Sequential epiretinal membrane removal with internal limiting membrane peeling in brilliant blue G-assisted macular surgery

Purpose To assess the selectivity of brilliant blue G (BBG) staining by analysing the morphological components of unstained and stained tissue obtained during epiretinal membrane (ERM) removal with internal limiting membrane (ILM) peeling in BBG-assisted macular surgery. Methods Twenty-six surgical specimens were removed from 13 eyes with epiretinal gliosis during vitrectomy using BBG for ERM and ILM peeling. We included eyes with idiopathic macular pucker, idiopathic macular hole and vitreomacular traction syndrome. The dye was injected into the fluid-filled globe. Unstained and stained epiretinal tissue was harvested consecutively and placed into separate containers. All specimens were processed for conventional transmission electron microscopy. Results The first surgical specimen of all eyes showed no intraoperative staining with BBG and corresponded to masses of cells and collagen. The second surgical specimen demonstrated good staining characteristics and corresponded to the ILM in all patients included. In seven eyes, the ILM specimens were seen with minor cell proliferations such as single cells or a monolayer of cells. Myofibroblasts, fibroblasts and astrocytes were present. In five cases, native vitreous collagen fibrils were found at the ILM. In six of the eyes, ILM specimens were blank. Conclusion Our clinicopathological correlation underlines the selective staining properties of BBG. The residual ILM is selectively stained by BBG even when a small amount of cells and collagen adheres to its vitreal side. To reduce the retinal exposure to the dye, the surgeon might choose to remove the ERM without using the dye, followed by a BBG injection to identify residual ILM

Quality of life in the follow-up of uveal melanoma patients after CyberKnife treatment

To assess quality of life in uveal melanoma patients within the first and second year after CyberKnife radiosurgery. Overall, 91 uveal melanoma patients were evaluated for quality of life through the Short-form (SF-12) Health Survey at baseline and at every follow-up visit over 2 years after CyberKnife radiosurgery. Statistical analysis was carried out using SF Health Outcomes Scoring Software and included subgroup analysis of patients developing secondary glaucoma and of patients maintaining a best corrected visual acuity (BCVA) of the treated eye of 0.5 log(MAR) or better. Analysis of variance, Greenhouse-Geisser correction, Student's t-test, and Fisher's exact test were used to determine statistical significance. Physical Functioning (PF) and Role Physical (RP) showed a significant decrease after CyberKnife radiosurgery, whereas Mental Health (MH) improved (P=0.007, P<0.0001 and P=0.023). MH and Social Functioning (SF) increased significantly (P=0.0003 and 0.026) in the no glaucoma group, MH being higher compared with glaucoma patients (P=0.02). PF and RP were significantly higher in patients with higher BCVA at the second follow-up (P=0.02). RP decreased in patients with BCVA<0.5 log(MAR) (P=0.013). Vitality (VT) increased significantly in patients whose BCVA could be preserved (P=0.031). Neither tumor localization nor size influenced the development of secondary glaucoma or change in BCVA. Although PF and RP decreased over time, MH improved continuously. Prevention of secondary glaucoma has a significant influence on both SF and MH, whereas preservation of BCVA affects VT. Emotional stability throughout follow-up contributes positively toward overall quality of life. CyberKnife radiosurgery may contribute to attenuation of emotional distress in uveal melanoma patients

Viability of Primary Human Pigment Epithelium Cells and Muller-Glia Cells after Intravitreal Ziv-Aflibercept and Aflibercept

Purpose: The aim of this study was to access the safety profiles of 2 fusion proteins with anti-vascular endothelial growth factor action (ziv-aflibercept and aflibercept) on retinal pigment epithelium cells and Muller-Glia cells in culture by assessing cell viability post drug exposure. Methods: Primary human retinal pigment epithelium cells (pRPE) and Muller-Glia cells (Mio-M1) were exposed to the clinical standardized concentrations of ziv-aflibercept (25 mg/mL) and aflibercept (40 mg/mL). Progressively higher concentrations of NaCI (300, 500, 1,000, 1,500, 2,000, 5,000, and 10,000 mosm/kg) were also applied to cells to assess the possibility of potentiating hyperosmotic cytotoxity effect. The study was applied to measure pRPE and Mio-M1 viability by a tetrazolium dye-reduction assay (XTT). Results: Cell viability of both pRPE and Mio-M1 presented no significant changes after exposure of ziv-aflibercept and aflibercept. Progressive NaCI concentrations did not significantly alter cell viability. The exposure to the negative control of 75 mu L/mL of dimethyl sulfoxide showed significant reduction in cell viability. Conclusions: At clinical doses, neither ziv-aflibercept nor aflibercept caused any significant reduction in cell viability in vitro. Furthermore, injection solutions of NaCI with higher osmolality caused no significant reduction in cell viability. (C) 2017 S. Karger AG, Base

Pharmacokinetics and biodistribution of Erufosine in nude mice - implications for combination with radiotherapy

<p>Abstract</p> <p>Background</p> <p>Alkylphosphocholines represent promising antineoplastic drugs that induce cell death in tumor cells by primary interaction with the cell membrane. Recently we could show that a combination of radiotherapy with Erufosine, a paradigmatic intravenously applicable alkylphosphocholine, <it>in vitro </it>leads to a clear increase of irradiation-induced cell death. In view of a possible combination of Erufosine and radiotherapy <it>in vivo </it>we determined the pharmacokinetics and bioavailability as well as the tolerability of Erufosine in nude mice.</p> <p>Methods</p> <p>NMRI (nu/nu) nude mice were treated by intraperitoneal or subcutaneous injections of 5 to 40 mg/kg body weight Erufosine every 48 h for one to three weeks. Erufosine-concentrations were measured in brain, lungs, liver, small intestine, colon, spleen, kidney, stomach, adipoid tissue, and muscle by tandem-mass spectroscopy. Weight course, blood cell count and clinical chemistry were analyzed to evaluate general toxicity.</p> <p>Results</p> <p>Intraperitoneal injections were generally well tolerated in all dose groups but led to a transient loss of the bodyweight (<10%) in a dose dependent manner. Subcutaneous injections of high-dose Erufosine caused local reactions at the injection site. Therefore, this regimen at 40 mg/kg body weight Erufosine was stopped after 14 days. No gross changes were observed in organ weight, clinical chemistry and white blood cell count in treated compared to untreated controls except for a moderate increase in lactate dehydrogenase and aspartate-aminotransferase after intensive treatment. Repeated Erufosine injections resulted in drug-accumulation in different organs with maximum concentrations of about 1000 nmol/g in spleen, kidney and lungs.</p> <p>Conclusion</p> <p>Erufosine was well tolerated and organ-concentrations surpassed the cytotoxic drug concentrations <it>in vitro</it>. Our investigations establish the basis for a future efficacy testing of Erufosine in xenograft tumor models in nude mice alone and in combination with chemo- or radiotherapy.</p

Minocycline is cytoprotective in human trabecular meshwork cells and optic nerve head astrocytes by increasing expression of XIAP, survivin, and Bcl-2

Marcus Kernt, Aljoscha S Neubauer, Kirsten H Eibl, Armin Wolf, Michael W Ulbig, Anselm Kampik, Cristoph HirneissDepartment of Ophthalmology, Ludwig-Maximilians-University, Munich, GermanyIntroduction: Primary open-angle glaucoma (POAG) is one of the leading causes of blindness. Activation of optic nerve head astrocytes (ONHA) and loss of trabecular meshwork cells (TMC) are pathognomonic for this neurodegenerative disease. Oxidative stress and elevated levels of transforming growth factor beta (TGF&amp;beta;) play an important role in the pathogenesis of POAG. This study investigates the possible antiapoptotic and cytoprotective effects of minocycline on TMC and ONHA under oxidative stress and increased TGF&amp;beta; levels.Methods: TMC and ONHA were treated with minocycline 1&amp;ndash;150 &amp;mu;M. Possible toxic effects and IC50 were evaluated after 48 hours. Cell proliferation and viability were examined in order to assess the protective effects of minocycline on TMC and ONHA. Expression of Bcl-2, XIAP, and survivin, as well as their mRNA expression, were assessed by real time polymerase chain reaction (RT-PCR) and Western Blot analysis 48 hours after treatment with minocycline alone and additional incubation with TGF&amp;beta;-2 or oxidative stress.Results: Minocycline 1&amp;ndash;75 &amp;mu;M showed no toxic effects on TMC and ONHA. Under conditions of oxidative stress, both TMC and ONHA showed an increase in viability and an ability to proliferate when treated with minocycline 20&amp;ndash;40 &amp;mu;M. RT-PCR and Western blotting yielded an overexpression of Bcl-2, XIAP, and survivin when TMC or ONHA were treated with minocycline 20&amp;ndash;40 &amp;mu;M under conditions of oxidative stress and when additionally incubated with TGF&amp;beta;-2.Conclusion: Minocycline up to 75 &amp;mu;M does not have toxic effects on TMC and ONHA. Treatment with minocycline 20&amp;ndash;40 &amp;mu;M led to increased viability and proliferation under oxidative stress and TGF&amp;beta;-2, as well as overexpression of Bcl-2, XIAP, and survivin. This protective pathway may help to prevent apoptotic cell death of TMC and ONHA and therefore be a promising approach to avoidance of progression of glaucomatous degeneration.Keywords: glaucoma, apoptosis, minocycline, trabecular meshwork, optic nerve head astrocyte

Alkylphosphocholines: a new therapeutic option in glaucoma filtration surgery

PURPOSE: To investigate the effect of alkylphosphocholines (APCs) on human Tenon fibroblast (HTF) proliferation, migration, and cell-mediated collagen gel contraction. METHODS: HTFs were isolated from tissue samples of three patients obtained during surgery and cultured in DMEM and 10% fetal calf serum (FCS). HTFs (passage 3-6) were treated with one APC in different concentrations spanning the 50% inhibitory concentration (IC(50)), as determined previously. Inhibition of cell proliferation was assessed by the tetrazolium dye reduction assay. Migration was determined in chemoattractant chambers with fibronectin-coated polycarbonated membranes. For inhibition of contraction, three-dimensional collagen gels were seeded with HTFs, and the gel size was measured. Cell viability was determined by the trypan blue exclusion assay. For analysis of the mechanism of action, protein kinase C (PKC) activity was measured. RESULTS: The IC(50) varied between 7.0 and 10.5 microM for all APCs tested. At this concentration, all four APCs inhibited HTF migration and cell-mediated collagen gel contraction in the presence of serum. The inhibitory effects on HTF proliferation, migration, and contraction were observed at nontoxic concentrations. PKC activity was reduced to 50% of control level at the IC(50) of all APCs applied. CONCLUSIONS: APCs are effective inhibitors of HTF proliferation, migration, and cell-mediated contraction of collagen gels at nontoxic concentrations. Their mechanism of action seems to involve the inhibition of the PKC pathway

Quality of life in the follow-up of uveal melanoma patients after CyberKnife treatment

To assess quality of life in uveal melanoma patients within the first and second year after CyberKnife radiosurgery. Overall, 91 uveal melanoma patients were evaluated for quality of life through the Short-form (SF-12) Health Survey at baseline and at every follow-up visit over 2 years after CyberKnife radiosurgery. Statistical analysis was carried out using SF Health Outcomes Scoring Software and included subgroup analysis of patients developing secondary glaucoma and of patients maintaining a best corrected visual acuity (BCVA) of the treated eye of 0.5 log(MAR) or better. Analysis of variance, Greenhouse-Geisser correction, Student's t-test, and Fisher's exact test were used to determine statistical significance. Physical Functioning (PF) and Role Physical (RP) showed a significant decrease after CyberKnife radiosurgery, whereas Mental Health (MH) improved (P=0.007, P<0.0001 and P=0.023). MH and Social Functioning (SF) increased significantly (P=0.0003 and 0.026) in the no glaucoma group, MH being higher compared with glaucoma patients (P=0.02). PF and RP were significantly higher in patients with higher BCVA at the second follow-up (P=0.02). RP decreased in patients with BCVA<0.5 log(MAR) (P=0.013). Vitality (VT) increased significantly in patients whose BCVA could be preserved (P=0.031). Neither tumor localization nor size influenced the development of secondary glaucoma or change in BCVA. Although PF and RP decreased over time, MH improved continuously. Prevention of secondary glaucoma has a significant influence on both SF and MH, whereas preservation of BCVA affects VT. Emotional stability throughout follow-up contributes positively toward overall quality of life. CyberKnife radiosurgery may contribute to attenuation of emotional distress in uveal melanoma patients

Alkylphosphocholines inhibit proliferation of human retinal pigment epithelial cells

PURPOSE: To investigate the effect and mechanism of action of alkylphosphocholines (APCs) on proliferation of human retinal pigment epithelium (RPE) cells and RPE-mediated collagen matrix contraction in vitro. METHODS: Cultured RPE cells of five human donors were treated with four APCs in the presence of fetal calf serum. Proliferation was assessed by the tetrazolium dye-reduction (MTT) assay and by counting the number of cells dividing in culture. The effect of APCs on RPE-mediated matrix contraction was determined in three-dimensional collagen gels. Cell viability was tested by the trypan blue exclusion assay. As a possible mechanism of APC action, protein kinase C (PKC) activity was quantified by scintillation counting of (32)P-labeled phosphate transferred to a PKC-specific substrate. RESULTS: All APCs inhibited RPE proliferation and RPE-mediated collagen matrix contraction in a dose-dependent manner in vitro. The antiproliferative and anticontractile effect of APCs increased with elongation of the fatty acid chain beyond C20. IC(50)s of all APCs varied between 8.5 micro M (erucyl-phosphocholine, C22:1-PC), 9.0 micro M (Z)-12-heneicosenyl-phosphocholine, C21:1-PC), 11.0 micro M (Z)-10-eicosenyl-phosphocholine, C20:1-PC), and 26.5 micro M (oleyl-phosphocholine, C18:1-PC). Trypan blue staining revealed a toxicity below 5% for all APCs within the concentration interval tested. PKC activity was significantly reduced by all four APCs, with C22:1-PC being the most effective. CONCLUSIONS: APCs inhibit proliferation of RPE cells and RPE-mediated matrix contraction in vitro at nontoxic concentrations. This effect seems to be exerted through inhibition of PKC activity. Therefore, APCs are promising candidates for treatment of RPE-mediated proliferative processes such as proliferative vitreoretinopathy