A2M inhibits inflammatory mediators of chondrocytes by blocking IL‐1β/NF‐κB pathway. Issue 1 (4th May 2022)

• Record Type: Journal Article
• Authors: Sun, Changqi; Cao, Can; Zhao, Ting; Guo, Hailing; Fleming, Braden C.; Owens, Brett; Beveridge, Jillian; McAllister, Scott; Wei, Lei
• Is Part Of: Journal of orthopaedic research. Volume 41:Issue 1(2023)
• Page Start: 241
• Page End: 248
• Publication Date: 2022-05-04
• Subjects: A2M -- A2M/IL‐1β binding -- inflammatory -- NF‐κB pathway -- posttraumatic osteoarthritis (PTOA); Orthopedics -- Periodicals; Musculoskeletal system -- Periodicals; 616.7
• DOI: 10.1002/jor.25348
• Languages: English
• ISSNs: 0736-0266

Abstract

Abstract: A hallmark of osteoarthritis (OA) is cartilage degeneration, which has been previously correlated with dramatic increases in inflammatory enzymes. Specifically, interleukin‐1β (IL‐1β) and subsequent upregulation of nuclear factor kappa B (NF‐κB) is implicated as an important player in the development of posttraumatic osteoarthritis (PTOA). Alpha 2‐macroglobulin (A2M) can inhibit this inflammatory pathway, making it a promising therapy for PTOA. Herein, we demonstrate that A2M binds and neutralizes IL‐1β, blocking downstream NF‐κB‐induced catabolism seen in in vitro. Human chondrocytes (cell line C28) were incubated with A2M protein and then treated with IL‐1β. A2M was labeled with VivoTag™ 680 to localize the protein postincubation. The degree of binding between A2M and IL‐1β was evaluated through immunoprecipitation (IP). Catabolic proteins, including IL‐1β and NF‐kB, were detected by Western blot. Pro‐inflammatory and chondrocyte‐related gene expression was examined by qRT‐PCR. VivoTag™ 680‐labeled A2M was observed in the cytoplasm of C28 human chondrocytes by fluorescence microscopy. IP experiments demonstrated that A2M could bind IL‐1β. Additionally, western blot analysis revealed that A2M neutralized IL‐1β and NF‐κB in a dose‐dependent manner. Moreover, A2M decreased levels of MMPs and TNF‐α and increased the expression of cartilage protective genes Col2, Type2, Smad4, and aggrecan. Mostly importantly, A2M was shown to directly neutralize IL‐1β to downregulate the pro‐inflammatory responses mediated by the NF‐kB pathway. These results demonstrate a mechanism by which A2M reduces inflammatory catabolic activity and protects cartilage after joint injury. Further in vivo studies are needed to fully understand the potential of A2M as a novel PTOA therapy.