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A.A. Urtti, E. Mannermaa, K.–S. Vellonen, K. Kaarniranta; Expression of Efflux Proteins in ARPE–19 Cells and in Primary RPE Cells . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2908. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Blood–Retinal barrier (BRB) restricts movements of drugs from blood circulation to retina. Efflux proteins of retinal pigment epithelium (RPE), which forms the outer part of BRB may have a role in this barrier, but they are only partly known. The aim of this study was to characterize efflux proteins in RPE cell culture model and also to find out whether growth conditions affect expression of efflux proteins.
RNA was extracted from ARPE–19 cells grown both in flasks and on filters (differentiated cells) in normal filter culture medium and in medium supplemented with basic fibroblast growth factor (bFGF) and bovine retina extract (BRE), and from commercial primary RPE cell line (hRPEpiC) from ScienCell Research Laboratories. Quantitative PCR (QPCR, TAQMAN) was used to evaluate transcript levels of p–glycoprotein (P–gp), multidrug resistance proteins 1–6 (MRPs) and breast cancer resistance protein (BCRP). The activity of efflux proteins was studied with calcein–AM uptake test using verapamil, MK571, cyclosporin–A and progesterone as efflux inhibitors.
QPCR results showed that at least MRP1, MRP4 and MRP5 mRNAs are expressed in ARPE–19 cells. The same efflux proteins were expressed in the primary cells. ARPE–19 cells showed higher RNA levels of MRP4 and MRP5 in filter than in flask cultures. Addition of BRE or bFGF to culture medium did not have significant effect. All efflux protein inhibitors increased the cellular fluorescence levels in calcein–AM test suggesting that efflux proteins are functional in ARPE–19 cells.
Particularly, we have shown pronounced expression of MRP4 and MRP5 in RPE cells. RPE expresses efflux proteins, but the expression profile is different from many other epithelia. Our findings may be important in the development of specific drug targeting to RPE cells or retina.
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