miR-145-5p combined with miR-140-5p regulates the differentiation of bone marrow mesenchymal stem cells into nucleus pulposus cells and its mechanism in the treatment of intervertebral disc degeneration.

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.