Abstract
Purpose :
In vitro assessment of bioengineered carriers and human anterior lens capsules (HALC) carrying cultivated porcine corneal endothelial cells (pCEC) as tissue-engineered grafts for Descemet membrane endothelial keratoplasty (DMEK).
Methods :
pCEC were isolated and cultured up to P2 before being seeded onto LinkCell™ bioengineered matrices of 20µm (LK20) or 100µm (LK100) thickness, and on HALC. During expansion on the carriers, pCEC viability and morphology were assessed by light microscopy. Expression of ZO-1 and Na+/K+ -ATPase was investigated by immunohistochemistry. pCEC viability was measured by glucose levels in the culture medium at different time points. Biomechanical properties of the pCEC-carrier constructs were evaluated by simulating DMEK surgery in vitro using an artificial anterior chamber (AC) and a human donor cornea denuded of DM and de-swelling after in vitro surgery was checked by optical coherence tomography (OCT).
Results :
pCEC seeded on LK20, LK100, and HALC, formed a uniform monolayer of hexagonal, tightly packed cells that expressed Na+/K+-ATPase and ZO-1. Glucose level in the culture medium decreased by an average of 53±33% and 62±10% for pCEC seeded on LK20 and HALC, respectively. During in vitro surgery the pCEC-LK20 and pCEC-LK100 constructs were handled like Descemet stripping endothelial keratoplasty (DSEK) grafts, i.e. folded like a ‘taco’ for insertion due to the challenges related to the rolling and sticking of the graft to the injector. The pCEC-HALC construct behaved similar to the DMEK reference model during implantation and unfolding in the artificial AC, showing good adhesion to the bare stroma. Furthermore, two days after the in vitro surgery, corneal thickness decreased by an average of 12% indicating deturgescence capacity of the pCEC-HALC constructs.
Conclusions :
HALC was the most suitable carrier for cultivated pCEC in in vitro DMEK surgery with good intraoperative graft handling and inducing postoperative corneal deturgescence, while the LK20 carrier showed good biocompatibility, but required an adapted surgical protocol, that is a DSEK protocol. Both carriers might be good candidates for potential future clinical applications.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.