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Qun Zeng, Tongalp H. Tezel, Andrea Gobin; Population Of An Injectable Hydrogel Surface With Retinal Pigment Epithelium (RPE): A Method For Repairing Human Bruch’S Membrane (bm) Defects. Invest. Ophthalmol. Vis. Sci. 2012;53(14):288. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To determine whether an injectable hydrogel developed to mend uneven defects of aged human BM can also support RPE repopulation in vitro.
A photopolymerizable biomimetic injectable hydrogel based on acrylate derivative of poly (ethylene glycol) (PEG, 0.1 g/ml) and hyaluronic acid was formulated to repair and permanently seal defects of the aged BM. The hydrogel was enriched with RGDS adhesive ligand peptide sequence to facilitate RPE reattachment and growth. The vertical stability of the polymerized hydrogel on human BM was determined, to decide whether the seal between the hydrogel and BM can withstand the contractile forces exerted by proliferating and migrating RPE cells. For this purpose, the hydrogel polymerized on human BM was clamped around its edge and deformed using a spherical load. The deformation displacement was used to quantify the mechanical and viscoelastic properties of the hydrogel. The effect of gel rigidity on RPE cell attachment and proliferation was tested by plating ARPE-19 cells (2% surface) on hydrogels containing different amounts of PEG (0.05-0.3 g/ml). RPE reattachment rate was calculated at time points varying between 1-96 hours to determine the reattachment kinetics. In order to test its ability to support RPE growth, the 6.0 mm hydrogel patches were polymerized on both 4% agarose and inner collagen layer of aged human BM. Plated synchronized ARPE-19 cells were fed with fluorescent beads for visualization and their growth was monitored daily with an inverted microscope.
The seal between the hydrogel and human BM was able to withstand a stress of 14 kN.m-2. Increase in the rigidity of the hydrogel resulted in a decrease in RPE cell attachment and proliferation. The optimum cell attachment (65.5 ± 13.5%) and growth was obtained on gels containing 0.1 g/ml of PEG. 87.1% of the RPE reattachment occurred within the first hour. On hydrogel patched-agarose and aged human BM, RPE attached and proliferation occurred only on the hydrogel patches. Within 10 days RPE cells reach confluence on all 6.0 mm hydrogel grafts.
After polymerization PEG attaches to the underlying BM with a force adequate to ascertain the vertical stability of the hydrogel graft. Formulated hydrogel can support RPE reattachment and repopulation on aged human BM in vitro. The physicochemical characteristics of the PEG hydrogel make it an ideal material for reconstructing the deranged subretinal milieu in age-related macular degeneration.
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