A sustainable tissue-specific hydrogel from decellularized corneal ECM for 3D In vitro ocular models

In vitro models are increasingly replacing animal testing due to their enhanced ability to mimic human physiology. In this study, we developed and characterized a hydrogel derived from decellularized extracellular matrix (C-dECM) of bovine corneas-a tissue-specific and sustainable biomaterial obtained from food industry waste. The hydrogel was produced through decellularization, enzymatic solubilization, and gelation, preserving key ECM components such as glycosaminoglycans and collagen. C-dECM hydrogels demonstrated stable gelation behavior (T50 = 51.6 ± 7.5 min), biocompatibility, and structural integrity across a range of concentrations. Scanning electron microscopy revealed a fibrillar structure with 0.2 μm average pore size and 78 μm thickness. In vitro assays with HaCaT keratinocytes and human corneal stromal cells (hCSC) showed >90 % cell viability and preserved morphology. The hydrogel supported the formation of a multilayered tissue of epithelial cells that resembles the corneal epithelium and was successfully integrated into a custom biotranswell platform. When challenged with an ocular irritant (2,6-dichlorobenzoyl chloride), the reconstructed tissue exhibited significant cytotoxic response (p < 0.0001), confirming the model's relevance for ocular toxicity screening. These findings highlight the potential of decellularized bovine cornea-derived hydrogel (C-dECM) as a sustainable and functional alternative for rat-derived matrices in 3D ocular models in preclinical research.