Scaffold-Free Extrusion-Based 3D Bioprinting of Cornea Constructs Using a Decellularized Corneal Extracellular Matrix Based Bioink and Human Placenta-Derived Mesenchymal Stem Cells.

Abstract

The development of bioinks tailored for corneal tissue engineering is crucial to replicating the native structure and function of the cornea. This study presents a scaffold-free extrusion-based 3D bioprinting (E3DB) approach to fabricate cornea constructs without support materials or molds. Bioinks composed of decellularized corneal extracellular matrix (dCECM), sodium alginate (SA), and type B gelatin (TBG) were formulated and evaluated for rheological performance, including viscosity, shear thinning, and viscoelasticity. Among the tested formulations, bioink 3G10 (SA: 3%, dCECM: 6/mL, TBG: 10%; 2:1:1 ratio) demonstrated optimal rheological and printability performance, enabling the fabrication of stable, curvature-preserving constructs. The printed constructs exhibited high shape fidelity, light transmittance comparable to native cornea, and Young's modulus values within the physiological range. Human placenta-derived mesenchymal stem cells (hPMSCs) encapsulated in bioink 3G10 showed high initial viability, a transient decline at day 7, and recovery by day 14, accompanied by morphological elongation. Gene expression analysis revealed marked upregulation of keratocyte-specific markers (KERA and ALDH) and suppression of ACTA2, indicating progression toward a keratocyte-like phenotype. These findings underscore the suitability of hPMSCs and dCECM-based bioinks for scaffold-free cornea bioprinting, providing a robust platform for the development of anatomically accurate and biologically functional corneal grafts.