RESUMO
BACKGROUND: Promoting the balance between bone formation and bone resorption is the main therapeutic goal for postmenopausal osteoporosis (PMOP), and bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation plays an important regulatory role in this process. Recently, several long non-coding RNAs (lncRNAs) have been reported to play an important regulatory role in the occurrence and development of OP and participates in a variety of physiological and pathological processes. However, the role of lncRNA tissue inhibitor of metalloproteinases 3 (lncTIMP3) remains to be investigated. METHODS: The characteristics of BMSCs isolated from the PMOP rat model were verified by flow cytometry assay, alkaline phosphatase (ALP), alizarin red and Oil Red O staining assays. Micro-CT and HE staining assays were performed to examine histological changes of the vertebral trabeculae of the rats. RT-qPCR and western blotting assays were carried out to measure the RNA and protein expression levels. The subcellular location of lncTIMP3 was analyzed by FISH assay. The targeting relationships were verified by luciferase reporter assay and RNA pull-down assay. RESULTS: The trabecular spacing was increased in the PMOP rats, while ALP activity and the expression levels of Runx2, Col1a1 and Ocn were all markedly decreased. Among the RNA sequencing results of the clinical samples, lncTIMP3 was the most downregulated differentially expressed lncRNA, also its level was significantly reduced in the OVX rats. Knockdown of lncTIMP3 inhibited osteogenesis of BMSCs, whereas overexpression of lncTIMP3 exhibited the reverse results. Subsequently, lncTIMP3 was confirmed to be located in the cytoplasm of BMSCs, implying its potential as a competing endogenous RNA for miRNAs. Finally, the negative targeting correlations of miR-214 between lncTIMP3 and Smad4 were elucidated in vitro. CONCLUSION: lncTIMP3 may delay the progress of PMOP by promoting the activity of BMSC, the level of osteogenic differentiation marker gene and the formation of calcium nodules by acting on the miR-214/Smad4 axis. This finding may offer valuable insights into the possible management of PMOP.
Assuntos
Diferenciação Celular , Células-Tronco Mesenquimais , MicroRNAs , Osteogênese , Osteoporose Pós-Menopausa , RNA Longo não Codificante , Proteína Smad4 , Animais , Feminino , Humanos , Ratos , Células da Medula Óssea/metabolismo , Diferenciação Celular/genética , Modelos Animais de Doenças , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Osteogênese/genética , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/patologia , Ratos Sprague-Dawley , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Proteína Smad4/metabolismo , Proteína Smad4/genética , Inibidor Tecidual de Metaloproteinase-3/genéticaRESUMO
BACKGROUND: Senile osteoporosis (SOP) is an age-related metabolic bone disease that currently lacks specific therapeutic interventions. Thus, this study aimed to investigate the effect of Astragaloside IV (AS-IV) on macrophage senescence, bone marrow mesenchymal stem cell (BMSC) osteogenesis, and SOP progression. METHODS: A senescent macrophage model was established and treated with varying concentrations of AS-IV. Cell activity was measured using the CCK8 assay. The senescence levels of macrophages were evaluated through ß-galactosidase staining, PCR, and immunofluorescence. Macrophage mitochondrial function was assessed using ROS and JC-1 staining. Macrophage polarization was evaluated through PCR, Western blot, and immunofluorescence. The inhibitory effects of AS-IV on macrophage senescence were investigated using Western blot analysis. Furthermore, the effects of macrophage conditioned medium (CM) on BMSCs osteogenic were detected using ALP, alizarin red, and PCR. RESULTS: AS-IV inhibited macrophage senescence and M1 polarization, alleviated mitochondrial dysfunction, and promoted M2 polarization. Mechanistically, it suppressed the STING/NF-κB pathway in H2O2-activated macrophages. Conversely, the STING agonist c-di-GMP reversed the effects of AS-IV on macrophage senescence. Additionally, AS-IV-induced macrophage CM promoted BMSC osteogenic differentiation. In vivo, AS-IV treatment ameliorated aberrant bone microstructure and bone mass loss in the SOP mouse model, inhibited macrophage senescence, and promoted M2 polarization. CONCLUSIONS: By modulating the STING/NF-κB signaling pathway, AS-IV potentially inhibited macrophage senescence and stimulated osteogenic differentiation of BMSCs, thus exerting an anti-osteoporotic effect. Consequently, AS-IV may serve as an effective therapeutic candidate for the treatment of osteoporosis.
