RESUMO
BACKGROUND: The development of continuous curvilinear capsulorrhexis (CCC) has contributed significantly to the tolerability and effectiveness of cataract extraction and intraocular lens implantation. Staining of the anterior capsule has become a popular method of increasing visibility when performing CCC. OBJECTIVE: The aim of this study was to determine, using scanning electron microscopy (SEM) and transmission electron microscopy (TEM), the highest concentrations of trypan blue dye that would not cause long-term toxicity after injection into the anterior chamber of rat eyes. METHODS: The eyes of healthy female albino Wistar rats were used in this investigator-masked, controlled, 2-period, experimental study conducted over 12 weeks at the Dicle University Experimental Animal Laboratory, Diyarbakir, Turkey. The rats were randomly divided into 5 groups of 4 using a random-number table. Each rat was administered a 0.05-mL injection of trypan blue into the right eye in 1 of the following concentrations: 0.4%, 0.2%, 0.1%, 0.05%, or 0.025%. A 0.05-mL pH-balanced saline solution was injected into the left eye of each rat to act as a control. At 1 day after injection and 4 weeks after injection (early period), 1 rat from each concentration group was euthanized and their eyes were enucleated. At 12 weeks after injection (late period) the remaining 2 rats from each group were euthanized and their eyes were enucleated. Corneal endothelial cells and the anterior lens capsule of the enucleated eyes were analyzed using SEM and TEM, and the results were compared with those of the control group. In the TEM analysis, the primary end point was the histopathologic changes in the cellular organelles when compared with those in the control group. In the SEM analysis, the primary end point was changes in cell shape, intracellular junctions, and density of the microvilli when compared with those in the control group. RESULTS: Forty eyes from 20 albino Wistar rats (mean [SD] age, 8.2 [1.6] weeks; mean [SD] weight, 175.6 [16.5] g) were used in the study. Each group of rats received a different concentration of trypan blue in the right eye. In the early period, both the 0.4% and 0.2% concentrations were associated with the impairment of the hexagonal structure of corneal endothelial cells and intercellular junctions. Those concentrations were also associated with an increased occurrence of cellular vacuolation, cytoplasmic edema, extensive granulation of the endoplasmic reticulum, pyknotic nuclei, and mitochondrial degeneration. In the late period, these changes were observed as persisting in a decreasing manner. With the 0.1% and 0.05% concentrations, the density of microvilli decreased, nuclei appeared normal, granulation of the endoplasmic reticulum and Golgi apparatus was active, and minimal levels of mitochondrial degeneration were observed. CONCLUSIONS: In this small experimental study in rat eyes, trypan blue at concentrations >0.025% was associated with impaired morphology and structure of corneal endothelial cells after short-term exposure. This effect continued in a decreasing fashion after long-term exposure. No significant changes were noted in the control group or the group administered the 0.025% concentration.
RESUMO
PURPOSE: To evaluate the effects of intracameral injection of ranibizumab and bevacizumab on the corneal endothelium by scanning electron microscopy (SEM). METHODS: Twenty-eight female rabbits were randomly divided into four equal groups. Rabbits in groups 1 and 2 underwent intracameral injection of 1 mg/0.1 mL and 0.5 mg/0.05 mL ranibizumab, respectively; group 3 was injected with 1.25 mg/0.05 mL bevacizumab. All three groups were injected with a balanced salt solution (BSS) into the anterior chamber of the left (fellow) eye. None of the rabbits in group 4 underwent an injection. Corneal thickness and intraocular pressure were measured before the injections, on the first day, and in the first month after injection. The rabbits were sacrificed and corneal tissues were excised in the first month after injection. Specular microscopy was used for the corneal endothelial cell count. Endothelial cell density was assessed and comparisons drawn between the groups and the control. Micrographs were recorded for SEM examination. The structure of the corneal endothelial cells, the junctional area of the cell membrane, the distribution of microvillus, and the cell morphology of the eyes that underwent intracameral injection of vascular endothelial growth factor (VEGF), BSS, and the control group were compared. RESULTS: Corneal thickness and intraocular pressure were not significantly different between the groups that underwent anti-VEGF or BSS injection and the control group on the first day and in the first month of injection. The corneal endothelial cell count was significantly diminished in all three groups; predominantly in group 1 and 2 (P<0.05). The SEM examination revealed normal corneal endothelial histology in group 3 and the control group. Eyes in group 1 exhibited indistinctness of corneal endothelial cell borders, microvillus loss in the luminal surface, excessive blebbing, and disintegration of intercellular junctions. In group 2, the cell structure of the corneal endothelium and intercellular junctions were normal. However, a relative reduction was observed in the microvillus density of endothelial cells. Although eyes in group 3 were morphologically similar to fellow eyes and the control group, disarrangement in endothelial cell borders was evident. CONCLUSION: The SEM examination pointed out deterioration in endothelial cell morphology after intracameral injection of 1 and 0.5 mg ranizumab. However, the effects of intracameral bevacizumab injection on corneal endothelial cells were similar to those found in fellow eyes and the control group. Further large-scale studies that examine the cellular changes by transmission electron microscopy are required to support the results of the present study that evaluates the structural changes in endothelial cells by SEM.