Neuropeptide Y1 receptor antagonist alleviated osteoarthritis by restoring chondrocyte autophagy through...

Osteoarthritis (OA) is a debilitating joint disorder affecting millions worldwide, characterized by progressive cartilage degradation and chronic pain. Emerging evidence suggests that neuropeptide Y (NPY) and its Y1 receptors are involved in OA pathogenesis, although the underlying molecular mechanisms remain poorly understood. This study investigates the role of NPY/Y1R signaling in OA progression through PI3K/AKT/mTOR-mediated regulation of chondrocyte autophagy.

Human cartilage samples were collected from ten OA patients (3 male,7 female, 63-75 years old) undergoing total knee arthroplasty and graded using the Kellgren-Lawrence system. Primary chondrocytes were isolated from neonatal C57BL/6 mice and treated with NPY (0.01-5 μM) or interleukin-1β (IL-1β, 10 ng/mL) to mimic OA-like degeneration. RNA sequencing (RNA-seq) and KEGG pathway analysis were performed to identify NPY-regulated signaling pathways. In vivo, OA was induced in 8-week-old male C57BL/6 mice via destabilization of the medial meniscus (DMM) or intra-articular injections of NPY (5 μM, every 4 weeks). Mice were treated with the Y1R antagonist (0.1 μM, weekly) or vehicle control. Pain behavior was assessed using von Frey filaments and CatWalk gait analysis. Cartilage degeneration was evaluated via histology (Safranin O/Fast Green, OARSI scoring), immunofluorescence (COLII, MMP13, LC3-II, p62), and micro-CT (subchondral bone remodeling, osteophyte formation). The activation status of the PI3K/AKT/mTOR pathway and autophagy-related markers was determined via Western blotting and immunofluorescence assays under both in vitro and in vivo conditions.

NPY and Y1R expression were significantly elevated in human OA cartilage compared to normal tissue. In vitro, NPY (5 μM) suppressed chondrocyte proliferation, reduced COLII expression, and increased MMP13 production. RNA-seq revealed NPY-mediated activation of the PI3K/AKT/mTOR pathway and inhibition of autophagy-related genes. NPY treatment enhanced the phosphorylation levels of PI3K, AKT, and mTOR, while concurrently decreasing LC3II expression and increasing p62 accumulation. The Y1R antagonist reversed these effects, restoring autophagy and attenuating cartilage degradation. In vivo, NPY injections induced OA-like changes, including cartilage thinning, osteophyte formation, and mechanical allodynia. Y1R antagonist treatment mitigated these effects, improving gait parameters and reducing subchondral bone sclerosis. Immunofluorescence confirmed that Y1R inhibition decreased PI3K/AKT/mTOR signaling and enhanced autophagy in chondrocytes.

This study demonstrates that NPY/Y1R signaling exacerbates OA progression through PI3K/AKT/mTOR-mediated suppression of chondrocyte autophagy. Pharmacological inhibition of Y1R emerges as a novel therapeutic strategy, effectively targeting both cartilage degeneration and pain, with potential disease-modifying effects on osteoarthritis progression.

This study highlights the NPY Y1 receptor as a promising therapeutic target for OA by demonstrating its role in modulating chondrocyte autophagy via the PI3K/AKT/mTOR pathway. The results support the development of Y1R antagonists as novel OA therapeutics. This work bridges molecular discovery to potential clinical application, offering hope for a transformative approach to OA management.