The IL-4/IL-4R/STAT6 signaling axis regulates macrophage efferocytosis in osteoarthritis.

BACKGROUND: Osteoarthritis (OA) is a leading cause of global disability, with no available therapies that effectively halt disease progression. Impairment of macrophage efferocytosis, the clearance of apoptotic cells, is implicated in OA pathogenesis, yet the underlying mechanisms remain poorly understood. This study investigates the role of the anti‑inflammatory cytokine interleukin‑4 (IL‑4) in regulating macrophage efferocytosis during OA. METHODS: Public single‑cell and bulk RNA‑seq datasets were analyzed to identify key cell populations and pathways. An OA model was established by destabilizing the medial meniscus (DMM) in C57BL/6 mice, followed by intra‑articular delivery of AAV‑IL‑4. Joint integrity was evaluated using micro‑CT, histology (H&E, Safranin O-Fast Green), immunohistochemistry, and immunofluorescence. Cytokine levels were measured by ELISA. Macrophage proliferation, polarization, phagocytosis, and efferocytosis were assessed by flow cytometry and fluorescence imaging. Molecular mechanisms were examined via Western blot, transcriptomic sequencing, and STAT6 knockdown. Pharmacological inhibition of AKT was used to assess the contribution of AKT signaling. RESULTS: IL‑4 expression was reduced in OA tissues, and low macrophage efferocytosis correlated with OA severity. In mice, IL‑4 supplementation attenuated cartilage degradation, bone loss, and synovial inflammation. In vitro, IL‑4 promoted macrophage proliferation, M2 polarization, IL‑10 secretion, and enhanced both phagocytosis and efferocytosis in both RAW264.7 cells and primary bone marrow-derived macrophages (BMMs). Mechanistically, IL‑4 increased phosphorylated STAT6 (pSTAT6) in macrophages, and STAT6 knockdown abolished the IL‑4‑induced functional enhancements. Transcriptomics and Western blotting revealed that IL-4/STAT6 signaling activates the PI3K/AKT pathway. Pharmacological inhibition of AKT reversed IL-4-enhanced efferocytosis and phagocytosis. CONCLUSION: The IL-4/IL-4R/STAT6 axis sustains macrophage function by activating the PI3K/AKT pathway, thereby boosting efferocytosis and mitigating OA progression. These findings provide a novel immunomodulatory framework for OA therapy, highlighting macrophage-directed strategies as a potential therapeutic avenue.