Abstract
P 081
Flat mounted internal limiting membrane specimens: An evaluation of cell proliferation in idiopathic macular holes
Ricarda Schumann1, Renate Scheler1, Markus Schaumberger2, Christos Haritoglou2, Anselm Kampik2, Arnd Gandorfer1
1Abteilung für Vitreoretinale Pathologie, Augenklinik der Ludwig-Maximilians-Universität München, München, 2Augenklinik der Ludwig-Maximilians-Universität München, München
Objective
To overcome limitations of conventional microscopy in evaluating cell density and cellular distribution we used a new preparation technique with flat mounted internal limiting membrane (ILM) specimens for interference microscopy and immunocytochemistry.
Methods
Fourty surgical specimens of the ILM were obtained from 40 eyes during pars plana vitrectomy for idiopathic macular holes (IMH) which comprised of 7 eyes with stage II IMH, 26 eyes with stage III IMH, and 7 eyes with stage IV IMH. Specimens were placed into 2% paraformaldehyde and 0.1% glutaraldehyde for fixation. After washing in phosphate-buffered saline 0.1M, the specimens were prepared as wholemounts on glass slides for interference microscopy. The mounting medium 4’-6-diamino-2-phenylindole (DAPI) was used to stain the cell nuclei. Cell quantification was documented using ImageJ with and without manual counting. Cellular distribution patterns were identified.
Results
Cellular proliferation was found in all ILM specimens. The total number of cells ranged from 18 to 6914 with a mean of 1663. The area of removed ILM ranged from 3,2 mm² to 22,6 mm² with a mean of 12,0 mm². Cellular distribution patterns varied between homogenous and inhomogenous with cell clusters. Cell density in ILM specimens with homogenous cell distribution was 95 Zellen / mm² and 365 Zellen / mm² in cell clusters. Specimens with cell clusters (n = 21) were shown with significant higher cell counts than specimens with homogenously distributed cells (n = 19). Comparing the stages of disease, no significant differences were seen in total cell count, peeled area of the ILM or cellular distribution. However, there was a tendency of cell count to increase with the stage of disease. Cluster formation was not correlated with the presence of metamorphopsia but increased with duration of symptoms before surgery.
Conclusions
This is the first report on cell proliferation in 100% of all evaluated eyes with IMH. Specimens with cell clusters show a higher cell count compared to specimens with homogenously distributed cells. There is a tendency of cell clusters and cell count to increase with the duration of symptoms and the stage of disease. Further investigation is needed to elucidate cell type and migration pattern in pathogenesis of macular holes.
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