Tissue engineering is one of the most promising areas for treatment of various ophthalmic diseases particularly for patients who suffer from limbal stem cell deficiency (LSCD) and this is due to the lack of existence of approriate matrix for stem cell regeneration. The aim of this research project is to design and fabricate triple layered electrospun nanofibers as a suitable corneal tissue engineering scaffold and the objective is to investigate and perform various in–vitro tests to find the most optimum and suitable scaffold for this purpose.

Electrospun Scaffolds were prepared in three layers. Poly (D, L-lactide-co-glycolide; PLGA, 50:50) nanofibers were electrospun as outer and inner layers of scaffold and aligned type I collagen nanofibers were electrospun in the middle layer. Furthermore, the scaffolds were crosslinked by 1-ethyl-3-(3 dimethylaminopropyl) carbodiimide hydrochloride (EDC) and Glutaraldehyde. Structural, physical, and mechanical properties of scaffolds were investigated by using runauer emmett teller (BET), Fourier transform infrared spectroscopy (FTIR), contact angle measurement, tensile test, degradation, shrinkage analysis, and scanning electron microscopy (SEM). Also, capability to support cell attachment and viability were characterized by SEM, MTT assay and DAPI staining.

According to the result of BET analysis, average pore size, correlation coefficient, and molecular cross-sectional area of electrospun scaffolds were about 6.44 nm, 0.82, and 0.16 nm2 respectively. Tensile tests on crosslinked scaffolds represented more suitable hydrophilicity and tensile behavior. In addition, degradation rate analysis indicated that non-crosslinked scaffolds degraded faster than crosslinked one and crosslinking led to controlled shrinkage in the scaffold. The SEM analysis depicted nano-sized fibers in good shape. Also, in-vitro study represented an improved cell attachment and proliferation in the presence of human endometrial stem cells (hEnSCs) for both cross linked and non-cross linked samples.

The current study suggests the possibility of producing an appropriate substrate for successful cornea tissue engineering with a novel design.