Optimization of decellularization and sterilization of porcine corneas for human corneal transplants

Abstract

Purpose : Corneal transplant is the main treatment against corneal blindness but the supply of human donor corneas (HC) is limited compared to the demand. Different artificial corneal substitutes have been generated, but so far have lacked biocompatibility or mechanical properties equivalence to human corneas. Here, we evaluated the potential of using decellularized porcine corneas (DPC) for human corneal transplants, including gamma irradiation (GI) as sterilization method for its future clinical translation.

Methods : Porcine corneas were decellularized using 0.1% sodium dodecyl sulfate (SDS). DPC were then sterilized with 25 kGy of GI. Histological evaluation was done to confirm the decellularization efficiency and TEM to evaluate the ultrastructure of DPC. Mechanical and optical characterization was performed comparing DPC, gamma irradiated DPCs (G-DPC), native porcine corneas (NPC) and HC. Water content, susceptibility to digestion with collagenase, and glucose permeability were also assessed. Recellularization of DPC and G-DPC was carried out using human corneal epithelial cells (HCEC) and human corneal fibroblast (HCF), independently, and live/dead cell assay and phenotypic evaluation with different corneal cell markers were performed. One way ANOVA was used for statistical analysis.

Results : Decellularization was able to remove cellular components from NPC, preserving the stromal ultrastructure. GI positively changed the mechanical properties of the DPC. Tensile strength of the HC was similar to G-DPC (p=0.0802) but significantly higher compared to DPC (p=0.0011) and NPC (p=0.0254). G-DPC completely blocked UV transmission. Glucose diffusion decreased non-significantly for G-DPC compared to HC (p=0.1657) but significantly from DPC (p=0.0076). G-DPC was more stable against collagenase degradation than DPC. Recellularization was possible for both DPC and G-DPC with HCEC and HCF, showing no cytotoxic effects. HCEC expressed cytokeratin 3+12 and mucin16 on both kinds of tissues, similar to HC.

Conclusions : Our results confirm that SDS effectively removes cells from NPC. GI changes mechanical properties of DPC, mimicking the HC. Biocompatibility studies suggest that decellularized porcine corneas can be repopulated. In vivo studies are in progress to further test the possibility of using decellularized, irradiated porcine corneas to treat human corneal disorders.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.