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A. Koutsonas, D. Carstesen, C. Henkel, P. Walter, A. W. Weinberger; Effect of Coagulation on the Autofluorescence Pattern of Arpe-19 Cells: An in vitro Study. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2158.
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© ARVO (1962-2015); The Authors (2016-present)
Patients with a normal retinal autofluorescence pattern in scanning laser ophthalmoscope imaging, who are treated with laser coagulation, show a focal increase in the autofluorescence (AF) similar to AMD related changes. In order to understand the possible mechanisms underlying this phenomenon, we conducted an in vitro study on the effect of coagulation on the autofluorescence pattern of ARPE-19 human cells.
Confluent monolayers of ARPE-19 cells (obtained from the ATCC depository) were cultured in 2-well chamber slides. A population of these cells was exposed to unlabelled POS (Photoreceptor outer segments) over a period of 4 weeks, as described by Dintelmann et al, in order to assimilate the in vivo biological activity of the cells. Throughout this period the AF of the POS treated and untreated cells was measured (Leica DM IRb fluorescence microscope). Then the POS treated cells underwent a focal electrocoagulation (each focus: 200-300 µm). The AF response of the coagulated cells was observed over a period of at least 4 weeks and compared to the other groups (statistical analysis with SPSS two-way ANOVA). Finally the isolated cell material was examined with 2D gel electrophoresis, followed by a mass spectrometry analysis.
The exposure of the ARPE-19 cells to POS did not lead to an increase in their AF. On the contrary, the POS treated cell group showed a statistical significant increase in the AF after the coagulation (p<0,05). The 2D electrophoresis revealed significant changes in the quantity of various cell proteins in the coagulated cell population. One of these proteins, which found to be decreased in the coagulated cell population, was identified in the mass spectrometry as the NHERF/EBP50 (ERM (ezrin, radixin, moesin)-binding phosphoprotein 50) protein. The EBP50 (NHERF) is an interacting partner of the cellular retinaldehyde-binding protein (CRALBP) - an important component of the visual cycle- at the apical RPE.
The clinical finding of the increase in the retinal AF after laser coagulation was confirmed in our in vitro study. The decrease of EBP50 after coagulation could play a significant role in the accumulation of destructive materials due to the resulting partial inhibition of the visual cycle. None the less, further research will be necessary in order to reveal the exact mechanisms of AF increase by laser coagulation.
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