Abstract

Introduction Senile osteoporosis (SOP) is an age-related skeletal disorder characterized by progressive bone mineral density loss and deteriorated bone microarchitecture, imposing significant burdens on aging populations. Impaired osteogenesis of mesenchymal stem cells (MSCs) is a critical feature of SOP, yet its intrinsic mechanisms remain incompletely understood. Objectives This study aimed to investigate the underlying mechanism responsible for the impaired osteogenesis of SOP-MSCs, and to explore the potential therapeutic target for SOP. Methods We integrated multi-omics sequencing including WGBS, CUT&Tag, scRNA-seq, and bulk RNA-seq to assess ubiquitin-like with PHD and RING finger domains 1 (UHRF1)-mediated alterations in the epigenetic landscape within SOP-MSCs. In vitro experiments including co-immunoprecipitation (co-IP), western blot, transmission electron microscopy (TEM), and immunofluorescence were employed for exploring the mechanism regulating autophagy. The therapeutic potential was evaluated in an SOP mouse model using a bone-targeting recombinant adeno-associated virus 9 (rAAV9). Results UHRF1, a DNA methylation regulator, was significantly downregulated in SOP-MSCs, which resulted in reduced DNA 5-mC levels and contributed to impaired osteogenesis in SOP. In terms of mechanism, UHRF1 deficiency-mediated lower DNA 5-mC levels impeded histone deacetylase 1 (HDAC1) recruitment, leading to elevated H3K27ac levels and the super-enhancers (SEs) redistribution. This epigenetic shift promoted aberrant SE formation and the overexpression of transglutaminase 2 (TGM2), which impaired autophagic flux by interfering with the interaction between Beclin1 and the endoplasmic reticulum and therefore impeded the osteogenic differentiation of SOP-MSCs. Targeting the UHRF1-TGM2 axis rescues senile osteoporosis in mice. Conclusion Our findings reveal a novel interaction among DNA 5-mC modification, SE landscapes and autophagic flux in MSC osteogenesis, and clarify th