Unraveling Key m6A Modification Regulators Signatures in Postmenopausal Osteoporosis through Bioinformatics and Experimental Verification.

OBJECTIVE: Bone marrow mesenchymal stem cells (BMSCs) show significant potential for osteogenic differentiation. However, the underlying mechanisms of osteogenic capability in osteoporosis-derived BMSCs (OP-BMSCs) remain unclear. This study aims to explore the impact of YTHDF3 (YTH N6-methyladenosine RNA binding protein 3) on the osteogenic traits of OP-BMSCs and identify potential therapeutic targets to boost their bone formation ability. METHODS: We examined microarray datasets (GSE35956 and GSE35958) from the Gene Expression Omnibus (GEO) to identify potential m6A regulators in osteoporosis (OP). Employing differential, protein interaction, and machine learning analyses, we pinpointed critical hub genes linked to OP. We further probed the relationship between these genes and OP using single-cell analysis, immune infiltration assessment, and Mendelian randomization. Our in vivo and in vitro experiments validated the expression and functionality of the key hub gene. RESULTS: Differential analysis revealed seven key hub genes related to OP, with YTHDF3 as a central player, supported by protein interaction analysis and machine learning methodologies. Subsequent single-cell, immune infiltration, and Mendelian randomization studies consistently validated YTHDF3's significant link to osteoporosis. YTHDF3 levels are significantly reduced in femoral head tissue from postmenopausal osteoporosis (PMOP) patients and femoral bone tissue from PMOP mice. Additionally, silencing YTHDF3 in OP-BMSCs substantially impedes their proliferation and differentiation. CONCLUSION: YTHDF3 may be implicated in the pathogenesis of OP by regulating the proliferation and osteogenic differentiation of OP-BMSCs.

A novel mechanism of Vildagliptin in regulating bone metabolism and mitigating osteoporosis.

Osteoporosis has become a global social problem with the tendency toward the aging population. The challenge in managing osteoporosis is to develop new anti-osteoporosis drugs that target bone anabolism. The purpose of this study was to uncover the novel mechanism of Vildagliptin on bone metabolism. We revealed that Vildagliptin significantly promoted osteogenic differentiation of precursor osteoblasts and bone marrow mesenchymal stem cells (BMSCs). At the same time, it significantly enhanced the polarization of RAW264.7 macrophages to the M2 type and the secretion of osteogenic factors BMP2 and TGF-ß1. This was confirmed by the increased osteogenic differentiation observed in the osteoblast-RAW264.7 co-culture system. Moreover, Vildagliptin significantly enhanced the transformation of BMSCs into the osteogenic morphology in the osteoblast-BMSC co-culture system. Finally, Vildagliptin also inhibited osteoclastic differentiation of RAW 264.7 cells. The potential mechanism underlying these effects involved targeting the GAS6/AXL/ERK5 pathway. In the in vivo study, Vildagliptin significantly alleviated postmenopausal osteoporosis in ovariectomized mice. These findings represent the first comprehensive revelation of the regulatory effect of Vildagliptin on bone metabolism. Specifically, Vildagliptin demonstrates the ability to promote bone anabolism and inhibit bone resorption by simultaneously targeting osteoblasts, BMSCs, and osteoclasts. The bone-protective effects of Vildagliptin were further confirmed in a postmenopausal osteoporosis model. The clinical significance of this study lies in laying a theoretical foundation for bone protection therapy in type-2 diabetes patients with compromised bone conditions or postmenopausal osteoporosis.

Defining the Role of Extrachromosomal DNA Amplifications in Medulloblastoma.

Circular extrachromosomal DNA (ecDNA), a common mechanism of oncogene amplification, has been identified as a major contributor to intratumoral heterogeneity and patient outcomes. In a recent publication in Nature Genetics, Chapman and colleagues further explored the role of ecDNA in the context of medulloblastoma. Using whole-genome sequencing, they found that 18% of the patients carry ecDNA amplification across a 468 medulloblastoma patient cohort. The presence of ecDNA was associated with worse survival. Single-cell FISH imaging and multiomic sequencing revealed that ecDNA copy number displayed a cell-to-cell variability within the sample, contributing to tumor heterogeneity. Furthermore, through sequencing and CRISPRi experiments, the authors uncovered frequent enhancer rewiring events on ecDNA that drive proliferation.

METTL3-mediated m6A modification of SOX4 regulates osteoblast proliferation and differentiation via YTHDF3 recognition.

N6-methyladenosine (m6A), the most prevalent internal modification in mRNA, is related to the pathogenesis of osteoporosis (OP). Although methyltransferase Like-3 (METTL3), an m6A transferase, has been shown to mitigate OP progression, the mechanisms of METTL3-mediated m6A modification in osteoblast function remain unclear. Here, fluid shear stress (FSS) induced osteoblast proliferation and differentiation, resulting in elevated levels of METTL3 expression and m6A modification. Through Methylated RNA Immunoprecipitation Sequencing (MeRIP-seq) and Transcriptomic RNA Sequencing (RNA-seq), SRY (Sex Determining Region Y)-box 4 (SOX4) was screened as a target of METTL3, whose m6A-modified coding sequence (CDS) regions exhibited binding affinity towards METTL3. Further functional experiments demonstrated that knockdown of METTL3 and SOX4 hampered osteogenesis, and METTL3 knockdown compromised SOX4 mRNA stability. Via RNA immunoprecipitation (RIP) assays, we further confirmed the direct interaction between METTL3 and SOX4. YTH N6-Methyladenosine RNA Binding Protein 3 (YTHDF3) was identified as the m6A reader responsible for modulating SOX4 mRNA and protein levels by affecting its degradation. Furthermore, in vivo experiments demonstrated that bone loss in an ovariectomized (OVX) mouse model was reversed through the overexpression of SOX4 mediated by adeno-associated virus serotype 2 (AAV2). In conclusion, our research demonstrates that METTL3-mediated m6A modification of SOX4 plays a crucial role in regulating osteoblast proliferation and differentiation through its recognition by YTHDF3. Our research confirms METTL3-m6A-SOX4-YTHDF3 as an essential axis and potential mechanism in OP.