EVALUATION OF CROSS-LINKING TIME FOR POROUS GELATIN HYDROGELS ON CELL SHEET DELIVERY PERFORMANCE

Abstract

To overcome the drawbacks posed by surgical manipulation of bioengineered corneal endothelial cell (CEC) sheets, a simple stirring process combined with freeze-drying method was recently developed for the production of cross-linked porous gelatin hydrogels that can provide the support structure and improve the aqueous humor circulation. In this study, we further evaluated the influence of cross-linking time (0–48 h) on the delivery performance of carbodiimide modified gelatin carriers. It was found that smaller pore size, lower porosity, and larger superficial area were associated with increasing extent of cross-linking of the carrier discs. Although the hydrogels treated for short reaction time (i.e., below 6 h) had low resistance to initial nutrient permeation, these materials exhibited rapid swelling, implying a potential anterior segment tissue squeezing effect for use as intraocular implants. In addition, the delivery carriers with limited extent of cross-linking degraded too fast to be effective for retention of cell sheet grafts at the site of injury. By contrast, the gelatin samples with cross-linking degrees greater than 50% showed slower degradation rates and smaller porous structure, thereby possibly causing a significant inhibition of CEC proliferation. Cell sheet transfer studies demonstrated that the carrier discs with a high cross-linking degree (59.4 ± 1.3%) were more difficult to achieve stable cell attachment than their counterparts with a low cross-linking degree (48.3 ± 1.5%). Our findings suggest that among the cross-linked porous samples studied, 12 h is the best cross-linking reaction time for preparation of cell sheet carriers with suitable delivery performance.