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Kwang-Soo Kim, Dhruvinkumar Patel, Kevin Holley, Qun Zeng, Andrea S. Gobin, Tongalp H. Tezel; Formulation of an Injectable Gel to Repair Uneven Defects of Aged Bruch’s membrane: An Initial Step of Tissue Reconstruction for Age-related Macular Degeneration. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3422.
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To formulate an injectable gel that can be used to fill the uneven defects of aged Bruch’s membrane and seal the subretinal space permanently upon polymerization while creating an even surface that can promote RPE attachment and repopulation.
A photopolymerizable biomimetic injectable gel was formulated as an initial attempt of reconstructive surgery for age-related macular degeneration. The hydrogel was based on acrylate derivative of poly-(ethylene glycol) (PEG, 0.1 g/ml) and hyaluronic acid. Varying concentrations of adhesive ligand peptide sequence (RGDS; 2.8-5.0 µmol/ml) was covalently attached to the PEG hydrogel through a PEG linker chain to enhance RPE attachment and survival. Attachment and proliferation rates, as well as the morphology of newly plated synchronized ARPE-19 cells (500 viable cells/mm2) was determined on the RGDS-enriched hydrogel in comparison with hydrogel alone and hydrogel mixed with a cocktail of laminin (330 µg/ml), fibronectin (250 µg/ml), and vitronectin (33 µg/ml). ARPE-19 cells plated on 4% agarose were used as negative control. Also, the effect UV exposure during polymerization (365 nm, 10 mW/cm2, for 30 seconds) on RPE and photoreceptor cell viability was assessed. The efficacy of Rose Bengal and Hematoxylin and Eosin for gel polymerization was also tested.
30 seconds of UV exposure during gel polymerization did not affect RPE (99.4±0.1% vs. 99.6±0.5 %) and photoreceptor (94.8±0.3% vs. 95.2±0.4 %) viability adversely (p=1.0). 24 hours after polymerization hydrophilic gels swelled 18%; but they maintained their integrity throughout the observation period of 3 months. Alternative ways of cross-linking with Rose Bengal and Hematoxylin and Eosin failed. RPE cell attachment was highest on RGDS-enriched hydrogel (65.5±13.5%), followed by PEG alone (23.3±9.0%) and agarose (10.8±0.8%, p<0.001). 5.0 µmol/ml of RGDS conjugation yielded the highest attachment and earliest spreading of RPE cells. Within 24 hours cells plated on agarose decreased in number, while they proliferated on PEG and PEG-RGDS (doubling times: 16.6 and 24.2 hrs, respectively). Adding extracellular matrix protein cocktail paradoxically lowered the attachment rate (0.8±1.9%).
Formulated biomimetic injectable PEG hydrogel can support RPE attachment and repopulation. It can be used to seal the subretinal space and aid in reconstructing the deranged subretinal milieu in age-related macular degeneration.
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