Effect of silk fibroin protein hydrolysis on biochemistry, gelation kinetics, and NF-kB bioactivity in vitro

Fibroin is a structural protein derived from silk cocoons, which may be used in a variety of biomedical applications due to its high biocompatibility and controllable material properties. Conversely, fibroin solution is inherently unstable in solution, which limits its potential utility. Fibroin hydrolysates possess enhanced aqueous solubility and stability, with known anti-inflammatory bioactivity. Here, silk-derived protein (SDP) was produced through controlled time, temperature, and pressure conditions to generate a novel and reproducible hydrolysate population. Both regenerated fibroin and SDP solution stability were characterized for MWD, amino acid content, solubility, viscosity, surface interaction, secondary structure formation, and in vitro assessment of NF-kB pathway activity. Mechanistic studies indicate that hydrolysis processing is required to enhance material stability by abolishing fibroin's ability to self-associate. In vitro assays using HCLE cells indicate SDP has dose dependent potency for inhibiting NF-kB driven gene expression of TNF-α and MMP-9. Collectively, the results support SDP's use as an anti-inflammatory wetting agent compatible with a wide range of both biomedical and industrial applications. Furthermore, the conditions used to generate SDP hydrolysates are readily accessible, produce a highly consistent material from batch-to-batch, and permit widespread investigation of this novel population for these purposes.