In the laboratory, we use stem-cell derived retina organoids to reproduce retina neuroepithelium and study the relation of RPE monolayer mechanobiology with photoreceptors cells and their interaction with other cell layers. The figure and video show the three-dimensional cellular organization of the retina organoids and their level of differentiation with inner nuclear layer cells (marked with Pax6) and photoreceptors cells (marked with opsin).
To study the effect biochemical and physical cues on RPE mechanics, we mimic BrM using hydrogels systems of controlled stiffness and functionalization. Hydrogels not only better mimic physiological stiffnesses, but also give us the possibility to employ mechanobiology techniques that allow both qualitative and quantitative evaluation of adhesive forces on the ECM (traction force microscopy) and at cell-cell junctions (monolayer stress microscopy). Different functionalization techniques can be used to study cell adhesion: full leght ECM proteins can be crosslinked on the gel surface or specific ECM-derived peptides can be bound on gold nanoparticles.
Mimicking Bruch's Membrane to Investigate RPE Mechanobiology
Stem-cell Based Retina Research Model
The retinal pigment epithelium (RPE) is a highly specialized cell layer located at the base of the retinal neuroepithelium. During development, the RPE is crucial for retina organization, while in the adult eye it defines the blood-retinal barrier, is crucial for light absorption and is responsible for the metabolic support of the light detecting rods and cones of the neuroepithelium.
RPE cells are tightly bound to the underlying basement membrane (BM), which is part of a thicker layer of extracellular matrix (ECM) called Bruch’s membrane (BrM). While the BM components carry the biochemical information for RPE adhesion, deeper layers of the BrM, composed of fibrillar collagen types I, III and V and elastin fibers, are proposed to determine physical properties of RPE.
BrM characteristics are very important for the retina homeostasis and RPE functions, not only providing support for RPE cells and acting as a physical separation between the RPE and the choroid blood vessels, but also being an integral part of the blood-retina barrier, thus, regulating the diffusion of biomolecules.