AbstractIn our previous study, IRX5 has been revealed a significant role in adipogenesis of hBMSCs. Considering the expansion of adipose tissue in bone marrow in aged and ovariectomy‐related osteoporosis, the effect of IRX5 on the osteogenesis of BMSCs still needs to be elucidated. In vivo, models of aging‐induced and ovariectomy‐induced osteoporotic mice, and in vitro studies of IRX5 gene gain‐ and loss‐of‐function in hBMSCs were employed. Histology, immunofluorescence, qRT‐PCR, and Western blot analysis were performed to detect the functions of IRX5 in hBMSCs osteogenic differentiation. RNA‐seq, transmission electron microscopy, Seahorse mito‐stress assay, and Surface Sensing of Translation assay were conducted to explore the effect of mammalian/mechanistic target of rapamycin (mTOR)‐mediated ribosomal translation and mitochondrial functions in the regulation of hBMSCs differentiation by IRX5. As a result, elevated IRX5 protein expression levels were observed in the bone marrow of osteoporotic mice compared to normal mice. IRX5 overexpression attenuated osteogenic processes, whereas IRX5 knockdown resulted in enhanced osteogenesis in hBMSCs. RNA‐seq and enrichment analysis unveiled that IRX5 overexpression exerted inhibitory effects on ribosomal translation and mitochondrial functions. Furthermore, the application of the mTOR activator, MHY1485, effectively reversed the inhibitory impact of IRX5 on osteogenesis and mitochondrial functions in hBMSCs. In summary, our findings suggest that IRX5 restricts mTOR‐mediated ribosomal translation, consequently impairing mitochondrial OxPhos, which in turn results in osteogenic dysfunction of hBMSCs.