Pre-B-cell leukemia homeobox 1 () is consistently upregulated in samples with low bone mass.
During osteogenic induction, PBX1 expression declines and inversely correlates with markers of bone formation.
Knockdown of PBX1 increases the expression of key osteogenic factors, enhances enzyme activity related to bone formation, and reduces cell death.
PBX1 binds to the promoter of High mobility group box 1 (), activating its expression.
Overexpression of HMGB1 can partially reverse the increased bone formation resulting from PBX1 knockdown.
Inhibition of PBX1 with TCRS-417 leads to a dose-dependent decrease in both PBX1 and HMGB1, while restoring osteogenic gene expression.
Simplified
BACKGROUND: Osteoporosis is a common skeletal disorder characterized by low bone mass and impaired bone formation, driven in part by defective of bone marrow mesenchymal stem cells (BMSCs).
METHODS: Transcriptomic profiling, molecular biology techniques, functional assays, and pharmacological inhibition were used to investigate the regulatory role and downstream effects of candidate genes during BMSC osteogenic differentiation.
RESULTS: Transcriptome-based screening identified Pre-B-cell leukemia homeobox 1 () as a candidate gene consistently upregulated in samples with low bone mass. During osteogenic induction, PBX1 expression declined progressively, inversely correlating with classical osteogenic markers. Functional experiments demonstrated that PBX1 knockdown enhanced expression of Runt-related transcription factor 2 (RUNX2), Osterix (OSX), and Osteocalcin (OCN), increased Alkaline phosphatase (ALP) activity, and reduced apoptosis, while PBX1 overexpression exerted the opposite effects. Mechanistically, PBX1 directly bound to the proximal promoter of High mobility group box 1 () and transcriptionally activated its expression. Rescue experiments revealed that overexpression of HMGB1 partially reversed the enhanced osteogenesis caused by PBX1 knockdown, confirming HMGB1 as a downstream effector. Time-course transcriptomic analysis and gene network correlation further linked PBX1 to pathways involving inflammation, cellular aging, and stemness maintenance. Pharmacological inhibition of PBX1 with TCRS-417 resulted in dose-dependent downregulation of both PBX1 and HMGB1, while significantly restoring osteogenic gene expression and mineralization capacity, underscoring its therapeutic potential for promoting bone formation in osteoporotic conditions.
CONCLUSION: PBX1 acts as an upstream transcriptional regulator that suppresses osteogenic differentiation in part by activating HMGB1, and pharmacological inhibition of PBX1 restores osteogenic potential.
Key numbers
P = 0.0038
Increase in expression
Statistical significance of expression increase after knockdown.
P < 0.0001
Increase in expression
Statistical significance of expression increase after knockdown.
P < 0.0001
Increase in expression
Statistical significance of expression increase after knockdown.
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Declarations. Ethics approval and consent to participate: Human bone marrow mesenchymal stem cells (hBMSCs) used in this study were obtained from commercial sources. According to the supplier’s documentation, all cells were ethically derived with informed donor consent. The experimental protocols were reviewed and approved by the Ethics Committee of Xiangtan Central Hospital (Project Title: Role of PBX1 in skeletal stem cell senescence and fracture healing in the elderly; Approval No.: 2025JJ81137; Date of Approval: 2025-01-26). All procedures were performed in accordance with the Declaration of Helsinki and relevant institutional guidelines and regulations. Consent for publication: All authors have read and approved the final manuscript, and consent to its publication. Competing interests: The authors declare no competing interests.
