Derivation and Differentiation of hESC-RPE

The pluripotent hESC line Regea15/025 (46,XX) was derived [37 (link)], characterized, and maintained on inactivated human foreskin fibroblasts (hFF) (CRL-2429, ATCC, Manassas, VA, USA) in serum-free hESC culture medium, as previously described [38 (link)]. The spontaneous differentiation of RPE from pluripotent hESCs in floating aggregates using the RPEbasic differentiation method and subsequent selection and enrichment were performed as previously described [38 (link),39 (link)]. The hESC-RPE cells were matured on Matrigel-coated inserts (13.44 µg Matrigel/insert, Millicell Hanging Inserts, MCRP24H48, seeding density 75,000 cells/insert) for 2 and 4 weeks. The tight junction formation was evaluated by a transepithelial electrical resistance measurement (TEER).

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Ramsay E., Raviña M., Sarkhel S., Hehir S., Cameron N.R., Ilmarinen T., Skottman H., Kjems J., Urtti A., Ruponen M, & Subrizi A. (2020). Avoiding the Pitfalls of siRNA Delivery to the Retinal Pigment Epithelium with Physiologically Relevant Cell Models. Pharmaceutics, 12(7), 667.

Publication 2020

CRL-2429

Cells Culture medium Electrical resistance Fibroblasts Foreskin Hesc Human Matrigel Serum Tight junction

Corresponding Organization : University of Eastern Finland

Other organizations : University of Helsinki, Institute of Technology Sligo, Monash University, Materials Science & Engineering, University of Warwick, Tampere University, St Petersburg University

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Variable analysis

independent variables

Differentiation of RPE from pluripotent hESCs in floating aggregates using the RPEbasic differentiation method
Subsequent selection and enrichment of hESC-RPE cells
Maturation of hESC-RPE cells on Matrigel-coated inserts for 2 and 4 weeks

dependent variables

Tight junction formation evaluated by transepithelial electrical resistance measurement (TEER)

control variables

Seeding density of 75,000 cells/insert
Matrigel coating (13.44 µg Matrigel/insert)

positive controls

None specified

negative controls

None specified

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