Bioinformatics Analysis and Experimental Validation of Differential Genes and Pathways in Bone Nonunions

Non-union fractures pose a significant clinical challenge, often leading to prolonged pain and disability. Understanding the molecular mechanisms underlying non-union fractures is crucial for developing effective therapeutic interventions. This study integrates bioinformatics analysis and experimental validation to unravel key genes and pathways associated with non-union fractures. We identified differentially expressed genes (DEGs) between non-union and fracture healing tissues using bioinformatics techniques. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were employed to elucidate the biological processes and pathways involved. Common DEGs were identified, and a protein–protein interaction (PPI) network was constructed. Fibronectin-1 (FN1), Thrombospondin-1 (THBS1), and Biglycan (BGN) were pinpointed as critical target genes for non-union fracture treatment. Experimental validation involved alkaline phosphatase (ALP) and Alizarin Red staining to confirm osteogenic differentiation. Our analysis revealed significant alterations in pathways related to cell behavior, tissue regeneration, wound healing, infection, and immune responses in non-union fracture tissues. FN1, THBS1, and BGN were identified as key genes, with their upregulation indicating potential disruptions in the bone remodeling process. Experimental validation confirmed the induction of osteogenic differentiation. The study provides comprehensive insights into the molecular mechanisms of non-union fractures, emphasizing the pivotal roles of FN1, THBS1, and BGN in extracellular matrix dynamics and bone regeneration. The findings highlight potential therapeutic targets and pathways for further investigation. Future research should explore interactions between these genes, validate results using in vivo fracture models, and develop tailored treatment strategies for non-union fractures, promising significant advances in clinical management.