Permeation across Retinal Pigment Epithelium barrier

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By Experimentica on August 24, 2020
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Summary: RPE cell monolayers make a tight paracellular barrier that can be used to screen early-stage drug candidates.

Model Description

Retinal pigment epithelium (RPE) is a tight cell monolayer between retina and choroidal blood. RPE form the outer blood-retinal barrier (oBRB) restricting the drug entry to retina after systemic administration. PCi-RPE1426 cell monolayers represent tight paracellular barrier, thus, this in vitro cell model can be used to screen drug molecules in early phase of drug development. 

Species RPE cells derived from human induced pluripotent stem cells (PCi-RPE1426, Phenocell, France) 
Assessment The permeability experiments across PCi-RPE1426 cell monolayers are conducted in Transwell® cell culture inserts (Corning Inc., Corning, NY, USA) in basolateral-to-apical direction (B-A, represent the transport from the choroid to the retina) to measure the transport of study molecules and standard molecules. The cumulative amount of study molecules in the receiver chamber versus time is measured. 
Standard molecules Low permeability standard: 6-carboxyfluorescein
High permeability standard: Rhodamine B 
Read-outs The rate (apparent permeability coefficient, Papp) of the study molecules is compared to the Papp values of the low and high permeability markers. 

Outcomes and Read-Outs 

Imaging

Fig. 1. Expression of tight junction proteins occludin (left) and ZO-1 (right) in human induced pluripotent stem cells (PCi-RPE1426) cultured on Transwell® cell culture inserts (28 days) detected by immunofluorescence staining (green). Scale bar 50 µm.
Fig. 1. Expression of tight junction proteins occludin (left) and ZO-1 (right) in human induced pluripotent stem cells (PCi-RPE1426) cultured on Transwell® cell culture inserts (28 days) detected by immunofluorescence staining (green). Scale bar = 50 µm. 

Permeability measurements

Fig. 2. The apparent permeability coefficient (Papp, cm/s) values for the standard molecules across human induced pluripotent stem cells (PCi-RPE1426).
Fig. 2. The apparent permeability coefficient (Papp, cm/s) values for the standard molecules across human induced pluripotent stem cells (PCi-RPE1426). 6-CF = 6-carboxyfluorescein (low permeability standard molecule); Rho 123 = rhodamine 123; Rho B = rhodamine B (high permeability standard molecule); B-A = basolateral-to-apical direction; A-B = apical-to-basolateral direction. 
Fig. 3. Transepithelial electrical resistance (TEER) values were measured before and after the permeability experiments from the PCi-RPE1426 cell monolayers cultured and differentiated in Transwell® cell culture inserts for 29 days.
Fig. 3. Transepithelial electrical resistance (TEER) values were measured before and after the permeability experiments from the PCi-RPE1426 cell monolayers cultured and differentiated in Transwell® cell culture inserts for 29 days. 

References

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