Abstract:

Objective To explore the facilitative effect of optimizing the mechanical microenvironment of the bone tunnel on tendon-bone healing after anterior cruciate ligament （ACL） reconstruction， and its regulatory correlation with the Transforming growth factor-β-activated kinase 1 （TAK1）/Nuclear factor kappa-B （NF-κB）/Mitogen-activated protein kinase （MAPK） signaling pathway. Methods A rabbit model of anterior cruciate ligament （ACL） reconstruction was established and randomly allocated into four groups： the normal control （NC） group， the conventional forward drilling without bone grafting （FD） group， the forward drilling plus bone grafting （FD + BG） group， and the reverse drilling plus bone grafting （RD + BG） group. Hematoxylin-eosin （HE） staining was carried out to observe the histomorphological changes at the tendon-bone interface. Quantitative real-time polymerase chain reaction （qRT-PCR） was utilized to detect the messenger ribonucleic acid （mRNA） expressions of inflammatory factors interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α）. Enzyme-linked immunosorbent assay （ELISA） was applied to measure the concentrations of the angiogenesis-related factor vascular endothelial growth factor （VEGF） and the tissue repair factor transforming growth factor-β1 （TGF-β1）. Western blot analysis was performed to evaluate the protein expressions of osteogenic factors osteocalcin （OCN）， osteopontin （OPN）， and key proteins in the TAK1/NF-κB/MAPK signaling pathway. Results Histomorphological findings indicated that the RD + BG group achieved the most optimal tendon-bone interface fusion， characterized by dense and mature newly-formed bone trabeculae. When compared with the FD + BG group， the RD + BG group showed significantly downregulated mRNA expressions of IL-6 and TNF-α， remarkably upregulated levels of VEGF， TGF-β1， along with protein expressions of OCN and OPN， and notably decreased phosphorylation ratios of key proteins in the TAK1/NF-κB/MAPK pathway （p-TAK1/TAK1， p-p65/p65， p-p38/p38） （P < 0.05）. Conclusion Optimization of the bone tunnel mechanical microenvironment （RD + BG） can effectively balance local inflammatory responses， precisely upregulate the expressions of repair-related and osteogenic factors， and consequently accelerate tendon-bone healing after ACL reconstruction. This process might be closely associated with the activation changes of the TAK1/NF-κB/MAPK signaling pathway， offering novel experimental evidence for the clinical refinement of ACL reconstruction surgery.

Key words: anterior cruciate ligament reconstruction, tendon-bone healing, mechanical microenvironment, TAK1/NF-κB/MAPK signaling pathway, osteogenic differentiation, inflammatory regulation