Characterization and application of a hydrogel derived from acellular porcine corneal stroma

Abstract

Purpose : Hydrogels may be an advantageous alternative to the clinically used fibrin gel in the treatment of corneal wounds. In this study, we created an acellular porcine corneal stroma hydrogel (APCS-gel) and evaluated its physical and regenerative properties.

Methods : The optimized APCS-gel was prepared by adjusting the concentration of acellular porcine corneal stroma (APCS). We evaluated it’s turbidimetric gelation kinetics, light transmittance and protein content using spectrophotometry, SDS-gel electrophoresis and mass spectrometry. Surface ultrastructure, biomaterial permeability and cellular infiltration of the APCS-gel were evaluated by scanning electron microscopy, permeability, H&E staining and tissue culture, respectively. Additionally, the cellular biocompatibility was evaluated by MTT, live/dead assays, and corneal epithelium reconstruction in vitro, using rabbit corneal epithelial (RCEC) and stromal keratocyte (RCSC) cells seeded on the APCS-gel. In vivo, the APCS-gel was transplanted onto the mouse cornea subjected to epithelium debridement and the injury recovery was evaluated by slit-lamp imaging and fluorescence immunostaining.

Results : The APCS-gel was successfully prepared by using APCS concentrations ranging from 10-30 mg/ml. The APCS-gel prepared from 20 mg/ml APCS demonstrated the highest light transmittance and fastest gelation kinetic properties. It retained most collagens, proteoglycans, glycoproteins and growth factors of native cornea. Compared with fibrin gel, the APCS-gel demonstrated a higher porosity ratio, increased permeability, and deeper RCSC cellular infiltration. Additionally, the APCS-gel showed better biocompatibility as evidenced by higher cellular survival and proliferation. After 7 days of culture, 3-4 layers of RCEC were formed on APCS-gel versus 1-2 layers on fibrin gel. We also found higher RCEC stem/progenitor cell phenotypes (K3-, p63+, ABCG2+) and proliferation phenotypes (Ki67+) on the APCS-gel. In vivo, the APCS-gel demonstrated faster corneal reepithelization after corneal epithelium debridement than both untreated mice and mice treated with fibrin gel.

Conclusions : APCS-gel demonstrated similar properties of native cornea, maintained corneal epithelial stemness, enhanced corneal epithelial proliferation in vitro and in vivo. It may be a promising therapeutic intervention for the treatment of corneal wounds in the future.

This is a 2021 ARVO Annual Meeting abstract.