fabrication-surgical-application-enriched-photo-crosslinked

Fabrication and Surgical Application of Enriched Photo-crosslinked Gelatin&#8211;Riboflavin Hydrogels for Corneal Wound Repair in Rabbits

This article presents a step&#8209;by&#8209;step protocol to fabricate, supplement, and surgically apply an in situ blue&#8209;light&#8211;crosslinkable ...

University of the Basque Country UPV/EHU

IIS Biobizkaia

The vision impairments suffered by millions of people worldwide and the shortage of corneal donors show the need of substitutes that mimic native tissue to promote cell growth and subsequent tissue regeneration. The current study focused on the in vitro assessment of protein-based biomaterials that could be a potential source for corneal scaffolds. Collagen, soy protein isolate (SPI), and gelatin films cross-linked with lactose or citric acid were prepared and physicochemical, transmittance, and degradation measurements were carried out. In vitro cytotoxicity, cell adhesion, and migration studies were performed with human corneal epithelial (HCE) cells and 3T3 fibroblasts for the films' cytocompatibility assessment. Transmittance values met the cornea's needs, and the degradation profile revealed a progressive biomaterials' decomposition in enzymatic and hydrolytic assays. Cell viability at 72 h was above 70% when exposed to SPI and gelatin films. Live/dead assays and scanning electron microscopy (SEM) analysis demonstrated the adhesion of both cell types to the films, with a similar arrangement to that observed in controls. Besides, both cell lines were able to proliferate and migrate over the films. Without ruling out any material, the appropriate optical and biological properties shown by lactose-crosslinked gelatin film highlight its potential for corneal bioengineering.

Cytocompatibility and Suitability of Protein-Based Biomaterials as Potential Candidates for Corneal Tissue Engineering.

Corneal persistent epithelial defects are common ophthalmic injuries that can cause significant visual and structural damage. While diagnosis is straightforward, treatment remains challenging. Noninvasive therapies like eye drops are preferred, but severe neurotrophic keratopathy may require surgical interventions. This study explores gelatin-based hydrogels as noninvasive alternatives for corneal repair. Four photo-cross-linkable hydrogels with gelatin and riboflavin phosphate (RFP) were evaluated: a control and variants incorporating 2.5% dextran (D), 0.4% hyaluronic acid (HA), or 1% methylcellulose (MC). In vitro assessments included physicochemical properties, biocompatibility, and release kinetics alongside ex vivo wound healing assays. The gelatin-RFP hydrogel maintained corneal transparency, while additives reduced it. Dextran slowed compound release, and HA and MC reduced the release rate of larger molecules. All hydrogels showed excellent biocompatibility, and ex vivo models confirmed re-epithelialization, though slower than controls. The unmodified gelatin-RFP hydrogel demonstrated the best potential for corneal tissue engineering, supporting its future clinical translation.

In Vitro Evaluation of Gelatin-Based Hydrogels as Potential Fillers for Corneal Wounds.

The objective of this study was to develop a reliable, cost-effective, and rapid in vitro model employing real-time PCR to assess inflammatory responses in hydrogel-based systems, and to comparatively evaluate the anti-inflammatory efficacy of human serum (HS), serum derived from plasma rich in growth factors (sPRGF), and human amniotic membrane extracts (HAMe) incorporated into gelatin-based hydrogels. An in vitro model of corneal inflammation was established by quantifying IL-1β expression via qPCR in TNFα-stimulated SV-40 immortalised human corneal epithelial (HCE) cells. Hydrogels functionalised with HS, sPRGF, or HAMe sourced from proximal, medial, distal, or pooled amniotic regions were evaluated for their anti-inflammatory potential. In vivo validation was conducted in a rabbit anterior stromal keratectomy model, assessing epithelial wound closure and clinical signs of irritation following application of unmodified hydrogels or hydrogels functionalised with autologous serum (AS) or HAMe. In vitro, hydrogels incorporating HS, followed by sPRGF and pooled HAMe, significantly attenuated IL-1β expression, whereas unmodified hydrogels exacerbated the inflammatory response; region-specific HAMe hydrogels demonstrated inconsistent effects. In vivo, AS-functionalised hydrogels facilitated complete re-epithelialisation by day 7 and achieved the lowest irritation scores, indicating both therapeutic efficacy and high tolerability. All hydrogel formulations were found to be biocompatible throughout the study period. These findings underscore the significant anti-inflammatory and regenerative potential of gelatin-based hydrogels functionalised with blood-derived products and support their development as bioactive platforms for ocular surface therapy. Furthermore, the in vitro model provides a robust preclinical screening tool, contributing to the refinement and reduction of animal use in biomedical research.

An in vitro pre-screening model to evaluate the corneal anti-inflammatory effect of human blood-derived products and amniotic membrane extracts incorporated into gelatine-based hydrogels.

Persistent epithelial defects (PEDs) and chronic corneal ulcers are lesions resistant to treatment for over 2 weeks, risking inadequate healing, reduced sensitivity, and corneal lysis or perforation. This study evaluates the regenerative potential of functionalized gelatin-based hydrogels for treating rabbit corneal wounds as a non-surgical alternative.

Article	Year
Cytocompatibility and Suitability of Protein-Based Biomaterials as Potential Candidates for Corneal Tissue Engineering. International journal of molecular sciences\| PubMed ID: 33807473	2021
In Vitro Evaluation of Gelatin-Based Hydrogels as Potential Fillers for Corneal Wounds. Biomacromolecules\| PubMed ID: 40079491	2025
An in vitro pre-screening model to evaluate the corneal anti-inflammatory effect of human blood-derived products and amniotic membrane extracts incorporated into gelatine-based hydrogels. Experimental eye research\| PubMed ID: 40889613	2025
Comparison of the regenerative potential of different functionalized gelatin-based hydrogels as fillers of rabbit corneal wounds. Frontiers in medicine\| PubMed ID: 41070062	2025

Cristina Romo-Valera

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