Optimization of polycaprolactone - based nanofiber matrices for the cultivation of corneal endothelial cells

Marcus Himmler,Fabian Garreis,Friedrich Paulsen,Thomas A Fuchsluger,Dirk W Schubert

doi:10.1038/s41598-021-98426-6

Abstract

Posterior lamellar transplantation of the eye’ s cornea (DSAEK, DMEK) currently is the gold standard for treating patients with corneal endothelial cell and back surface pathologies resulting in functional impairment. An artificial biomimetic graft carrying human corneal endothelium could minimize the dependency on human donor corneas giving access to this vision-restoring surgery to large numbers of patients, thus reducing current long waiting lists. In this study, four groups of electrospun nanofibrous scaffolds were compared: polycaprolactone (PCL), PCL/collagen, PCL/gelatin and PCL/chitosan. Each of the scaffolds were tissue-engineered with human corneal endothelial cells (HCEC-B4G12) and analyzed with regard to their potential application as artificial posterior lamellar grafts. Staining with ZO-1 and Na+/K+-ATPase antibodies revealed intact cell functionalities. It could be shown, that blending leads to decreasing contact angle, whereby a heterogeneous blend morphology could be revealed. Scaffold cytocompatibility could be confirmed for all groups via live/dead staining, whereby a significant higher cell viability could be observed for the collagen and gelatine blended matrices with 97 ± 3% and 98 ± 2% living cells respectively. TEM images show the superficial anchoring of the HCECs onto the scaffolds. This work emphasizes the benefit of blended PCL nanofibrous scaffolds for corneal endothelial keratoplasty.

Highlights

Transparency of the cornea, the window to the eye, is a key feature for clear vision
In addition to adhesive contacts and gap junctions, the cells have tight junctions, which means that the endothelium forms a diffusion barrier to the directly adjacent aqueous humour that helps to maintain the intraocular pressure of the anterior chamber
Water uptake into the stroma impairs the regular arrangement of the collagen fibrils, subsequent stromal swelling impairs optic properties of the cornea which clinically shows haziness, results in blindness of the patient due to an opaque c ornea[1,4].Treatments include the transplantation of the whole cornea or rather of single corneal layers, so-called lamellar keratoplasty[8]

Summary

Introduction

Transparency of the cornea, the window to the eye, is a key feature for clear vision. The cells’ basement membrane rests on the Bowman’s membrane It is separating the epithelial cell layer from the corneal stroma, the main layer of a cornea, consisting of highly aligned collagen fibrils. Human corneal endothelial cells (HCEC) arrange in a highly ordered hexagonally shaped monolayer on top of the Descemet’s membrane. Water uptake into the stroma impairs the regular arrangement of the collagen fibrils, subsequent stromal swelling impairs optic properties of the cornea which clinically shows haziness, results in blindness of the patient due to an opaque c ornea[1,4].Treatments include the transplantation of the whole cornea (penetrating keratoplasty, PK) or rather of single corneal layers, so-called lamellar (endothelial) keratoplasty[8]. In comparison to films or hydrogels, nanofibrous matrices mimic the native tissue In this context, electrospun nanofibrous scaffolds, resembling the fibrous structure of the natural Descemet’s membrane seem to be a promising approach towards artificial posterior lamellar grafts to be used in DMEK surgeries. PCL was blended with collagen, gelatine and chitosan and the resulting scaffolds were examined for fiber morphology, scaffold characteristics and biocompatibility

Methods

Results

Conclusion