<i>In Situ</i> Ca ²⁺ -Reinforced Full-Active Hydrogels for the Treatment of Rheumatoid Arthritis by Modulating the NF-κB and NLRP3 Pathways

Rheumatoid arthritis (RA) tissues exhibit high levels of reactive oxygen species (ROS) and free Ca²⁺, which contribute to a vicious cycle of sustained inflammatory responses. This study developed an <i>in situ</i> Ca²⁺-reinforced injectable full-active HPAP hydrogel. The full-active hydrogel is formed by cross-linking phenylboronic acid-functionalized hyaluronic acid (HA-PBA) with sodium alginate (ALG) and paeoniflorin (PF) through boronate ester bonds, all of which play significant roles in RA treatment. HPAP hydrogel exhibits relatively low strength <i>in vitro</i>, allowing for easy injection into the joint cavity. Then, ALG can trigger secondary cross-linking in the RA joint cavity using Ca²⁺, forming the HPAP/Ca²⁺ hydrogel <i>in situ</i> and enhancing drug retention (with significant signals detectable <i>in vivo</i> even 17 days postinjection). Furthermore, a decrease in the free Ca²⁺ concentration can inhibit the activation of the NLRP3 inflammasome pathway. HA-PBA participates in the formation of ROS-responsive boronate ester bonds in hydrogel, thereby reducing ROS levels in RA tissues. Additionally, the released PF inhibits the activation of the NF-κB pathway, mitigating the inflammatory response. <i>In vitro</i> and <i>in vivo</i> experiments demonstrate that the HPAP hydrogel can effectively scavenge ROS, inhibit the NF-κB and NLRP3 pathways, reprogram macrophages, and reduce osteoclast differentiation and cartilage matrix degradation. In the RA animal models, HPAP hydrogel significantly improved arthritis scores, bone erosion, and inflammation levels, showing better treatment efficacy compared to commercial dexamethasone. Given the simplicity of the components of this full-active hydrogel, all of which are natural and safe compounds, it exhibits promising clinical translation potential in RA treatment.