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A. Oelker, M. W. Grinstaff; Epithelialization of Collagen-Modified Poly(2-Hydroxyethyl Methacrylate) Hydrogels for Corneal Implants. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5714.
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To design and characterize poly(2-hydroxyethyl methacrylate) (polyHEMA) hydrogels for part of a multi-component, partially-regenerative synthetic corneal implant.
PolyHEMA hydrogels were formed by mixing 60% monomer mixture including various ratios of (2-hydroxyethyl methacrylate), ethyleneglycol dimethacrylate (EGDMA), and methacrylic acid (MA) with 40% solvent. Gelation occured upon mixing with radical initiator (ammonium persulfate) and catalyst (tetramethylethylene diamine). Modification of the hydrogels was achieved by activating the surfaces with 2% carbonyldiimidazole in water (1 hour, 20 °C) then reacting with type I collagen (0.1 mg/mL, pH 9, 20 °C, 12 hours). Hydrogel formulations containing a range of cross-link densities (0.01, 0.1, 1 % EGDMA) and collagen coating densities (9, 16, and 64 µg/cm2) were then prepared for in vitro cell studies. Hydrogels were sterilized in 70% ethanol, dialyzed in buffer, and seeded with primary rabbit corneal epithelial cells that were isolated and cultured using conventional methods. The number of cells and average cell surface area on the hydrogels was monitored over a 24-hour period using MTS and fluorescein diacetate assays, respectively.
We evaluated several hydrogel formulations with varying cross-link density and collagen coating densities for support of in vitro adhesion and spreading of primary rabbit corneal epithelial cells over a 24-hour period. Epithelial cell adhesion was found to remain relatively constant over the range of polyHEMA cross-linking densities studied, but was found to increase with increasing collagen coating density. Similarly, cell surface area was comparable on polyHEMA hydrogels of various cross-link densities but increased on hydrogels with increasing collagen coating density. As expected, unmodified hydrogels did not support significant cell adhesion or spreading. Modified hydrogels seeded with cells and cultured past the 24-hour experiment were found to achieve full surface epithelialization within 1 week.
Preliminary in vitro evaluation of the polyHEMA hydrogels showed that epithelialization was enabled by surface modification with type I collagen. Epithelialization was found to be dependent upon collagen coating density but not upon hydrogel cross-linking density. Future work will include design of a soft hydrogel scaffold for stromal regeneration as well as integration of the polyHEMA hydrogel with the soft hydrogel to form a multi-component corneal implant.
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