Abstract
Purpose: :
Transplantation of retinal pigment epithelium (RPE) is a promising approach for the treatment of retinal diseases including dry age-related macular degeneration (AMD). Past replacement strategies using RPE cell suspensions or sheets have failed largely due to poor adhesion or lack of organization on a Bruch’s Membrane (BrM) that is aged or itself diseased. In this study, we developed and tested a thin film, porous poly(ε-caprolactone) (PCL) scaffold to act as a biomimetic BrM substitute for RPE culture and potential implantation into the sub-retinal space.
Methods: :
A nanopatterned mold was created using a combined process of photolithography and reactive ion etching. Porous PCL scaffolds were then produced from the mold by spin-assisted templating and mounted on transwell inserts. Fetal human RPE cells (fhRPE) were maintained at confluence on the scaffolds for up to 8 weeks and compared to non-porous PCL and porous polyester (PET) transwells (Costar) using transepithelial resistance, gene expression, phagocytic capacity, and immunohistochemistry.
Results: :
fhRPE transepithelial resistance significantly increased on porous PCL compared to PET transwells beginning at 5 days and persisting through the end of the 8 week experiment. Cells on porous PCL also displayed significant upregulation of key RPE proteins including a 6.45-fold increase in RPE65 and 2.19-fold increase in CRALBP compared to the PET control at 4 weeks. Alternately, cells on non-porous PCL fail to show a significant change in major RPE markers compared to the control. fhRPE cultured on porous PCL and PET displayed similar levels of binding and phagocytosis of fluorescently-labeled photoreceptor outer segments after four weeks of culture. In addition, the greatest degree of pigmentation was observed in cells on the porous PCL followed by non-porous PCL, and finally PET.
Conclusions: :
Both material and porosity are important factors in RPE maturation and function in vitro. Our porous PCL thin film scaffold exploits both characteristics to promote the development of a mature RPE monolayer as evaluated by gene expression profile, organization, and polarization while allowing transport across the membrane.
Keywords: retinal pigment epithelium • Bruch's membrane • age-related macular degeneration