Promotes Osteoblast Differentiation Research Articles

Electrical Stimulation Generates Induced Tumor-Suppressing Cells, Offering a Potential Option for Combatting Breast Cancer and Bone Metastasis

Treating advanced metastatic cancer, particularly with bone metastasis, remains a significant challenge. In previous studies, induced tumor-suppressing (iTS) cells were successfully generated through genetic, chemical, and mechanical interventions. This study investigates the potential of electrical stimulation to generate iTS cells. Using a custom electrical stimulator with platinum electrodes, mesenchymal stem cells (MSCs) and Jurkat T cells were stimulated under optimized conditions (50 mV/cm, 10–100 Hz, 1 h). Conditioned medium (CM) from electrically stimulated cells demonstrated tumor-suppressing capabilities, inhibiting tumor cell migration, 3D spheroid growth, and cancer tissue fragment viability. Additionally, the CM reduced osteoclast maturation while promoting osteoblast differentiation. Proteomic analysis revealed enrichment of tumor-suppressing proteins, including histone H4, in the CM. Functional studies identified Piezo1 as a key mediator, as its knockdown significantly impaired the tumor-suppressive effects. Mechanistically, the process was distinct from other methods, such as mechanical vibration, with SUN1 inhibition showing no effect on iTS cell generation by electrical stimulation. These findings demonstrate the efficacy of electrical stimulation in enhancing the antitumor capabilities of MSCs and T cells, offering a novel approach to cancer therapy. Further exploration of this strategy could provide valuable insights into developing new treatments for metastatic cancer.

Epiregulin ameliorates ovariectomy-induced bone loss through orchestrating the differentiation of osteoblasts and osteoclasts.

Epiregulin plays a role in a range of biological activities including malignancies. This study aims to investigate the potential contribution of epiregulin to bone cell differentiation and bone homeostasis. The data showed that epiregulin expression was upregulated during osteogenesis but downregulated during adipogenesis. Functionally, epiregulin promoted osteoblast differentiation while inhibiting adipocyte differentiation from mesenchymal progenitor cells. Epidermal growth factor receptor (EGFR), one of the two known receptors for epiregulin, exerted opposing effects compared to epiregulin. Intriguingly, silencing EGFR almost completely abolished the dysregulation of osteoblast and adipocyte differentiation induced by epiregulin, suggesting that EGFR is indispensable for mediating epiregulin function. Further mechanistic exploration indicated that epiregulin/EGFR signaled via the inactivation of mechanistic target of rapamycin complex 1 (mTORC1) pathway. Moreover, epiregulin downregulated receptor activator of nuclear factor-κB ligand (RANKL) expression in BMSCs and inhibited the differentiation of bone marrow osteoclast precursor cells into osteoclasts. Treatment of ovariectomized female mice with recombinant epiregulin increased osteoblasts and bone formation, decreased osteoclasts and bone resorption, and ameliorated cancellous bone loss. Consistently, epiregulin treatment improved the potential of BMSCs to differentiate into osteoblasts. Collectively, this study has identified a critical role of epiregulin in regulating osteoblast differentiation through EGFR-mediated inactivation of mTORC1 pathway, as well as osteoclast differentiation via a mechanism associated with RANKL signaling. Additionally, it highlights the potential of epiregulin as a strategy for combating osteoporosis.

Shengxue Busui Decoction activates the PI3K/Akt and VEGF pathways, enhancing vascular function and inhibiting osteocyte apoptosis to combat steroid-induced femoral head necrosis

IntroductionSteroid-induced osteonecrosis of the femoral head (SONFH) is a debilitating condition with no specific treatment. Inhibiting osteocyte apoptosis may be a promising therapeutic approach. Shengxue Busui Decoction (SBD) has shown protective effects against SONFH, but its mechanisms are not fully understood. This study aims to investigate the effects of SBD on SONFH in rats, identifying its key active components and regulatory mechanisms using network pharmacology, bioinformatics, machine learning, and experimental validation.MethodsKey active components and disease targets of SBD were identified through network pharmacology and bioinformatics. GO/KEGG enrichment and ssGSEA analyses were performed to identify critical pathways. Cytoscape and machine learning (SVM) were used for target prediction and molecular docking validation. A dexamethasone (Dex)-induced SONFH rat model was established, and SBD was administered for 60 days. Histological changes were assessed via HE staining, osteoclast activity through TRAP staining, apoptosis levels with TUNEL assays, and vascular function through hematological tests. ELISA was used to measure ALP and OCN levels. In vitro, Dex-induced osteoblast apoptosis in MC3T3-E1 cells was examined to assess SBD’s effect on osteoblast proliferation, apoptosis, and signaling. Western blotting analyzed Caspase-9, Caspase-3, Bax, Bcl-2, and pathway-related proteins. ALP and Alizarin Red staining evaluated osteoblast differentiation and mineralization.ResultsNetwork pharmacology identified curcumin, berberine, and diosgenin as key active components of SBD, with the PI3K/Akt and VEGFR pathways as critical targets, and RAF1, FOXO3, and BRAF as hub genes. In vivo, SBD intervention significantly reduced bone structural damage and apoptosis, decreasing the rate of empty bone lacunae. SBD also increased osteogenic markers ALP and OCN in SONFH rats. In vitro, SBD inhibited osteoblast apoptosis, promoted PI3K/Akt and VEGF pathway expression, and enhanced osteoblast differentiation and mineralization.ConclusionThis study integrates network pharmacology with experimental validation, showing that SBD protects against SONFH by inhibiting osteoblast apoptosis via PI3K/Akt and VEGFR pathways. SBD promotes osteoblast differentiation and mineralization, improving bone structure and vascular function. Curcumin, berberine, and diosgenin are likely key contributors to these effects, highlighting SBD as a potential therapeutic strategy for SONFH.

Dental pulp stem cell-derived intracellular vesicles prevent orthodontic relapse by inhibiting PI3K/Akt/NF-κB-mediated osteoclast activity

BackgroundOrthodontic relapse, the undesired deviation of teeth from their corrected positions, remains a significant challenge in clinical orthodontics. Incomplete periodontal bone remodeling has been identified as a key factor in this process. Despite decades of research, currently there are no effective strategies to prevent relapse.MethodsWe isolated and identified dental pulp stem cell-derived intracellular vesicles (DPSC-IV) from human dental pulp tissue. To investigate its effect, DPSC-IV was added to osteoblast or osteoclast differentiation medium. During the orthodontic retention period, DPSC-IV was administrated to rats by subgingival injection. Relapse distance and relapse rate were calculated to evaluate DPSC-IV’s ability to prevent relapse. Additionally, Western blot analysis were used to examine DPSC-IV’s inhibitory effect on osteoclast differentiation.ResultsDPSC-IV significantly promoted osteoblast differentiation and inhibited osteoclast differentiation. Application of DPSC-IV during retention resulted in a significant reduction in both relapse distance and relapse rate, with improved periodontal structure and decreased osteoclast activity. This effect was mediated by the PI3K/Akt/NF-κB signaling pathway and could be reversed by the PI3K activator insulin-like growth factor-1 (IGF-1).ConclusionThis study highlights the potential of DPSC-IV as a novel preventive approach against orthodontic relapse, offering a novel strategy for maintaining long-term orthodontic stability.Graphical

Effect of photobiological regulation of green laser on orthodontic tooth retention in rats

Green lasers have a stronger effect on promoting osteoblast differentiation, which is critical for orthodontic tooth retention. This study investigated the impact of green laser photobiomodulation on orthodontic tooth retention in rats. A total of 100 male Sprague–Dawley rats were divided into two groups: Group A (control) and Group B (green laser irradiation). The left upper first molar was moved using a 0.20-mm nickel-titanium coil spring applying a force of 50 g for 3 weeks. The coil spring was then replaced with a 0.25-mm ligature wire to establish an orthodontic tooth retention model. Group B received green laser irradiation on the periodontium surrounding the molars. Retention devices were removed on days 1, 4, 10, 13, and 21. After 3 days of recurrence, the rats were sacrificed on days 4, 7, 13, 16, and 24. The left maxillary molar region was scanned using 3Shape to assess recurrence, and micro-computed tomography was used to evaluate alveolar bone density. Tissue staining was performed to observe periodontal remodeling and bone morphogenetic protein-2 (BMP-2) expression. Over time, the recurrence rate of the molar decreased significantly in both groups (P < 0.01), while alveolar bone density and BMP-2 expression increased (P < 0.01). Group B showed a lower recurrence rate and higher bone density, BMP-2 expression, and osteoblast counts than Group A. Green laser photobiomodulation promoted periodontal tissue remodeling, increased osteoblast numbers, stimulated new bone formation, and reduced the recurrence rate during orthodontic tooth retention in rats.

Loss of KAT6B causes premature ossification and promotes osteoblast differentiation during development.

The MYST family histone acetyltransferase gene, KAT6B (MYST4, MORF, QKF) is mutated in two distinct human congenital disorders characterised by intellectual disability, facial dysmorphogenesis and skeletal abnormalities; Say-Barber-Biesecker-Young-Simpson variant of Ohdo syndrome and Genitopatellar syndrome. Despite its requirement in normal skeletal development, the cellular and transcriptional effects of KAT6B in skeletogenesis have not been thoroughly studied. Here, we show that germline deletion of the Kat6b gene in mice causes premature ossification in vivo, resulting in shortened craniofacial elements and increased bone density, as well as shortened tibias with an expanded pre-hypertrophic layer, as compared to wild type controls. Mechanistically, we show that the loss of KAT6B in mesenchymal progenitor cells promotes transition towards an osteoblast-progenitor state with upregulation of gene targets of RUNX2, a master regulator of osteoblast development and concomitant downregulation of SOX9, a critical gene in chondrocyte development. Moreover, we find that compound heterozygosity at Kat6b and Runx2 loci partially rescues the reduction in ossification of Runx2 heterozygous, but not homozygous mice, suggesting that KAT6B may limit the action of RUNX2, possibly through a role in maintaining progenitors in an undifferentiated state. Moreover, our results show that KAT6B has essential roles in regulating the expression of a large number of genes involved in skeletogenesis and bone development.

The mechanism of action of indole-3-propionic acid on bone metabolism.

Indole-3-propionic acid (IPA), a metabolite produced by gut microbiota through tryptophan metabolism, has recently been identified as playing a pivotal role in bone metabolism. IPA promotes osteoblast differentiation by upregulating mitochondrial transcription factor A (Tfam), contributing to increased bone density and supporting bone repair. Simultaneously, it inhibits the formation and activity of osteoclasts, reducing bone resorption, possibly through modulation of the nuclear factor-κB (NF-κB) pathway and downregulation of osteoclast-associated factors, thereby maintaining bone structural integrity. Additionally, IPA provides indirect protection to bone health by regulating host immune responses and inflammation via activation of receptors such as the Aryl hydrocarbon Receptor (AhR) and the Pregnane X Receptor (PXR). This review summarizes the roles and signaling pathways of IPA in bone metabolism and its impact on various bone metabolic disorders. Furthermore, we discuss the therapeutic potential and limitations of IPA in treating bone metabolic diseases, aiming to offer novel strategies for clinical management.

DEC1 deficiency promotes osteoclastic activity by augmenting NFATc1 signaling via transactivation and the Ca2+/calcineurin pathway.

We have previously demonstrated that DEC1 promotes osteoblast differentiation. This study aims to evaluate the impact of DEC1 knockout on osteopenic activities, such as osteoclast differentiation and the expression of bone-degrading genes. To gain mechanistic insights, we employed both in vivo and in vitro experiments, utilizing cellular and molecular approaches, including osteoclast differentiation assays and RNA-seq in combination with ChIP-seq. Our results showed that NFATc1, a master regulator of osteoclast differentiation, and PPP3CB, a member of the calcineurin family, were significantly upregulated in DEC1-/- mice. In vitro experiments revealed that osteoclast differentiation significantly increased both the number and size of osteoclasts in DEC1-/- bone marrow macrophages (BMMs) compared to DEC1+/+ BMMs. Additionally, NFATc1 expression was notably higher in DEC1-/- BMMs than in DEC1+/+ BMMs. Overexpression of DEC1 reduced NFATc1 promoter activity, while knockout increased it. Furthermore, intracellular free Ca2+ levels and calcineurin activity were elevated (∼150 %) in DEC1-/- BMMs compared to DEC1+/+ BMMs. Importantly, the use of calcineurin inhibitors and calcium channel blockers effectively abolished the increased osteoclast differentiation observed in DEC1-/- BMMs. In summary, DEC1 deficiency promotes osteoclast differentiation by enhancing NFATc1 signaling through transcriptional regulation and the Ca2+/calcineurin pathway. Clinically, the mRNA levels of DEC1 were reduced by up to 75 % in patients with osteoporosis. The findings of this study establish that inducing DEC1 expression, alongside attenuators of the Ca2+/calcineurin pathway, offers a molecular basis for preventing and treating osteoporosis associated with DEC1 deficiency.

RUNX2 Phase Separation Mediates Long-Range Regulation Between Osteoporosis-Susceptibility Variant and XCR1 to Promote Osteoblast Differentiation.

GWASs have identified many loci associated with osteoporosis, but the underlying genetic regulatory mechanisms and the potential drug target need to be explored. Here, a new regulatory mechanism is found that a GWAS intergenic SNP (rs4683184) functions as an enhancer to influence the binding affinity of transcription factor RUNX2, whose phase separation can mediate the long-range chromatin interaction between enhancer and target gene XCR1 (a member of the GPCR family), leading to changes of XCR1 expression and osteoblast differentiation. Bone-targeting AAV of Xcr1 can improve bone formation in osteoporosis mice, suggesting that XCR1 can be a new susceptibility gene for osteoporosis. This study is the first to link non-coding SNP with phase separation, providing a new insight into long-range chromatin regulation mechanisms with susceptibility to complex diseases, and finding a potential target for the development of osteoporosis drugs and corresponding translational research.

The Role of Formononetin in Osteoblast Function and Mineralization Potential with Deproteinized Bovine Bone Material.

Dental bone formation involves various cellular and molecular mechanisms, and phytoestrogens such as formononetin (FORM) are promising because of their estrogenic, anti-inflammatory, and antioxidant effects. This study investigated the effect of FORM on osteoblast proliferation, differentiation, and mineralization in combination with spongiosa granulates (BO) in vitro. Human fetal osteoblast cells (hFOB1.19) were treated with increasing concentrations of FORM (1, 10, and 100 µg/mL), BO, or their combination. Cell proliferation was assessed using a MTT assay. Alkaline phosphatase (ALP) activity, intracellular Ca2+, and Pi levels were measured using ELISA. Vascular endothelial growth factor (VEGF) and osteocalcin expression levels were analyzed by western blotting. Cell proliferation increased with FORM, with or without BO, after 6 days (p < 0.001). FORM and BO had a synergistic effect on ALP activity (p < 0.001). Intracellular Ca2+ and Pi levels were highest in the BO-FORM group, suggesting superior mineralization (p < 0.05). VEGF and osteocalcin expression was significantly upregulated with FORM, alone and with BO (p < 0.05), indicating improved angiogenesis and bone maturation over 9 days. FORM enhances osteoblast proliferation, differentiation, and mineralization potential, particularly in BO spongiosa granulates. These data support the in vitro potential of formononetin-phytoestrogen in promoting osteoblast differentiation and mineralization potential with BO. These findings suggest that FORM, combined with BO, could improve bone augmentation in clinical applications such as maxillofacial surgery. FORM shows valuable potential for clinical applications, such as maxillofacial surgery, by promoting faster and more effective healing.

The natural flavone quercetin: An extensive analysis of its pharmacological mechanisms and medicinal prospects

Natural flavonoid quercetin is widely distributed in fruits, vegetables, and medicinal plants. It has attracted much interest due to its wide range of biological advantages and potential as a treatment for several illnesses. To better understand the mechanisms behind the therapeutic of effects quercetin in cancer, obesity, diabetes, and osteoporosis, this review attempts to present a thorough overview of these mechanisms. By promoting osteoblast differentiation, reducing osteoclast activity, and increasing bone mineral density, quercetin shows promise as a protective agent against bone loss in osteoporosis. Its anti-inflammatory properties also help to reduce the risk of fracture and bone resorption linked to osteoporosis. Through its targeting of various signalling pathways involved in cell proliferation, apoptosis, angiogenesis, and metastasis, quercetin shows promising anti-cancer properties in cancer. Moreover, its effectiveness in reducing the progression of cancer is attributed to its capacity to regulate inflammation and oxidative stress. Through its effects on adipocyte differentiation, glucose uptake, insulin signalling, and lipid metabolism, quercetin shows anti-obesity and anti-diabetic effects in obesity and diabetes. Furthermore, the antioxidant and anti-inflammatory qualities of quercetin are essential in reducing the negative effects of obesity on insulin resistance and the complications associated with diabetes. In summary, this review offers valuable implications for the development of quercetin-based therapeutics and nutraceuticals for disease management by shedding light on the complex mechanisms through which quercetin exerts its biological benefits in different disease contexts.

Dehydroepiandrosterone and Bone Health: Mechanisms and Insights.

Dehydroepiandrosterone (DHEA), a steroid hormone produced by the adrenal glands, plays a key role in various physiological processes, including bone health. Its age-related decline is linked to reduced bone density, though the mechanisms by which DHEA affects bone metabolism remain complex. This review summarises the diverse effects of DHEA on bone metabolism and density, highlighting its therapeutic potential; Methods: A literature search on the effects of DHEA on bone-related parameters was conducted from PubMed and Scopus using a specific search string, and after removing duplicates and irrelevant articles, 36 relevant full-text studies were included; Results: DHEA promotes osteoblast differentiation and proliferation, regulates the RANKL/OPG ratio, and inhibits osteoclastogenesis and bone resorption. Its osteogenic effects are mediated through multiple signalling pathways. In ovariectomised rat models, DHEA enhances trabecular bone volume, stimulates osteoblast proliferation, and increases oestradiol production and aromatase activity. In elderly individuals with low androgen levels, DHEA supplementation increases sulphated DHEA and oestradiol levels and improves bone mineral density, particularly in the ultra-distal radius of women and the femoral neck of men. However, the clinical use of DHEA remains debated due to inconsistent study results. Its effects on bone health may vary based on factors such as age, gender, and health conditions, emphasising the need for further research to clarify its mechanisms and optimise its use; Conclusions: In conclusion, while DHEA shows potential as a modulator of bone health, comprehensive clinical trials are required to assess its efficacy and safety, particularly in at-risk populations.

Preferred Orientation of Hydroxyapatite Ceramics Along the c-Axis Promotes Osteoblast Differentiation.

Hydroxyapatite (HAp) is similar to the main inorganic components of bone and tooth enamel. Furthermore, it possesses biocompatibility, making it suitable for clinical use in artificial bones. This study aimed to verify whether the preferred orientation of HAp influences osteogenesis. Using the templated grain growth method, we successfully fabricated HAp ceramics with a preferred orientation to m (a)-planes (aHAp) and examined the effects of this orientation on bone differentiation. Osteosarcoma-derived osteoblasts (MG-63) were cultured on aHAp and HAp ceramics made from commercially available powder (iHAp). Electron backscatter diffraction analysis revealed the crystal orientation distribution of HAp ceramics and the numerous exposed a-planes of aHAp. The MG-63 cultured on aHAp exhibited significantly higher alkaline phosphatase activity, a marker of early bone differentiation, compared to iHAp. Furthermore, the two-dimensional electrophoresis results indicated that the expressed proteins differed between aHAp and iHAp. These results indicate that controlling HAp's crystal structure may promote the osteogenic potential of osteoblasts. In this study, we propose that the a-plane of HAp promotes bone differentiation during the early stages, presenting a promising approach for novel biomaterials, such as high-performance artificial bones.

Diffusion model assisted designing self-assembling collagen mimetic peptides as biocompatible materials.

Collagen self-assembly supports its mechanical function, but controlling collagen mimetic peptides (CMPs) to self-assemble into higher-order oligomers with numerous functions remains challenging due to the vast potential amino acid sequence space. Herein, we developed a diffusion model to learn features from different types of human collagens and generate CMPs; obtaining 66% of synthetic CMPs could self-assemble into triple helices. Triple-helical and untwisting states were probed by melting temperature (Tm); hence, we developed a model to predict collagen Tm, achieving a state-of-art Pearson's correlation (PC) of 0.95 by cross-validation and a PC of 0.8 for predicting Tm values of synthetic CMPs. Our chemically synthesized short CMPs and recombinantly expressed long CMPs could self-assemble, with the lowest requirement for hydrogel formation at a concentration of 0.08% (w/v). Five CMPs could promote osteoblast differentiation. Our results demonstrated the potential for using computer-aided methods to design functional self-assembling CMPs.

Exosomal miR-590-3p derived from bone marrow mesenchymal stem cells promotes osteoblast differentiation and osteogenesis by targeting TGFBR1.

Bone marrow mesenchymal stem cells (BMSCs) have been verified to be essential factors regulating osteogenic functions, which is mainly attributed to their secretion of extracellular vesicles. Exosomes derived from BMSCs (BMSCs-Exo) contribute to osteoblast functions that are critical for improving bone defect. Our current study aims to investigate the molecular mechanism dominated by BMSCs-Exo that affects osteoblast differentiation and osteogenesis. The first step this study validated that BMSCs co-culture enhanced the differentiation ability of osteoblast and promoted bone mineralization, while these tendencies were abolished after GW4869 treatment. Next, the BMSCs-Exo was isolated and identified by TEM observation, insight detection, and western blot analysis. Furthermore, BMSCs-Exo treatment could efficiently promote the differentiation ability and the bone mineralization of osteoblasts, decrease the mRNA levels of Collagen I and Collagen III, and increase the levels of osteogenic proteins, including alkaline phosphatase (ALP), Turning Bone Morphogenetic Protein 2 (BMP2), Bone sialoprofein (BSP), osteocalcin (OCN), and osterix (OSX). However, the abovementioned effects of BMSCs-Exo could be abolished by miR-590-3p silencing. Mechanistic analysis unmasked the negative regulation of miR-590-3p on its downstream target TGFBR1. Finally, the effects of miR-590-3p/TGFBR1 axis on the differentiation and osteogenesis of osteoblasts were validated by rescue assays. In conclusion, the present study demonstrates that exosomal miR-590-3p secreted by BMSCs can induce osteoblast differentiation and osteogenesis.

D-limonene suppresses RANKL-induced osteoclast differentiation and promotes osteoblast activity in vitro.

Treatments for osteoporosis are typically given postfracture. Therefore, identifying safe prophylactic interventions to reduce fracture risk would be beneficial. One approach is to utilize the bioactive properties of natural compounds to modify osteoclast and osteoblast activity. d-limonene a well-tolerated, anti-inflammatory monoterpene found in citrus fruits holds promise due to its suppressive effect on NFκB, a key regulator of bone cell activity. We found that limonene promoted osteoblast differentiation and bone nodule formation and inhibited RANKL-induced osteoclast formation and bone resorption in vitro. Limonene also reduced the proresorptive signal provided by osteoblast, augmenting markers of osteoblast differentiation (alkaline phosphatase, osterix, and osteocalcin) and significantly decreasing osteoclastogenic cytokine production (PTHrP, IL-1β, and TNF-α). Therefore, limonene supplementation represents a potential route in combination with current interventions to optimize bone cell activity to maintain or enhance bone mass.

Sulfated galactofucan from Sargassum fusiforme protects against postmenopausal osteoporosis by regulating bone remodeling

Osteoporosis is a degenerative bone disease highly prevalent in older women, causing high morbidity and mortality rates. Fourteen kinds of fucoidan were isolated from Sargassum fusiforme through acid (named as SFS), alkaline (SFJ) and water (SFW). SFW was passed through an anion exchange column to obtain SFW-0, SFW-0.5 and SFW-2. SFW-0.5 and SFW-2 were degraded to obtain different sulfate group contents SFW-x-M/S/O (x for 0.5 or 2). We further confirmed SFW-0.5-O was the most effective fraction of SFW. SFW-0.5-O may have alternating backbones of (Gal)n and (Fuc)n, and the main sulfation may be at C2/C3 of the Fuc/Gal residues. SFW-0.5-O inhibition of OC differentiation was associated with IRF-8 signaling; meanwhile, SFW-0.5-O promoted osteoblast differentiation and bone mineral nodule formation. SFW-0.5-O also effectively ameliorated osteoporosis symptom caused by estrogen deprivation in vivo. We uncovered that the fucoidan active fraction SFW-0.5-O demonstrated effective bone protection, may be exploited for osteoporosis therapy.

CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells.

We investigated the regulation of histone deacetylases (HDACs) by miR-2861 in the osteoblastic differentiation of human mesenchymal stem cells (MSCs) and miR-2861 binding site by CRISPR activation (CRISPRa). Transfection of miR-2861 into human MSCs was performed and the effect on osteoblast differentiation was analyzed. Using catalytically inactive Cas12a, the CRISPRa system induced targeted overexpression of endogenous miRNA and repressed the luciferase activities of reporters that contained functional miRNA target sites. The delivery of miR-2861 into MSCs enhanced osteoblast differentiation by decreased expressions of the HDAC1, 4 and 5 genes. The mechanism of HDAC5 repression by miR-2861 in humans has not been fully elucidated. To this end, the HDAC5 mRNA sequence was analyzed and a putative primate-specific miR-2861 binding site was identified in the 3' untranslated region (3'-UTR). CRISPRa was applied to validate the putative binding site and an increase in endogenous miR-2861 was found to repress the expression of a reporter that contained the novel miR-2861 binding site. The delivery of miR-2861 to human MSCs enhanced osteoblast differentiation. In the 3'-UTR, the HDAC5 repression was mediated by the miR-2861 binding site, and miR-2861 promoted osteoblast differentiation via the inhibition of HDAC5 through a primate-specific miRNA binding site. Therefore, miRNAmiR-2861 with the CRISPRa methods might be a good biomaterial for osteogenesis augmentation.

Composite Scaffold Materials of Nanocerium Oxide Doped with Allograft Bone: Dual Optimization Based on Anti-Inflammatory Properties and Promotion of Osteogenic Mineralization.

Spinal fusion technique is widely used in the treatment of lumbar degeneration, cervical instability, disc injury, and spinal deformity. However, it is usually accompanied by a high incidence of fusion failure and pseudoarthrosis, placing higher demands on bone implants. Therefore, materials with good biocompatibility, osteoconductivity, and even induce bone ingrowth, which can be used to improve spinal fusion rate and bone regeneration, have become a hot research topic. Here, ultra-small cerium oxide nanoparticles (CeO2 NPs) are prepared and loaded onto the surface of the homograft bone surface to prepare a composite scaffold AB@PLGA/CeO2. The composite scaffold shows the competitive ability to promote osteoblast differentiation in vitro. In vivo experiments show that AB@PLGA/CeO2 has a good bone enhancement effect. In particular, good biological effects of collagen fiber formation, osteogenic mineralization, and tissue repair are shown in intervertebral implant fusion. Further, transcriptome sequencing confirms that CeO2 NPs promote osteogenic differentiation and mineralization by regulating extracellular matrix (ECM) and collagen formation. Meanwhile, CeO2 NPs can regulate the function of the PI3K-Akt signaling pathway to exert its ability to promote osteogenic differentiation and mineralization and affect p53 and cell cycle signaling pathway to regulate osteogenic differentiation and mineralization. Hence, the proposed scaffold is a promising strategy for intervertebral fusion in the clinic.

The MCP-3/Ccr3 axis contributes to increased bone mass by affecting osteoblast and osteoclast differentiation

Several CC subfamily chemokines have been reported to regulate bone metabolism by affecting osteoblast or osteoclast differentiation. However, the role of monocyte chemotactic protein 3 (MCP-3), a CC chemokine, in bone remodeling is not well understood. Here, we show that MCP-3 regulates bone remodeling by promoting osteoblast differentiation and inhibiting osteoclast differentiation. In a Ccr3-dependent manner, MCP-3 promoted osteoblast differentiation by stimulating p38 phosphorylation and suppressed osteoclast differentiation by upregulating interferon beta. MCP-3 increased bone morphogenetic protein 2-induced ectopic bone formation, and mice with MCP-3-overexpressing osteoblast precursor cells presented increased bone mass. Moreover, MCP-3 exhibited therapeutic effects by abrogating receptor activator of nuclear factor kappa-B ligand-induced bone loss. Therefore, MCP-3 has therapeutic potential for diseases involving bone loss due to its positive role in osteoblast differentiation and negative role in osteoclast differentiation.