RESUMO
This study aimed to explore how mechanical stress affects osteogenic differentiation via the miR-187-3p/CNR2 pathway. To conduct this study, 24 female C57BL/6 mice, aged 8 weeks, were used and divided into four groups. The Sham and OVX groups did not undergo treadmill exercise, while the Sham + EX and OVX + EX groups received a 8-week treadmill exercise. Post-training, bone marrow and fresh femur samples were collected for further analysis. Molecular biology analysis, histomorphology analysis, and micro-CT analysis were conducted on these samples. Moreover, primary osteoblasts were cultured under osteogenic conditions and divided into GM group and CTS group. The cells in the CTS group underwent a sinusoidal stretching regimen for either 3 or 7 days. The expression of early osteoblast markers (Runx2, OPN, and ALP) was measured to assess differentiation. The study findings revealed that mechanical stress has a regulatory impact on osteoblast differentiation. The expression of miR-187-3p was observed to decrease, facilitating osteogenic differentiation, while the expression of CNR2 increased significantly. These observations suggest that mechanical stress, miR-187-3p, and CNR2 play crucial roles in regulating osteogenic differentiation. Both in vivo and in vitro experiments have confirmed that mechanical stress downregulates miR-187-3p and upregulates CNR2, which leads to the restoration of distal femoral bone mass and enhancement of osteoblast differentiation. Therefore, mechanical stress promotes osteoblasts, resulting in improved osteoporosis through the miR-187-3p/CNR2 signaling pathway. These findings have broad prospect and provide molecular biology guidance for the basic research and clinical application of exercise in the prevention and treatment of PMOP.
Assuntos
Diferenciação Celular , MicroRNAs , Osteoblastos , Osteogênese , Osteoporose Pós-Menopausa , Estresse Mecânico , Animais , Feminino , Humanos , Camundongos , Células Cultivadas , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Osteoblastos/metabolismo , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/terapia , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/patologia , Transdução de SinaisRESUMO
Rationale: NLRP3 inflammasome is critical in the development and progression of many metabolic diseases driven by chronic inflammation, but its effect on the pathology of postmenopausal osteoporosis (PMOP) remains poorly understood. Methods: We here firstly examined the levels of NLRP3 inflammasome in PMOP patients by ELISA. Then we investigated the possible mechanisms underlying the effect of NLRP3 inflammasome on PMOP by RNA sequencing of osteoblasts treated with NLRP3 siRNA and qPCR. Lastly, we accessed the effect of decreased NLRP3 levels on ovariectomized (OVX) rats. To specifically deliver NLRP3 siRNA to osteoblasts, we constructed NLRP3 siRNA wrapping osteoblast-specific aptamer (CH6)-functionalized lipid nanoparticles (termed as CH6-LNPs-siNLRP3). Results: We found that the levels of NLRP3 inflammasome were significantly increased in patients with PMOP, and were negatively correlated with estradiol levels. NLRP3 knock-down influenced signal pathways including immune system process, interferon signal pathway. Notably, of the top ten up-regulated genes in NLRP3-reduced osteoblasts, nine genes (except Mx2) were enriched in immune system process, and five genes were related to interferon signal pathway. The in vitro results showed that CH6-LNPs-siNLRP3 was relatively uniform with a dimeter of 96.64 ± 16.83 nm and zeta potential of 38.37 ± 1.86 mV. CH6-LNPs-siNLRP3 did not show obvious cytotoxicity and selectively delivered siRNA to bone tissue. Moreover, CH6-LNPs-siNLRP3 stimulated osteoblast differentiation by activating ALP and enhancing osteoblast matrix mineralization. When administrated to OVX rats, CH6-LNPs-siNLRP3 promoted bone formation and bone mass, improved bone microarchitecture and mechanical properties by decreasing the levels of NLRP3, IL-1ß and IL-18 and increasing the levels of OCN and Runx2. Conclusion: NLRP3 inflammasome may be a new biomarker for PMOP diagnosis and plays a key role in the pathology of PMOP. CH6-LNPs-siNLRP3 has potential application for the treatment of PMOP.
Assuntos
Inflamassomos , Lipossomos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Nanopartículas , Osteoblastos , Osteoporose Pós-Menopausa , Animais , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Feminino , Humanos , Ratos , Inflamassomos/metabolismo , Nanopartículas/química , Osteoporose Pós-Menopausa/metabolismo , Regulação para Baixo/efeitos dos fármacos , Ratos Sprague-Dawley , RNA Interferente Pequeno/administração & dosagem , Aptâmeros de Nucleotídeos/farmacologia , Aptâmeros de Nucleotídeos/administração & dosagem , Modelos Animais de Doenças , Pessoa de Meia-Idade , OvariectomiaRESUMO
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 AND AIMS: Postmenopausal osteoporosis (PMO) and type 2 diabetes mellitus (T2DM) frequently coexist in postmenopausal women. The study aimed to explore metabolic variations linked to these circumstances and their simultaneous presence through proton nuclear magnetic resonance metabolomics (1H NMR). MATERIALS AND METHODS: Serum samples from 80 postmenopausal women, including 20 PMO individuals, 20 T2DM, 20 T2DM + PMO, and 20 healthy postmenopausal women, were analyzed using 1H NMR spectroscopy. RESULTS: Our study revealed significant metabolic profile differences among the four groups. Notably, the T2DM + PMO group showed elevated levels of alanine, pyruvate, glutamate, lactate, and aspartate, indicating their involvement in lipid metabolism, energy, and amino acids. Importantly, our multivariate statistical analysis identified a metabolite set that accurately distinguished the groups, suggesting its potential as an early diagnostic marker. CONCLUSION: The 1H NMR metabolomics approach uncovered metabolic biomarkers intricately linked to postmenopausal osteoporosis (PMO), type 2 diabetes mellitus (T2DM), and their concurrent presence. Among these biomarkers, alanine emerged as a pivotal player, showing its significant role in the metabolic landscape associated with PMO and T2DM. These findings shed light on the pathophysiological mechanisms underlying these conditions and underscore alanine's potential as a diagnostic biomarker.
Assuntos
Biomarcadores , Diabetes Mellitus Tipo 2 , Metabolômica , Osteoporose Pós-Menopausa , Humanos , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/diagnóstico , Feminino , Pessoa de Meia-Idade , Biomarcadores/sangue , Metabolômica/métodos , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/sangue , Osteoporose Pós-Menopausa/diagnóstico , Idoso , Espectroscopia de Ressonância Magnética/métodos , MetabolomaRESUMO
OBJECTIVE: To explore the multi-component synergistic mechanism of Zuogui Wan (, ZGW) in treating postmenopausal osteoporosis (PMOP). METHODS: The main components and target genes of ZGW were screened via the Traditional Chinese Medicine Systems Pharmacology (TCMSP). In addition, the target gene sets of PMOP were derived from the GeneCards and Online Mendelian Inheritance in Man databases. The search tool for recurring instances of neighbouring genes (STRING) 11.0 software was used to analyze the interaction among intersecting genes. Cytoscape 3.6.1 software and the Matthews correlation coefficient (MCC) algorithm were used to screen the core genes. Fifty Sprague-Dawley female rats were randomly divided into the sham-operated (Sham) group and the four ovariectomized (OVX) subgroups. Rats subjected to Sham or OVX were administered with the vehicle (OVX, 1 mL water/100 g weight), 17ß-estradiol (E2, 50 µg·kg-1·d-1), and lyophilized powder of ZGW at a low dose of 2.3 (ZGW-L) and high dose of 4.6 (ZGW-H) g·kg-1·d-1 for three months. The bone density and bone strength were assessed using dual-energy X-ray and three-point bending tests, respectively. Furthermore, enzyme-linked immun-osorbent assay, Hematoxylin-eosin staining, and western blot analysis were used to determine the potential pharmacological mechanisms of action of ZGW in PMOP. RESULTS: A total of 117 active compounds of ZGW were screened from the TCMSP. Furthermore, 108 intersecting genes of drugs and diseases were identified. Using STRING software and the MCC algorithm, ten core genes, including C-X-C chemokine living 8 (CXCL8), C-C chemokine receptor type 2 (CCR2), alpha-2a active receptor (ADRA2A), melatonin receptor type 1B (MTNR1B), and amyloid-beta A4 protein (APP), were identified. The anti-osteoporosis regulation network of ZGW was constructed using the Cytoscape software. The animal experiments demonstrated that ZGW groups significantly reduced the serum levels of ß-C-terminal telopeptide of type I collagen (ß-CTX) and increased serum levels of bone-specific alkaline phosphatase (BALP) (P < 0.05, P < 0.01). The OVX group exhibited a significant decrease in bone mineral density and bone strength compared with the Sham group (P < 0.01). Moreover, treatment with ZGW resulted in increased trabecular thickness, improved arrangement of trabecular structure, and reduced empty bone lacunae. Furthermore, treatment with ZGW significantly increased the protein expression of CXCL8, ADRA2A, and CCR2 (P < 0.05, P < 0.01), and significantly decreased the protein expression of Runx2 (P < 0.01). Furthermore, the ZGW and E2 groups demonstrated significantly increased BMD (P < 0.05, P < 0.01), improved bone strength (P < 0.05, P < 0.01), reduced expression of CXCL8, ADRA2A, and CCR2, and increased runt-related transcription factor 2 levels in bone tissue (P < 0.05, P < 0.01) compared with the OVX group. However, there were no significant differences in MTNR1B and APP expression among the groups. CONCLUSION: ZGW shows synergistic mechanisms in PMOP through multiple components, targets, and pathways.
Assuntos
Densidade Óssea , Medicamentos de Ervas Chinesas , Osteoporose Pós-Menopausa , Ratos Sprague-Dawley , Medicamentos de Ervas Chinesas/administração & dosagem , Feminino , Animais , Osteoporose Pós-Menopausa/tratamento farmacológico , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/fisiopatologia , Osteoporose Pós-Menopausa/metabolismo , Ratos , Humanos , Densidade Óssea/efeitos dos fármacosRESUMO
Menopause is a normal physiological process accompanied by changes in various physiological states. The incidence of vascular calcification (VC) increases each year after menopause and is closely related to osteoporosis (OP). Although many studies have investigated the links between VC and OP, the interaction mechanism of the two under conditions of estrogen loss remains unclear. MicroRNAs (miRNAs), which are involved in epigenetic modification, play a critical role in estrogen-mediated mineralization. In the past several decades, miRNAs have been identified as biomarkers or therapeutic targets in diseases. Thus, we hypothesize that these small molecules can provide new diagnostic and therapeutic approaches. In this review, we summarize the close interactions between VC and OP and the role of miRNAs in their interplay.
Assuntos
MicroRNAs , Pós-Menopausa , Calcificação Vascular , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Feminino , Calcificação Vascular/genética , Calcificação Vascular/metabolismo , Pós-Menopausa/genética , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/metabolismo , Estrogênios/metabolismo , Biomarcadores/metabolismo , Osteoporose/genética , Osteoporose/metabolismo , Epigênese GenéticaRESUMO
Postmenopausal osteoporosis (PMOP) is a major public health problem worldwide, afflicting many postmenopausal women. Although many studies have focused on the biological role of individual lipids in osteoporosis, no studies have systematically elucidated the lipid profile of osteoporosis. In this study, liquid chromatography-tandem mass spectrometry (LC-MS/MS) technology based on multiple reaction monitoring (MRM) method was used to compare the levels of lipid molecules in bone marrow cells of osteoporotic mice (OVX) group and sham-operation (Sham) group. Principal component analysis (PCA) was used for multivariate statistics. Differential lipids were obtained by bar graph, heatmap and volcano map. A total of 400 lipid molecules were identified. A total of 199 lipid molecules were identified to be associated with PMOP, including 6 phospholipids and 3 sphingolipids. These differential lipid molecules provide a systematic lipid profile for osteoporosis, which helps to discover new candidate osteoporosis biomarkers, and their changes at the molecular level can be used as new targets for diagnosis or prevention.
Assuntos
Modelos Animais de Doenças , Lipidômica , Lipídeos , Osteoporose Pós-Menopausa , Espectrometria de Massas em Tandem , Animais , Lipidômica/métodos , Feminino , Camundongos , Osteoporose Pós-Menopausa/metabolismo , Espectrometria de Massas em Tandem/métodos , Lipídeos/análise , Medula Óssea/metabolismo , Cromatografia Líquida/métodos , Análise de Componente Principal , Biomarcadores/análise , Camundongos Endogâmicos C57BL , Humanos , Ovariectomia , Células da Medula Óssea/metabolismoRESUMO
Osteoporosis (OP) is a metabolic bone disease that can lead to major health challenges. The theory of Traditional Chinese medicine believes that kidney-Yin deficiency (KYD) is the main cause of postmenopausal osteoporosis. This study was aimed to investigate the effect of EZW on anti-osteoporosis with KYD, and explore potential mechanisms from the perspective of the kidney, bone and bone marrow through analysis of metabolomics and proteomics. The model of OP with KYD was established by rats treated with bilateral ovariectomy (OVX), and then given intragastric administration of thyroid and reserpine to induce. Micro-CT was applied to determine the microstructures of bone. Serum levels associated with bone turnover markers and kidney-Yin deficiency were detected by enzyme-linked immunosorbent (ELISA) assay. The differential metabolites in the kidney, bone and bone marrow were analyzed by metabolomics. The differentially expressed proteins in these three tissues were detected via proteomics. The findings suggested that EZW could alleviate a variety of metabolites and proteins among the kidney, bone and bone marrow, primarily in amino acid metabolism, carbohydrate metabolism, nucleotide metabolism and lipid metabolism, thus leading to improvements of OP with KYD, which provided theoretical basis for clinical treatment of EZW on OP with KYD.
Assuntos
Medula Óssea , Medicamentos de Ervas Chinesas , Rim , Metabolômica , Osteoporose , Ovariectomia , Proteômica , Ratos Sprague-Dawley , Deficiência da Energia Yin , Animais , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Feminino , Ratos , Medula Óssea/efeitos dos fármacos , Medula Óssea/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Proteômica/métodos , Metabolômica/métodos , Deficiência da Energia Yin/tratamento farmacológico , Osteoporose/tratamento farmacológico , Osteoporose/metabolismo , Osso e Ossos/metabolismo , Osso e Ossos/efeitos dos fármacos , Modelos Animais de Doenças , Osteoporose Pós-Menopausa/tratamento farmacológico , Osteoporose Pós-Menopausa/metabolismo , Microtomografia por Raio-X/métodos , Medicina Tradicional Chinesa/métodos , MultiômicaRESUMO
Senescence in bone marrow mesenchymal stem cells (BMSCs), triggered by excessive oxidative stress, plays a crucial role in the onset of postmenopausal osteoporosis. Recent studies underscore the importance of mitochondrial rehabilitation and quality control as key determinants in the modulation of oxidative stress and cellular senescence. MitoTEMPO, a mitochondria-targeted antioxidant, has been shown to mitigate the heightened levels of reactive oxygen species (ROS). In our research, we observed that BMSCs from ovariectomized (OVX) rats displayed premature senescence, which was attributed to combined mitochondrial and lysosomal dysfunction, a condition that worsens with extended estrogen deprivation. Treatment with MitoTEMPO effectively reversed these effects, reinstating lysosomal functionality and suppressing the mitochondrial unfolded protein response (UPRmt). Subsequent in vivo experiments corroborated these observations, revealing that MitoTEMPO administration in OVX rats curtailed trabecular bone loss and reduced the expression of p53, HSP60, and CLPP in the trabecular bone region of the proximal tibia. Overall, our findings suggest that MitoTEMPO holds promise as a therapeutic agent to counteract senescence in OVX-BMSCs, offering a potential strategy for treating postmenopausal osteoporosis.
Assuntos
Antioxidantes , Senescência Celular , Células-Tronco Mesenquimais , Mitocôndrias , Ovariectomia , Estresse Oxidativo , Espécies Reativas de Oxigênio , Animais , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Feminino , Senescência Celular/efeitos dos fármacos , Antioxidantes/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Ratos , Ratos Sprague-Dawley , Compostos Organotiofosforados/farmacologia , Lisossomos/metabolismo , Lisossomos/efeitos dos fármacos , Resposta a Proteínas não Dobradas/efeitos dos fármacos , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/patologia , Proteína Supressora de Tumor p53/metabolismo , Humanos , Compostos Organofosforados , PiperidinasRESUMO
OBJECTIVE: We aimed to investigate the effect of Lycium barbarum polysaccharide (LBP) on the proliferation and differentiation of osteoblasts in postmenopausal individuals with osteoporosis using in vitro cell experiments. METHODS: We assessed the effect of long-term LBP consumption on the intestinal metabolites of individuals using a simulation of the human intestinal microbiota ecosystem. We also tested the capacity of LBP in proliferating MC3T3-E1 cells using the cell counting kit-8 (CCK-8) method and analyzed the effect of intestinal metabolites on the osteogenic differentiation of MC3T3-E1 cells by testing bone metabolism viability with relevant indicators. RESULTS: The level of short-chain fatty acids (SCFAs) significantly increased (p < 0.05), and the concentrations of acetic acid, propionic acid, and butyric acid all showed an upward trend after the treatment using LBP. At appropriate concentrations, the fermentation supernatant can enhance osteoblast proliferation by significantly increasing the active expression of bone-alkaline phosphatase (B-ALP) and osteocalcin (OCN) in osteoblasts (p < 0.05). CONCLUSION: By modulating the metabolites of intestinal microbiota, production of SCFAs, the prebiotic properties of LBP can enhance osteoblast differentiation through in vitro simulation experiment and cell-based assay.
Assuntos
Diferenciação Celular , Proliferação de Células , Medicamentos de Ervas Chinesas , Osteoblastos , Osteoporose Pós-Menopausa , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Humanos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Camundongos , Animais , Osteoporose Pós-Menopausa/tratamento farmacológico , Osteoporose Pós-Menopausa/metabolismo , Ácidos Graxos Voláteis/metabolismo , Osteogênese/efeitos dos fármacos , Microbioma Gastrointestinal/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Linhagem Celular , Osteocalcina/metabolismoRESUMO
The role of monocytes in postmenopausal osteoporosis is widely recognized; however, the mechanisms underlying monocyte reprogramming remain unknown. In this study, single-cell RNA sequencing (scRNA-seq) was conducted on CD14+ bone marrow monocytes obtained from 3 postmenopausal women with normal BMD and 3 women with postmenopausal osteoporosis (PMOP). Monocle2 was used to classify the monocytes into 7 distinct clusters. The proportion of cluster 1 significantly decreased in PMOP patients, while the proportion of cluster 7 increased. Further analysis via GSEA, transcription factor activity analysis, and sc-metabolic analysis revealed significant differences between clusters 1 and 7. Cluster 7 exhibited upregulated pathways associated with inflammation, immunity, and osteoclast differentiation, whereas cluster 1 demonstrated the opposite results. Monocle2, TSCAN, VECTOR, and scVelo data indicated that cluster 1 represented the initial subset and that cluster 7 represents one of the terminal subsets. BayesPrism and ssGSEA were employed to analyze the bulk transcriptome data obtained from the GEO database. The observed alterations in the proportions of 1 and 7 were validated and found to have diagnostic significance. CD16 serves as the marker gene for cluster 7, thus leading to an increased proportion of CD16+ monocytes in women with PMOP. Flow cytometry was used to assess the consistency of outcomes with those of the bioinformatic analysis. Subsequently, an additional scRNA-seq analysis was conducted on bone marrow mononuclear cells obtained from 3 patients with PMOP and 3 postmenopausal women with normal BMD. The differential proportions of cluster 1 and cluster 7 were once again confirmed, with the pathological effect of cluster 7 may attribute to cell-cell communication. The scRNA-seq findings suggest that an imbalance in monocyte subsets is a characteristic feature of PMOP. These findings elucidate the limitations of utilizing bulk transcriptome data for detecting alterations in monocytes, which may influence novel research inquiries.
Monocytes are a type of white blood cell that plays a role in postmenopausal osteoporosis (PMOP), a condition where bones become weak and brittle after menopause. However, how monocytes change in this condition is not fully understood. In this study, single-cell RNA sequencing was used to analyze bone marrow monocytes from postmenopausal women with normal bone density and those with osteoporosis. Two distinct types of monocytes were identified, which were called clusters 1 and 7. In women with PMOP, there was a decrease in cluster 1 monocytes and an increase in cluster 7 monocytes. This change was validated in external datasets and in peripheral blood. Further analysis showed that cluster 7 monocytes positively correlated with inflammation, immunity, and osteoclast differentiation (a process that leads to bone resorption). Cluster 1 monocytes were found to be the initial subset, while cluster 7 monocytes were one of the terminal subsets. Overall, this study suggests that an imbalance in monocyte subsets is a characteristic feature of postmenopausal osteoporosis. These findings have important implications for understanding the role of monocytes in bone health.
Assuntos
Monócitos , Osteoporose Pós-Menopausa , Análise de Sequência de RNA , Análise de Célula Única , Humanos , Feminino , Monócitos/metabolismo , Monócitos/patologia , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/sangue , Pessoa de Meia-Idade , Idoso , Regulação da Expressão Gênica , Transcriptoma , Receptores de Lipopolissacarídeos/metabolismoRESUMO
Postmenopausal osteoporosis (OP) is a prevalent skeletal disease with not fully understood molecular mechanisms. This study aims to investigate the role of circular RNA (circRNA) in postmenopausal OP and to elucidate the potential mechanisms of the circRNA-miRNA-mRNA regulatory network. We obtained circRNA and miRNA expression profiles from postmenopausal OP patients from the Gene Expression Omnibus database. By identifying differentially expressed circRNAs and miRNAs, we constructed a circRNA-miRNA-mRNA network and identified key genes associated with OP. Further, through a range of experimental approaches, including dual-luciferase reporter assays, RNA pull-down experiments, and qRT-PCR, we examined the roles of circ_0134120, miR-590-5p, and STAT3 in the progression of OP. Our findings reveal that the interaction between circ_0134120 and miR-590-5p in regulating STAT3 gene expression is a key mechanism in OP, suggesting the circRNA-miRNA-mRNA network is a potential therapeutic target for this condition.
Assuntos
Redes Reguladoras de Genes , MicroRNAs , Osteoporose Pós-Menopausa , RNA Circular , Fator de Transcrição STAT3 , Humanos , RNA Circular/genética , MicroRNAs/genética , Feminino , Osteoporose Pós-Menopausa/genética , Osteoporose Pós-Menopausa/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Regulação da Expressão Gênica , Perfilação da Expressão Gênica , Pessoa de Meia-IdadeRESUMO
By 2060, an estimated one in four Americans will be elderly. Consequently, the prevalence of osteoporosis and fragility fractures will also increase. Presently, no available intervention definitively prevents or manages osteoporosis. This study explores whether Pool 7 Compound 3 (P7C3) reduces progressive bone loss and fragility following the onset of ovariectomy (OVX)-induced osteoporosis. Results confirm OVX-induced weakened, osteoporotic bone together with a significant gain in adipogenic body weight. Treatment with P7C3 significantly reduced osteoclastic activity, bone marrow adiposity, whole-body weight gain, and preserved bone area, architecture, and mechanical strength. Analyses reveal significantly upregulated platelet derived growth factor-BB and leukemia inhibitory factor, with downregulation of interleukin-1 R6, and receptor activator of nuclear factor kappa-B (RANK). Together, proteomic data suggest the targeting of several key regulators of inflammation, bone, and adipose turnover, via transforming growth factor-beta/SMAD, and Wingless-related integration site/be-catenin signaling pathways. To the best of the knowledge, this is first evidence of an intervention that drives against bone loss via RANK. Metatranscriptomic analyses of the gut microbiota show P7C3 increased Porphyromonadaceae bacterium, Candidatus Melainabacteria, and Ruminococcaceae bacterium abundance, potentially contributing to the favorable inflammatory, and adipo-osteogenic metabolic regulation observed. The results reveal an undiscovered, and multifunctional therapeutic strategy to prevent the pathological progression of OVX-induced bone loss.
Assuntos
Modelos Animais de Doenças , Osteoporose Pós-Menopausa , Ovariectomia , Animais , Feminino , Osteoporose Pós-Menopausa/metabolismo , Ratos , Humanos , Ratos Sprague-DawleyRESUMO
Purpose: Metabolic and immune changes in the early stages of osteoporosis are not well understood. This study aimed to explore the changes in bone metabolites and bone marrow lymphocyte subsets and their relationship during the osteoporosis onset. Methods: We established OVX and Sham mouse models. After 5, 15, and 40 days, five mice in each group were sacrificed. Humeri were analyzed by microCT. The bone marrow cells of the left femur and tibia were collected for flow cytometry analysis. The right femur and tibia were analyzed by LC-MS/MS for metabolomics analysis. Results: Bone microarchitecture was significantly deteriorated 15 days after OVX surgery. Analysis of bone metabolomics showed that obvious metabolite changes had happened since 5 days after surgery. Lipid metabolism was significant at the early stage of the osteoporosis. The proportion of immature B cells was increased, whereas the proportion of mature B cells was decreased in the OVX group. Metabolites were significantly correlated with the proportion of lymphocyte subsets at the early stage of the osteoporosis. Conclusion: Lipid metabolism was significant at the early stage of the osteoporosis. Bone metabolites may influence bone formation by interfering with bone marrow lymphocyte subsets.
Assuntos
Osteoporose Pós-Menopausa , Osteoporose , Humanos , Feminino , Camundongos , Animais , Osteoporose Pós-Menopausa/etiologia , Osteoporose Pós-Menopausa/metabolismo , Cromatografia Líquida , Espectrometria de Massas em Tandem , Osteoporose/etiologia , Osteoporose/metabolismo , Modelos Animais de Doenças , Subpopulações de Linfócitos/metabolismoRESUMO
Postmenopausal osteoporosis (PMOP) is a metabolic bone disease that results from overproduction and hyperactivation of osteoclasts caused by insufficient estrogen in women after menopause. Current therapeutic strategies are mainly focused on treating PMOP patients who have already developed severe bone loss or even osteoporotic fractures. Obviously, a better strategy is to prevent PMOP from occurring in the first place. However, such reagents are largely lacking. Piperlongumine (PLM), an amide alkaloid extracted from long pepper Piper longum, exhibits the anti-osteoclastogenic effect in normal bone marrow macrophages (BMMs) and the protective effect against osteolysis induced by titanium particles in mice. This study examined the preventive effect of PLM on PMOP and explored the potential mechanism of this effect using both ovariectomized mice and their primary cells. The result showed that PLM (5 and 10 mg kg-1) administered daily for 6 weeks ameliorated ovariectomy-induced bone loss and osteoclast formation in mice. Further cell experiments showed that PLM directly suppressed osteoclast formation, F-actin ring formation, and osteoclastic resorption pit formation in BMMs derived from osteoporotic mice, but did not obviously affect osteogenic differentiation of bone marrow stromal cells (BMSCs) from these mice. Western blot analysis revealed that PLM attenuated maximal activation of p38 and JNK pathways by RANKL stimulation without affecting acute activation of NF-κB, AKT, and ERK signaling. Furthermore, PLM inhibited expression of key osteoclastogenic transcription factors NFATc1/c-Fos and their target genes (Dcstamp, Atp6v0d2, Acp5, and Oscar). Taken together, our findings suggest that PLM inhibits osteoclast formation and function by suppressing RANKL-induced activation of the p38/JNK-cFos/NFATc1 signaling cascade, thereby preventing ovariectomy-induced osteoporosis in mice. Thus, PLM can potentially be used as an anti-resorption drug or dietary supplement for the prevention of PMOP.
Assuntos
Alcaloides , Benzodioxóis , Reabsorção Óssea , Osteoporose Pós-Menopausa , Osteoporose , Humanos , Feminino , Animais , Camundongos , Osteogênese , Sistema de Sinalização das MAP Quinases , Osteoclastos , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/prevenção & controle , Osteoporose/etiologia , Osteoporose/genética , Diferenciação Celular , NF-kappa B/metabolismo , Osteoporose Pós-Menopausa/metabolismo , Ovariectomia/efeitos adversos , Alcaloides/metabolismo , Ligante RANK/metabolismoRESUMO
Postmenopausal osteoporosis (PMOP) is a common kind of osteoporosis that is associated with excessive osteocyte death and bone loss. Previous studies have shown that TNF-α-induced osteocyte necroptosis might exert a stronger effect on PMOP than apoptosis, and TLR4 can also induce cell necroptosis, as confirmed by recent studies. However, little is known about the relationship between TNF-α-induced osteocyte necroptosis and TLR4. In the present study, we showed that TNF-α increased the expression of TLR4, which promoted osteocyte necroptosis in PMOP. In patients with PMOP, TLR4 was highly expressed at skeletal sites where exists osteocyte necroptosis, and high TLR4 expression is correlated with enhanced TNF-α expression. Osteocytes exhibited robust TLR4 expression upon exposure to necroptotic osteocytes in vivo and in vitro. Western blotting and immunofluorescence analyses demonstrated that TNF-α upregulated TLR4 expression in vitro, which might further promote osteocyte necroptosis. Furthermore, inhibition of TLR4 by TAK-242 in vitro effectively blocked osteocyte necroptosis induced by TNF-α. Collectively, these results suggest a novel TLR4-mediated process of osteocyte necroptosis, which might increase osteocyte death and bone loss in the process of PMOP.
Assuntos
Osteócitos , Osteoporose Pós-Menopausa , Receptor 4 Toll-Like , Fator de Necrose Tumoral alfa , Feminino , Humanos , Necroptose , Osteócitos/metabolismo , Osteoporose Pós-Menopausa/metabolismo , Receptor 4 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Probiotics have been found to have beneficial effects on bone metabolism. In this randomized, double-blind, placebo-controlled trial, the effects of multispecies probiotic supplementation on bone turnover markers were evaluated after 12 weeks. Forty postmenopausal women with osteopenia were included and randomly divided into two groups. The intervention group received multispecies probiotics, while the control group received identical placebo sachets daily. The baseline characteristics of both groups were similar. Still, the median serum bone resorption marker C-terminal telopeptide of type I collagen (CTX) was slightly higher in the multispecies probiotic group than in the placebo group (0.35 (0.12, 0.53) vs. 0.16 (0.06, 0.75); p-value = 0.004). After 12 weeks, the mean difference in serum CTX at baseline versus 12 weeks was significantly different between the multispecies probiotic and placebo groups (-0.06 (-0.29, 0.05) vs. 0.04 (-0.45, 0.67); p-value < 0.001). The multispecies probiotic group showed a significant decrease in serum CTX at 12 weeks compared with baseline (p-value 0.026). However, the placebo group showed no significant change in serum CTX (p-value 0.18). In conclusion, multispecies probiotics may have a preventive effect on bone through their antiresorptive effect in osteopenic postmenopausal women.
Assuntos
Doenças Ósseas Metabólicas , Osteoporose Pós-Menopausa , Probióticos , Humanos , Feminino , Pós-Menopausa , Biomarcadores , Remodelação Óssea , Método Duplo-Cego , Osteoporose Pós-Menopausa/prevenção & controle , Osteoporose Pós-Menopausa/metabolismo , Densidade ÓsseaRESUMO
Ferroptosis is a necrotic form of iron-dependent regulatory cell death. Estrogen withdrawal can interfere with iron metabolism, which is responsible for the pathogenesis of postmenopausal osteoporosis (PMOP). Here, it is demonstrated that estrogen withdrawal induces iron accumulation in the skeleton and the ferroptosis of osteocytes, leading to reduced bone mineral density. Furthermore, the facilitatory effect of ferroptosis of osteocytes is verified in the occurrence and development of postmenopausal osteoporosis is associated with over activated osteoclastogenesis using a direct osteocyte/osteoclast coculture system and glutathione peroxidase 4 (GPX4) knockout ovariectomized mice. In addition, the nuclear factor erythroid derived 2-related factor-2 (Nrf2) signaling pathway is confirmed to be a crucial factor in the ferroptosis of osteocytic cells. Nrf2 regulates the expression of nuclear factor kappa-B ligand (RANKL) by regulating the DNA methylation level of the RANKL promoter mediated by DNA methyltransferase 3a (Dnmt3a), which is as an important mechanism in osteocytic ferroptosis-mediated osteoclastogenesis. Taken together, this data suggests that osteocytic ferroptosis is involved in PMOP and can be targeted to tune bone homeostasis.
Assuntos
Ferroptose , Osteoporose Pós-Menopausa , Camundongos , Humanos , Animais , Feminino , Osteócitos/metabolismo , Osteoporose Pós-Menopausa/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estrogênios/metabolismo , Ferro/metabolismoRESUMO
BACKGROUND: Postmenopausal osteoporosis (PMO) is associated with dysregulation of bone metabolism and gut microbiota. Quinoa is a grain with high nutritional value, and its effects and potential mechanisms on PMO have not been reported yet. Therefore, the purpose of this study is to investigate the bone protective effect of quinoa on ovariectomy (OVX) rats by regulating bone metabolism and gut microbiota. RESULTS: Quinoa significantly improved osteoporosis-related biochemical parameters of OVX rats and ameliorated ovariectomy-induced bone density reduction and trabecular structure damage. Quinoa intervention may repair the intestinal barrier by upregulating the expression of tight junction proteins in the duodenum. In addition, quinoa increased the levels of Firmicutes, and decreased the levels of Bacteroidetes and Prevotella, reversing the dysregulation of the gut microbiota. This may be related to estrogen signaling pathway, secondary and primary bile acid biosynthesis, benzoate degradation, synthesis and degradation of ketone bodies, NOD-like receptor signaling pathway and biosynthesis of tropane, piperidine and pyridine alkaloids. Correlation analysis showed that there is a strong correlation between gut microbiota with significant changes in abundance and parameters related to osteoporosis. CONCLUSION: Quinoa could significantly reverse the high intestinal permeability and change the composition of gut microbiota in OVX rats, thereby improving bone microstructure deterioration and bone metabolism disorder, and ultimately protecting the bone loss of OVX rats. © 2024 Society of Chemical Industry.
Assuntos
Densidade Óssea , Chenopodium quinoa , Microbioma Gastrointestinal , Ovariectomia , Ratos Sprague-Dawley , Animais , Ratos , Feminino , Chenopodium quinoa/química , Densidade Óssea/efeitos dos fármacos , Humanos , Bactérias/classificação , Bactérias/metabolismo , Bactérias/isolamento & purificação , Bactérias/genética , Osteoporose/metabolismo , Osteoporose/prevenção & controle , Osteoporose Pós-Menopausa/metabolismo , Osteoporose Pós-Menopausa/prevenção & controle , Osteoporose Pós-Menopausa/microbiologiaRESUMO
Insufficient bone formation and excessive bone resorption caused by estrogen deficiency are the major factors resulting in the incidence of postmenopausal osteoporosis (PMOP). The existing drugs usually fail to re-establish the osteoblast/osteoclast balance from both sides and generate side-effects owing to the lack of bone-targeting ability. Here, engineered cell-membrane-coated nanogels PNG@mR&C capable of scavenging receptor activator of nuclear factor-κB ligand (RANKL) and responsively releasing therapeutic PTH 1-34 in the bone microenvironment are prepared from RANK and CXCR4 overexpressed bone mesenchymal stem cell (BMSC) membrane-coated chitosan biopolymers. The CXCR4 on the coated-membranes confer bone-targeting ability, and abundant RANK effectively absorb RANKL to inhibit osteoclastogenesis. Meanwhile, the release of PTH 1-34 triggered by osteoclast-mediated acid microenvironment promote osteogenesis. In addition, the dose and frequency are greatly reduced due to the smart release property, prolonged circulation time, and bone-specific accumulation. Thus, PNG@mR&C exhibits satisfactory therapeutic effects in the ovariectomized (OVX) mouse model. This study provides a new paradigm re-establishing the bone metabolic homeostasis from multitargets and shows great promise for the treatment of PMOP.