Potato protein isolate hydrogels as potential scaffolds for bone tissue engineering

Introduction/Objectives:
Potato protein isolate (PPI), enriched with patatin, shows comparable gelation ability to egg protein. This study aimed to investigate the potential of PPI hydrogels as a scaffold for bone tissue engineering.

Methods:
Potato protein isolate powder, Solanic 200, was provided by Royal Avebe U.A. PPI films were generated in multiwell plates by oven-baking a 5% w/w PPI suspension at 70 °C for 3 hours. MG-63 cells were used as a model cell line and cultured on PPI films with adhesion and cell proliferation measured using the resazurin assay. Cell viability was measured on day 10 post-seeding using LIVE/DEAD stain and quantified using FIJI software. PPI hydrogels were formed by heating 10%, 15%, or 20% w/w PPI in a 90 °C water bath for 30 mins. For crosslinked gels, 5% w/w 1-ethyl-3-(3-
dimethylaminopropyl)carbodiimide hydrochloride (EDC) and 5% w/w Nhydroxysuccinimide (NHS) relative to PPI were added to the solutions immediately prior to heating. The water-holding capacity of hydrogels was evaluated by weighing the gel mass before and after soaking in water at room temperature for 24 hours. Hydrogel degradation was examined by incubating 20% w/w PPI hydrogels in 0.1% v/v trypsin solution at 37 °C and weighing the gels at 1, 3, 6, and 10 days. Young’s modulus was calculated based on the stress-strain curve of both natural and crosslinked PPI gels under compression.

Results:
The adhesion of MG-63 cells to PPI films relative to tissue culture plastic was over 95% and cells showed steady growth over time. At 10 days post-seeding, the cell viability on PPI films was approximately 100%. For hydrogels, when soaked in water, the mass of all concentrations of natural PPI gels increased no more than 5%. Crosslinked hydrogels increased by 16.63% and 8.63% for 10 and 20% crosslinked hydrogels, respectively. In the degradation test, both natural and crosslinked PPI gels steadily decreased in mass from day 1 to day 10. Crosslinked hydrogels increased in mass on day 1 due to water absorption. From day 1 to day 6, crosslinked gels had higher mass percentages than natural gels. However, on day 10, natural PPI gels retained 70% of the original mass while crosslinked gels were 60%, which was significantly lower. The Young’s modulus of hydrogels increased with increasing protein concentration, reaching 0.2 MPa in 20% gels. The stress-strain curve of crosslinked PPI gels showed a non-linear relationship, suggesting viscoelastic properties, which will be further investigated.

Conclusions:
PPI films were cytocompatible with MG-63 cells. The crosslinked PPI hydrogels showed promising biodegradable properties and water-holding ability. The viscoelasticity, porosity, interconnectivity, and biocompatibility of crosslinked hydrogels with MG63 cells will be evaluated in the future. Overall, heat-induced PPI hydrogel is a simple prepared, inexpensive, and renewable material for bone tissue engineering applications.