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Coordinating the immune response and bioenergy metabolism in bone defect environments is essential for promoting bone regeneration. Mitochondria are important organelles that control internal balance and metabolism. Repairing dysfunctional mitochondria has been proposed as a therapeutic approach for disease intervention. Here, an engineered hierarchical hydrogel with immune responsiveness can adapt to the bone regeneration environment and mediate the targeted mitochondria transfer between cells. The continuous supply of mitochondria by macrophages can restore the mitochondrial bioenergy of bone marrow mesenchymal stem cells (BMSC). Fundamentally solving the problem of insufficient energy support of BMSCs caused by local inflammation during bone repair and regeneration. This discovery provides a new therapeutic strategy for promoting bone regeneration and repair, which has research value and practical application prospects in the treatment of various diseases caused by mitochondrial dysfunction.
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Background and aim: A surplus of glucocorticoids (GC) is a main cause of non-traumatic osteonecrosis of the femoral head (ONFH), and Jintiange (JTG), as one of the traditional Chinese medicines (TCM), also plays an instrumental role in the alleviation of bone loss simultaneously. Therefore, JTG was thought to be able to reverse GC-induced ONFH (GC-ONFH) to a certain extent. Experimental procedure: In vivo, the effect of JTG on trabeculae in the subchondral bone of the femoral head was investigated using micro-computed tomography (micro-CT), TdT-mediated dUTP nick end labeling (TUNEL) and histological staining; in vitro, proliferation, viability, apoptosis, and senescence of purified bone mesenchymal stem cells (BMSCs) were examined to demonstrate the direct impact of JTG on these cells. Meanwhile after using a series of interventions, the function of JTG on BMSC differentiation could be assessed by measuring of osteogenic and adipogenic markers at levels of protein and mRNA. Results: Our final results demonstrated that with the involvement of Wnt/ß-catenin pathway, JTG was able to significantly promote osteogenesis, restrain adipogenesis, delay senescence in BMSCs, reduce osteoclast number, weaken apoptosis, and enhance proliferation of osteocytes, all of which could mitigate the progression of subchondral osteonecrosis. Conclusion: According to the results of experiments in vitro and vivo, JTG was deemed to relieve the early GC-ONFH using the prevention of destruction of subchondral bone, which was contributed to regulating the differentiation of BMSCs and the number of osteoclasts.
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Osteogenesis is tightly coupled with angiogenesis spatiotemporally. Previous studies have demonstrated that type H blood vessel formed by endothelial cells with high expression of CD31 and Emcn (CD31hi Emcnhi ECs) play a crucial role in bone regeneration. The mechanism of the molecular communication around CD31hi Emcnhi ECs and bone mesenchymal stem cells (BMSCs) in the osteogenic microenvironment is unclear. This study indicates that exosomes from bone mesenchymal stem cells with 7 days osteogenic differentiation (7D-BMSCs-exo) may promote CD31hi Emcnhi ECs angiogenesis, which was verified by tube formation assay, qRT-PCR, Western blot, immunofluorescence staining and µCT assays etc. in vitro and in vivo. Furthermore, by exosomal miRNA microarray and WGCNA assays, we identified downregulated miR-150-5p as the most relative hub gene coupling osteogenic differentiation and type H blood vessel angiogenesis. With bioinformatics assays, dual luciferase reporter experiments, qRT-PCR and Western blot assays, SOX2(SRY-Box Transcription Factor 2) was confirmed as a novel downstream target gene of miR-150-5p in exosomes, which might be a pivotal mechanism regulating CD31hi Emcnhi ECs formation. Additionally, JC-1 immunofluorescence staining, Western blot and seahorse assay results showed that the overexpression of SOX2 could shift metabolic reprogramming from oxidative phosphorylation (OXPHOS) to glycolysis to enhance the CD31hi Emcnhi ECs formation. The PI3k/Akt signaling pathway might play a key role in this process. In summary, BMSCs in osteogenic differentiation might secrete exosomes with low miR-150-5p expression to induce type H blood vessel formation by mediating SOX2 overexpression in ECs. These findings might reveal a molecular mechanism of osteogenesis coupled with type H blood vessel angiogenesis in the osteogenic microenvironment and provide a new therapeutic target or cell-free remedy for osteogenesis impaired diseases.
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Diferenciación Celular , Células Endoteliales , Exosomas , Células Madre Mesenquimatosas , MicroARNs , Neovascularización Fisiológica , Osteogénesis , MicroARNs/genética , MicroARNs/metabolismo , Exosomas/metabolismo , Osteogénesis/genética , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Diferenciación Celular/genética , Neovascularización Fisiológica/genética , Animales , Células Endoteliales/metabolismo , Células Endoteliales/citología , Ratones , Humanos , Células Cultivadas , Transducción de Señal , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/genética , Factores de Transcripción SOXB1/metabolismo , Factores de Transcripción SOXB1/genética , Reprogramación Metabólica , AngiogénesisRESUMEN
Articular cartilage defect therapy is still dissatisfactory in clinic. Direct cell implantation faces challenges, such as tumorigenicity, immunogenicity, and uncontrollability. Extracellular vesicles (EVs) based cell-free therapy becomes a promising alternative approach for cartilage regeneration. Even though, EVs from different cells exhibit heterogeneous characteristics and effects. The aim of the study was to discover the functions of EVs from the cells during chondrogenesis timeline on cartilage regeneration. Here, bone marrow mesenchymal stem cells (BMSCs)-EVs, juvenile chondrocytes-EVs, and adult chondrocytes-EVs were used to represent the EVs at different differentiation stages, and fibroblast-EVs as surrounding signals were also joined to compare. Fibroblasts-EVs showed the worst effect on chondrogenesis. While juvenile chondrocyte-EVs and adult chondrocyte-EVs showed comparable effect on chondrogenic differentiation as BMSCs-EVs, BMSCs-EVs showed the best effect on cell proliferation and migration. Moreover, the amount of EVs secreted from BMSCs were much more than that from chondrocytes. An injectable decellularized extracellular matrix (dECM) hydrogel from small intestinal submucosa (SIS) was fabricated as the EVs delivery platform with natural matrix microenvironment. In a rat model, BMSCs-EVs loaded SIS hydrogel was injected into the articular cartilage defects and significantly enhanced cartilage regeneration in vivo. Furthermore, protein proteomics revealed BMSCs-EVs specifically upregulated multiple metabolic and biosynthetic processes, which might be the potential mechanism. Thus, injectable SIS hydrogel loaded with BMSCs-EVs might be a promising therapeutic way for articular cartilage defect.
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The therapy of large defects in peripheral nerve injury (PNI) suffers from several drawbacks, especially the lack of autologous nerve donors. Nerve conduits are considered as a solution for nerve injury treatment, but biocompatibility improvements is still required for conduits prepared with synthetic materials. Cell-derived extracellular matrix (ECM) has drawn attention due to its lower risk of immunogenic response and independence from donor availability. The goal of this study is to coat bone mesenchymal stem cell-derived ECMs on poly(lactic-co-glycolic) acid (PLGA) conduits to enhance their ability to support neural growth and neurite extensions. The ECM-coated conduits have better hydrophilic properties than the pure PLGA conduits. A marked increase on PC12 and RSC96 cells' viability, proliferation and dorsal root ganglion neurite extension was observed. Quantitative PCR analysis exhibited a significant increase in markers for cell proliferation (GAP43), neurite extension (NF-H, MAP2, andßIII-tubulin) and neural function (TREK-1). These results show the potential of ECM-coated PLGA conduits in PNI therapy.
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Proliferación Celular , Supervivencia Celular , Matriz Extracelular , Células Madre Mesenquimatosas , Regeneración Nerviosa , Neuritas , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Animales , Ratas , Neuritas/metabolismo , Células PC12 , Matriz Extracelular/metabolismo , Células Madre Mesenquimatosas/citología , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/química , Regeneración Nerviosa/efectos de los fármacos , Andamios del Tejido/química , Materiales Biocompatibles Revestidos/química , Materiales Biocompatibles Revestidos/farmacología , Ganglios Espinales , Traumatismos de los Nervios Periféricos/terapia , Ingeniería de Tejidos/métodos , Polímeros/química , Ensayo de MaterialesRESUMEN
Postmenopausal osteoporosis (PMO) is characterized by bone loss and microstructural damage, and it is most common in older adult women. Currently, there is no cure for PMO. The flavonoid chemical 7,8-dihydroxyflavone (7,8-DHF) specifically activates tropomyosin receptor kinase B (TRKB). Furthermore, 7,8-DHF has various biological characteristics, including anti-inflammatory and antioxidant effects. However, the specific implications and fundamental mechanisms of 7,8-DHF in PMO remain unclear. We used protein imprinting, flow cytometry, tissue staining, and other methods to estimate the preventive mechanisms of 7,8-DHF against hydrogen peroxide (H2O2)-induced apoptosis in primary mouse bone marrow mesenchymal stem cells (BMSCs), osteogenic differentiation ability, and bone mass in ovariectomized (OVX) mice. We found that 7,8-DHF effectively prevented H2O2-induced reductions in the viability and osteogenic differentiation capacity of primary BMSCs. Mechanistically, 7,8-DHF induced the TRKB to activate the PI3K/AKT/NRF2 pathway. In vivo experiments with the OVX mouse model confirmed that 7,8-DHF can inhibit oxidative stress and promote bone formation, indicating that 7,8-DHF improves the viability and osteogenic differentiation ability of BMSCs stimulated via H2O2 by activating the TRKB/PI3K/AKT and NRF2 pathways, thereby improving PMO.
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Flavonas , Peróxido de Hidrógeno , Células Madre Mesenquimatosas , Factor 2 Relacionado con NF-E2 , Osteogénesis , Osteoporosis Posmenopáusica , Estrés Oxidativo , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , Receptor trkB , Transducción de Señal , Animales , Estrés Oxidativo/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Ratones , Femenino , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Flavonas/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Osteoporosis Posmenopáusica/metabolismo , Osteoporosis Posmenopáusica/patología , Osteoporosis Posmenopáusica/tratamiento farmacológico , Osteogénesis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Peróxido de Hidrógeno/metabolismo , Humanos , Receptor trkB/metabolismo , Receptor trkB/genética , Diferenciación Celular/efectos de los fármacos , Apoptosis/efectos de los fármacos , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/genética , Ovariectomía , Ratones Endogámicos C57BLRESUMEN
The skeleton is a metabolically active organ undergoing continuous remodeling initiated by bone marrow stem cells (BMSCs). Recent research has demonstrated that BMSCs adapt the metabolic pathways to drive the osteogenic differentiation and bone formation, but the mechanism involved remains largely elusive. Here, using a comprehensive targeted metabolome and transcriptome profiling, we revealed that one-carbon metabolism was promoted following osteogenic induction of BMSCs. Methotrexate (MTX), an inhibitor of one-carbon metabolism that blocks S-adenosylmethionine (SAM) generation, led to decreased N6-methyladenosine (m6A) methylation level and inhibited osteogenic capacity. Increasing intracellular SAM generation through betaine addition rescued the suppressed m6A content and osteogenesis in MTX-treated cells. Using S-adenosylhomocysteine (SAH) to inhibit the m6A level, the osteogenic activity of BMSCs was consequently impeded. We also demonstrated that the pro-osteogenic effect of m6A methylation mediated by one-carbon metabolism could be attributed to HIF-1α and glycolysis pathway. This was supported by the findings that dimethyloxalyl glycine rescued the osteogenic potential in MTX-treated and SAH-treated cells by upregulating HIF-1α and key glycolytic enzymes expression. Importantly, betaine supplementation attenuated MTX-induced m6A methylation decrease and bone loss via promoting the abundance of SAM in rat. Collectively, these results revealed that one-carbon metabolite SAM was a potential promoter in BMSC osteogenesis via the augmentation of m6A methylation, and the cross talk between metabolic reprogramming, epigenetic modification, and transcriptional regulation of BMSCs might provide strategies for bone regeneration.
The bone is a self-renewing tissue that continues to reshape throughout life. Bone marrow mesenchymal stem cells (BMSCs) are essential for bone homeostasis as they are capable of osteogenic differentiation. Recent evidence suggests that BMSCs drive the osteogenic differentiation through metabolic reprogramming, but the mechanism remains unclear. In this paper, we explored the metabolic alteration following osteogenic induction of BMSCs and found that one-carbon metabolism was obviously promoted in this process. The underlining mechanisms of the osteogenic potential driven by one-carbon metabolism seem to be its contribution on N6-methyladenosine (m6A) methylation and consequent glycolysis level by providing methyl donor. We demonstrated that one-carbon metabolism-mediated m6A methylation was a potential promoter in BMSC osteogenesis, and metabolic-epigenetic coupling might provide novel therapeutic targets for bone regeneration.
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Adenosina , Carbono , Osteogénesis , Ratas Sprague-Dawley , S-Adenosilmetionina , Animales , S-Adenosilmetionina/metabolismo , S-Adenosilmetionina/farmacología , Osteogénesis/efectos de los fármacos , Adenosina/análogos & derivados , Adenosina/farmacología , Adenosina/metabolismo , Metilación/efectos de los fármacos , Carbono/metabolismo , Carbono/farmacología , Ratas , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/citología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Masculino , Metotrexato/farmacología , Glucólisis/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/efectos de los fármacosRESUMEN
Following the publication of the above paper, it has been drawn to the Editor's attention by a concerned reader that the immunohistochemical assay data shown in Fig. 4B on p. 245 were strikingly similar to data appearing in different form in another article written by different authors at different research institutes that had already been published in the journal International Journal of Biological Sciences prior to the submission of this paper to International Journal of Molecular Medicine. In view of the fact that the contentious data had already apparently been published previously, the Editor of International Journal of Molecular Medicine has decided that this paper should be retracted from the Journal. After having been in contact with the authors, they agreed with the decision to retract the paper. The Editor apologizes to the readership for any inconvenience caused. [International Journal of Molecular Medicine 46: 239-251, 2020; DOI: 10.3892/ijmm.2020.4595].
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OBJECTIVE: Bone mesenchymal stem cells (BMSCs) have been tentatively applied in the treatment of glucocorticoid-induced osteoporosis (GIOP) and systemic lupus erythematosus (SLE). However, the effects of BMSCs on osteoporosis within the context of glucocorticoid (GC) application in SLE remain unclear. Our aim was to explore the roles of BMSCs and different doses of GC interventions on osteoporosis in SLE murine models. METHODS: MRL/MpJ-Faslpr mice were divided into eight groups with BMSC treatment and different dose of GC intervention. Three-dimensional imaging analysis and hematoxylin and eosin (H&E) staining were performed to observe morphological changes. The concentrations of osteoprotegerin (OPG) and receptor activator of nuclear factor κB ligand (RANKL) in serum were measured by enzyme-linked immunosorbent assay (ELISA). The subpopulation of B cells and T cells in bone marrows and spleens were analyzed by flow cytometry. Serum cytokines and chemokines were assessed using Luminex magnetic bead technology. RESULTS: BMSCs ameliorated osteoporosis in murine SLE models by enhancing bone mass, improving bone structure, and promoting bone formation through increased bone mineral content and optimization of trabecular morphology. BMSC and GC treatments reduced the number of B cells in bone marrows, but the effect was not significant in spleens. BMSCs significantly promoted the expression of IL-10 while reducing IL-18. Moreover, BMSCs exert immunomodulatory effects by reducing Th17 expression and rectifying the Th17/Treg imbalance. CONCLUSION: BMSCs effectively alleviate osteoporosis induced by SLE itself, as well as osteoporosis resulting from SLE combined with various doses of GC therapy. The therapeutic effects of BMSCs appear to be mediated by their influence on bone marrow B cells, T cell subsets, and associated cytokines. High-dose GC treatment exerts a potent anti-inflammatory effect but may hinder the immunotherapeutic potential of BMSCs. Our research may offer valuable guidance to clinicians regarding the use of BMSC treatment in SLE and provide insights into the judicious use of GCs in clinical practice.
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Modelos Animales de Enfermedad , Glucocorticoides , Lupus Eritematoso Sistémico , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Osteoporosis , Animales , Lupus Eritematoso Sistémico/complicaciones , Lupus Eritematoso Sistémico/terapia , Lupus Eritematoso Sistémico/tratamiento farmacológico , Lupus Eritematoso Sistémico/inmunología , Ratones , Osteoporosis/etiología , Osteoporosis/tratamiento farmacológico , Osteoporosis/terapia , Glucocorticoides/administración & dosificación , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/metabolismo , Femenino , Ratones Endogámicos MRL lpr , Citocinas/metabolismoRESUMEN
Periodontal disease ranks third among noncommunicable illnesses, behind cancer and cardiovascular disease, and is closely related to the occurrence and progression of various systemic diseases. However, elucidating the processes of periodontal disease and promoting periodontal bone regeneration remains a challenge. Here, quercetin is demonstrated to reduce the oxidative stress state of orofacial mesenchymal stem cells (OMSCs) in vitro and to affect the osteogenic growth of OMSCs through molecular mechanisms that mediate the m6A change in Per1. Nevertheless, the limited therapeutic efficacy of systemic medication and the limitations of local medication resulting from the small, moist, and highly dynamic periodontal environment make it challenging to treat periodontal tissues with medication. Herein, a biosafe injectable hydrogel drug-controlled delivery system is constructed as a bone-enhancing factory and loaded with quercetin to treat oxidative stress injury in periodontal tissues. This drug-carrying system made up of nanoscale bioglass microspheres and a light-cured injectable hydrogel, allows effective drug particle loading and cementation in the dynamic and moist periodontal environment. Furthermore, the system demonstrates the ability to stimulate OMSCs osteogenic differentiation in a Per1-dependent manner, which ultimately promotes periodontal bone repair, suggesting that this system has potential for clinical periodontal therapy.
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Cerámica , Hidrogeles , Células Madre Mesenquimatosas , Estrés Oxidativo , Quercetina , Estrés Oxidativo/efectos de los fármacos , Cerámica/farmacología , Animales , Quercetina/farmacología , Quercetina/administración & dosificación , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Regeneración Ósea/efectos de los fármacos , Humanos , Osteogénesis/efectos de los fármacos , Modelos Animales de Enfermedad , Ratones , Pérdida de Hueso Alveolar/tratamiento farmacológicoRESUMEN
BACKGROUND: Osteoporosis (OP) has become a major public health problem worldwide. Most OP treatments are based on the inhibition of bone resorption, and it is necessary to identify additional treatments aimed at enhancing osteogenesis. In the bone marrow (BM) niche, bone mesenchymal stem cells (BMSCs) are exposed to a hypoxic environment. Recently, a few studies have demonstrated that hypoxia-inducible factor 2alpha (HIF-2α) is involved in BMSC osteogenic differentiation, but the molecular mechanism involved has not been determined. AIM: To investigate the effect of HIF-2α on the osteogenic and adipogenic differentiation of BMSCs and the hematopoietic function of hematopoietic stem cells (HSCs) in the BM niche on the progression of OP. METHODS: Mice with BMSC-specific HIF-2α knockout (Prx1-Cre;Hif-2αfl/fl mice) were used for in vivo experiments. Bone quantification was performed on mice of two genotypes with three interventions: Bilateral ovariectomy, semilethal irradiation, and dexamethasone treatment. Moreover, the hematopoietic function of HSCs in the BM niche was compared between the two mouse genotypes. In vitro, the HIF-2α agonist roxadustat and the HIF-2α inhibitor PT2399 were used to investigate the function of HIF-2α in BMSC osteogenic and adipogenic differentiation. Finally, we investigated the effect of HIF-2α on BMSCs via treatment with the mechanistic target of rapamycin (mTOR) agonist MHY1485 and the mTOR inhibitor rapamycin. RESULTS: The quantitative index determined by microcomputed tomography indicated that the femoral bone density of Prx1-Cre;Hif-2αfl/fl mice was lower than that of Hif-2αfl/fl mice under the three intervention conditions. In vitro, Hif-2αfl/fl mouse BMSCs were cultured and treated with the HIF-2α agonist roxadustat, and after 7 d of BMSC adipogenic differentiation, the oil red O staining intensity and mRNA expression levels of adipogenesis-related genes in BMSCs treated with roxadustat were decreased; in addition, after 14 d of osteogenic differentiation, BMSCs treated with roxadustat exhibited increased expression of osteogenesis-related genes. The opposite effects were shown for mouse BMSCs treated with the HIF-2α inhibitor PT2399. The mTOR inhibitor rapamycin was used to confirm that HIF-2α regulated BMSC osteogenic and adipogenic differentiation by inhibiting the mTOR pathway. Consequently, there was no significant difference in the hematopoietic function of HSCs between Prx1-Cre;Hif-2αfl/fl and Hif-2αfl/fl mice. CONCLUSION: Our study showed that inhibition of HIF-2α decreases bone mass by inhibiting the osteogenic differentiation and increasing the adipogenic differentiation of BMSCs through inhibition of mTOR signaling in the BM niche.
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Age-related osteoporosis is characterized by an imbalance between osteogenic and adipogenic differentiation in bone mesenchymal stem cells (BMSCs). Forkhead box O 3 (FoxO3) transcription factor is involved in lifespan and cell differentiation. In this study, we explore whether FoxO3 regulates age-related bone loss and marrow fat accumulation. The expression levels of FoxO3 in BMSCs during aging were detected in vivo and in vitro. To explore the role of FoxO3 in osteogenic and adipogenic differentiation, primary BMSCs were isolated from young and aged mice. FoxO3 expression was modulated by adenoviral vector transfection. The role of FoxO3 in bone-fat balance was evaluated by alizarin red S staining, oil red O staining, quantitative reverse transcription-polymerase chain reaction, Western blot, and histological analysis. Age-related bone loss and fat deposit are associated with downregulation of FoxO3. Overexpression of FoxO3 alleviated age-related bone loss and marrow fat accumulation in aged mice. Mechanistically, FoxO3 reduced adipogenesis and enhanced osteogenesis of BMSCs via downregulation of PPAR-γ and Notch signaling, respectively. In conclusion, FoxO3 is an essential factor controlling the fate of BMSCs and is a potential target for the prevention of age-related osteoporosis.
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Adipogénesis , Envejecimiento , Proteína Forkhead Box O3 , Células Madre Mesenquimatosas , Osteogénesis , Animales , Proteína Forkhead Box O3/metabolismo , Proteína Forkhead Box O3/genética , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Envejecimiento/genética , Envejecimiento/metabolismo , Osteogénesis/genética , Ratones , Adipogénesis/genética , Diferenciación Celular/genética , Ratones Endogámicos C57BL , Osteoporosis/metabolismo , Osteoporosis/patología , Osteoporosis/genética , Huesos/metabolismo , Transducción de Señal , PPAR gamma/metabolismo , PPAR gamma/genética , Masculino , Células Cultivadas , Receptores Notch/metabolismo , Receptores Notch/genéticaRESUMEN
BACKGROUND: Regular physical activity during childhood and adolescence is beneficial to bone development, as evidenced by the ability to increase bone density and peak bone mass by promoting bone formation. AIM: To investigate the effects of exercise on bone formation in growing mice and to investigate the underlying mechanisms. METHODS: 20 growing mice were randomly divided into two groups: Con group (control group, n = 10) and Ex group (treadmill exercise group, n = 10). Hematoxylin-eosin staining, immunohistochemistry, and micro-CT scanning were used to assess the bone formation-related indexes of the mouse femur. Bioinformatics analysis was used to find potential miRNAs targets of long non-coding RNA H19 (lncRNA H19). RT-qPCR and Western Blot were used to confirm potential miRNA target genes of lncRNA H19 and the role of lncRNA H19 in promoting osteogenic differentiation. RESULTS: Compared with the Con group, the expression of bone morphogenetic protein 2 was also significantly increased. The micro-CT results showed that 8 wk moderate-intensity treadmill exercise significantly increased bone mineral density, bone volume fraction, and the number of trabeculae, and decreased trabecular segregation in the femur of mice. Inhibition of lncRNA H19 significantly upregulated the expression of miR-149 and suppressed the expression of markers of osteogenic differentiation. In addition, knockdown of lncRNA H19 significantly downregulated the expression of autophagy markers, which is consistent with the results of autophagy-related protein changes detected in mouse femurs by immunofluorescence. CONCLUSION: Appropriate treadmill exercise can effectively stimulate bone formation and promote the increase of bone density and bone volume in growing mice, thus enhancing the peak bone mass of mice. The lncRNA H19/miR-149 axis plays an important regulatory role in osteogenic differentiation.
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BACKGROUND: Large bone defects pose a clinical treatment challenge; inhibiting transferrin receptor 2 (TfR2), which is involved in iron metabolism, can promote osteogenesis. Iron-based metal-organic frameworks (MOF-Fe) particles not only inhibit TfR2 but also serve as biomimetic catalysts to remove hydrogen peroxide in reactive oxygen species (ROS); excess ROS can disrupt the normal functions of osteoblasts, thereby hindering bone regeneration. This study explored the potential effects of MOF-Fe in increasing osteogenic activity and clearing ROS. METHODS: In vitro experiments were performed to investigate the osteogenic effects of MOF-Fe particles and assess their impact on cellular ROS levels. To further validate the role of MOF-Fe in promoting bone defect repair, we injected MOF-Fe suspensions into the femoral defects of SD rats and implanted MOF-Fe-containing hydrogel scaffolds in rabbit cranial defect models and observed their effects on bone healing. RESULTS: In vitro, the presence of MOF-Fe significantly increased the expression levels of osteogenesis-related genes and proteins compared to those in the control group. Additionally, compared to those in the untreated control group, the cells treated with MOF-Fe exhibited a significantly increased ability to remove hydrogen peroxide from ROS and generate oxygen and water within the physiological pH range. In vivo experiments further confirmed the positive effect of MOF-Fe in promoting bone defect repair. CONCLUSION: This study supports the application of MOF-Fe as an agent for bone regeneration, particularly for mitigating ROS and activating the bone morphogenetic protein (BMP) pathway, demonstrating its potential value.
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Proteína Morfogenética Ósea 2 , Regeneración Ósea , Osteogénesis , Ratas Sprague-Dawley , Animales , Proteína Morfogenética Ósea 2/metabolismo , Proteína Morfogenética Ósea 2/genética , Ratas , Regeneración Ósea/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Conejos , Estructuras Metalorgánicas/química , Receptores de Transferrina/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Peroxidasa/metabolismo , Materiales Biomiméticos/química , Materiales Biomiméticos/farmacología , Transducción de Señal/efectos de los fármacos , Peróxido de Hidrógeno , MasculinoRESUMEN
After epiphyseal fracture, the epiphyseal plate is prone to ischemia and hypoxia, leading to the formation of bone bridge and deformity. However, the exact mechanism controlling the bone bridge formation remains unclear. Notch/RBPJ signaling axis has been indicated to regulate angiogenesis and osteogenic differentiation. Our study aims to investigate the mechanism of bone bridge formation after epiphyseal plate injury, and to provide a theoretical basis for new therapeutic approaches to prevent the bone bridge formation. The expression of DLL4 and RBPJ was significantly up-regulated in HUVECs after ischemia and hypoxia treatment. Notch/RBPJ pathway positively regulated the osteogenic differentiation of BMSCs. HUVECs can induce osteogenic differentiation of BMSCs under ischemia and hypoxia. Notch/RBPJ pathway is involved in the regulation of the trans-epiphyseal bridge formation. Notch/RBPJ in HUVECs is associated with osteogenic differentiation of BMSCs and may participate in the regulation of the bone bridge formation across the epiphyseal plate.
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Diferenciación Celular , Células Endoteliales de la Vena Umbilical Humana , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas , Neovascularización Fisiológica , Osteogénesis , Receptores Notch , Transducción de Señal , Humanos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Receptores Notch/metabolismo , Receptores Notch/genética , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/metabolismo , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/genética , Hipoxia de la Célula , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Células Cultivadas , Proteínas de Unión al Calcio/metabolismo , Proteínas de Unión al Calcio/genética , AngiogénesisRESUMEN
Asthma, a disease intricately linked to immune inflammation, is significantly influenced by the immune regulatory effect of bone mesenchymal stem cells (BMSCs). This study aims to investigate changes in the homing of BMSCs in bronchial asthma, focusing on the Notch homolog (Notch)1/Jagged1 signaling pathway's role in regulating T helper 1(Th1)/T helper 2(Th2) drift. Additionally, we further explore the effects and mechanisms of homologous BMSCs implantation in asthma-related immune inflammation. Following intervention with BMSCs, a significant improvement in the pathology of rats with asthma was observed. Simultaneously, a reduction in the expression of inflammatory cells and inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin(IL)-4, and IL-13 was observed in bronchoalveolar lavage fluid (BALF). Furthermore, there was an increase in the expression of Th1 cytokine Interferon-γï¼IFN-γï¼and the transcription factor T-box expressed in T cell (T-bet), while the expression of Th2 cytokine IL-13 and transcription factor GATA binding protein (GATA)-3 decreased in lung tissue. This indicates that the Th1/Th2 drift leans towards Th1, which a crucial in ameliorating asthma inflammation. Importantly, inhibition of the Notch1 signaling pathway led to an increased expression of the Stromal cell-derived factor-1(SDF-1)/C-X-C motif chemokine receptor (CXCR)4 chemokine axis. Consequently, the homing ability of bone marrow mesenchymal stem cells to asthma-affected lung tissue was significantly enhanced. BMSCs demonstrated heightened efficacy in regulating the cytokine/chemokine network and Th1/Th2 balance, thereby restoring a stable state during the immune response process in asthma. In conclusion, inhibiting the Notch signaling pathway enhances the expression of the SDF-1 and CXCR4 chemokine axis, facilitating the migration of allogeneic BMSCs to injured lung tissues. This, in turn, promotes immune regulation and improves the Th1/Th2 imbalance, thereby enhancing the therapeutic effect on asthmatic airway inflammation.
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Asma , Células Madre Mesenquimatosas , Ratas , Animales , Ratones , Interleucina-13/metabolismo , Asma/tratamiento farmacológico , Pulmón/patología , Citocinas/metabolismo , Transducción de Señal , Factores de Transcripción/metabolismo , Inflamación/metabolismo , Quimiocinas/metabolismo , Células Madre Mesenquimatosas/metabolismo , Células Th2 , Modelos Animales de Enfermedad , Ratones Endogámicos BALB C , Receptor Notch1/metabolismoRESUMEN
BACKGROUND: Exosomes (Exos) generated from bone mesenchymal stem cells (BMSCs) are elucidated to enhance cutaneous wound healing in mice models of diabetes mellitus (DM). While underlying mechanisms remain unknown. METHODS: Next-generation sequencing (NGS) was used to examine changes in circRNA expression levels following Exo treatment. Luciferase assays were used to determine the interactions between RNAs. Immunofluorescence staining was used to examine reactive oxygen species (ROS) in endothelial progenitor cells (EPCs) cultured in high glucose (HG) conditions. Therapeutic effects regarding Exos were also examined by immunofluorescence. RESULTS: We found that Exo treatment enhanced cutaneous wound healing significantly. NGS indicated that circ-Snhg11 was involved in Exo-mediated tissue repairing. Downregulation of circ-Snhg11 decreased Exo-mediated therapy responses during wound healing in diabetic mouse. Our luciferase reporter data confirmed that SLC7A11 and miR-144-3p were circ-Snhg11 downstream targets. miR-144-3p overexpression or SLC7A11 knockdown altered the protective effects of circ-Snhg11 upon EPCs exposed to HG conditions. Upregulation of circ-Snhg11 incremented therapy effects of Exo treatment during wound healing in DM mice through enhanced angiogenesis along with a reduction in GPX4-mediated ferroptosis. CONCLUSIONS: circ-Snhg11 in BMSC-Exos enhanced SLC7A11/GPX4-mediated anti-ferroptosis signals via miR-144-3p sponging resulting in enhanced diabetic wound healing and improved angiopoiesis.
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BACKGROUND: Rotator cuff tears (RCTs) are culprit of shoulder pain and dysfunction. Tendon-bone interface (TBI) mal-healing is an essential contributor to retear after RCTs. Consequently, present project was conducted to investigate the role of bone marrow mesenchymal stem cells (BMSCs)-derived exosomes on TBI healing. METHOD: Young BMSCs (Y-BMSCs) and Aged BMSCs (A-BMSCs) were isolated from Young (3-month-old) and old (24-month-old) SD rats, and their-derived exosomes (A-BMSCs-exo and Y-BMSCs-exo) were identified. RCTs model was established, and A-BMSCs-exo and Y-BMSCs-exo were injected at the rotator cuff using hydrogel as a vehicle. Pathological changes of TBI were observed by HE, Sirius Red and Oil Red O staining. Western blotting and RT-qPCR were applied to assess the expression of extracellular matrix (ECM)-, tendon cell (TCs)-, osteogenic-, tendon-derived stem cell (TDSCs)- and angiogenic-associated proteins and mRNAs in TBI. RESULT: Y-BMSCs exhibited increased activity, osteogenic and lipogenic abilities than A-BMSCs. After A-BMSCs-exo and Y-BMSCs-exo treatment, TBI displayed massive sharpey's fibers growing along the tendon longitudinally, and a collagen fiber-chondrocyte migration zone forming a typical tendon-noncalcified fibrocartilage-calcified fibrocartilage-bone structure. A-BMSCs-exo and Y-BMSCs-exo significantly upregulated the expression of collagen Col I/II/III, Aggrecan, TNMD, SCX, Runx2, OPN, CD45, Sox2, CD31 and VEGFR2 in TBI. In vitro, A-BMSCs-exo and Y-BMSCs-exo significantly enhanced the activity of TCs and TDSCs, TDSCs stemness, and reduced the osteogenic and lipogenic capacity of TDSCs. The effect of Y-BMSCs-exo was significantly stronger than that of A-BMSCs-exo. CONCLUSION: BMSCs-derived exosomes facilitate ECM remodeling, osteogenic differentiation, angiogenesis, and stemness of TDSCs, thereby accelerating TBI healing in RCTs, with better outcomes using young individual-derived BMSCs.
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Exosomas , Células Madre Mesenquimatosas , Lesiones del Manguito de los Rotadores , Ratas , Animales , Lesiones del Manguito de los Rotadores/terapia , Ratas Sprague-Dawley , Tendones , Colágeno Tipo I/genéticaRESUMEN
BACKGROUND: Bone fracture is a common orthopedic disease that needs over 3 months to recover. Promoting the osteogenic differentiation of bone mesenchymal stem cells (BMSCs) is beneficial for fracture healing. Therefore, this research aimed to study the roles of long non-coding RNA (lncRNA) KCNQ10T1 in osteogenic differentiation of BMSCs. METHODS: BMSCs were treated with osteogenic medium and assessed by CCK-8 and flow cytometry assays. Alkaline phosphatase (ALP) staining, alizarin red staining (ARS), as well as concentration of osteoblast markers were measured to evaluate osteogenic differentiation of BMSCs. Western blot was employed to detect proteins; while, qRT-PCR was for mRNA levels. Additionally, targeted relationships between KCNQ10T1 and miR-19a-3p, as well as miR-19a-3p and SMAD5 were verified by dual luciferase reporter gene assay along with RNA pull-down method. RESULTS: Upregulation of KCNQ10T1 promoted the ALP staining and ARS intensity, increased the cell viability and decreased the apoptosis rate of BMSCs. Besides, KCNQ10T1 overexpression increased the ALP, OPG, OCN and OPN protein levels. KCNQ10T1 sponges miR-19a-3p, which targets Smad5. Upregulated miR-19a-3p reversed the overexpressed KCNQ10T1-induced effects, and depletion of SMAD5 reversed the miR-19a-3p inhibitor-induced effects on osteogenic medium-treated BMSCs. CONCLUSIONS: Upregulation of KCNQ10T1 promoted osteogenic differentiation of BMSCs through miR-19a-3p/SMAD5 axis in bone fracture.
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Fracturas Óseas , Células Madre Mesenquimatosas , MicroARNs , ARN Largo no Codificante , Animales , Ratones , Diferenciación Celular/genética , Células Cultivadas , Fracturas Óseas/metabolismo , Células Madre Mesenquimatosas/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Osteogénesis/genética , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismoRESUMEN
Introduction: The incidence and mortality rates of sepsis-induced acute kidney injury (SAKI) remain high, posing a substantial healthcare burden. Studies have implicated a connection between the development of SAKI and inflammation response, apoptosis, and autophagy. Moreover, evidence suggests that manipulating autophagy could potentially influence the prognosis of this condition. Notably, exosomes derived from bone mesenchymal stem cells (BMSCs-Exo) have exhibited promise in mitigating cellular damage by modulating pathways associated with inflammation, apoptosis, and autophagy. Thus, this study aims to investigate the influence of BMSCs-Exo on SAKI and the potential mechanisms that drive this impact. Methods: The SAKI model was induced in HK-2 cells using lipopolysaccharide (LPS), while rats underwent cecal ligation and puncture (CLP) to simulate the condition. Cell viability was assessed using the CCK-8 kit, and kidney damage was evaluated through HE staining, blood urea nitrogen (BUN), and serum creatinine (SCr) measurements. Inflammatory-related RNAs and proteins were quantified via qPCR and ELISA, respectively. Apoptosis was determined through apoptosis-related protein levels, flow cytometry, and TUNEL staining. Western blot analysis was utilized to measure associated protein expressions. Results: In vivo, BMSCs-Exo ameliorated kidney injury in CLP-induced SAKI rats, reducing inflammatory cytokine production and apoptosis levels. Fluorescence microscope observed the absorption of BMSCs-Exo by renal cells following injection via tail vein. In the SAKI rat kidney tissue, there was an upregulation of LC3-II/LC3-I, p62, and phosphorylated AMP-activated protein kinase (p-AMPK) expressions, indicating blocked autophagic flux, while phosphorylated mammalian target of rapamycin (p-mTOR) expression was downregulated. However, BMSCs-Exo enhanced LC3-II/LC3-I and p-AMPK expression, concurrently reducing p62 and p-mTOR levels. In vitro, BMSCs-Exo enhanced cell viability in LPS-treated HK-2 cells, and exerted anti-inflammation and anti-apoptosis effects which were consistent with the results in vivo. Similarly, rapamycin (Rapa) exhibited a protective effect comparable to BMSCs-Exo, albeit partially abrogated by 3-methyladenine (3-MA). Conclusion: BMSCs-Exo mitigate inflammation and apoptosis through autophagy in SAKI, offering a promising avenue for SAKI treatment.