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Bjørg Skjøth Lunding, Maja Søberg Udsen, Simon J Clark, Tina Jehs, Julie Friis Weber, Mogens Holst Nissen; The effect of stimulation with pro-inflammatory cytokines on uveal choroidal melanocytes. Invest. Ophthalmol. Vis. Sci. 2020;61(7):2234.
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© ARVO (1962-2015); The Authors (2016-present)
The aim of this study was to examine the immunological role of uveal choroidal melanocytes (UCM) in relation to retinal inflammation. UCM are the most abundant cell type in the choroid, but their functions have yet to be fully elucidated. This knowledge will be useful in a number of research areas particularly concerning age-related retinal diseases.
UCM were isolated from one human donor. The UCM were cultured in a direct culture system at 5% CO2, 37°C. The cells were stimulated with the pro-inflammatory cytokines TNFα, IFNγ and IL1β, alone or in combination. The effect of this stimulation was evaluated on different levels. Morphological changes were assessed via light microscopy. Cell viability of the cultures was evaluated using MTT and LDH assays. Changes at gene level were assessed via an Affymetrix GeneChip array performed on harvested cells, while the presence of selected proteins in the cell culture medium was determined by an MSD U-PLEX immunoassay. The MSD kit contained assays for eight chemokines and two pro-inflammatory cytokines.
We observed that stimulation with pro-inflammatory cytokines was damaging to the UCM, particularly when two or three cytokines were combined. This was apparent as severe morphological changes and as increased cytotoxicity in the cultures. We found that several genes were upregulated, and this upregulation resulted in increased production of proteins that were secreted. Several of the most upregulated genes code for chemokines or other inflammation-related proteins. Some genes were upregulated regardless of which cytokine treatment was applied, while others were only upregulated with particular combinations of cytokines. All the proteins assessed with the MSD assay, were present at higher concentrations in cell culture supernatant from stimulated UCM compared to unstimulated UCM.
These results indicate that inflammatory conditions can have detrimental effects on UCM. Several of the upregulated proteins are associated with inflammation. When they are secreted, they may enhance inflammatory conditions. We have previously shown that UCM are immunologically active by secreting chemokines and that they are able to attract monocytes and inhibit T-cell proliferation. The upregulation of chemokines genes seen in this study supports the role of UCM in migration and activation of phagocytic cells and lymphocytes to the posterior of the eye.
This is a 2020 ARVO Annual Meeting abstract.
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