Engineered Basement Membranes for Regenerating the Corneal Endothelium

Abstract

Basement membranes are protein-rich extracellular matrices (ECM) that are essential for epithelial and endothelial tissue structure and function. Aging and disease cause changes in the physical properties and ECM composition of basement membranes, which has spurred research to develop methods to repair and/or regenerate these tissues. An area of critical clinical need is the cornea, where failure of the endothelium leads to stromal edema and vision loss. Here we developed an engineered basement membrane (EBM) that consists of a dense layer of collagen IV and/or laminin approximately 5–10nm thick, created using surface-initiated assembly, conformally attached to a collagen I film. These EBMs were used to engineer a corneal endothelium (CE) that mimicked the structure of Descemet’s membrane with a thin stromal layer, towards use as a graft for lamellar keratoplasty. Results showed that bovine and human CE cells formed confluent monolayers on the EBM, expressed ZO-1 at the cell-cell borders and achieved a density of ~1600 cells mm ⁻² for 28 and 14 days, respectively. These results demonstrated that our technique was capable of fabricating EBMs with structural and compositional properties that mimic native basement membranes and that our EBM may be a suitable carrier for engineering transplant quality CE grafts.

Graphical abstract

Human and bovine corneal endothelial cells are cultured on engineered basement membranes (EBM) designed to mimic the structure of the native Descemet’s membrane. Culturing these cells on the EBMs increases monolayer cell density compared to standard compressed collagen I gels. Additionally the cells on the EBMs have more robust ZO-1 and more cortical F-actin stress fibers.