Tomasz Choragiewicz

Toxicity testing of the VEGF inhibitors bevacizumab, ranibizumab and pegaptanib in rats both with and without prior retinal ganglion cell damage

Purpose:  To evaluate the effects of intravitreally introduced vascular endothelial growth factor (VEGF) inhibitors in rat eyes with healthy retinal ganglion cells (RGC) and into others with N‐methyl‐D‐aspartate (NMDA)‐induced RGC damage.

In vivo toxicity study of rhodamine 6G in the rat retina.

PURPOSE To investigate the intraocular effect of rhodamine 6G (R6G) on retinal structures and function in an in vivo rat model and to develop an in vivo method for accurate evaluation of new dyes for intraocular surgery. METHODS R6G in physiologic saline solution (PSS) was injected into the vitreous of adult Brown Norway rats at concentrations of 0.0002%, 0.002%, 0.02%, 0.2%, and 0.5%. Control animals received only PSS. Retinal toxicity was assessed by retinal ganglion cell (RGC) counts, light microscopy 7 days later, photopic electroretinography (ERG), and measurement of scotopic sensitivity and recovery of dark adaptation 48 hours and 7 days after intravitreous injection. RESULTS R6G at concentrations of 0.2% and 0.5% led to a dose-dependent loss of RGC. The most significant loss occurred at 0.5%. Lower concentrations (0.0002%, 0.002%, and 0.02%) produced no statistically significant retinal ganglion cell loss. Analysis of the eyes by light microscopy showed no structural changes in the central retina, although injections of 0.5% R6G were followed by impressive degenerative changes adjacent to the injection sites. ERGs showed no effects of the highest R6G concentration on rods, kinetics of rhodopsin recovery after bleaching, or cone-driven responses. CONCLUSIONS R6G can be safely injected in doses of up to 0.02% in rats, but has a toxic effect on retinal ganglion cells at higher concentrations. Accumulation of R6G may be a problem at higher concentrations, particularly at the injection site.

Neuroprotective effects of tempol on retinal ganglion cells in a partial optic nerve crush rat model with and without iron load.

Iron overload can contribute to oxidative stress in many tissues. We studied the effects of pretreatment with iron dextran on RGC loss in a calibrated partial optic nerve crush (PONC) model in rats, along with the protection offered by tempol (4-hydroxy-2,2,6,6-tetramethylpiperidinyl-1-oxyl, a membrane-permeable superoxide dismutase mimetic and free-radical scavenger), in the same experimental paradigm. A total of 40 rats in 6 groups of 5-8 animals each underwent PONC in one eye and sham crush in the other. Animals were pretreated with a single iron dextran load 24 h prior to PONC, and treated with tempol 6 h before and then once daily after PONC. Control animals were treated with PBS. RGC were retrogradely labeled with a fluorescent marker; all data are expressed in percent of the RGC count in the respective sham-treated eye. Immunohistochemistry was performed to visualize 3-nitrotyrosine, a marker of nitroxidative stress. PONC without iron pretreatment resulted in the survival of only 31.4% of labeled RGC after 7 days. Even fewer RGC (12.7%) survived after PONC with iron pretreatment. However, tempol in doses of 20 mg/kg of body weight (BW) significantly attenuated this effect when given as described above; in the group without iron pretreatment the number of surviving RGC doubled from 31.4% to 62.1%. In the group with iron pretreatment the survival rate of RGC increased even more pronouncedly, from 12.7% without tempol to 46.2% with tempol. Tempol in doses of 1 mg/kg BW and 5 mg/kg BW showed no significant rescue of RGC. Immunostaining showed nitrotyrosine-positive RGCs in PONC but not in sham-treated eyes and an increase in positive cells after iron load. Tempol treatment reduced nitrotyrosine staining in both the iron and non-iron groups. Our results demonstrate that PONC results in significantly greater RGC damage when iron pretreatment is performed, and that the compound tempol may provide additional protection for RGC in cases of neuronal damage both with and without prior iron treatment.

Caspase inhibitors protect against NMDA-mediated retinal ganglion cell death

Background:  Apoptosis is a major mechanism of cell death in glutamate‐induced excitotoxicity and caspases as the executors of apoptosis play an important role in the development of various central nervous system and eye diseases. We studied the involvement of certain caspases in excitotoxic retinal ganglion cell death, which was experimentally induced in Brown Norway Rats by application of the glutamate receptor agonist N‐methyl‐D‐aspartate (NMDA).

In vivo toxicity testing of methyl blue and aniline blue as vital dyes for intraocular surgery.

Purpose: To investigate the biocompatibility of methyl blue and aniline blue as vital dyes for vitreoretinal surgery in an in vivo rat model and to evaluate the effect of these dyes on retinal structure and function. Methods: Adult Brown–Norway rats received intravitreal injections of 0.1%, 0.2%, and 2% methyl blue or aniline blue dissolved in balanced salt solution with balanced salt solution serving as a control. Retinal toxicity was assessed 7 days thereafter by means of retinal ganglion cell counts, light microscopy, and electroretinography. Results: No significant decrease in retinal ganglion cell counts at concentrations up to 0.2% was observed. At 2%, however, a significant retinal ganglion cell loss was detected with both dyes (more pronounced for aniline blue). Light microscopy showed no structural changes in the central retina for concentrations up to 0.2%. Electroretinographies detected no adverse effects of methyl blue or aniline blue on rod- or cone-driven responses at concentrations up to 0.2%. Conclusion: Methyl blue and aniline blue are very biocompatible and may, therefore, be usable for intraocular surgery. Further testing with other animal models will be necessary to confirm this. The safety margin of methyl blue is possibly higher than that of aniline blue.

Outcomes of Sutureless Iris-Claw Lens Implantation

Purpose. To evaluate the indications, refraction, and visual and safety outcomes of iris-claw intraocular lens implanted retropupillary with sutureless technique during primary or secondary operation. Methods. Retrospective study of case series. The Haigis formula was used to calculate intraocular lens power. In all cases the wound was closed without suturing. Results. The study comprised 47 eyes. The mean follow-up time was 15.9 months (SD 12.2). The mean preoperative CDVA was 0.25 (SD 0.21). The final mean CDVA was 0.46 (SD 0.27). No hypotony or need for wound suturing was observed postoperatively. Mean postoperative refractive error was −0.27 Dsph (−3.87 Dsph to +2.85 Dsph; median 0.0, SD 1.28). The mean postoperative astigmatism was −1.82 Dcyl (min −0.25, max −5.5; median −1.25, SD 1.07). Postoperative complications were observed in 10 eyes. The most common complication was ovalization of the iris, which was observed in 8 eyes. The mean operation time was 35.9 min (min 11 min, max 79 min; median 34, SD 15.4). Conclusion. Retropupilary iris-claw intraocular lens (IOL) implantation with sutureless wound closing is an easy and fast method, ensuring good refractive outcome and a low risk of complication. The Haigis formula proved to be predictable in postoperative refraction.

Surgical Management of Traumatic Retinal Detachment with Primary Vitrectomy in Adult Patients

Purpose. To evaluate functional and anatomical results of pars plana vitrectomy (PPV) in the retinal detachment (RD) followed by severe eye trauma. Methods. Retrospective analysis of medical records of forty-one consecutive patients treated with 23-gauge PPV due to traumatic RD. Age, gender, timing of PPV, visual acuity, and presence of intraocular foreign body (IOFB) and proliferative vitreoretinopathy (PVR) were included in the analysis. Results. Mean age of patients was 47 years; the majority of patients were men (88%). Closed globe injury was present in 21 eyes and open globe injury in 20 eyes (IOFB in 13 eyes, penetration injury in 4 eyes, and eye rupture in 3 eyes). Mean follow-up period was 14 months; mean timing of PPV was 67 days. Twenty-seven (66%) eyes had a functional success; 32 eyes (78%) had anatomical success. As a tamponade silicone oil was used in 33 cases and SF6 gas in 8 cases. Conclusions. Severe eye injuries are potentially devastating for vision, but vitreoretinal surgery can improve anatomical and functional outcomes. Among analysed pre- and intra- and postoperative factors, absence of PVR, postoperative retinal attachment, and silicone oil as a tamponade were related to significantly improved visual acuity.

Presence and distribution of L-kynurenine aminotransferases immunoreactivity in human cataractous lenses.

Purpose:  To investigate the presence and distribution of l‐kynurenine aminotransferases immunoreactivity in human and animal lenses during cataract formation.

Intraoperative Macula Protection by Perfluorocarbon Liquid for the Metallic Intraocular Foreign Body Removal during 23-Gauge Vitrectomy

Purpose. To evaluate visual and safety outcomes of 23-gauge (G) pars plana vitrectomy (PPV) with application of perfluorocarbon liquid (PFCL) for intraoperative protection of the macula during intraocular foreign body (IOFB) removal. Methods. Retrospective study of 42 patients who underwent 23 G PPV for IOFB removal from posterior segment with intraoperative PFCL application for the macula shielding. Collected data included corrected distance visual acuity (CDVA), size of IOFB, and complication rate. The mean follow-up period was 12 months. Results. The mean preoperative CDVA was 0.54 logMAR (SD 0.46), and the final mean CDVA was 0.68 logMAR (SD 0.66). All IOFBs were metallic with mean dimensions of 4.6 mm × 2.1 mm. Twenty-two IOFBs were removed through the corneal tunnel and 20 IOFBs through the sclerotomy. No intraoperative iatrogenic lesion of the macula was observed. As a tamponade, silicon oil was applied in 31 eyes, SF6 gas in 5 eyes, air in 4 eyes, and 2 eyes required no tamponade. Secondary retinal detachment was observed in 17% of cases, but at the end of the follow-up, all the retinas were attached. Conclusion. PFCL application during PPV is a safe method of protecting the macula from unexpected falling of the metallic IOFB during its removal.

Ketogenic Diet Attenuates NMDA-Induced Damage to Rat's Retinal Ganglion Cells in an Age-Dependent Manner

Objective: This study was conducted to investigate neuroprotective effects of a high fat/low carbohydrate and protein diet (ketogenic diet, KD) in a model of N-methyl D-aspartate (NMDA)-induced retinal ganglion cell (RGC) damage in juvenile and young adult rats. Methods: Juvenile (30-35 days old) and young adult (56-70 days old) female Brown Norway rats were fed the KD for 21 days; rats exposed to a standard rodent diet (SRD) served as controls. The main constituents of the KD used in the present study were approximately 80% fats, 8% proteins, and less than 1% carbohydrates. On day 14 of exposure to the KD (or the SRD in the control group), each rat received a single intravitreal injection of NMDA; RGCs were then retrogradely labelled by hydroxystilbamidine on day 19 and collected on day 21 to assess the degree of damage induced by NMDA. Blood biomarkers to confirm the expected metabolic response to the KD (i.e. ketosis and hypoglycaemia) were also assessed. Results: Although both the juvenile and young adult rats developed comparable ketosis and hypoglycaemia when fed the KD, NMDA-induced loss in RGCs was significantly attenuated only in juvenile rats exposed to the KD in comparison with those fed the SRD; exposure to the KD had no protective effect in young adult rats. In summary, exposure to the KD had a neuroprotective effect in NMDA-induced RGC damage in juvenile rats, but not in young adult rats. Conclusion: These results support further exploration of metabolic interventions to treat optic neuropathies associated with neurodegeneration.