Integrin-linked kinase-mediated promotion of osteogenic differentiation in bone marrow mesenchymal stem cells: A driver of heterotopic ossification in ankylosing spondylitis

Excessive osteogenesis in bone marrow mesenchymal stem cells (BMSCs) contributes to the ectopic ossification associated with ankylosing spondylitis (AS), yet the underlying mechanisms are not fully understood. Integrin-linked kinase (ILK) plays an important role in the inflammatory process of AS, but its expression and effects on osteophytogenesis require further evaluation. Hence, we aimed to explore the role and mechanisms of ILK in the syndesmophyte formation of AS. After establishing the BMSC lines, the mineralization potential of BMSCs from AS patients (AS-BMSCs) was found to be greater than BMSCs of healthy volunteers (HV-BMSCs). The expression of ILK was consistent with the osteogenic hyperactivity of AS-BMSCs. Additionally, knockdown of ILK using small interfering ribonucleic acid suppressed osteogenic differentiation in BMSCs. Conversely, ILK upregulation via lentiviral transfection promoted their osteogenesis. The activity of protein kinase B (Akt)/ glycogen synthase kinase-3β (GSK-3β)/ β-catenin pathway in AS-BMSCs was higher than HV-BMSCs after osteogenic induction, while ILK overexpression further activated this axis. Besides, the osteogenic medium enhanced the nuclear translocation of β-catenin only in AS-BMSCs. Animal experiments revealed that the size and number of osteophytes progressively increased in a time-dependent manner in ankylosing enthesitis mice. Moreover, the expression of ILK in entheseal BMSCs was higher at week 24 and week 32 than at week 8, and this elevated expression positively correlated with osteophyte development. These findings indicate that increased ILK leads to excessive mineralization in AS-BMSCs via the activation of the Akt/GSK-3β/β-catenin pathway, resulting in ectopic ossification in AS patients.