RESUMEN
Implant-associated infection (IAI) has become an intractable challenge in clinic. The healing of IAI is a complex physiological process involving a series of spatiotemporal connected events. However, existing titanium-based implants in clinic suffer from poor antibacterial effect and single function. Herein, a versatile surface platform based on the presentation of sequential function is developed. Fabrication of titania nanotubes and poly-γ-glutamic acid (γ-PGA) achieves the efficient incorporation of silver ions (Ag+) and the pH-sensitive release in response to acidic bone infection microenvironment. The optimized PGA/Ag platform exhibits satisfactory biocompatibility and converts macrophages from pro-inflammatory M1 to pro-healing M2 phenotype during the subsequent healing stage, which creates a beneficial osteoimmune microenvironment and promotes angio/osteogenesis. Furthermore, the PGA/Ag platform mediates osteoblast/osteoclast coupling through inhibiting CCL3/CCR1 signaling. These biological effects synergistically improve osseointegration under bacterial infection in vivo, matching the healing process of IAI. Overall, the novel integrated PGA/Ag surface platform proposed in this study fulfills function cascades under pathological state and shows great potential in IAI therapy.
Asunto(s)
Antibacterianos , Ácido Poliglutámico , Plata , Titanio , Animales , Titanio/química , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Ratones , Ácido Poliglutámico/química , Ácido Poliglutámico/análogos & derivados , Plata/química , Plata/farmacología , Propiedades de Superficie , Nanotubos/química , Células RAW 264.7 , Infecciones Relacionadas con Prótesis/tratamiento farmacológico , Oseointegración/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Osteoblastos/efectos de los fármacos , Osteoblastos/citología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Cicatrización de Heridas/efectos de los fármacos , Prótesis e ImplantesRESUMEN
Bone tissue engineering addresses the limitations of autologous resources and the risk of allograft disease transmission in bone diseases. In this regard, engineered three-dimensional (3D) models emerge as biomimetic alternatives to natural tissues, replicating intracellular communication. Moreover, the unique properties of super-paramagnetic iron oxide nanoparticles (SPIONs) were shown to promote bone regeneration via enhanced osteogenesis and angiogenesis in bone models. This study aimed to investigate the effects of SPION on both osteogenesis and angiogenesis and characterized a co-culture of Human umbilical vein endothelial cells (HUVEC) and MG-63 cells as a model of bone microtissue. HUVECs: MG-63s with a ratio of 4:1 demonstrated the best results among other cell ratios, and 50 µg/mL of SPION was the optimum concentration for maximum survival, cell migration and mineralization. In addition, the data from gene expression illustrated that the expression of osteogenesis-related genes, including osteopontin, osteocalcin, alkaline phosphatase, and collagen-I, as well as the expression of the angiogenesis-related marker, CD-31, and the tube formation, is significantly elevated when the 50 µg/mL concentration of SPION is applied to the microtissue samples. SPION application in a designed 3D bone microtissue model involving a co-culture of osteoblast and endothelial cells resulted in increased expression of specific markers related to angiogenesis and osteogenesis. This includes the design of a novel biomimetic model to boost blood compatibility and biocompatibility of primary materials while promoting osteogenic activity in microtissue bone models. Moreover, this can improve interaction with surrounding tissues and broaden the knowledge to promote superior-performance implants, preventing device failure.
Asunto(s)
Regeneración Ósea , Técnicas de Cocultivo , Células Endoteliales de la Vena Umbilical Humana , Osteogénesis , Ingeniería de Tejidos , Humanos , Regeneración Ósea/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Ingeniería de Tejidos/métodos , Nanopartículas de Magnetita/química , Neovascularización Fisiológica/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Nanopartículas Magnéticas de Óxido de Hierro/química , Supervivencia Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Osteoblastos/metabolismo , Osteoblastos/efectos de los fármacos , Osteoblastos/citologíaRESUMEN
The aim of this study was to develop multifunctional magnetic poly(ε-caprolactone) (PCL) mats with antibacterial properties for bone tissue engineering and osteosarcoma prevention. To provide good dispersion of magnetic iron oxide nanoparticles (IONs), they were first grafted with PCL using a novel three-step approach. Then, a series of PCL-based mats containing a fixed amount of ION@PCL particles and an increasing content of ascorbic acid (AA) was prepared by electrospinning. AA is known for increasing osteoblast activity and suppressing osteosarcoma cells. Composites were characterized in terms of morphology, mechanical properties, hydrolytic stability, antibacterial performance, and biocompatibility. AA affected both the fiber diameter and the mechanical properties of the nanocomposites. All produced mats were nontoxic to rat bone marrow-derived mesenchymal cells; however, a composite with 5 wt.% of AA suppressed the initial proliferation of SAOS-2 osteoblast-like cells. Moreover, AA improved antibacterial properties against Staphylococcus aureus and Escherichia coli compared to PCL. Overall, these magnetic composites, reported for the very first time, can be used as scaffolds for both tissue regeneration and osteosarcoma prevention.
Asunto(s)
Ácido Ascórbico , Poliésteres , Staphylococcus aureus , Ingeniería de Tejidos , Poliésteres/química , Ácido Ascórbico/química , Ácido Ascórbico/farmacología , Humanos , Ratas , Animales , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/crecimiento & desarrollo , Escherichia coli/efectos de los fármacos , Antibacterianos/química , Antibacterianos/farmacología , Nanopartículas de Magnetita/química , Osteoblastos/metabolismo , Osteoblastos/citología , Línea Celular Tumoral , Osteosarcoma/patología , Huesos , Nanocompuestos/química , Andamios del Tejido/química , Ensayo de MaterialesRESUMEN
Diet-induced obesity is associated with enhanced systemic inflammation that limits bone regeneration. HDAC inhibitors are currently being explored as anti-inflammatory agents. Prior reports show that myeloid progenitor-directed Hdac3 ablation enhances intramembranous bone healing in female mice. In this study, we determined if Hdac3 ablation increased intramembranous bone regeneration in mice fed a high-fat/high-sugar (HFD) diet. Micro-CT analyses demonstrated that HFD-feeding enhanced the formation of periosteal reaction tissue of control littermates, reflective of suboptimal bone healing. We confirmed enhanced bone volume within the defect of Hdac3-ablated females and showed that Hdac3 ablation reduced the amount of periosteal reaction tissue following HFD feeding. Osteoblasts cultured in a conditioned medium derived from Hdac3-ablated cells exhibited a four-fold increase in mineralization and enhanced osteogenic gene expression. We found that Hdac3 ablation elevated the secretion of several chemokines, including CCL2. We then confirmed that Hdac3 deficiency increased the expression of Ccl2. Lastly, we show that the proportion of CCL2-positve cells within bone defects was significantly higher in Hdac3-deficient mice and was further enhanced by HFD. Overall, our studies demonstrate that Hdac3 deletion enhances intramembranous bone healing in a setting of diet-induced obesity, possibly through increased production of CCL2 by macrophages within the defect.
Asunto(s)
Dieta Occidental , Histona Desacetilasas , Osteogénesis , Animales , Femenino , Histona Desacetilasas/metabolismo , Histona Desacetilasas/genética , Histona Desacetilasas/deficiencia , Ratones , Dieta Occidental/efectos adversos , Osteoblastos/metabolismo , Dieta Alta en Grasa/efectos adversos , Periostio/metabolismo , Periostio/patología , Quimiocina CCL2/metabolismo , Quimiocina CCL2/genética , Regeneración Ósea , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/metabolismo , Obesidad/etiología , Obesidad/patologíaRESUMEN
BACKGROUND: Bone loss caused by microgravity exposure presents a serious threat to the health of astronauts, but existing treatment strategies have specific restrictions. This research aimed to investigate whether salidroside (SAL) can mitigate microgravity-induced bone loss and its underlying mechanism. METHODS: In this research, we used hindlimb unloading (HLU) and the Rotary Cell Culture System (RCCS) to imitate microgravity in vivo and in vitro. RESULTS: The results showed that salidroside primarily enhances bone density, microstructure, and biomechanical properties by stimulating bone formation and suppressing bone resorption, thereby preserving bone mass in HLU rats. In MC3T3-E1 cells cultured under simulated microgravity in rotary wall vessel bioreactors, the expression of osteogenic genes significantly increased after salidroside administration, indicating that salidroside can promote osteoblast differentiation under microgravity conditions. Furthermore, the Nrf2 inhibitor ML385 diminished the therapeutic impact of salidroside on microgravity-induced bone loss. Overall, this research provides the first evidence that salidroside can mitigate bone loss induced by microgravity exposure through stimulating the Nrf2/HO-1 pathway. CONCLUSION: These findings indicate that salidroside has great potential for treating space-related bone loss in astronauts and suggest that Nrf2/HO-1 is a viable target for counteracting microgravity-induced bone damage.
Asunto(s)
Glucósidos , Factor 2 Relacionado con NF-E2 , Fenoles , Simulación de Ingravidez , Glucósidos/farmacología , Glucósidos/uso terapéutico , Animales , Fenoles/farmacología , Fenoles/uso terapéutico , Factor 2 Relacionado con NF-E2/metabolismo , Ratones , Simulación de Ingravidez/efectos adversos , Ratas , Masculino , Hemo-Oxigenasa 1/metabolismo , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Ingravidez/efectos adversos , Osteogénesis/efectos de los fármacos , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Suspensión Trasera , Resorción Ósea/prevención & control , Resorción Ósea/etiología , Resorción Ósea/metabolismo , Densidad Ósea/efectos de los fármacos , Proteínas de la MembranaRESUMEN
The periosteum contains skeletal stem/progenitor cells that contribute to bone fracture healing. However, the in vivo identity of periosteal skeletal stem cells (P-SSCs) remains unclear, and membrane protein markers of P-SSCs that facilitate tissue engineering are needed. Here, we identified integral membrane protein 2A (Itm2a) enriched in SSCs using single-cell transcriptomics. Itm2a+ P-SSCs displayed clonal multipotency and self-renewal and sat at the apex of their differentiation hierarchy. Lineage-tracing experiments showed that Itm2a selectively labeled the periosteum and that Itm2a+ cells were preferentially located in the outer fibrous layer of the periosteum. The Itm2a+ cells rarely expressed CD34 or Osx, but expressed periosteal markers such as Ctsk, CD51, PDGFRA, Sca1, and Gli1. Itm2a+ P-SSCs contributed to osteoblasts, chondrocytes, and marrow stromal cells upon injury. Genetic lineage tracing using dual recombinases showed that Itm2a and Prrx1 lineage cells generated spatially separated subsets of chondrocytes and osteoblasts during fracture healing. Bone morphogenetic protein 2 (Bmp2) deficiency or ablation of Itm2a+ P-SSCs resulted in defects in fracture healing. ITM2A+ P-SSCs were also present in the human periosteum. Thus, our study identified a membrane protein marker that labels P-SSCs, providing an attractive target for drug and cellular therapy for skeletal disorders.
Asunto(s)
Curación de Fractura , Proteínas de la Membrana , Periostio , Animales , Periostio/metabolismo , Periostio/citología , Ratones , Curación de Fractura/genética , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Humanos , Células Madre/metabolismo , Células Madre/citología , Proteína Morfogenética Ósea 2/metabolismo , Proteína Morfogenética Ósea 2/genética , Fracturas Óseas/patología , Fracturas Óseas/metabolismo , Fracturas Óseas/terapia , Fracturas Óseas/genética , Osteoblastos/metabolismo , Osteoblastos/citología , Diferenciación Celular , Condrocitos/metabolismo , Condrocitos/citología , Masculino , Linaje de la CélulaRESUMEN
Mesenchymal stromal stem cells (MSCs) or skeletal stem cells (SSCs) play a major role in tissue repair due to their robust ability to differentiate into osteoblasts, chondrocytes, and adipocytes. Complex cell signaling cascades tightly regulate this differentiation. In osteogenic differentiation, Runt-related transcription factor 2 (RUNX2) and ALP activity are essential. Furthermore, during the latter stages of osteogenic differentiation, mineral formation mediated by the osteoblast occurs with the secretion of a collagenous extracellular matrix and calcium deposition. Activation of nuclear factor erythroid 2-related factor 2 (NRF2), an important transcription factor against oxidative stress, inhibits osteogenic differentiation and mineralization via modulation of RUNX2 function; however, the exact role of NRF2 in osteoblastogenesis remains unclear. Here, we demonstrate that NRF2 activation in human bone marrow-derived stromal cells (HBMSCs) suppressed osteogenic differentiation. NRF2 activation increased the expression of STRO-1 and KITLG (stem cell markers), indicating NRF2 protects HBMSCs stemness against osteogenic differentiation. In contrast, NRF2 activation enhanced mineralization, which is typically linked to osteogenic differentiation. We determined that these divergent results were due in part to the modulation of cellular calcium flux genes by NRF2 activation. The current findings demonstrate a dual role for NRF2 as a HBMSC maintenance factor as well as a central factor in mineralization, with implications therein for elucidation of bone formation and cellular Ca2+ kinetics, dystrophic calcification and, potentially, application in the modulation of bone formation.
Asunto(s)
Diferenciación Celular , Células Madre Mesenquimatosas , Factor 2 Relacionado con NF-E2 , Osteoblastos , Osteogénesis , Humanos , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Osteogénesis/fisiología , Diferenciación Celular/fisiología , Osteoblastos/metabolismo , Osteoblastos/citología , Calcificación Fisiológica/fisiología , Células Cultivadas , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/citología , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genéticaRESUMEN
Osteosarcoma (OS) is the mostly commonly occurring primary bone cancer. Despite comprehensive treatment programs including neoadjuvant chemotherapy and tumour resection, survival rates have not improved significantly since the 1970s. Survival rates are dramatically reduced for patients who suffer a local recurrence. Furthermore, primary bone cancer patients are at increased risk of bone fractures. Consequently, there is an urgent need for alternative treatment options. In this paper we report the development of novel gallium doped bioactive glass that selectively kill bone cancer cells whilst simultaneously stimulating new bone growth. Here we show, using a combination of 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide, LIVE/DEAD assays and image analysis, that bioactive glasses containing gallium oxide are highly toxic and reduce both the proliferation and migration of bone cancer cells (Saos-2) in a dose dependant manner. Glasses containing 5 mol% gallium oxide reduced the viability of OS cells by 99% without being cytotoxic to the non-cancerous normal human osteoblasts (NHOst) control cells. Furthermore, Fourier transform infrared and energy-dispersive x-ray spectroscopy results confirmed the formation of an amorphous calcium phosphate/hydroxyapatite like layer on the surface of the bioactive glass particulates, after 7 d incubating in simulated body fluid, indicating the early stages of bone formation. These materials show significant potential for use in bone cancer applications as part of a multimodal treatment.
Asunto(s)
Antineoplásicos , Neoplasias Óseas , Proliferación Celular , Supervivencia Celular , Galio , Vidrio , Osteosarcoma , Humanos , Galio/química , Osteosarcoma/tratamiento farmacológico , Antineoplásicos/farmacología , Antineoplásicos/química , Vidrio/química , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Neoplasias Óseas/tratamiento farmacológico , Espectroscopía Infrarroja por Transformada de Fourier , Osteoblastos/efectos de los fármacos , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Movimiento Celular/efectos de los fármacos , Sistemas de Liberación de Medicamentos , Ensayo de MaterialesRESUMEN
BACKGROUND: Obesity poses a significant global health challenge, given its association with the excessive accumulation of adipose tissue (AT) and various systemic disruptions. Within the adipose microenvironment, expansion and enrichment with immune cells trigger the release of inflammatory mediators and growth factors, which can disrupt tissues, including bones. While obesity's contribution to bone loss is well established, the direct impact of obese AT on osteoblast maturation remains uncertain. This study aimed to explore the influence of the secretomes from obese and lean AT on osteoblast differentiation and activity. METHODS: SAOS-2 cells were exposed to the secretomes obtained by culturing human subcutaneous AT from individuals with obesity (OATS) or lean patients, and their effects on osteoblasts were evaluated. RESULTS: In the presence of the OATS, mature osteoblasts underwent dedifferentiation, showing an increased proliferation accompanied by a morphological shift towards a mesenchymal phenotype, with detrimental effects on osteogenic markers and the calcification capacity. Concurrently, the OATS promoted the expression of mesenchymal and adipogenic markers, inducing the formation of cytoplasmic lipid droplets in SAOS-2 cells exposed to an adipogenic differentiation medium. Additionally, TGF-ß1 emerged as a key mediator of these effects, as the OATS was enriched with this growth factor. CONCLUSIONS: Our findings demonstrate that obese subcutaneous AT promotes the dedifferentiation of osteoblasts and increases the adipogenic profile in these cells.
Asunto(s)
Adipogénesis , Tejido Adiposo , Desdiferenciación Celular , Obesidad , Osteoblastos , Fenotipo , Transducción de Señal , Factor de Crecimiento Transformador beta1 , Humanos , Osteoblastos/metabolismo , Osteoblastos/patología , Obesidad/patología , Obesidad/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Tejido Adiposo/metabolismo , Tejido Adiposo/patología , Secretoma/metabolismo , Diferenciación Celular , Proliferación Celular , Osteogénesis , MasculinoRESUMEN
Fibrous dysplasia (FD) is a mosaic skeletal disorder involving the development of benign, expansile fibro-osseous lesions during childhood that cause deformity, fractures, pain, and disability. There are no well-established treatments for FD. Fibroblast activation protein (FAPα) is a serine protease expressed in pathological fibrotic tissues that has promising clinical applications as a biomarker and local pro-drug activator in several pathological conditions. In this study, we explored the expression of FAP in FD tissue and cells through published genetic expression datasets and measured circulating FAPα in plasma samples from patients with FD and healthy donors. We found that FAP genetic expression was increased in FD tissue and cells, and present at higher concentrations in plasma from patients with FD compared to healthy donors. Moreover, FAPα levels were correlated with skeletal disease burden in patients with FD. These findings support further investigation of FAPα as a potential imaging and/or biomarker of FD, as well as a pro-drug activator specific to FD tissue.
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Endopeptidasas , Displasia Fibrosa Ósea , Gelatinasas , Proteínas de la Membrana , Serina Endopeptidasas , Humanos , Serina Endopeptidasas/metabolismo , Serina Endopeptidasas/genética , Femenino , Masculino , Endopeptidasas/metabolismo , Endopeptidasas/genética , Gelatinasas/metabolismo , Gelatinasas/genética , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Displasia Fibrosa Ósea/metabolismo , Displasia Fibrosa Ósea/genética , Displasia Fibrosa Ósea/patología , Adulto , Adolescente , Niño , Biomarcadores/metabolismo , Biomarcadores/sangre , Osteoblastos/metabolismo , Osteoblastos/patología , Persona de Mediana EdadRESUMEN
BACKGROUND: Bone regeneration is a well-regulated dynamic process, of which the prominent role of the immune system on bone homeostasis is more and more revealed by recent research. Before fully activation of the bone remodeling cells, the immune system needs to clean up the microenvironment in facilitating the bone repair initiation. Furthermore, this microenvironment must be maintained properly by various mechanisms over the entire bone regeneration process. OBJECTIVE: This review aims to summarize the role of the T-helper 17/Regulatory T cell (Th17/Treg) balance in bone cell remodeling and discuss the relevant progress in bone tissue engineering. RESULTS: The role of the immune response in the early stages of bone regeneration is crucial, especially the impact of the Th17/Treg balance on osteoclasts, mesenchymal stem cells (MSCs), and osteoblasts activity. By virtue of these knowledge advancements, innovative approaches in bone tissue engineering, such as nano-structures, hydrogel, and exosomes, are designed to influence the Th17/Treg balance and thereby augment bone repair and regeneration. CONCLUSION: Targeting the Th17/Treg balance is a promising innovative strategy for developing new treatments to enhance bone regeneration, thus offering potential breakthroughs in bone injury clinics.
Asunto(s)
Regeneración Ósea , Huesos , Linfocitos T Reguladores , Células Th17 , Ingeniería de Tejidos , Humanos , Linfocitos T Reguladores/inmunología , Ingeniería de Tejidos/métodos , Regeneración Ósea/inmunología , Animales , Células Th17/inmunología , Huesos/inmunología , Células Madre Mesenquimatosas/inmunología , Células Madre Mesenquimatosas/metabolismo , Remodelación Ósea/inmunología , Osteoblastos/inmunología , Osteoclastos/inmunología , Osteoclastos/metabolismoRESUMEN
Maxillofacial bone defects can severely impact quality of life by impairing physiological functions such as chewing, breathing, swallowing, and pronunciation. Polyether ether ketone (PEEK) is commonly used for the repair of maxillofacial defects due to its mechanical adaptability, while its osteogenic properties still need refinement. Herein, we have utilized the piezoelectric effect exhibited by barium titanate (BTO) under low-intensity pulsed ultrasound (LIPUS) to develop an ultrasound responsive PEEK (PDA@BTO-SPEEK, PBSP) through the mediating effect of polydopamine (PDA), for repairing maxillofacial bone defects. After modification by PDA@BTO, PBSP possesses better hydrophilicity, which is conducive to cell growth and adhesion. Simultaneously, by virtue of the piezoelectric characteristics of BTO, PBSP obtains a piezoelectric coefficient that matches the bone cortex. Notably, when PBSP is stimulated by LIPUS, it can generate stable electricity and effectively accelerate the osteogenic differentiation of osteoblasts through the regulation of the Piezo1-induced calcium (Ca2+) influx and Akt/GSK3ß/ß-catenin pathway. In addition, PBSP presents satisfactory therapeutic effects in rat skull defect models, and its osteogenic efficiency can be further improved under LIPUS stimulation with high tissue penetration. Collectively, PBSP + LIPUS exhibits great potential as a promising alternative strategy for the repair of maxillofacial bone defects.
Asunto(s)
Benzofenonas , Glucógeno Sintasa Quinasa 3 beta , Cetonas , Osteogénesis , Polietilenglicoles , Polímeros , Proteínas Proto-Oncogénicas c-akt , Ratas Sprague-Dawley , beta Catenina , Animales , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Polímeros/química , Osteogénesis/efectos de los fármacos , Ratas , Polietilenglicoles/química , Proteínas Proto-Oncogénicas c-akt/metabolismo , Cetonas/química , Cetonas/farmacología , beta Catenina/metabolismo , Diferenciación Celular/efectos de los fármacos , Osteoblastos/efectos de los fármacos , Ondas Ultrasónicas , Indoles/química , Indoles/farmacología , Masculino , Transducción de Señal/efectos de los fármacos , Cráneo/efectos de los fármacos , Titanio/química , Titanio/farmacología , Regeneración Ósea/efectos de los fármacosRESUMEN
Developing bones can adapt their shape in response to mechanical stresses from neighbouring growing organs. In a new study, Koichi Matsuo and colleagues examine how bone-forming osteoblasts and bone-resorbing osteoclasts coordinate growth in the mouse fibula. They describe the process called 'endo-forming trans-pairing', where bone resorption by osteoclasts in the outer periosteum is paired with bone formation by osteoblasts in the inner endosteum to shape the growing bone. To learn more about the story behind the paper, we caught up with first author Yukiko Kuroda and the corresponding author Koichi Matsuo, Professor at the School of Medicine, Keio University, Japan.
Asunto(s)
Huesos , Microscopía/métodos , Huesos/citología , Osteoclastos/citología , Osteoclastos/metabolismo , Osteoblastos/citología , Osteoblastos/metabolismo , Nervio Ciático , Desarrollo Óseo , Animales , MorfogénesisRESUMEN
Diabetes-related bone loss represents a significant complication that persistently jeopardizes the bone health of individuals with diabetes. Primary cilia proteins have been reported to play a vital role in regulating osteoblast differentiation in diabetes-related bone loss. However, the specific contribution of KIAA0753, a primary cilia protein, in bone loss induced by diabetes remains unclear. In this investigation, we elucidated the pivotal role of KIAA0753 as a promoter of osteoblast differentiation in diabetes. RNA sequencing demonstrated a marked downregulation of KIAA0753 expression in pro-bone MC3T3 cells exposed to a high glucose environment. Diabetes mouse models further validated the downregulation of KIAA0753 protein in the femur. Diabetes was observed to inhibit osteoblast differentiation in vitro, evidenced by downregulating the protein expression of OCN, OPN and ALP, decreasing primary cilia biosynthesis, and suppressing the Hedgehog signalling pathway. Knocking down KIAA0753 using shRNA methods was found to shorten primary cilia. Conversely, overexpression KIAA0753 rescued these changes. Additional insights indicated that KIAA0753 effectively restored osteoblast differentiation by directly interacting with SHH, OCN and Gli2, thereby activating the Hedgehog signalling pathway and mitigating the ubiquitination of Gli2 in diabetes. In summary, we report a negative regulatory relationship between KIAA0753 and diabetes-related bone loss. The clarification of KIAA0753's role offers valuable insights into the intricate mechanisms underlying diabetic bone complications.
Asunto(s)
Diferenciación Celular , Proteínas Asociadas a Microtúbulos , Osteoblastos , Transducción de Señal , Animales , Humanos , Masculino , Ratones , Línea Celular , Cilios/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Experimental/genética , Proteínas Hedgehog/metabolismo , Proteínas Hedgehog/genética , Ratones Endogámicos C57BL , Osteoblastos/metabolismo , Osteogénesis/genética , Proteínas Asociadas a Microtúbulos/metabolismoRESUMEN
Plastin-3 (PLS3) encodes T-plastin, an actin-bundling protein mediating the formation of actin filaments by which numerous cellular processes are regulated. Loss-of-function genetic defects in PLS3 are reported to cause X-linked osteoporosis and childhood-onset fractures. However, the molecular etiology of PLS3 remains elusive. Functional compensation by actin-bundling proteins ACTN1, ACTN4, and FSCN1 was investigated in zebrafish following morpholino-mediated pls3 knockdown. Primary dermal fibroblasts from six patients with a PLS3 variant were also used to examine expression of these proteins during osteogenic differentiation. In addition, Pls3 knockdown in the murine MLO-Y4 cell line was employed to provide insights in global gene expression. Our results showed that ACTN1 and ACTN4 can rescue the skeletal deformities in zebrafish after pls3 knockdown, but this was inadequate for FSCN1. Patients' fibroblasts showed the same osteogenic transdifferentiation ability as healthy donors. RNA-seq results showed differential expression in Wnt1, Nos1ap, and Myh3 after Pls3 knockdown in MLO-Y4 cells, which were also associated with the Wnt and Th17 cell differentiation pathways. Moreover, WNT2 was significantly increased in patient osteoblast-like cells compared to healthy donors. Altogether, our findings in different bone cell types indicate that the mechanism of PLS3-related pathology extends beyond actin-bundling proteins, implicating broader pathways of bone metabolism.
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Diferenciación Celular , Glicoproteínas de Membrana , Proteínas de Microfilamentos , Osteogénesis , Pez Cebra , Pez Cebra/metabolismo , Pez Cebra/genética , Animales , Osteogénesis/genética , Humanos , Proteínas de Microfilamentos/metabolismo , Proteínas de Microfilamentos/genética , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/genética , Ratones , Fibroblastos/metabolismo , Osteoblastos/metabolismo , Proteínas de Pez Cebra/metabolismo , Proteínas de Pez Cebra/genética , Técnicas de Silenciamiento del GenRESUMEN
Osteoporosis and other degenerative bone diseases pose significant challenges to global healthcare systems due to their prevalence and impact on quality of life. Current treatments often alleviate symptoms without fully restoring damaged bone tissue, highlighting the need for innovative approaches like stem cell therapy. Adipose-derived mesenchymal stem cells (ADMSCs) are particularly promising due to their accessibility, abundant supply, and strong differentiation potential. However, ADMSCs tend to favor adipogenic pathways, necessitating the use of differentiation inducers (DIs), three-dimensional (3D) hydrogel environments, and photobiomodulation (PBM) to achieve targeted osteogenic differentiation. This study investigated the combined effects of osteogenic DIs, a fast-dextran hydrogel matrix, and PBM at specific wavelengths and fluences on the proliferation and differentiation of immortalized ADMSCs into osteoblasts. Near-infrared (NIR) and green (G) light, as well as their combination, were used with fluences of 3 J/cm2, 5 J/cm2, and 7 J/cm2. The results showed statistically significant increases in alkaline phosphatase levels, a marker of osteogenic differentiation, with G light at 7 J/cm2 demonstrating the most substantial impact on ADMSC differentiation. Calcium deposits, visualized by Alizarin red S staining, appeared as early as 24 h post-treatment in PBM groups, suggesting accelerated osteogenic differentiation. ATP luminescence assays indicated increased proliferation in all experimental groups, particularly with NIR and NIR-G light at 3 J/cm2 and 5 J/cm2. MTT viability and LDH membrane permeability assays confirmed enhanced cell viability and stable cell health, respectively. In conclusion, PBM significantly influences the differentiation and proliferation of hydrogel-embedded immortalized ADMSCs into osteoblast-like cells, with G light at 7 J/cm2 being particularly effective. These findings support the combined use of 3D hydrogel matrices and PBM as a promising approach in regenerative medicine, potentially leading to innovative treatments for degenerative bone diseases.
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Diferenciación Celular , Terapia por Luz de Baja Intensidad , Células Madre Mesenquimatosas , Osteogénesis , Osteogénesis/efectos de la radiación , Diferenciación Celular/efectos de la radiación , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/efectos de la radiación , Humanos , Terapia por Luz de Baja Intensidad/métodos , Técnicas de Cultivo Tridimensional de Células/métodos , Proliferación Celular/efectos de la radiación , Tejido Adiposo/citología , Hidrogeles/química , Osteoblastos/citología , Osteoblastos/metabolismo , Osteoblastos/efectos de la radiación , Fosfatasa Alcalina/metabolismo , Células CultivadasRESUMEN
Bone has the ability to heal itself; however, bone defects fail to heal once the damage exceeds a critical size. Bone regeneration remains a significant clinical challenge, with autograft considered the ideal bone graft material due to its sufficient porosity, osteogenic cells, and biological growth factors. However, limitations to bone grafting, such as limited bone stock and high resorption rates, have led to a great deal of research into developing bone graft substitutes. The P28 peptide is a small molecule bioactive biomimetic alternative to mimic the bone morphogenetic protein 2 (BMP-2). In this study, we investigated the potential of P28-loaded hybrid scaffolds to mimic the natural bone structure for enhancing the bone regeneration process. We hypothesized that the peptide-loaded scaffolds and nude scaffolds both have the potential to promote bone healing, and the bone healing process is accelerated by the release of the peptide. To verify our hypothesis, C2C12 cells were evaluated for the presence of calcium deposits by histological stain at 7 and 14 days in cultures with hybrid scaffolds. Total RNA was isolated from C2C12 cells cultured with hybrid scaffolds for 7 and 14 days to assess osteoblast differentiation. The project findings demonstrated that the hybrid scaffold could enhance osteoblast differentiation and significantly improve the therapeutic effects of the scaffold in bone regeneration.
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Regeneración Ósea , Diferenciación Celular , Cerámica , Quitosano , Andamios del Tejido , Regeneración Ósea/efectos de los fármacos , Andamios del Tejido/química , Quitosano/química , Quitosano/farmacología , Animales , Ratones , Cerámica/química , Cerámica/farmacología , Diferenciación Celular/efectos de los fármacos , Osteoblastos/efectos de los fármacos , Osteoblastos/citología , Osteoblastos/metabolismo , Línea Celular , Osteogénesis/efectos de los fármacos , Proteína Morfogenética Ósea 2/farmacología , Ingeniería de Tejidos/métodos , Péptidos/química , Péptidos/farmacología , HumanosRESUMEN
Chitosan is a promising natural polymer for coatings, it combines intrinsic antibacterial and pro-osteoblastic properties, but the literature still has a gap from the development to behavior of these coatings, so this systematic review aimed to answer, "What is the relationship between the physical and chemical properties of polymeric chitosan coatings on titanium implants on antibacterial activity and osteoblast viability?". PRISMA guidelines was followed, the search was applied into 4 databases and grey literature, without the restriction of time and language. The selection process occurred in 2 blinded steps by the authors. The criteria of eligibility were in vitro studies that evaluated the physical, chemical, microbiological, and biological properties of chitosan coatings on titanium surfaces. The risk of bias was analyzed by the specific tool. Of 734 potential articles 10 were included; all had low risk of bias. The coating was assessed according to the technique of fabrication, FT-IR, thickness, adhesion, roughness, wettability, antibacterial activity, and osteoblast viability. The analyzed coatings showed efficacy on antibacterial activity and cytocompatibility dependent on the class of material incorporated. Thus, this review motivates the development of time-dependent studies to optimize manufacturing and allow for an increase in patents and availability on the market.
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Antibacterianos , Quitosano , Materiales Biocompatibles Revestidos , Osteoblastos , Titanio , Quitosano/química , Quitosano/farmacología , Titanio/química , Titanio/farmacología , Materiales Biocompatibles Revestidos/química , Materiales Biocompatibles Revestidos/farmacología , Humanos , Antibacterianos/farmacología , Antibacterianos/química , Osteoblastos/efectos de los fármacos , Osteoblastos/citología , Propiedades de Superficie , Prótesis e Implantes , Animales , Supervivencia Celular/efectos de los fármacosRESUMEN
BACKGROUND: Mechanical unloading-induced bone loss threatens prolonged spaceflight and human health. Recent studies have confirmed that osteoporosis is associated with a significant reduction in bone microvessels, but the relationship between them and the underlying mechanism under mechanical unloading are still unclear. METHODS: We established a 2D clinostat and hindlimb-unloaded (HLU) mouse model to simulate unloading in vitro and in vivo. Micro-CT scanning was performed to assess changes in the bone microstructure and mass of the tibia. The levels of CD31, Endomucin (EMCN) and histone deacetylase 6 (HDAC6) in tibial microvessels were detected by immunofluorescence (IF) staining. In addition, we established a coculture system of microvascular endothelial cells (MVECs) and osteoblasts, and qRTâPCR or western blotting was used to detect RNA and protein expression; cell proliferation was detected by CCKâ8 and EdU assays. ChIP was used to detect whether HDAC6 binds to the miRNA promoter region. RESULTS: Bone mass and bone microvessels were simultaneously significantly reduced in HLU mice. Furthermore, MVECs effectively promoted the proliferation and differentiation of osteoblasts under coculture conditions in vitro. Mechanistically, we found that the HDAC6 content was significantly reduced in the bone microvessels of HLU mice and that HDAC6 inhibited the expression of miR-375-3p by reducing histone acetylation in the miR-375 promoter region in MVECs. miR-375-3p was upregulated under unloading and it could inhibit MVEC proliferation by directly targeting low-density lipoprotein-related receptor 5 (LRP5) expression. In addition, silencing HDAC6 promoted the miR-375-3p/LRP5 pathway to suppress MVEC proliferation under mechanical unloading, and regulation of HDAC6/miR-375-3p axis in MVECs could affect osteoblast proliferation under coculture conditions. CONCLUSION: Our study revealed that disuse-induced bone loss may be closely related to a reduction in the number of bone microvessels and that the modulation of MVEC function could improve bone loss induced by unloading. Mechanistically, the HDAC6/miR-375-3p/LRP5 pathway in MVECs might be a promising strategy for the clinical treatment of unloading-induced bone loss.
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Proliferación Celular , Células Endoteliales , Epigénesis Genética , Suspensión Trasera , Histona Desacetilasa 6 , MicroARNs , Microvasos , Osteoblastos , Animales , MicroARNs/metabolismo , MicroARNs/genética , Células Endoteliales/metabolismo , Histona Desacetilasa 6/metabolismo , Histona Desacetilasa 6/genética , Microvasos/patología , Osteoblastos/metabolismo , Ratones Endogámicos C57BL , Ratones , Técnicas de Cocultivo , Diferenciación Celular , Masculino , Resorción Ósea/patología , Secuencia de Bases , Inhibidores de Histona Desacetilasas/farmacologíaRESUMEN
Diamond-like carbon (DLC) wear debris, which is often composed of different types of structures, is generated from DLC-modified artificial joints in the human body, and its biocompatibility evaluation is especially important to prevent wear-debris-induced implant failure. Here, RAW 264.7 macrophages (inflammatory-reaction assay) and primary mouse osteoblasts (osteoblastogenesis assay) were employed to investigate the toxicity of DLC wear particles (DWPs) by evaluation of cell viability and morphology, enzyme-linked immunosorbent assays, and quantitative reverse-transcription polymerase chain reaction (PCR). Relevant histopathological analysis of rat joints was also performed in vivo. We found that DWPs with a relatively high sp2/sp3 ratio (graphite-phase tendency) manifested a higher cytotoxicity and significant inhibition of osteoblastogenesis. DWPs with a relatively low sp2/sp3 ratio (diamond-phase tendency) showed good biocompatibility in vivo. The DWPs exhibiting a low sp2/sp3 ratio demonstrated reduced secretion of TNF-α and IL-6, along with increased secretion of TIMP-1, resulting in the downregulation of MMP-2 and MMP-9 and upregulation of interleukin-10 (IL-10), thereby attenuating the inflammatory response. Moreover, coculturing osteoblasts with DWPs exhibiting a low sp2/sp3 ratio resulted in an elevated OPG/RANKL ratio and increased expression of OPG mRNA. Because of the absence of electrostatic repulsion, DWPs with a relatively low sp2/sp3 ratio enhanced bovine serum albumin adsorption, which favored cellular activities. Cytotoxicity assessment of DWPs can help establish an evaluation system for particle-related joint disease and can facilitate the clinical application of DLC-coated prostheses.