# Comparative Analysis of Genipin-Crosslinked Carp Collagen Gels and Their Effects on L929 Fibroblast Growth and Differentiation

## TL;DR

This study shows how to create stable carp collagen gels using genipin crosslinking, which support fibroblast growth and could be useful for biomedical applications.

## Contribution

A new method for stabilizing carp collagen gels using phosphate buffer-induced gelation and genipin crosslinking is introduced.

## Key findings

- Genipin-crosslinked carp collagen gels are thermally stable and structurally strong.
- Gel microstructure influences fibroblast adhesion, spreading, and proliferation.
- Carp collagen gels stimulate hyaluronic acid production by fibroblasts.

## Abstract

Fish collagen exhibits lower denaturation temperatures and reduced mechanical stability than mammalian collagen, limiting its biomedical applicability and motivating the development of effective stabilization strategies. In this study, we address this challenge by establishing a simple and effective strategy to fabricate stable collagen gels from carp-derived collagen using phosphate buffer-induced gelation followed by genipin crosslinking. Gel properties were regulated by adjusting the gelation time prior to crosslinking. After 3 h of gelation followed by 24 h crosslinking, gels derived from carp skin, carp scale, and carp swim bladder exhibited relatively compact surfaces with irregular fibrils. Conversely, gels subjected to 24 h of gelation before crosslinking exhibited longer, thicker fibrils with less compact arrangement. Rheological and differential scanning calorimetry analyses showed that the crosslinked gels were thermally stable (40.84–47.17 °C) and structurally strong (G’: 11–50 kPa; G”: 1–8 kPa; tan δ: 0.05–0.17) gels. L929 fibroblasts cultured on these gels displayed distinct adhesion, spreading, and proliferation behaviors depending on gel microstructure. Furthermore, hyaluronic acid (HA) quantification showed that cells on the carp collagen gels stimulated HA production (25.05–26.15 ng/well). These results demonstrate that genipin-crosslinked carp collagen gels constitute tunable, cytocompatible collagen-based gels, with surface microstructure influencing fibroblast behavior, offering useful design insights for the development of stabilized fish-collagen materials for biomaterial applications.