Abstract
BACKGROUND: Intervertebral disc degeneration (IDD) is closely related to the dysfunction of nucleus pulposus cells (NPCs) and the imbalance of their microenvironment. MicroRNAs play a key role in cell differentiation and homeostasis regulation, but the mechanisms underlying the differentiation of bone marrow mesenchymal stem cells (BMSCs) into NPCs are still unclear. METHODS: Using BMSCs and NPCs, this study performed qPCR to measure the expression of miR-140-5p, miR-145-5p, and cartilage-related genes; dual-luciferase reporter assays to verify miR-145-5p targeting SOX9; After inducing BMSC differentiation through miRNA, cell proliferation and migration were analyzed using CCK-8 and Transwell assays, while ELISA and oxidative stress kits were used to detect inflammation factors and oxidative stress levels. RESULTS: Compared to BMSCs, NPCs exhibited significantly upregulated expression of miR-140-5p, SOX9, COL2A1 and Aggrecan, alongside decreased miR-145-5p levels, reduced ROS and enhanced SOD activity. Mechanistically, SOX9 was confirmed as a direct target of miR-145-5p, while network analysis revealed its functional connection to miR-140-5p targets. Furthermore, dual miRNA modulation synergistically suppressed Notch signaling, promoting the expression of nucleus pulposus-associated markers (KRT19, CA12, HIF-1α), enhancing anabolic genes while suppressing catabolic factors. This coordinated regulation attenuated proliferation and migration while improving oxidative stress and inflammatory microenvironments, collectively promoting chondrogenic differentiation. CONCLUSIONS: The synergistic action of miR-145-5p (via SOX9 targeting) and miR-140-5p promotes BMSC differentiation toward an NPC-like phenotype in vitro, providing mechanistic insight and identifying a potential therapeutic target for disc degeneration that requires future in vivo validation.