PORES OF THE INTERNAL LIMITING MEMBRANE: A Common Finding in Vitreomaculopathies.

PURPOSE: To describe presence and distribution of pores of the inner limiting membrane (ILM) in eyes with vitreomaculopathies. METHODS: Inner limiting membrane specimens were harvested from 117 eyes of 117 patients during vitrectomy with membrane peeling from eyes with vitreomacular traction syndrome, idiopathic and secondary epiretinal gliosis, and idiopathic full-thickness macular hole. All specimens were processed as flat-mounts for immunocytochemistry and examined by phase-contrast, interference, and fluorescence microscopy. Demographic and clinical data were correlated. RESULTS: Inner limiting membrane pores were found in all vitreomaculopathies. They were identified in 47 (40.2%) of 117 eyes being most evident with antilaminin. In eyes with full-thickness macular hole >400 µ m, pores were seen in more than half of all eyes. They occur as numerous and uniformly distributed defects of the flat-mounted ILM with a mean diameter of 9.5 ± 2.4 µ m. Edges of ILM pores are round with an irregular contour and no specific cellular pattern. Pores were distinguished from retinal vessel thinning and iatrogenic artefacts. CONCLUSION: Contrary to previous reports, ILM pores are a common finding in vitreomaculopathies easily visible with antilaminin staining. Further studies are needed to clarify whether their presence correlates with differences in disease progression or imaging before and after vitrectomy with ILM peeling.

Scleral Thickness as a Risk Factor for Central Serous Chorioretinopathy and Pachychoroid Neovasculopathy.

In the pathophysiology of central serous chorioretinopathy (CSC), scleral changes inducing increased venous outflow resistance are hypothesized to be involved. This work aims to investigate anterior scleral thickness (AST) as a risk factor for pachychoroid disorders. A randomized prospective case-control study was performed at the Ludwig Maximilians University, Department of Ophthalmology. In patients with CSC or pachychoroid neovasculopathy (PNV) and in an age- and refraction-matched control group, swept source optical coherence tomography (SS-OCT) was used to measure anterior scleral thickness (AST). Subfoveal choroidal thickness (SFCT) was assessed using enhanced depth imaging OCT (EDI-OCT). In total, 46 eyes of 46 patients were included in this study, with 23 eyes in the CSC/PNV and 23 eyes in the control group. A significantly higher AST was found in the CSC/PNV compared with the control group (403.5 ± 68.6 (278 to 619) vs. 362.5 ± 62.6 (218 to 498) µm; p = 0.028). Moreover, the CSC/PNV group showed a higher SFCT (392.8 ± 92.8 (191-523) vs. 330.95 ± 116.5 (167-609) µm, p = 0.004). Compared with the age- and refraction-matched controls, patients with CSC and PNV showed a significantly thicker anterior sclera. Scleral thickness might contribute to the venous overload hypothesized to induce pachychoroid phenotypes.

Long-Term Results of Adjunct Autologous Platelet-Rich Plasma in Lamellar Macular Hole Surgery Showing Lasting Restoration of Foveal Anatomy.

The aim of this study was to evaluate the long-time results of highly concentrated autologous platelet-rich plasma (PRP) used as an adjunct in lamellar macular hole (LMH) surgery. Nineteen eyes of nineteen patients with progressive LMH were enrolled in this interventional case series, on which 23/25-gauge pars plana vitrectomy was performed and 0.1 mL of highly concentrated autologous platelet-rich plasma was applied under air tamponade. Posterior vitreous detachment was induced, and the peeling of tractive epiretinal membranes, whenever present, was performed. In cases of phakic lens status, combined surgery was carried out. Postoperatively, all patients were instructed to remain in a supine position for the first two postoperative hours. Best-corrected visual acuity (BCVA) testing, microperimetry, and spectral domain optical coherence tomography (SD-OCT) were carried out preoperatively and at minimum 6 months (in median 12 months) postoperatively. Foveal configuration was postoperatively restored in 19 of 19 patients. Two patients who had not undergone ILM peeling showed a recurring defect at 6-month follow-up. Best-corrected visual acuity improved significantly from 0.29 ± 0.08 to 0.14 ± 0.13 logMAR (p = 0.028, Wilcoxon signed-rank test). Microperimetry remained unchanged (23.38 ± 2.53 preoperatively; 23.0 ± 2.49 dB postoperatively; p = 0.67). No patients experienced vision loss after surgery, and no significant intra- or postoperative complications were observed. Using PRP as an adjunct in macular hole surgery significantly improves morphological and functional outcomes. Additionally, it might be an effective prophylaxis to further progression and also the formation of a secondary full-thickness macular hole. The results of this study might contribute to a paradigm shift in macular hole surgery towards early intervention.

Cell composition at the vitreomacular interface in traumatic macular holes.

PURPOSE: To describe characteristics of the vitreomacular interface (VMI) in traumatic macular holes (TMH) compared to idiopathic macular holes (IMH) using immunofluorescence and electron microscopy, and to correlate with clinical data. METHODS: For immunocytochemical and ultrastructural analyses, premacular tissue with internal limiting membrane (ILM) and epiretinal membrane (ERM) was harvested during vitrectomy from 5 eyes with TMH and 5 eyes with IMH. All specimens were processed as flat mounts for phase-contrast microscopy, interference and fluorescence microscopy, and transmission electron microscopy (TEM). Primary antibodies were used against microglial and macroglial cells. Clinical data was retrospectively evaluated. RESULTS: Surgically excised premacular tissue of eyes with TMH showed a less pronounced positive immunoreactivity for anti-glutamine synthetase, anti-vimentin and anti-IBA1 compared to eyes with IMH. Cell nuclei staining of the flat-mounted specimens as well as TEM presented a lower cell count in eyes with TMH compared to IMH. All detected cells were found on the vitreal side of the ILM. No collagen fibrils were seen in specimens of TMH. According to patients' age, intraoperative data as well as spectral-domain optical coherence tomography (SD-OCT) analysis revealed an attached posterior vitreous in the majority of TMH cases (60%), whereas all eyes with IMH presented posterior vitreous detachment. CONCLUSION: The vitreomacular interface in TMH and IMH shows significant differences. In TMH, glial cells are a rare finding on the vitreal side of the ILM.

Improving morphological outcome in lamellar macular hole surgery by using highly concentrated autologous platelet-rich plasma.

PURPOSE: To evaluate the use of highly concentrated autologous platelet-rich plasma (PRP) in lamellar macular hole (LMH) surgery with regard to function and morphology. METHODS: We included 12 eyes of 12 patients with progressive LMH in this interventional case series. After 23/25-gauge pars plana vitrectomy, 0.1ml highly concentrated autologous platelet-rich plasma was applied under air tamponade. Induction of posterior vitreous detachment and peeling of tractive epiretinal membranes were performed whenever present. Phacovitrectomy was undertaken in cases of phakic lens status. Postoperatively, all patients were instructed to rest in a supine position for the first two postoperative hours. Best-corrected visual acuity (BCVA) testing, microperimetry, spectral-domain optical coherence tomography (SD-OCT), and fundus photography were carried out preoperatively and 6 months postoperatively. RESULTS: Foveal configuration was restored in 10 of 12 patients (83.3%) at 6 months postoperatively. Two patients who had not undergone ILM peeling showed a recurring defect at 6-month follow-up. Best-corrected visual acuity improved significantly from 0.29 ± 0.08 to 0.14 ± 0.13 logMAR (Wilcoxon: p=0.028). Microperimetry remained unchanged (23.38 ± 2.53 preoperatively; 23.0 ± 2.49 dB postoperatively; p=0.67). No patient experienced vision loss after surgery, and no significant intra- or postoperative complications occurred. CONCLUSION: The application of PRP in the surgical therapy of LMH results in good morphological and functional outcomes. Additional peeling of the ILM seems to be mandatory when using PRP to prevent the recurrence of LMH. Strict postoperative supine positioning for 2 h avoids PRP dislocation. Larger sample sizes are needed to confirm the results.

Highly Concentrated Autologous Platelet-Rich Plasma Restores Foveal Anatomy in Lamellar Macular Hole Surgery.

BACKGROUND: Lamellar macular holes (LMHs) are an entity of a progressive disease in which the efficacy of the therapy of choice, vitrectomy, seems to be reduced. It is unknown whether highly concentrated autologous platelet-rich plasma (PRP) is of value in the therapy of LMHs. The purpose of this study was to gauge the potential of highly concentrated PRP to restore foveal anatomy in LMH surgery. PATIENTS AND METHODS: In this interventional case series, eight eyes of eight patients with progressive LMH were included. All patients underwent a 23-gauge pars plana vitrectomy with induction of a posterior vitreous detachment and peeling of tractive epiretinal membranes whenever present. Under air tamponade, 0.1 mL of highly concentrated autologous PRP was applied. Subsequently, a gas or air tamponade was performed. All patients were instructed to rest in the supine position for the first 1 to 2 postoperative hours. Best-corrected visual acuity (BCVA) testing, microperimetry, spectral-domain optical coherence tomography (SD-OCT) and fundus photography were performed prior to and 3 months after surgery. RESULTS: SD-OCT showed closure of the macular defect with restoration of a normal foveal configuration in all (8 of 8) patients 3 months postoperatively. BCVA improved significantly, from 0.28 ± 0.08 to 0.12 ± 0.14 logMAR (Wilcoxon: p = 0.03). Microperimetry remained unchanged (24.13 ± 1.96 vs. 23.7 ± 1.54 dB; p = 0.46). No clinically significant intra- or postoperative complications were observed. CONCLUSION: The use of highly concentrated PRP enables excellent anatomical and functional outcomes in the surgical therapy of LMH. Further prospective comparative trials are warranted to compare this promising technique with existing surgical strategies.

Comparison of Vitreomacular Interface Changes in Myopic Foveoschisis and Idiopathic Epiretinal Membrane Foveoschisis.

PURPOSE: To compare characteristics of the vitreomacular interface (VMI) in myopic foveoschisis (mFS) and idiopathic epiretinal membrane foveoschisis (iERM-FS), and to correlate with optical coherence tomography and clinical data. DESIGN: Clinicopathologic study. METHODS: Epiretinal membrane and internal limiting membrane (ILM) specimens were removed from eyes with mFS (5 eyes) and iERM-FS (5 eyes). Harvested tissue was processed for immunocytochemistry and prepared by ultrathin series sectioning for transmission electron microscopy. Cell and collagen compositions were compared and correlated with clinical data. RESULTS: All eyes presented fibrocellular membranes irrespective if associated with mFS or iERM-FS. Cell and collagen types and distribution on the vitreal side of the ILM were similar in both groups, consistent with presence of tractional membranes on optical coherence tomography images. Immunostaining of all specimens were positive for glial cells, microglia, and hyalocytes. Electron microscopy revealed evidence of epiretinal cell multilayers with masses of vitreous collagen and signs of vitreous remodeling in both groups. Three eyes with mFS but none of the eyes with iERM-FS showed massive thinning of the ILM with prominent retinal undulations and presence of retinal nerve fiber layer fragments. CONCLUSION: Whereas fibrocellular components of premacular tissue in mFS are similar to iERM-FS, pathologic abnormalities of the ILM were exclusively present in high myopia. Although peeling of the ILM appears important to completely remove tractional components of the VMI, histopathologic findings emphasize the risk for retinal damage in these highly myopic eyes, indicating that individual preoperative assessment and modification of surgical techniques require further investigation.

Premacular Cells as Source of Neurotrophic Factors in Idiopathic Macular Holes.

Purpose: To describe the presence of neurotrophic growth factors and histopathologic characteristics of internal limiting membrane (ILM) specimens obtained from large idiopathic full-thickness macular holes (FTMH). Methods: In 24 eyes of 24 patients with FTMH of diameter >400 µm, ILM specimens were harvested directly at the edge surrounding the macular hole during vitrectomy with peeling. We performed interference and phase contrast microscopy of flat mounts followed by immunostaining and transmission electron microscopy. Primary antigens directed against neurotrophic growth factors as well as antigens to glial and ganglion cells were used. Topographic relationship of cells and collagen was demonstrated by serial ultrathin sectioning. Results: Immunofluorescence microscopy demonstrated the presence of glial-derived neurotrophic factor and ciliary neurotrophic factor. Expression of vimentin, glial fibrillary acidic protein (GFAP), neurofilament, calretinin, and melanopsin was seen positive too. Cellular retinaldehyde-binding protein was seen positive in half of the specimens. Co-localisation of anti-GFAP as well as anti-vimentin with neurotrophic factors was found. Electron microscopy revealed cells exclusively on the vitreal side of the ILM. Cell fragments on the retinal side were rarely seen. Conclusion: In large FTMH, ILM specimens present positive immunolabelling of neurotrophic factors. The co-localization with macroglial cell markers suggests a premacular cell composition as a source of the neurotrophic factors. Ultrastructurally, premacular cells were found on the vitreal side of the ILM and not within the collagen network of the ILM itself.

Response of neovascular central serous chorioretinopathy to an extended upload of anti-VEGF agents.

PURPOSE: To determine the anatomical and functional outcomes of an extended 6-month intravitreal anti-vascular endothelial growth factor (anti-VEGF) upload in choroidal neovascularization (CNV) secondary to chronic central serous chorioretinopathy (CSCR). METHODS: A retrospective database analysis was performed applying the following inclusion criteria: (1) diagnosis of CSCR, (2) diagnosis of secondary CNV, and (3) treatment of at least six consecutive injections of anti-VEGF. Outcome measures included the change of central retinal subfield thickness, remodeling of the pigment epithelium detachments, and change in visual function. RESULTS: Twenty-one eyes of 21 patients were included. Mean patient age was 65 ± 8.3 years, and 35% of the patients (n = 8) were female. Mean disease duration before diagnosis of CNV was 48 ± 25.3 months. Mean central retinal thickness decreased from 346 ± 61 to 257 ± 57 µm (p < 0.01) after the sixth injection while mean visual acuity improved from 0.65 ± 0.35 to 0.49 ± 0.29 (logMAR; p < 0.01). Of note, an extended upload of six as opposed to three injections yielded an additional mean central retinal thickness reduction (280 ± 46 µm vs. 257 ± 57 µm, p = 0.038). Significant CNV remodeling was observed as a decrease in pigment epithelium detachment (PED) vertical (p = 0.021) and horizontal diameter (p = 0.024) as well as PED height (p < 0.01). CONCLUSION: An extended anti-VEGF upload of six consecutive injections seems to be effective in inducing CNV remodeling and fluid resorption in CNV complicating chronic CSCR.

Premacular Cell Proliferation Profiles in Tangential Traction Vitreo-Maculopathies Suggest a Key Role for Hyalocytes.

PURPOSE: To compare immunocytochemical and ultrastructural features of premacular tissue surgically removed from eyes with tangential traction vitreo-maculopathies. METHODS: By spectral-domain optical coherence tomography (SD-OCT), premacular tissue was differentiated into premacular proliferation and premacular membrane (PMM). Specimens were harvested during vitrectomy from 10 eyes with macular pucker, lamellar macular hole (LMH) and full-thickness macular hole, and prepared for immunocytochemistry and transmission electron microscopy. RESULTS: All specimens showed positive autofluorescence consistent with the yellow colour of peeled tissue. Glial cells were predominantly positive in premacular proliferation. Hyalocytes were the main cell type in PMM. Electron microscopy revealed densely packed premacular glial cells neighbouring hyalocytes and vitreous collagen strands. Myofibroblasts with features indicative of contractile properties were found in PMM, exclusively. Cell composition of premacular proliferation was free of contractile elements. CONCLUSION: All three types of vitreo-maculopathy have similar cell constituents in their premacular tissue. Cell population of premacular proliferation is not unique for LMHs. Corresponding to SD-OCT, electron microscopy demonstrates hyalocytes and vitreous collagen in PMMs both directly adjacent to the cellular complex of premacular proliferation. Study results point to the vitreous as one important pathogenic player potentially driving the degenerative cellular process at the vitreoretinal interface in tangential traction vitreo-maculopathies.

Comparison of Surgically Excised Premacular Membranes in Eyes with Macular Pucker and Proliferative Vitreoretinopathy.

PURPOSE: To describe and compare characteristics of premacular membranes in idiopathic macular pucker (iMP) and proliferative vitreoretinopathy (PVR) using immunofluorescence and transmission electron microscopy. MATERIALS AND METHODS: For immunocytochemical and ultrastructural analyses, premacular membranes were harvested during vitrectomy from 16 eyes with iMP and 12 eyes with PVR. All specimens were processed as flat mounts for phase-contrast and fluorescence microscopy. We used 19 different primary antibodies such as anti-α-smooth muscle actin (α-SMA), anti-integrin-αv, anti-galectin, anti-IBA-1, anti-EMMPRIN (CD147), anti-ricinus (RCS) and anti-collagen-type I. Eight of 28 eyes were also prepared for transmission electron microscopy. RESULTS: In all eyes with iMP and PVR, positive immunoreactivity of integrins, especially αvß3 was found. There was also a strong staining of anti-α-SMA, anti-galectin, anti-EMMPRIN, anti-RCS, anti-IBA1 and anti-collagen-type I. Transmission electron microscopy showed that premacular membrane of iMP composed of myofibroblasts, glial cells and fibroblasts. In eyes with PVR, retinal pigment epithelial cells and myofibroblasts were seen as predominant cell types. CONCLUSION: Premacular membranes of iMP and PVR presented with similarities in cell distribution and immunoreactivity, but showed differences in cell composition. Herein, we demonstrate immunocytochemical characteristics involved in fibrotic processes. Cell transdifferentiation into myofibroblasts represents an important process in pathogenesis of both entities. In order to address future anti-fibrotic treatment strategies, we emphasize that both fibrotic diseases share distinct immunocytochemical and ultrastructural features.

Vitrectomy for Diabetic Macular Edema: Optical Coherence Tomography Criteria and Pathology of the Vitreomacular Interface.

PURPOSE: To correlate spectral-domain optical coherence tomography (SDOCT) criteria and clinical data with pathology of the vitreomacular interface (VMI) in eyes with diabetic macular edema (DME). DESIGN: Retrospective cross-sectional study and laboratory investigation. METHODS: We included specimens of 27 eyes of 26 patients with center-involved DME that underwent vitrectomy with peeling of the internal limiting membrane (ILM). Selection of specimens was consecutive and in retrospect using our register of the Vitreoretinal Pathology Unit. Clinical data and SDOCT examinations were correlated to immunocytochemistry and transmission electron microscopy. Classification of DME comprised sponge-like diffuse retinal thickening, cystoid macular edema, and serous retinal detachment. VMI was evaluated for presence of epiretinal membrane (ERM) and thickened vitreous cortex (tVC). RESULTS: ERMs and tVC were found in all DME types. Diffuse DME showed tVC more often than cystoid DME. Hyalocytes, contractile myofibroblasts, glial cells, matrix metalloproteinases-2 and -9, and collagen type I, II, and III were positive tested irrespective of DME type. There were no significant cell fragments at the retinal side of the ILM. Visual acuity improved in the majority of cases and macular thickness decreased significantly during mean follow-up of 17 ± 10 months. CONCLUSIONS: All eyes presented pathologic VMI changes irrespective of the OCT classification of DME type or presence of ERM. Composition of fibrocellular membranes at the VMI indicated remodeling of vitreous cortex and transdifferentiation of hyalocytes into myofibroblasts. Our findings might argue for an early surgical intervention in eyes with DME irrespective of the presence of traction formation imaged by SDOCT.

Premacular membranes in tissue culture.

PURPOSE: To investigate integrity and characteristics of human premacular membranes (PMM) with and without standard tissue culturing using mechanical traction. METHODS: Premacular membranes were harvested from 32 eyes of 32 patients with idiopathic macular pucker during standard vitrectomy. By flat-mount preparation with phase contrast and interference microscopy, specimens were prepared for time-lapse microscopy, immunocytochemistry, and transmission electron microscopy. Sixteen of 32 specimens were held in tissue culture with tangential traction by using entomological pins. Of these, specimens of 7 eyes were analyzed with and without tissue culturing for comparison. Primary antibodies were used for myofibroblasts, hyalocytes, macro-/microglial cells, and retinal pigment epithelial and immune cells. RESULTS: Hyalocytes, macroglia, and microglia composed the main cell composition of surgically removed PMM. Correlation of time-lapse microscopy with immunofluorescence microscopy identified fast and unidirectional moving small round cells as microglia. Slowly moving elongated large cells were characterized as alpha-smooth muscle actin (α-SMA)-positive myofibroblasts. Following tissue culturing with tangential stretch, enhanced positive immunolabelling for α-SMA and integrins-αv was seen. All other labelling results were demonstrated to be similar with pre-culture conditions. Ultrastructural analysis revealed fibroblasts, myofibroblasts, and proliferation of glial cells following tissue culture. CONCLUSION: This study demonstrates abundance of fibroblasts, myofibroblasts, and glial cells in PMM from idiopathic macular pucker following tissue culture with tangential stretch application. We found enhanced contractive properties of the cultured PPM that appear to indicate transdifferentiation of the cell composition. This in vitro model may improve understanding of pathogenesis in traction maculopathies and help to establish further anti-fibrosis treatment strategies.

VITRECTOMY FOR INTERMEDIATE AGE-RELATED MACULAR DEGENERATION ASSOCIATED WITH TANGENTIAL VITREOMACULAR TRACTION: A CLINICOPATHOLOGIC CORRELATION.

PURPOSE: To describe the morphologic characteristics of the vitreomacular interface in intermediate age-related macular degeneration associated with tangential traction due to premacular membrane formation and to correlate with optical coherence tomography (OCT) findings and clinical data. METHODS: Premacular membrane specimens were removed sequentially with the internal limiting membrane from 27 eyes of 26 patients with intermediate age-related macular degeneration during standard vitrectomy. Specimens were processed for immunocytochemical staining of epiretinal cells and extracellular matrix components. Ultrastructural analysis was performed using transmission electron microscopy. Spectral domain optical coherence tomography images and patient charts were evaluated in retrospect. RESULTS: Immunocytochemistry revealed hyalocytes and myofibroblasts as predominant cell types. Ultrastructural analysis demonstrated evidence of vitreoschisis in all eyes. Myofibroblasts with contractile properties were observed to span between folds of the internal limiting membrane and vitreous cortex collagen. Retinal pigment epithelial cells or inflammatory cells were not detected. Mean visual acuity (Snellen) showed significant improvement from 20/72 ± 20/36 to 20/41 ± 20/32 (P < 0.001) after a mean follow-up period of 19 months (median, 17 months). During this period, none of the eyes required anti-vascular endothelial growth factor therapy. CONCLUSION: Fibrocellular premacular proliferation in intermediate age-related macular degeneration predominantly consists of vitreous collagen, hyalocytes, and myofibroblasts with contractile properties. Vitreoschisis and vitreous-derived cells appear to play an important role in traction formation of this subgroup of eyes. In patients with intermediate age-related macular degeneration and contractile premacular membrane, release of traction by vitrectomy with internal limiting membrane peeling results in significantly functional and anatomical improvement.

Correlative Microscopy of Lamellar Hole-Associated Epiretinal Proliferation.

Purpose. To describe morphology of lamellar hole-associated epiretinal proliferation (LHEP) removed from eyes with lamellar macular holes (LMH). Methods. Based on optical coherence tomography data, 10 specimens of LHEP were removed from 10 eyes with LMH during standard vitrectomy. Specimens were prepared for correlative light and electron microscopy (CLEM) using an immunonanogold particle of 1.4 nm diameter that was combined with a fluorescein moiety, both having been attached to a single antibody fragment. As primary antibodies, we used antiglial fibrillary acidic protein (GFAP), anti-CD45, anti-CD64, anti-α-smooth muscle actin (α-SMA), and anticollagen type I and type II. Results. In LHEP, GFAP-positive cells possess ultrastructural characteristics of fibroblasts and hyalocytes. They represent the major cell types and were densely packed in cell agglomerations on vitreous collagen strands. Epiretinal cells of LHEP rarely demonstrated contractive properties as α-SMA-positive myofibroblasts were an infrequent finding. Conclusion. CLEM indicates that epiretinal cells in LHEP might originate from the vitreous and that remodelling processes of vitreous collagen may play an important role in pathogenesis of eyes with LMH.

Pathology of Internal Limiting Membrane Specimens Following Intravitreal Injection of Ocriplasmin.

PURPOSE: To describe ultrastructure and immunocytochemistry of internal limiting membrane peelings after unsuccessful treatment with ocriplasmin, to compare with untreated eyes, and to correlate with clinical imaging data. DESIGN: Interventional comparative case series. METHODS: Internal limiting membrane specimens were removed from 10 eyes with small macular holes and vitreomacular traction during vitrectomy after intravitreal ocriplasmin injection without release of traction or closure of macular holes during follow-up. Based on optical coherence tomography analysis, specimens from 10 other eyes without ocriplasmin treatment served as controls. All specimens were processed as flat mounts for phase-contrast microscopy followed by immunolabeling for fluorescence microscopy and embedding in epoxy resin with serial sectioning for transmission electron microscopy. RESULTS: Despite the absence of contractive epiretinal membranes on optical coherence tomography, we found epiretinal cells and vitreous collagen fibrils on the internal limiting membrane in specimens removed from eyes with and without previous pharmacologic vitreolysis. Immunolabeling revealed glial cells and hyalocytes in macular holes, whereas myofibroblasts were predominant in vitreomacular traction. There was no apparent damage of the vitreoretinal interface after unsuccessful pharmacologic vitreolysis compared to untreated controls. CONCLUSIONS: Epiretinal cell proliferation and vitreous collagen fibrils with close adhesions to the internal limiting membrane are not always detectable by optical coherence tomography or may not have been recognized. Since they are associated with unsuccessful ocriplasmin treatment, presence and topography of epiretinal cells and vitreous collagen remnants on the internal limiting membrane should be further elucidated in order to refine criteria and indications for case selection in pharmacologic vitreolysis.