In vitro model of the outer blood-retina barrier.
The outer blood-retina barrier (BRB) is formed by the retinal pigment epithelium (rpe) and functions similarly to the blood-brain barrier (BBB). In contrast to the BBB, which is composed of a myriad of capillaries, the rpe can in principle be prepared as an intact planar tissue sheet without disruption of its barrier and carrier functions. Both a rapid and gentle procedure to isolate porcine rpe and a method to implement the harvested rpe in drug penetration testing are presented. Enucleated eyes were flat-mounted and the RPE/choroid tissue sheets with or without the retina were isolated. Fluorescence microscopy based on double-labeling with propidium iodide/calcein and scanning electron microscopy revealed well-preserved cell and tissue architecture. For drug evaluation, specimens were immobilized as the interface between test compartments in a dual-chamber device. Ten different test agents were added to one chamber at defined concentrations. After an incubation time of 30 min at 37 degrees C permeated drug levels in both compartments were quantified by HPLC-tandem mass spectrometry or HPLC with fluorescence detection. Sodium fluorescein used as a barrier marker indicated that the rpe model had excellent seal integrity. The use of a representative subset of pharmaceuticals with known BBB permeability characteristics demonstrated that the rpe model had a large permeability dynamic range (factor >350). These findings showed that the model represents a valuable tool for the investigation of the blood barrier penetration of test compounds.
Steuer
H
H
Natural and Medical Sciences Institute (NMI) at the University Tübingen, Markwiesenstr. 55, D-72770 Reutlingen, Germany.
Jaworski
A
A
Stoll
D
D
Schlosshauer
B
B
D000818:Animals; D016477:Artifacts; D001813:Blood-Retinal Barrier / Q000187:drug effects / Q000502:physiology*; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology*; D002829:Choroid / Q000098:blood supply / Q000187:drug effects / Q000502:physiology; D015194:Diffusion Chambers, Culture / Q000295:instrumentation / Q000379:methods; D004210:Dissection / Q000379:methods; D019793:Fluorescein / Q000493:pharmacokinetics; D008954:Models, Biological; D009924:Organ Culture Techniques / Q000295:instrumentation / Q000379:methods*; D010599:Pharmacokinetics; D010857:Pigment Epithelium of Eye / Q000098:blood supply / Q000187:drug effects / Q000502:physiology*; D015203:Reproducibility of Results; D034421:Sus scrofa
D019793:Fluorescein