In Vitro Evaluation and Transplantation of Human Corneal Endothelial Cells Cultured on Biocompatible Carriers

Abstract

Corneal transplantation is currently the only effective treatment option for dysfunctional corneal endothelial cells (CEC). In this study, we test in vitro the surgical potential of cultivated human corneal endothelial cells (hCEC) on human anterior lens capsule (HALC), LinkCell™ bioengineered collagen sheets of 20-µm thickness (LK20), and denuded Descemet membrane (dDM) as tissue-engineered grafts for Descemet membrane (DM) endothelial keratoplasty (DMEK) to bypass the problem of donor tissue availability. Primary hCEC cultured on all carriers formed a monolayer of tightly packed cells with a high cell viability rate (96% ± 4%). hCEC on HALC and LK20 showed unremarkable expression of zonula occludens-1 (ZO-1) and Na+/K+-adenosine triphosphatase (ATPase), while Na+/K+-ATPase expression of cells seeded on dDM was mainly cytoplasmic. All hCEC–carrier constructs were evaluated by simulating DMEK surgery in vitro using a human donor cornea without DM mounted on an artificial anterior chamber (AC) and a regular DMEK-graft used as a surgical reference model. During in vitro surgery, hCEC–HALC constructs behaved most similarly to a DMEK-graft during implantation and unfolding, showing good adhesion to the bare stroma. On the other hand, hCEC–LK20 and hCEC–dDM constructs required some additional handling because of challenges related to the surgical procedure, although they were both successfully unfolded and implanted in the artificial AC. The hCEC–dDM constructs showed similar graft adherence as hCEC–HALC constructs, while adherence of hCEC–LK20 constructs was less effective. After the in vitro surgery, the estimated area populated by viable cells on the hCEC–HALC and hCEC–LK20 constructs was ∼83% and ∼67%, respectively. Overall, hCEC–HALC constructs behaved most similarly to a DMEK-graft during in vitro DMEK surgery, while graft adhesion and surgical handling, respectively, are parameters still requiring optimization for hCEC–LK20 and hCEC–dDM constructs.