NLK knockdown in hBMSCs enhance repair of critical-size bone defects by modulating neurogenic and osteogenic differentiation

Nemo-like kinase (NLK), an evolutionarily conserved MAP kinase-related kinase, is highly expressed in neural tissues and critically regulates cell proliferation, migration, and apoptosis by regulating numerous transcriptional molecules. Despite the widespread application of mesenchymal stem cells (MSCs) in regenerative medicine, the functional role and molecular mechanisms of NLK in MSC-mediated tissue repair remained poorly understood. Here, the dual regulatory effects of NLK on both neurogenic and osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) were investigated. The results showed that NLK acted as a potent inhibitor of hBMSC neurogenesis in vitro and suppressed osteogenesis both in vitro and in vivo. Mechanistically, NLK downregulated the transcriptional coactivators LEF1 and TCF4, thereby impairing their pro-differentiation functions during neural and bone formation. These findings suggested that NLK-mediated suppression of LEF1/TCF4 signaling might hinder endogenous bone repair by dual inhibition of hBMSC neurogenic and osteogenic capacities. Targeting this pathway could offer novel therapeutic strategies for enhancing bone defect regeneration and inform the design of advanced biomaterials for bone tissue engineering.