follow-up of 12 months.

Bevacizumab as adjuvant for neovascular glaucoma.

PURPOSE: We aimed to evaluate the longterm effects of intraocular bevacizumab (Avastin) injections as adjuvant treatment in patients with neovascular glaucoma. METHODS: Twenty eyes of 18 consecutive patients with secondary neovascular glaucoma caused by proliferative diabetic retinopathy (n = 7), ischaemic central retinal vein occlusion (n = 7), ischaemic ophthalmopathy (n = 2) and retinal ischaemia resulting from persistent detachment (n = 2) were treated with intraocular bevacizumab injections (1.25 mg/0.05 ml) in addition to other treatments. The main outcome measure was the change in degree of iris rubeosis. Secondary outcomes included intraocular pressure (IOP), best corrected visual acuity (BCVA) and numbers of additional interventions or antiglaucoma medications administered after injection. RESULTS: Mean (+/- standard deviation) follow-up was 67.7 +/- 13.8 weeks (range 50-93 weeks). At the last follow-up, complete regression of rubeosis was detectable in five (20%) eyes, incomplete regression in seven (35%), stabilization in six (30%), and an increase in two (10%) eyes. Mean IOP was 26.0 +/- 8.9 mmHg at baseline and significantly decreased to 14.75 +/- 5.3 mmHg at the last follow-up visit (p = 0.000005). Mean baseline BCVA (logMAR [logarithm of the minimum angle of resolution] 1.43 +/- 0.89) was stabilized during the follow-up period (logMAR 1.5 +/- 0.98). Patients received an average of 2.75 injections. Additional treatments were laser photocoagulation in 13 (65%) eyes, cyclodestructive procedure in 14 (70%), cryopexy in six (30%), drainage procedures in two (10%), and vitrectomy in five (25%) eyes. CONCLUSIONS: Bevacizumab may be beneficial as adjuvant treatment in neovascular glaucoma because of its anti-angiogenic properties and its ability to prevent establishment or progression of angular obstruction. The causative disease inducing the angiogenic process requires treatment in all cases. Antiglaucoma treatment is needed in cases of persistent elevated IOP.