Bone regeneration and repair are crucial to ambulation and quality of life. Factors such as poor general health, serious medical comorbidities, chronic inflammation, and ageing can lead to delayed healing and nonunion of fractures, and persistent bone defects. Bioengineering strategies to heal bone often involve grafting of autologous bone marrow aspirate concentrate (BMAC) or mesenchymal stem cells (MSCs) with biocompatible scaffolds. While BMAC shows promise, variability in its efficacy exists due to discrepancies in MSC concentration and robustness, and immune cell composition. Understanding the mechanisms by which macrophages and lymphocytes – the main cellular components in BMAC – interact with MSCs could suggest novel strategies to enhance bone healing. Macrophages are polarized into pro-inflammatory (M1) or anti-inflammatory (M2) phenotypes, and influence cell metabolism and tissue regeneration via the secretion of cytokines and other factors. T cells, especially helper T1 (Th1) and Th17, promote inflammation and osteoclastogenesis, whereas Th2 and regulatory T (Treg) cells have anti-inflammatory pro-reconstructive effects, thereby supporting osteogenesis. Crosstalk among macrophages, T cells, and MSCs affects the bone microenvironment and regulates the local immune response. Manipulating the proportion and interactions of these cells presents an opportunity to alter the local regenerative capacity of bone, which potentially could enhance clinical outcomes. Cite this article:
To investigate the effects of senescent osteocytes on bone homeostasis in the progress of age-related osteoporosis and explore the underlying mechanism. In a series of in vitro experiments, we used tert-Butyl hydroperoxide (TBHP) to induce senescence of MLO-Y4 cells successfully, and collected conditioned medium (CM) and senescent MLO-Y4 cell-derived exosomes, which were then applied to MC3T3-E1 cells, separately, to evaluate their effects on osteogenic differentiation. Furthermore, we identified differentially expressed microRNAs (miRNAs) between exosomes from senescent and normal MLO-Y4 cells by high-throughput RNA sequencing. Based on the key miRNAs that were discovered, the underlying mechanism by which senescent osteocytes regulate osteogenic differentiation was explored. Lastly, in the in vivo experiments, the effects of senescent MLO-Y4 cell-derived exosomes on age-related bone loss were evaluated in male SAMP6 mice, which excluded the effects of oestrogen, and the underlying mechanism was confirmed.Aims
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Transcription factor nuclear factor kappa B (NF-κB) plays a major role in the pathogenesis of chronic inflammatory diseases in all organ systems. Despite its importance, NF-κB targeted drug therapy to mitigate chronic inflammation has had limited success in preclinical studies. We hypothesized that sex differences affect the response to NF-κB treatment during chronic inflammation in bone. This study investigated the therapeutic effects of NF-κB decoy oligodeoxynucleotides (ODN) during chronic inflammation in male and female mice. We used a murine model of chronic inflammation induced by continuous intramedullary delivery of lipopolysaccharide-contaminated polyethylene particles (cPE) using an osmotic pump. Specimens were evaluated using micro-CT and histomorphometric analyses. Sex-specific osteogenic and osteoclastic differentiation potentials were also investigated in vitro, including alkaline phosphatase, Alizarin Red, tartrate-resistant acid phosphatase staining, and gene expression using reverse transcription polymerase chain reaction (RT-PCR).Aims
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Currently, the effect of drug treatment for osteoporosis is relatively poor, and the side effects are numerous and serious. Melatonin is a potential drug to improve bone mass in postmenopausal women. Unfortunately, the mechanism by which melatonin improves bone metabolism remains unclear. The aim of this study was to further investigate the potential mechanism of melatonin in the treatment of osteoporosis. The effects of melatonin on mitochondrial apoptosis protein, bmal1 gene, and related pathway proteins of RAW264.7 (mouse mononuclear macrophage leukaemia cells) were analyzed by western blot. Cell Counting Kit-8 was used to evaluate the effect of melatonin on cell viability. Flow cytometry was used to evaluate the effect of melatonin on the apoptosis of RAW264.7 cells and mitochondrial membrane potential. A reactive oxygen species (ROS) detection kit was used to evaluate the level of ROS in osteoclast precursors. We used bmal1-small interfering RNAs (siRNAs) to downregulate the Aims
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Astragalus polysaccharide (APS) participates in various processes, such as the enhancement of immunity and inhibition of tumours. APS can affect osteoporosis (OP) by regulating the osteogenic differentiation of human bone mesenchymal stem cells (hBMSCs). This study was designed to elucidate the mechanism of APS in hBMSC proliferation and osteoblast differentiation. Reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting were performed to determine the expression of microRNA (miR)-760 and ankyrin repeat and FYVE domain containing 1 (ANKFY1) in OP tissues and hBMSCs. Cell viability was measured using the Cell Counting Kit-8 assay. The expression of cyclin D1 and osteogenic marker genes (osteocalcin (OCN), alkaline phosphatase (ALP), and runt-related transcription factor 2 (RUNX2)) was evaluated using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Mineral deposits were detected through Alizarin Red S staining. In addition, Western blotting was performed to detect the ANKFY1 protein levels following the regulation of miR-760. The relationship between miR-760 and ANKFY1 was determined using a luciferase reporter assay.Aims
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Long non-coding RNAs (lncRNAs) act as crucial regulators in osteoporosis (OP). Nonetheless, the effects and potential molecular mechanism of lncRNA PCBP1 Antisense RNA 1 (PCBP1-AS1) on OP remain largely unclear. The aim of this study was to explore the role of lncRNA PCBP1-AS1 in the pathogenesis of OP. Using quantitative real-time polymerase chain reaction (qRT-PCR), osteogenesis-related genes (alkaline phosphatase (ALP), osteocalcin (OCN), osteopontin (OPN), and Runt-related transcription factor 2 (RUNX2)), PCBP1-AS1, microRNA (miR)-126-5p, group I Pak family member p21-activated kinase 2 (PAK2), and their relative expression levels were determined. Western blotting was used to examine the expression of PAK2 protein. Cell Counting Kit-8 (CCK-8) assay was used to measure cell proliferation. To examine the osteogenic differentiation, Alizarin red along with ALP staining was used. RNA immunoprecipitation assay and bioinformatics analysis, as well as a dual-luciferase reporter, were used to study the association between PCBP1-AS1, PAK2, and miR-126-5p.Aims
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It has been established that mechanical stimulation benefits tendon-bone (T-B) healing, and macrophage phenotype can be regulated by mechanical cues; moreover, the interaction between macrophages and mesenchymal stem cells (MSCs) plays a fundamental role in tissue repair. This study aimed to investigate the role of macrophage-mediated MSC chondrogenesis in load-induced T-B healing in depth. C57BL/6 mice rotator cuff (RC) repair model was established to explore the effects of mechanical stimulation on macrophage polarization, transforming growth factor (TGF)-β1 generation, and MSC chondrogenesis within T-B enthesis by immunofluorescence and enzyme-linked immunosorbent assay (ELISA). Macrophage depletion was performed by clodronate liposomes, and T-B healing quality was evaluated by histology and biomechanics. In vitro, bone marrow-derived macrophages (BMDMs) were stretched with CELLOAD-300 load system and macrophage polarization was identified by flow cytometry and quantitative real-time polymerase chain reaction (qRT-PCR). MSC chondrogenic differentiation was measured by histochemical analysis and qRT-PCR. ELISA and qRT-PCR were performed to screen the candidate molecules that mediated the pro-chondrogenic function of mechanical stimulated BMDMs.Aims
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This study examined whether systemic administration of melatonin would have different effects on osseointegration in ovariectomized (OVX) rats, depending on whether this was administered during the day or night. In this study, a titanium rod was implanted in the medullary cavity of one femoral metaphysis in OVX rats, and then the rats were randomly divided into four groups: Sham group (Sham, n = 10), OVX rat group (OVX, n = 10), melatonin day treatment group (OVX + MD, n = 10), and melatonin night treatment group (OVX + MN, n = 10). The OVX + MD and OVX + MN rats were treated with 30 mg/kg/day melatonin at 9 am and 9 pm, respectively, for 12 weeks. At the end of the research, the rats were killed to obtain bilateral femora and blood samples for evaluation.Aims
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The involvement of cyclin D1 in the proliferation of microglia, and the generation and maintenance of bone cancer pain (BCP), have not yet been clarified. We investigated the expression of microglia and cyclin D1, and the influences of cyclin D1 on pain threshold. Female Sprague Dawley (SD) rats were used to establish a rat model of BCP, and the messenger RNA (mRNA) and protein expression of ionized calcium binding adaptor molecule 1 (IBA1) and cyclin D1 were detected by reverse transcription-polymerase chain reaction (RT-PCR) and western blot, respectively. The proliferation of spinal microglia was detected by immunohistochemistry. The pain behaviour test was assessed by quantification of spontaneous flinches, limb use, and guarding during forced ambulation, mechanical paw withdrawal threshold, and thermal paw withdrawal latency.Aims
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Aims. Tissue inhibitors of metalloproteinases (TIMPs) are the endogenous inhibitors of the zinc-dependent matrix metalloproteinases (MMP) and A disintegrin and metalloproteinases (ADAM) involved in extracellular matrix modulation. The present study aims to develop the TIMPs as
Aims. We aimed to develop a gene signature that predicts the occurrence of postmenopausal osteoporosis (PMOP) by studying its genetic mechanism. Methods. Five datasets were obtained from the Gene Expression Omnibus database. Unsupervised consensus cluster analysis was used to determine new PMOP subtypes. To determine the central genes and the core modules related to PMOP, the weighted gene co-expression network analysis (WCGNA) was applied. Gene Ontology enrichment analysis was used to explore the
This study investigated the effects of β-caryophyllene (BCP) on protecting bone from vitamin D deficiency in mice fed on a diet either lacking (D-) or containing (D+) vitamin D. A total of 40 female mice were assigned to four treatment groups (n = 10/group): D+ diet with propylene glycol control, D+ diet with BCP, D-deficient diet with control, and D-deficient diet with BCP. The D+ diet is a commercial basal diet, while the D-deficient diet contains 0.47% calcium, 0.3% phosphorus, and no vitamin D. All the mice were housed in conditions without ultraviolet light. Bone properties were evaluated by X-ray micro-CT. Serum levels of klotho were measured by enzyme-linked immunosorbent assay.Aims
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The association of auraptene (AUR), a 7-geranyloxycoumarin, on osteoporosis and its potential pathway was predicted by network pharmacology and confirmed in experimental osteoporotic mice. The network of AUR was constructed and a potential pathway predicted by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) terms enrichment. Female ovariectomized (OVX) Institute of Cancer Research mice were intraperitoneally injected with 0.01, 0.1, and 1 mM AUR for four weeks. The bone mineral density (BMD) level was measured by dual-energy X-ray absorptiometry. The bone microstructure was determined by histomorphological changes in the femora. In addition, biochemical analysis of the serum and assessment of the messenger RNA (mRNA) levels of osteoclastic markers were performed.Aims
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The decrease in the number of satellite cells (SCs), contributing to myofibre formation and reconstitution, and their proliferative capacity, leads to muscle loss, a condition known as sarcopenia. Resistance training can prevent muscle loss; however, the underlying mechanisms of resistance training effects on SCs are not well understood. We therefore conducted a comprehensive transcriptome analysis of SCs in a mouse model. We compared the differentially expressed genes of SCs in young mice (eight weeks old), middle-aged (48-week-old) mice with resistance training intervention (MID+ T), and mice without exercise (MID) using next-generation sequencing and bioinformatics.Aims
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Despite the interest in the association of gut microbiota with bone health, limited population-based studies of gut microbiota and bone mineral density (BMD) have been made. Our aim is to explore the possible association between gut microbiota and BMD. A total of 3,321 independent loci of gut microbiota were used to calculate the individual polygenic risk score (PRS) for 114 gut microbiota-related traits. The individual genotype data were obtained from UK Biobank cohort. Linear regressions were then conducted to evaluate the possible association of gut microbiota with L1-L4 BMD (n = 4,070), total BMD (n = 4,056), and femur total BMD (n = 4,054), respectively. PLINK 2.0 was used to detect the single-nucleotide polymorphism (SNP) × gut microbiota interaction effect on the risks of L1-L4 BMD, total BMD, and femur total BMD, respectively.Aims
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Acquired heterotopic ossification (HO) is a debilitating disease characterized by abnormal extraskeletal bone formation within soft-tissues after injury. The exact pathogenesis of HO remains unknown. It was reported that Achilles tendon puncture (ATP) mouse model was performed on ten-week-old male C57BL/6J mice. One week after ATP procedure, the mice were given different treatments (e.g. JQ1, shMancr). Achilles tendon samples were collected five weeks after treatment for RNA-seq and real-time quantitative polymerase chain reaction (RT-qPCR) analysis; the legs were removed for micro-CT imaging and subsequent histology. Human bone marrow mesenchymal stem cells (hBMSCs) were isolated and purified bone marrow collected during surgeries by using density gradient centrifugation. After a series of interventions such as knockdown or overexpressing Aims
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Aims. Accumulated evidence indicates that local cell origins may ingrain differences in the phenotypic activity of human osteoblasts. We hypothesized that these differences may also exist in osteoblasts harvested from the same bone type at periarticular sites, including those adjacent to the fixation sites for total joint implant components. Methods. Human osteoblasts were obtained from the acetabulum and femoral neck of seven patients undergoing total hip arthroplasty (THA) and from the femoral and tibial cuts of six patients undergoing total knee arthroplasty (TKA). Osteoblasts were extracted from the usually discarded bone via enzyme digestion, characterized by flow cytometry, and cultured to passage three before measurement of metabolic activity, collagen production, alkaline phosphatase (ALP) expression, and mineralization. Results. Osteoblasts from the acetabulum showed lower proliferation (p = 0.034), cumulative collagen release (p < 0.001), and ALP expression (p = 0.009), and produced less mineral (p = 0.006) than those from the femoral neck. Osteoblasts from the tibia produced significantly less collagen (p = 0.021) and showed lower ALP expression than those from the distal femur. Conclusion. We have demonstrated for the first time an anatomical regional variation in the
To investigate whether idiopathic osteonecrosis of the femoral head (ONFH) is related to impaired osteoblast activities. We cultured osteoblasts isolated from trabecular bone explants taken from the femoral head and the intertrochanteric region of patients with idiopathic ONFH, or from the intertrochanteric region of patients with osteoarthritis (OA), and compared their viability, mineralization capacity, and secretion of paracrine factors.Aims
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Circular RNAs (circRNAs) are a novel type of non-coding RNA that plays major roles in the development of diverse diseases including osteonecrosis of the femoral head (ONFH). Here, we explored the impact of hsa_circ_0066523 derived from forkhead box P1 (FOXP1) (also called circFOXP1) on bone mesenchymal stem cells (BMSCs), which is important for ONFH development. RNA or protein expression in BMSCs was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot, respectively. Cell Counting Kit 8 (CCK8) and 5-ethynyl-2’-deoxyuridine (EdU) were used to analyze cell proliferation. Alkaline phosphatase (ALP) activity, ALP staining, and Alizarin Red S staining were employed to evaluate the osteoblastic differentiation. Chromatin immunoprecipitation (ChIP), luciferase reporter, RNA pull down, and RNA immunoprecipitation (RIP) assays were combined for exploring molecular associations.Aims
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LY3023414 is a novel oral phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) dual inhibitor designed for advanced cancers, for which a phase II clinical study was completed in March 2020; however, little is known about its effect on bone modelling/remodelling. In this study, we aimed to explore the function of LY3023414 in bone modelling/remodelling. The function of LY3023414 was explored in the context of osteogenesis (bone formation by osteoblasts) and osteoclastogenesis (osteoclast formation and bone resorption). Murine preosteoblast MC3T3-E1 cell line and murine bone marrow-derived macrophage cells (BMMs) were subjected to different treatments. An MTS cell proliferation assay was used to examine the cytotoxicity. Thereafter, different induction conditions were applied, such as MCSF and RANKL for osteoclastogenesis and osteogenic media for osteogenesis. Specific staining, a bone resorption assay, and quantitative real-time polymerase chain reaction (qRT-PCR) were subsequently used to evaluate the effect of LY3023414. Moreover, small interfering RNA (siRNA) was applied to knockdown Akt1 or Akt2 for further validation. Lastly, western blot was used to examine the exact mechanism of action.Aims
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