Three-Dimensional Analysis of Peeled Internal Limiting Membrane Using Focused Ion Beam/Scanning Electron Microscopy

PURPOSE: To reevaluate the effect of internal limiting membrane peeling during vitrectomy on the Müller cell damage, we examined the ultrastructure of the internal limiting membrane by using focused ion beam/scanning electron microscopy (FIB/SEM). METHODS: A total of 12 internal limiting membranes obtained during surgery in both the macular hole and the idiopathic epiretinal membrane groups were processed for observation by FIB/SEM. Three-dimensional structures of the internal limiting membrane were analyzed. RESULTS: The number of cell fragments in the macular hole group was 5.07 ± 1.03 per unit area of internal limiting membrane (100 μm(2)). The total volume of cell fragments was 3.54 ± 1.24 μm(3)/100 μm(2). In contrast, the number of cell fragments in the epiretinal membrane group was 12.85 ± 3.45/100 μm(2), and the total volume of cell fragments was 10.45 ± 2.77 μm(3)/100 μm(2). Data for both values were significantly higher than those observed in the macular hole group (P = 0.0024 and P = 0.0022, respectively, Mann-Whitney U test). No statistical difference was found for the mean volume of the cell fragment between the two groups. CONCLUSIONS: All of the internal limiting membrane examined in this study showed cell fragments on the retinal surface of the internal limiting membrane. As compared with macular hole, epiretinal membrane exhibited a higher number and total volume of cell fragments, indicating that internal limiting membrane peeling for epiretinal membrane might have a higher risk of causing inner retinal damage. TRANSLATIONAL RELEVANCE: FIB/SEM was a useful tool for three-dimensional quantitative analysis of the internal limiting membrane.