This study proposes a biodegradable nerve conduit containing genipin-cross-linked gelatin annexed with tricalcium phosphate ceramic particles (genipin-gelatin-tricalcium phosphate, GGT) in peripheral nerve regeneration. Firstly, cytotoxicity tests revealed that the GGT-extracts were not toxic, and promoted the proliferation and neuronal differentiation of adipose tissue-derived stem cells (ADSCs). Secondly, the GGT composite film effectively supported ADSCs attachment and growth. Additionally, the GGT substrate was biocompatible with the neonatal rat sciatic nerve and produced a beneficial effect on peripheral nerve repair through in vitro tissue culture. Finally, the experiments in this study confirmed the effectiveness of a GGT/ADSCs nerve conduit as a guidance channel for repairing a 10-mm gap in a rat sciatic nerve. Eight weeks after implantation, the mean recovery index of compound muscle action potentials (CMAPs) was significantly different between the GGT/ADSCs and autografts groups (p < 0.05), both of which were significantly superior to the GGT group (p < 0.05). Furthermore, walking track analysis also showed a significantly higher sciatic function index (SFI) score (p < 0.05) and better toe spreading development in the GGT/ADSCs group than in the autograft group. Histological observations and immunohistochemistry revealed that the morphology and distribution patterns of nerve fibers in the GGT/ADSCs nerve conduits were similar to those of the autografts. The GGT nerve conduit offers a better scaffold for the incorporation of seeding undifferentiated ADSCs, and opens a new avenue to replace autologous nerve grafts for the rapid regeneration of damaged peripheral nerve tissues and an improved approach to patient care.

A biodegradable composite (GGT) containing tricalcium phosphate ceramic particles and genipin crosslinked gelatin was developed for use as a bone substitute. The objective of this study was to assess the biocompatibility and the osteoconductivity of the GGT composite on new bone formation in vitro. Additionally, biodegradation and biocompatibility of the GGT composite in animals were investigated. Results of the GGT composites cocultured with osteoblasts showed that the concentration of genipin used as a crosslinking agent should be <0.5 wt % to avoid cytotoxicity. For in vivo degradation studies, we found that when the concentration of genipin in the composite <0.5 wt % was not enough to fully crosslink the gelatin, it results in a rapid degradation of the gelatin-genipin mixture. However, we also found that the foreign body capsule surrounding the GGT composite containing 1.0 wt % of the genipin was much thicker than that in the other three groups, that is, the composites containing 0.05, 0.1, and 0.5 wt % of the genipin. We therefore concluded that the ideal concentration of genipin used in the GGT was 0.5 wt %. Finally, we examined the organ culture units, which were maintained in cultured medium for 5 weeks. Morphology of tissue was observed and the quantitative evaluation of the regenerated bone was determined. We found that the GGT composites containing 0.5 wt % of the genipin had an excellent biocompatibility and could produce osteoconduction for the regenerating bone tissues.

The purpose of this study was to prepare and evaluate in vitro the feasibility and cytocompatibility of a novel composite (GGT) as a large defect bone substitute. The composite is tricalcium phosphate ceramic particles combined with genipin crosslinked gelatin. After soaking the GGT composites in Ringer solutions at 37 degrees C for 7, 14, 28, 42, 56, and 84 days, the in vitro biologic degradation rate and biocompatibility were determined. Substances released from soaked GGT composites were analyzed with an ultraviolet visible light spectrophotometer. In addition, the solution soaking the GGT was co-cultured with osteoblasts to determine whether or not the released substances from GGT could facilitate the growth of bone cells. After they had been cultured for 2 days, the osteoblasts were tested for differentiation and proliferation by alkaline phosphatase (ALP) activity and a MTT assay. Results indicate that the concentration of the genipin solution is a critical factor in deciding the crosslinking degree of the GGT composite. Complete crosslinking reaction in the GGT composite occurred when 0.5 wt % of genipin had been added. Cytotoxic testing revealed that 80 ppm of the genipin in the culture medium served as the level over which cytotoxicity to osteoblasts could be produced. In addition, we found that gelatin and calcium continuously were released from the GGT composite in the soaking solution, which promoted differentiation and proliferation of the osteoblasts.

This study investigates the feasibility of a novel nanocomposite (GC/Ag) of a genipin-crosslinked chitosan (GC) film in which was embedded various amounts of Ag nanoparticles for wound-dressing applications. In situ UV-vis results revealed that adding chitosan solution did not affect the characteristics of Ag nanoparticles. The water uptake ratios and surface hydrophilicity of the GC/Ag nanocomposite were better and the degradation rates slightly lower than those of the pure GC film. The presence of Ag nanoparticles enhanced L929 cell attachment and growth. Its function as an anti-microbial agent in a GC/Ag nanocomposite was assessed for Ag contents of over 100 ppm. In conclusion, silver ions had dual functions--structural reinforcement and provision of antimicrobial properties to a biocompatible polymer.

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