Evaluation of the Suitability of Biocompatible Carriers as Artificial Transplants Using Cultured Porcine Corneal Endothelial Cells

ABSTRACT

Purpose/Aim: Evaluating the suitability of bioengineered collagen sheets and human anterior lens capsules (HALCs) as carriers for cultivated porcine corneal endothelial cells (pCECs) and in vitro assessment of the cell-carrier sheets as tissue-engineered grafts for Descemet membrane endothelial keratoplasty (DMEK).

Materials and Methods: pCECs were isolated, cultured up to P2 and seeded onto LinkCell™ bioengineered matrices of 20 µm (LK20) or 100 µm (LK100) thickness, and on HALC. During expansion, pCEC viability and morphology were assessed by light microscopy. ZO-1 and Na⁺/K⁺-ATPase expression was investigated by immunohistochemistry. Biomechanical properties of pCEC-carrier constructs were evaluated by simulating DMEK surgery in vitro using an artificial anterior chamber (AC) and a human donor cornea without Descemet membrane (DM).

Results: During in vitro expansion, cultured pCECs retained their proliferative capacity, as shown by the positive staining for proliferative marker Ki67, and a high cell viability rate (96 ± 5%). pCECs seeded on all carriers formed a monolayer of hexagonal, tightly packed cells that expressed ZO-1 and Na⁺/K⁺-ATPase. During in vitro surgery, pCEC-LK20 and pCEC-LK100 constructs were handled like Descemet stripping endothelial keratoplasty (DSEK) grafts, i.e. folded like a “taco” for insertion because of challenges related to rolling and sticking of the grafts in the injector. pCEC-HALC constructs behaved similar to the DMEK reference model during implantation and unfolding in the artificial AC, showing good adhesion to the bare stroma.

Conclusions: In vitro DMEK surgery showed HALC as the most suitable carrier for cultivated pCECs with good intraoperative graft handling. LK20 carrier showed good biocompatibility, but required a DSEK-adapted surgical protocol. Both carriers might be notional candidates for potential future clinical applications.

KEYWORDS:

• Porcine endothelial cells
• cell culture
• donor material
• endothelial cell transplantation
• cell carriers

Disclosure Statement

Dr. Melles is a consultant for DORC International/Dutch Ophthalmic USA and SurgiCube International. Dr. Dapena is a consultant for DORC International/Dutch Ophthalmic USA.

Dr. Rafat serves on the Board of Directors of the company LinkoCare Life Sciences AB, which is a spin-off firm developing products related to the research being reported, and holds relevant patents. Dr. Rafat’s terms of arrangements have been reviewed and approved by Linköping University in accordance with its policy on objectivity in research. The other authors have no conflicting relationship to disclose.

Ethical statement

The study was carried out following the tenets of the declaration of Helsinki.

Additional information

Funding

The project leading to this study has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 667400 (ARREST BLINDNESS Consortium).