Abstract
Purpose :
Significant shortage of viable transplantable donor corneas necessitates engineering of an appropriate replacement. Alternative technologies to repair or replace damaged cornea must involve materials that closely match the native tissue. We have prepared a novel biomaterial (CSEM) whose composition approximates the composition of corneal stroma (patent pending). The current study involves the utilization of this biomaterial for the fabrication of a cornea-construct and its characterization.
Methods :
CSEM was chemically cross-linked with glutaraldehyde to prepare constructs of desired thickness (μM) and diameter. Following this process, the constructs were washed thoroughly with PBS to remove residual chemicals, equilibrated in serum-free DMEM/F12 medium and stored aseptically in the same medium at 4 °C until further use. To assess the ability of the construct to support cell growth, the constructs were seeded with primary human or porcine corneal epithelial cells or fibroblasts. Further, a suspension of unmodified CSEM containing human/porcine corneal epithelial cells was layered on de-epithelialized human donor corneas.
Results :
CSEM contains a mixture of extracellular matrix proteins and mucopolysaccharides. CSEM-construct built to a desired consistency (soft-to-stiff) exhibited characteristics required for a corneal implant/inlay. It is transparent, stable at 60 °C, non-toxic and resistant to trypsin digestion. Microenvironment of the three-dimensional construct facilitated excellent growth of fibroblasts. Acellular CSEM construct when implanted into the cornea integrated well with the stroma extracellular matrix. The construct also supported the growth of corneal epithelial cells and formation of a multilayered (2-3 cellular layers) epithelium. Epithelial growth on the construct was confined to the surface and sustained beyond 30 days. Further, CSEM suspension containing epithelial cells generated a dense monolayer of epithelial sheet on ex vivo de-epithelialized human corneas in 24 hours. Epithelial marker protein cytokeratin 3 was expressed by regenerated corneal epithelial cells.
Conclusions :
This study shows that CSEM is biocompatible and provides an environment native to cornea to facilitate tissue regeneration. CSEM can be fabricated into a transplantable construct to repair corneal damage that causes significant loss of native tissue (partial or full-thickness trauma) or for permanent corneal replacement.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.