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A. Chassignol, E. Brasnu, C. Baudouin, L. Riancho, J.-M. Warnet, F. Brignole-Baudouin; Interactions Between Peripheral Blood Lymphocytes and Wong-Kilbourne Derivative of Chang Conjunctival Cells in vitro. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4629.
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Conjunctival epithelium plays a central role in the ocular surface immune system, in which lymphocytes and epithelial cells are actively involved. The purpose of this study was to investigate the interactions between both cell types in vitro, and to determine whether the Wong-Kilbourne derivative of Chang conjunctiva (WKD) cells could express E-cadherin that is the natural ligand for the human mucosal lymphocyte antigen (HML-1) expressed on intraepithelial lymphocytes (IEL).
Peripheral blood lymphocytes (PBL) were cocultured for 7 days with non-treated and trichostatin A-treated WKD cells using cell-contact or transwell cultures. Scanning electron microscopy (SEM), flow cytometry (FCM), standard immunofluorescence and specific membrane stainings (PKH26/PKH67) were used to assess the morphologic, apoptotic (sub-G1 peak) and inflammatory changes on both cell types.
Morphological interactions with plasmic membrane exchanges between epithelial cells and lymphocytes were observed using PKH26/PKH67 stainings in confocal microscopy characterized by an obvious integration of lymphocyte membrane in epithelial cells’ membrane after a 90mn-co-culture. SEM confirmed tight connections between both cell types. FCM tests showed a significant increase of the expressions of E-cadherin on trichostatin A-treated WKD cells, as well as HLA DR and apoptosis on WKD cells cultured with PBL compared with controls.
In this co-culture model, tight in vitro interactions between lymphocytes and WKD cells were shown. Further experiments are required to assess the ability of PBL to develop an IEL phenotype after co-culture with E-cadherin positive WKD cells. This model could constitute new interesting ways to explore for physiopathological and toxicological in vitro studies on ocular surface.
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