Intraarticularly injectable silk hydrogel microspheres with enhanced mechanical and structural stability to attenuate osteoarthritis

A silk fibroin (silk) hydrogel was prepared by using diglycidyl ether (BDDE), a chemical crosslinker commonly used to generate Food and Drug Administration (FDA)-approved hyaluronic acid (HA) medical products. The silk/BDDE hydrogels exhibited high elasticity (compressive modulus of 166 ± 15.0 kPa), anti-fatigue properties, and stable structure and mechanical strength in aqueous solution. Chemical crosslinking was conducted in a high concentration (9.3 M) of lithium bromide (LiBr) solution, a salt that is commonly used to dissolve degummed silk fibers during silk solubilization. The unfolded and extended structure of silk molecules with these reaction conditions, as well as the unique ionic environment provided by LiBr facilitated a high degree of crosslinking in the hydrogel. Similar hydrogels were not obtained when the silk was dissolved in other silk fiber-dissolving reagents (e.g., Ajisawa's, formic acid (FA)/LiBr, FA/CaCl2 solutions), likely because partially folded silk structures and the ionic conditions with these reagents were less favorable for the crosslinking reaction. Based on these findings, silk/BDDE hydrogel spheres were prepared using an oil/water (o/w) emulsification method and biocompatibility and biodegradation were evaluated in vivo, along with other silk gel control systems (e.g., enzyme-catalyzed di-tyrosine and pulverized silk/BDDE gel particles with irregular shapes). Histological and immunohistochemical analyses demonstrated that the silk/BDDE hydrogel spheres were biocompatible and served as a bio-lubricant to treat osteoarthritis (OA). The intra-articular injection of the gel spheres reduced pain as measured with OA rats, reduced cartilage damage and resisted the digestive environment in the articular cavity for extended time frames (>4 weeks), suggesting utility for pain relief and sustained drug release for future OA treatments.