ALDH5A1 as a key in osteonecrosis of the femoral head: Insights from bioinformatics and experimental validation.

The present study aims to identify key genes and elucidate the molecular mechanisms underlying osteonecrosis of the femoral head (ONFH), with the goal of discovering effective diagnostic and therapeutic targets. Microarray data were sourced from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) were identified using the Limma method. Feature genes were selected through Support Vector Machine, Random Forest and Least Absolute Shrinkage and Selection Operator algorithms. Diagnostic performance was assessed using receiver operating characteristic (ROC) analysis. Functional enrichment and immune infiltration analyses were performed. Experimental validation included RT-qPCR, western blotting and immunofluorescence in ONFH bone tissue. Additionally, a competing endogenous RNA (ceRNA) network was constructed using Cytoscape (v3.10.1). A total of 51 DEGs were identified, comprising 33 downregulated and 18 upregulated genes. Aldehyde dehydrogenase 5 family member A1 (ALDH5A1) consistently emerged as a key gene across all machine learning models, demonstrating high diagnostic value according to ROC analysis. Enrichment analysis indicated that ALDH5A1-related DEGs were predominantly involved in 'Toll-like receptor signaling', 'RIG-like receptor signaling', 'Leishmania infection' and 'Cytosolic DNA sensing'. Immune analysis revealed associations between ONFH and HLA/MHC-I molecules, neutrophils and regulatory T cells, with ALDH5A1 showing a negative correlation with activated CD4