Abstract
Large bone defects caused by trauma or disease needs extra intervention. Gelatin/chitosan complex is one of the most valuable compositions for bone healing, but fast degradation in aqueous solution and low mechanical properties increase the need for cross-linking agent. The cross-linker concentration and cross-linking method had a significant effect on the properties of fabricated scaffolds. Here, three different cross-linking methods of Glutaraldehyde (GA), including addition to the solution, vapor exposure, and immersion, were studied by different in-vitro analyses to find the best GA cross-linker concentration and cross-linking method. Scanning electron microscopy showed homogeneous microstructures in all samples. Fourier transform infrared spectrophotometry revealed cross-linking reactions in all samples. Swelling ratio and biodegradation ratio was reduced by increasing cross-linking concentration and exposure time. Nonetheless, higher cross-linker concentration and exposure time improved mechanical properties, while it seems the cross-linking exposure time had more effect than concentration. Accordingly, GA (1 wt%) cross-linked scaffold with solution addition method showed suitable performance with 39.3° contact angle, 1.45±0.05 MPa compressive strength, 22.31±1.3 (%) swelling ratio, and 26.33±4.47 (%) biodegradation ratio. In-vitro experiments indicated cells were spread all over the scaffolds with higher than 80 (%) cell viability in all time points. The expression of alkaline phosphatase (ALP) and osteo-related genes (osteocalcin and related transcription factor 2) were improved during 14 days of cell incubation and showed the high capacity of the scaffold support in mineralization and osto-differentiation. Therefore GA (1 wt% ) cross-linked scaffold with solution addition was introduced as the best candidate for bone repair and further studies.
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