In vitro study demonstrates successful 3D bioprinting of quiescent corneal keratocyte constructs using an amniotic extract hydrogel, suggesting strong potential for corneal tissue engineering.
Corneal stromal bioprinting requires bioinks that combine optical transparency, structural stability, printability, and a biologically instructive microenvironment capable of maintaining keratocyte quiescence. Here, we report the development of GAME, a composite bioink integrating gelatin methacryloyl (GelMA) with murine amniotic membrane extract (AME), and comprehensively characterize its biochemical composition, physicochemical properties, bioprintability, and ability to support human induced pluripotent stem cell-derived corneal stromal keratocyte-like cells (hiPSC-CSKs). Quantitative proteomic analysis demonstrated that AME possesses a conserved proteome enriched in collagens, small leucine-rich proteoglycans, fibronectin, nidogens, and growth factor-associated regulatory proteins, with compositional consistency across independently prepared batches. Incorporation of AME into GelMA generated a transparent hydrogel with an interconnected porous microarchitecture, sustained structural stability, and elastic-dominant viscoelastic behavior within the range reported to support corneal stromal cell function. The GAME bioink also tolerated surgical suturing and enabled reproducible extrusion bioprinting of corneal-scale constructs with high printability and geometric fidelity following photocrosslinking. Bioprinted hiPSC-CSKs embedded in GAME exhibited high viability throughout 21 days of culture while maintaining expression of the keratocyte-associated markers keratocan, ALDH3A1, and CD34, with minimal α-smooth muscle actin (α-SMA) expression, indicating preservation of a quiescent, non-fibrotic stromal phenotype after bioprinting. These findings establish GAME as a compositionally defined, mechanically robust, and cytocompatible bioink for corneal stromal tissue engineering. More broadly, this work demonstrates that proteomic characterization of biologically derived supplements provides a rational framework for the development of reproducible and biologically instructive bioinks for corneal tissue engineering.
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Monla et al. (2026) studied this question.