RESUMEN
Osteoarthritis (OA) is a common aging-related disease affecting entire joint structures, encompassing articular cartilage and subchondral bone. Although senescence and dysfunction of chondrocytes are considered crucial factors in the occurrence of OA, the exact pathogenesis remains to be investigated. In our study, chondrocytes were incubated with a conditioned medium obtained from osteoclasts at different differentiation stages, suggesting that osteoclasts and osteoclast precursors suppressed anabolism and promoted the catabolism of chondrocytes in vitro. In contrast, the function of osteoclasts was more significant than osteoclast precursors. Further blocking of osteoclast exosome secretion by using GW4869 abolished the effect of osteoclasts on chondrocytes. Functionally, exosomal transfer of osteoclast-derived miR-212-3p inhibited Smad2 to mediate chondrocyte dysfunction, thus accelerating cartilage matrix degradation in OA via TGF-ß1/Smad2 signaling. The mechanism was also confirmed within the articular cartilage in OA patients and surgery-induced OA mice. Our study provides new information on intercellular interactions in the bone microenvironment within articular cartilage and subchondral bone during OA progression. The miR-212-3p/Smad2 axis is a potential target for the prevention and therapy of OA.
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Cartílago Articular , MicroARNs , Osteoartritis , Animales , Humanos , Ratones , Cartílago Articular/metabolismo , Condrocitos/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Osteoartritis/metabolismo , Osteoclastos/metabolismo , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
Osteoarthritis (OA) is a prevalent degenerative disease of the joint, featured by articular cartilage destruction and subchondral bone marrow lesions. Articular cartilage and subchondral bone constitute an osteochondral unit that guarantees joint homeostasis. During OA initiation, activated osteoclasts in subchondral bone ultimately result in impaired capacities of the subchondral bone in response to mechanical stress, followed by the degradation of overlying articular cartilage. Thus, targeting osteoclasts could be a potential therapeutic option for treating OA. Here, we observed that farnesoid X receptor (FXR) expression and osteoclast fusion and activity in subchondral bone were concomitantly changed during early-stage OA in the OA mouse model established by anterior cruciate ligament transection (ACLT). Then, we explored the therapeutic effects of FXR agonist GW4064 on the osteochondral pathologies in ACLT mice. We showed that GW4064 obviously ameliorated subchondral bone deterioration, associated with reduction in tartrate-resistant acid phosphatase (TRAP) positive multinuclear osteoclast number, as well as articular cartilage degradation, which were blocked by the treatment with FXR antagonist Guggulsterone. Mechanistically, GW4064 impeded osteoclastogenesis through inhibiting subchondral bone osteoclast fusion via suppressing c-Jun N-terminal kinase (JNK) 1/2/nuclear factor of activated T-cells 1 (NFATc1) pathway. Taken together, our results present evidence for the protective effects of GW4064 against OA by blunting osteoclast-mediated aberrant subchondral bone loss and subsequent cartilage deterioration. Therefore, GW4064 demonstrates the potential as an alternative therapeutic option against OA for further drug development.
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Resorción Ósea/prevención & control , Regulación de la Expresión Génica/efectos de los fármacos , Isoxazoles/farmacología , Osteoartritis/prevención & control , Osteoclastos/efectos de los fármacos , Osteogénesis , Proteínas de Unión al ARN/agonistas , Animales , Remodelación Ósea , Resorción Ósea/etiología , Resorción Ósea/metabolismo , Resorción Ósea/patología , Femenino , Ratones , Ratones Endogámicos C57BL , Proteína Quinasa 8 Activada por Mitógenos/genética , Proteína Quinasa 8 Activada por Mitógenos/metabolismo , Proteína Quinasa 9 Activada por Mitógenos/genética , Proteína Quinasa 9 Activada por Mitógenos/metabolismo , Factores de Transcripción NFATC/genética , Factores de Transcripción NFATC/metabolismo , Osteoartritis/etiología , Osteoartritis/metabolismo , Osteoartritis/patología , Osteoclastos/metabolismo , Osteoclastos/patologíaRESUMEN
The repair of bone defects, especially for the large segment of bone defects, has always been an urgent problem in orthopedic clinic and attracted researchers' attention. Nowadays, the application of tissue engineering bone in the repair of bone defects has become the research hotspot. With the rapid development of tissue engineering, the novel and functional scaffold materials for bone repair have emerged. In this review, we have summarized the multi-functional roles of osteoclasts in bone remodeling. The development of matrix-based tissue engineering bone has laid a theoretical foundation for further investigation about the novel bone regeneration materials which could perform high bioactivity. From the point of view on preserving pre-osteoclasts and targeting mature osteoclasts, this review introduced the novel matrix-based tissue engineering bone based on osteoclasts in the field of bone tissue engineering, which provides a potential direction for the development of novel scaffold materials for the treatment of bone defects.
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Osteoclastos , Ingeniería de Tejidos , Regeneración Ósea , Huesos , HumanosRESUMEN
Osteoarthritis (OA) is a degenerative joint disease in the elderly. Although OA has been considered as primarily a disease of the articular cartilage, the participation of subchondral bone in the pathogenesis of OA has attracted increasing attention. This review summarises the microstructural and histopathological changes in subchondral bone during OA progression that are due, at the cellular level, to changes in the interactions among osteocytes, osteoblasts, osteoclasts (OCs), endothelial cells and sensory neurons. Therefore, we focus on how pathological cellular interactions in the subchondral bone microenvironment promote subchondral bone destruction at different stages of OA progression. In addition, the limited amount of research on the communication between OCs in subchondral bone and chondrocytes (CCs) in articular cartilage during OA progression is reviewed. We propose the concept of 'OC-CC crosstalk' and describe the various pathways by which the two cell types might interact. Based on the 'OC-CC crosstalk', we elaborate potential therapeutic strategies for the treatment of OA, including restoring abnormal subchondral bone remodelling and blocking the bridge-subchondral type H vessels. Finally, the review summarises the current understanding of how the subchondral bone microenvironment is related to OA pain and describes potential interventions to reduce OA pain by targeting the subchondral bone microenvironment.
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Cartílago Articular , Osteoartritis , Anciano , Huesos/metabolismo , Cartílago Articular/metabolismo , Células Endoteliales/metabolismo , Humanos , Osteoartritis/patología , DolorRESUMEN
Targeted lipid nanobubbles as theranostic ultrasound molecular probes with both targeted contrast-enhanced ultrasound molecular imaging and synergistic treatment capabilities are expected to overcome severe challenges in the diagnosis and treatment of refractory triple-negative breast cancer (TNBC). In this study, AS1411 aptamer-functionalised nucleolin-targeted doxorubicin-loaded lipid nanobubbles (AS1411-DOX-NBs) were constructed, and their physicochemical properties as well as anti-tumour and cardioprotective efficacies were systematically tested and evaluated. The results showed that AS1411-DOX-NBs can carry and maintain the physicochemical and pharmacodynamic properties of doxorubicin (DOX) and show stronger tumour cell-killing abilityin vitroby increasing the active uptake of drugs. AS1411-DOX-NBs also significantly inhibited the growth of TNBC xenografts while maintaining the weight and health of the mice. Echocardiography and pathological examination further confirmed that AS1411-DOX-NBs effectively caused tumour tissue apoptosis and necrosis while reducing DOX-induced cardiotoxicity. The AS1411-DOX-NBs constructed in this study enable both targeted contrast-enhanced ultrasound molecular imaging and synergistic therapeutic efficacy and can be used as safe and efficient theranostic ultrasound molecular probes for the diagnosis and treatment of TNBC.
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Aptámeros de Nucleótidos/administración & dosificación , Cardiotónicos/administración & dosificación , Doxorrubicina/administración & dosificación , Oligodesoxirribonucleótidos/administración & dosificación , Fosfoproteínas/metabolismo , Proteínas de Unión al ARN/metabolismo , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Animales , Aptámeros de Nucleótidos/química , Aptámeros de Nucleótidos/farmacología , Cardiotónicos/química , Cardiotónicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Doxorrubicina/efectos adversos , Doxorrubicina/química , Ecocardiografía , Femenino , Humanos , Liposomas , Ratones , Nanopartículas , Nanoestructuras , Oligodesoxirribonucleótidos/química , Oligodesoxirribonucleótidos/farmacología , Resultado del Tratamiento , Neoplasias de la Mama Triple Negativas/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto , NucleolinaRESUMEN
Cancer metastasis is a unique feature of malignant tumours. Even bone can become a common colonization site due to the tendency of solid tumours, including breast cancer (BCa) and prostate cancer (PCa), to metastasize to bone. Currently, a previous concept in tumour metabolism called tumour dormancy may be a promising target for antitumour treatment. When disseminated tumour cells (DTCs) metastasize to the bone microenvironment, they form a flexible regulatory network called the "bone-tumour-inflammation network". In this network, bone turnover as well as metabolism, tumour progression, angiogenesis and inflammatory responses are highly unified and coordinated, and a slight shift in this balance can result in the disruption of the microenvironment, uncontrolled inflammatory responses and excessive tumour growth. The purpose of this review is to highlight the regulatory effect of the "bone-tumour-inflammation network" in tumour dormancy. Osteoblast-secreted factors, bone turnover and macrophages are emphasized and occupy in the main part of the review. In addition, the prospective clinical application of tumour dormancy is also discussed, which shows the direction of future research.
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Neoplasias Óseas/metabolismo , Neoplasias de la Mama/metabolismo , Inflamación/metabolismo , Neoplasias de la Próstata/metabolismo , Neoplasias Óseas/genética , Neoplasias Óseas/patología , Neoplasias Óseas/secundario , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Femenino , Humanos , Inflamación/genética , Inflamación/patología , Masculino , Metástasis de la Neoplasia , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patología , Osteoblastos/metabolismo , Osteoblastos/patología , Neoplasias de la Próstata/patología , Microambiente Tumoral/genéticaRESUMEN
The invasion of osteoclasts into the cartilage via blood vessels advances the process of endochondral ossification, and dysregulation of dynamic intercellular interactions results in skeletal dysplasias. Although the regulation of osteoclasts by growth plate chondrocytes has been reported in detail, the effect of osteoclasts on chondrocytes remains to be determined. In this study, ATDC5 cells and bone marrow mesenchymal stem cells were differentiated into chondrocytes and treated with conditioned medium obtained from bone marrow macrophages differentiated to osteoclast precursors and osteoclasts. Exosomes were inhibited in conditioned medium or were isolated directly from osteoclasts to further determine whether osteoclast-derived exosomes play an important role in chondrocyte hypertrophy. Additionally, exosomal miRNAs were detected, and let-7a-5p was selected as an miRNA with significantly increased expression in osteoclast-derived exosomes. Experiments were performed to verify the potential target Smad2 and investigate how let-7a-5p affected chondrocytes. The results suggest that both osteoclast precursors and osteoclasts promote chondrocyte hypertrophy and that the promotive effect of osteoclasts is more significant than that of osteoclast precursors. Osteoclast-derived exosomes promote the hypertrophic differentiation of chondrocytes. Moreover, osteoclast-derived exosomal let-7a-5p inhibits Smad2 to decrease the transforming growth factor-ß-induced inhibition of chondrocyte hypertrophy. Our research reveals the role of osteoclasts in the regulation of chondrocytes and provides insights into the highly coordinated intercellular process of endochondral ossification.
RESUMEN
Sphingosine-1-phosphate (S1P) is a natural bioactive lipid molecule and a common first or second messenger in the cardiovascular and immune systems. By binding with its receptors, S1P can serve as mediator of signalling during cell migration, differentiation, proliferation and apoptosis. Although the predominant role of S1P in bone regeneration has been noted in many studies, this role is not as well-known as its roles in the cardiovascular and immune systems. In this review, we summarize previous research on the role of S1P receptors (S1PRs) in osteoblasts and osteoclasts. In addition, S1P is regarded as a bridge between bone resorption and formation, which brings hope to patients with bone-related diseases. Finally, we discuss S1P and its receptors as therapeutic targets for treating osteoporosis, inflammatory osteolysis and bone metastasis based on the biological effects of S1P in osteoclastic/osteoblastic cells, immune cells and tumour cells.
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Neoplasias Óseas/genética , Resorción Ósea/genética , Lisofosfolípidos/genética , Receptores de Esfingosina-1-Fosfato/genética , Esfingosina/análogos & derivados , Neoplasias Óseas/patología , Neoplasias Óseas/secundario , Resorción Ósea/patología , Humanos , Metástasis de la Neoplasia , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteólisis/genética , Osteólisis/patología , Osteoporosis/genética , Osteoporosis/patología , Esfingosina/genética , Receptores de Esfingosina-1-Fosfato/metabolismoRESUMEN
In bone remodeling, after a lifespan of â¼2 weeks, osteoclasts undergo apoptosis in each bone turnover cycle, resulting in generation of a large number of apoptotic bodies (ABs). However, the biological roles of osteoclast-derived ABs (OC-ABs) in bone remodeling have not been investigated and remain unknown. In this study, we stimulated bone marrow macrophages with receptor activator of NF-κB ligand (RANKL) to obtain both preosteoclasts and mature osteoclasts (mOCs). We then used alendronate to induce apoptosis in preosteoclasts and mOCs and generate the respective ABs and used flow cytometry and immunoblotting to characterize the sizes and immunogenic characteristics of the extracted ABs. We show that mOC-ABs are engulfed by preosteoblastic MC3T3-E1 cells and promote the viability of these cells. Among all osteoclast-derived extracellular vesicles, mOC-ABs had the highest osteogenic potency. We further observed that mOC-ABs had the highest vesicular receptor activator of NF-κB (RANK) levels among all types of osteoclast-derived extracellular vesicles. Of note, masking of vesicular RANK by soluble RANKL strongly abolished the osteogenic potency of osteoclast-derived ABs. Mechanistically, we found that mOC-ABs induce osteoblast differentiation by activatingPI3K/AKT/mechanistic target of rapamycin (mTOR)/ribosomal protein S6 kinase signaling. In conclusion, OC-ABs promote osteogenic differentiation by stimulating osteoblast differentiation via activation of RANKL reverse signaling. These findings provide important insights into the reversal phase between the bone resorption and formation stages during bone remodeling and identify an AB-dependent cellular signaling mechanism in osteoclast-osteoblast coupling.
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Diferenciación Celular , Vesículas Extracelulares/metabolismo , Osteoclastos/metabolismo , Ligando RANK/metabolismo , Células 3T3 , Animales , Células de la Médula Ósea/citología , Remodelación Ósea , Macrófagos/metabolismo , Ratones , OsteogénesisRESUMEN
Central ischemic necrosis is one of the biggest obstacles in the clinical application of traditional tissue-engineered bone (TEB) in critical-sized bone defect regeneration. Because of its ability to promote vascular invasion, endochondral ossification-based TEB has been applied for bone defect regeneration. However, inadequate chondrocyte hypertrophy can hinder vascular invasion and matrix mineralization during endochondral ossification. In light of recent studies suggesting that ceria nanoparticles (CNPs) improve the blood vessel distribution within TEB, we modified TEB scaffold surfaces with CNPs and investigated the effect and mechanism of CNPs on endochondral ossification-based bone regeneration. The CNPs used in this study were synthesized by the microemulsion method and modified with alendronate-anchored polyethylene glycol 600. We showed that CNPs accelerated new bone formation and enhanced endochondral ossification-based bone regeneration in both a subcutaneous ectopic osteogenesis model and a mouse model of critical-sized bone defects. Mechanistically, CNPs significantly promoted endochondral ossification-based bone regeneration by ensuring sufficient hypertrophic differentiation via the activation of the RNA helicase, DEAH (Asp-Glu-Ala-His) box helicase 15, and its downstream target, p38 MAPK. These results suggested that CNPs could be applied as a biomaterial to improve the efficacy of endochondral ossification-based bone regeneration in critical-sized bone defects.-Li, J., Kang, F., Gong, X., Bai, Y., Dai, J., Zhao, C., Dou, C., Cao, Z., Liang, M., Dong, R., Jiang, H., Yang, X., Dong, S. Ceria nanoparticles enhance endochondral ossification-based critical-sized bone defect regeneration by promoting the hypertrophic differentiation of BMSCs via DHX15 activation.
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Células de la Médula Ósea/metabolismo , Regeneración Ósea/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Cerio , Fémur , Células Madre Mesenquimatosas/metabolismo , Nanopartículas/química , Osteogénesis/efectos de los fármacos , ARN Helicasas/metabolismo , Animales , Células de la Médula Ósea/patología , Cerio/química , Cerio/farmacología , Fémur/lesiones , Fémur/metabolismo , Fémur/patología , Células Madre Mesenquimatosas/patología , Ratones , Ratones Endogámicos BALB C , Ratones DesnudosRESUMEN
PURPOSE: To construct aptamer AS1411-functionalized targeted lipid nanobubbles that could simultaneously target abnormally highly expressed nucleolin (NCL) on tumor tissue and neovasculature. Additionally, the study of their contrast-enhanced ultrasound molecular imaging capabilities in vitro and in vivo to explore new methods and approaches for the early and accurate diagnosis of triple-negative breast cancer (TNBC). METHODS: First, the targeted lipid-nucleic acid molecules were constructed by an amide reaction. Then, the targeted lipid nanobubbles (AS1411-NBs) and nontargeted lipid nanobubbles (NBs) were prepared by membrane hydration, mechanical vibration and centrifugal floatation. The physicochemical characteristics and contrast-enhanced ultrasound imaging capabilities of AS1411-NBs and NBs were compared and analyzed in vitro and in vivo. RESULTS: There were no significant differences between the AS1411-NBs and NBs in their concentration, average particle size or ultrasound imaging capabilities in vitro (P > 0.05). However, AS1411-NBs could simultaneously target NCL in tumor tissue and neovasculature to effectively prolong the duration of contrast-enhanced ultrasound imaging compared to NBs in vivo. The area under the time-intensity curve was significantly different between AS1411-NBs and NBs (P < 0.001), and the drug loading capacity of the AS1411-NBs was also significantly higher than that of the NBs (P < 0.05). CONCLUSIONS: Aptamer AS1411-functionalized targeted lipid nanobubbles could significantly prolong the duration of contrast-enhanced ultrasound imaging to achieve dual-targeted ultrasound molecular imaging of tumor tissue and neovasculature. AS1411-NBs also have higher drug loading and targeted drug delivery capabilities compared with NBs, which can provide new methods and approaches for the early accurate diagnosis and effective treatment of TNBC.
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Neoplasias de la Mama/diagnóstico por imagen , Medios de Contraste/química , Lípidos/química , Microburbujas , Fosfoproteínas/efectos de los fármacos , Proteínas de Unión al ARN/efectos de los fármacos , Neoplasias de la Mama Triple Negativas/diagnóstico por imagen , Animales , Línea Celular Tumoral , Sistemas de Liberación de Medicamentos , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Imagen Molecular/métodos , Neovascularización Patológica/diagnóstico por imagen , Tamaño de la Partícula , Ultrasonografía , Ensayos Antitumor por Modelo de Xenoinjerto , NucleolinaRESUMEN
Excessive osteoclast formation is one of the important pathological features of inflammatory bone destruction. Interleukin-37 (IL-37) is an anti-inflammatory agent that is present throughout the body, but it displays low physiological retention. In our study, high levels of the IL-37 protein were detected in clinical specimens from patients with bone infections. However, the impact of IL-37 on osteoclast formation remains unclear. Next, IL-37 alleviated the inflammatory bone destruction in the mouse in vivo. We used receptor activator of nuclear factor-κB ligand and lipopolysaccharide to trigger osteoclastogenesis under physiological and pathological conditions to observe the role of IL-37 in this process and explore the potential mechanism of this phenomenon. In both induction models, IL-37 exerted inhibitory effects on osteoclast differentiation and bone resorption. Furthermore, IL-37 decreased the phosphorylation of inhibitor of κBα and p65 and the expression of nuclear factor of activated T cells c1, while the dimerization inhibitor of myeloid differentiation factor 88 reversed the effects. These data provide evidence that IL-37 modulates osteoclastogenesis and a theoretical basis for the clinical application of IL-37 as a treatment for bone loss-related diseases.
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Resorción Ósea/metabolismo , Huesos/metabolismo , Inflamación/metabolismo , Interleucina-1/metabolismo , Osteoclastos/metabolismo , Osteogénesis/fisiología , Animales , Diferenciación Celular/fisiología , Línea Celular , Humanos , Inflamación/inducido químicamente , Ligandos , Lipopolisacáridos/farmacología , Ratones , Factor 88 de Diferenciación Mieloide/metabolismo , FN-kappa B/metabolismo , Células RAW 264.7 , Transducción de Señal/fisiologíaRESUMEN
The movements of life at every level from organs, tissues, cells to sub-cells, are all conducted in certain physical environments. In the human body, skeletal tissue among all connective tissues is influenced the most by physical forces. Studying the biological behavior of bone cells under different physical environments is helpful in further understanding bone homeostasis and metabolism. Among all bone cells, osteoclast (OC) and OC steered bone remodeling is one of the key points in bone metabolism. In the past few decades, people's understanding of OC was mostly limited to its involvement of bone resorption under physiological and pathological conditions. However, more and more studies started to focus on how physical forces affect the formation and differentiation of OC. This review tries to illustrate the knowledge up to date about how osteoclastogenesis is regulated by physical forces through direct and indirect ways, including fluid shear force, compressive force, and microgravity. The direct way describes the straightforward effects produced by different forces in osteoclastogenesis, whereas the indirect way describes the effects of different forces in osteoclastogenesis through regulation of other bone cells when a certain force is applied. Molecular mechanisms were analyzed and reviewed in both direct and indirect regulation by different forces. Finally, we discussed the status quo and tendency of related research, as well as other unresolved issues, and some future prospects.
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Adaptación Fisiológica , Huesos/metabolismo , Osteogénesis/fisiología , Fenómenos Biomecánicos , HumanosRESUMEN
Osteoclasts derived from the monocyte/macrophage hematopoietic lineage regulate bone resorption, a process balanced by bone formation in the continual renewal of the skeletal system. As dysfunctions of these cells result in bone metabolic diseases such as osteoporosis and osteopetrosis, the exploration of the mechanisms regulating their differentiation is a priority. A potential mechanism may involve long noncoding RNAs (lncRNAs), which are known to regulate various cell biology activities, including proliferation, differentiation, and apoptosis. The expression of the lncRNA AK077216 (Lnc-AK077216) is significantly upregulated during osteoclastogenesis identified by microarray and verified by qPCR. Up- and downregulation of Lnc-AK077216, respectively promotes and inhibits osteoclast differentiation, bone resorption, and the expression of related genes on the basis of tartrate-resistant acid phosphatase staining, qPCR, and western blot results. In addition, Lnc-AK077216 suppresses NIP45 expression and promotes the expression of NFATc1, an essential transcription factor during osteoclastogenesis. Besides, it was found that the expression of Lnc-AK077216 and Nfatc1 is upregulated, whereas Nip45 expression is downregulated in bone marrow and spleen tissues of ovariectomized mice. The results suggest that Lnc-AK077216 regulates NFATc1 expression and promotes osteoclast formation and function, providing a novel mechanism of osteoclastogenesis and a potential biomarker or a new drug target for osteoporosis.
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Resorción Ósea , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Macrófagos/efectos de los fármacos , Factores de Transcripción NFATC/metabolismo , Osteoclastos/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Osteoporosis Posmenopáusica/enzimología , Ligando RANK/farmacología , ARN Largo no Codificante/metabolismo , Animales , Apoptosis/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Macrófagos/enzimología , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Factores de Transcripción NFATC/genética , Osteoclastos/enzimología , Osteoclastos/patología , Osteoporosis Posmenopáusica/genética , Osteoporosis Posmenopáusica/patología , Ovariectomía , Células RAW 264.7 , ARN Largo no Codificante/genética , Transducción de SeñalRESUMEN
OBJECTIVE: Chondrocyte hypertrophy, a terminal stage of chondrocyte differentiation, is essential to the endochondral bone formation and is one of the major pathological factors in osteoarthritis. This study investigated the role of microRNA-29b (miR-29b), which is involved in chondrogenesis, in the regulation of hypertrophy in chondrocytes. METHODS: miR-29b expression was assessed during murine mesenchymal stem cells (mMSCs) chondrogenesis. To detect whether miR-29b affects chondrocyte hypertrophy, the mMSCs induced toward chondrogenesis were transfected with miR-29b or its antisense inhibitor (antagomiR-29b). Finally, the differential effects of antagomiR-29b on chondrocytes at different differentiation stages were evaluated by loss-of-function experiments. RESULTS: miR-29b expression was low-level during the early chondrogenic differentiation, however, it was changed to high level during hypertrophy. Subsequently, the gain-of-function and loss-of-function experiments had confirmed that miR-29b promoted hypertrophy in mMSC-derived chondrocytes. In addition, we confirmed that on day 7, when cells were treated with antagomiR-29b, was the optimal intervention time for preventing hypertrophic phenotype of mMSCs in vitro. CONCLUSION: miR-29b regulated chondrogenesis homeostasis and enhance hypertrophic phenotype. These data suggest that miR-29b is a key regulator of the chondrocyte phenotype derived from mMSCs and it might be a potential target for articular cartilage repair.
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Tissue-engineered constructs (TECs) hold great promise for treating large bone defects. Incorporated mesenchymal stem cells (MSCs) can facilitate the vascularization of TECs. Nevertheless, the underlying mechanism remains ambiguous. Here we analyzed the roles of C-X-C chemokine receptor 2 (CXCR2) and its downstream signal pathways in MSC-induced endothelial progenitor cell (EPC) migration. Transwell assays and immunofluorescence staining were performed for cell migration analysis in vitro and in vivo, respectively. A series of signal inhibitors and short hairpin RNA was used for screening essential signaling molecules. We found that blockade of CXCR2 abolished the migration of EPCs toward MSCs as well as subsequent vascularization and bone repair in TECs. Moreover, screening results suggested that steroid receptor coactivator (Src) acted as a predominant downstream effector of CXCR2. Further molecular biologic and histomorphological experiments revealed that the action of Src required the phosphorylation of ras-related C3 botulinum toxin substrate 1 (Rac1), which was pivotal for the development of lamellipodia and filopodia. The phosphorylation and colocalization of paxillin kinase linker (PKL) and vav guanine nucleotide exchange factor 2 (Vav2) were essential for the activation of Rac1. Therefore, we demonstrated that MSCs promoted EPC migration via activating CXCR2 and its downstream Src-PKL/Vav2-Rac1 signaling pathway. These findings unveiled the molecular mechanism in the vascularization of TECs and were expected to provide novel targets for efficacy improvement.-Li, Z., Yang, A., Yin, X., Dong, S., Luo, F., Dou, C., Lan, X., Xie, Z., Hou, T., Xu, J., Xing, J. Mesenchymal stem cells promote endothelial progenitor cell migration, vascularization, and bone repair in tissue-engineered constructs via activating CXCR2-Src-PKL/Vav2-Rac1.
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Regeneración Ósea , Movimiento Celular , Células Progenitoras Endoteliales/metabolismo , Células Madre Mesenquimatosas/metabolismo , Transducción de Señal , Ingeniería de Tejidos/métodos , Animales , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Células Progenitoras Endoteliales/citología , Células Progenitoras Endoteliales/fisiología , Proteínas Activadoras de GTPasa , Humanos , Péptidos y Proteínas de Señalización Intercelular , Ratones , Neovascularización Fisiológica , Coactivadores de Receptor Nuclear/metabolismo , Fosfoproteínas/metabolismo , Proteínas Proto-Oncogénicas c-vav/metabolismo , Receptores de Interleucina-8B/metabolismo , Proteína de Unión al GTP rac1/metabolismoRESUMEN
Endochondral ossification is crucial for bone formation in both adult bone repair process and embryo long-bone development. In endochondral ossification, bone marrow-derived mesenchymal stem cells (BMSCs) first differentiate to chondrocytes, then BMSC-derived chondrocytes endure a hypertrophic process to generate new bone. Endochondral ossification-based bone repair is a promising strategy to cure massive bone defect, which is a major clinical issue in orthopedics. However, challenges still remain for this novel strategy. One challenge is to ensure the sufficient hypertrophic differentiation. Another is to maintain the survival of the above hypertrophic chondrocytes under the hypoxic environment of massive bone defect. To solve this issue, mangiferin (MAG) was introduced to endochondral ossification-based bone repair. In this report, we proved MAG to be a novel autophagy inducer, which promoted BMSC-derived hypertrophic chondrocyte survival against hypoxia-induced injury through inducing autophagy. Furthermore, MAG enhances hypertrophic differentiation of BMSC-derived chondrocytes via upregulating key hypertrophic markers. Mechanistically, MAG induced autophagy in BMSC-derived chondrocytes by promoting AMPKα phosphorylation. Additionally, MAG balanced the expression of sex-determining region Y-box 9 and runt-related transcription factor 2 to facilitate hypertrophic differentiation. These results indicated that MAG was a potential drug to improve the efficacy of endochondral ossification-based bone repair in massive bone defects.-Bai, Y., Liu, C., Fu, L., Gong, X., Dou, C., Cao, Z., Quan, H., Li, J., Kang, F., Dai, J., Zhao, C., Dong, S. Mangiferin enhances endochondral ossification-based bone repair in massive bone defect by inducing autophagy through activating AMP-activated protein kinase signaling pathway.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Autofagia/efectos de los fármacos , Huesos/diagnóstico por imagen , Osteogénesis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Xantonas/farmacología , Animales , Huesos/metabolismo , Diferenciación Celular/efectos de los fármacos , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Condrogénesis/efectos de los fármacos , Femenino , Masculino , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Ratones Endogámicos BALB C , Fosforilación/efectos de los fármacosRESUMEN
Cartilage is a kind of special connective tissue which does not contain neither blood vessels nor lymphatics and nerves. Therefore, the damage in cartilage is difficult to be repaired spontaneously. Constructing tissue engineered cartilage provides a new technique for cartilage repairing. Mesenchymal stem cells (MSCs) possess a unique capability of self-renew and can differentiate into pre-chondrocytes which are frequently applied as seed cells in tissue engineering. However, in regenerated cartilage the chondrocytes derived from MSCs can hardly maintain homeostasis and preferentially present hypertrophic like phenotype. We investigated the effects of cyanidin, a natural organic compound, on chondrogenic and subsequent hypertrophic differentiation of MSCs in order to seek approaches to inhibit chondrocyte hypertrophy. We evaluated the effects of cyanidin on expression of chondrogenic and hypertrophic marker genes through RT-PCR, Western blot, alcian blue staining, and immunocytochemistry. The results showed that both chondrogenic related genes Sox9, Col2a1, and hypertrophic marker genes Runx2, Col10a1 were inhibited by cyanidin. In addition, we found that cyanidin promoted Nrf2 and p62 expression and suppressed LC3B expression during chondrogenic stage of MSCs. Meanwhile phosphorylation of IκBα and autophagosome related protein LC3B were inactivated by cyanidin during chondrocyte hypertrophic stage. Furthermore, rapamycin, an autophagy activator, abrogated the inhibitory effect of cyanidin on chondrogenic, and hypertrophic differentiation of MSCs. In conclusion, one potential mechanism of cyanidin, by which the chondrogenic and hypertrophic differentiation of MSCs were inhibited, was due to decreased autophagy activity. Our results indicated that cyanidin was a potential therapeutic agent for keeping mature chondrocyte functions.
Asunto(s)
Antocianinas/farmacología , Autofagia/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Condrocitos/efectos de los fármacos , Condrogénesis/efectos de los fármacos , Células Madre Mesenquimatosas/efectos de los fármacos , Animales , Autofagosomas/efectos de los fármacos , Autofagosomas/metabolismo , Autofagosomas/patología , Línea Celular , Condrocitos/metabolismo , Condrocitos/patología , Colágeno Tipo II/genética , Colágeno Tipo II/metabolismo , Colágeno Tipo X/genética , Colágeno Tipo X/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Relación Dosis-Respuesta a Droga , Regulación de la Expresión Génica , Glicosaminoglicanos/metabolismo , Hipertrofia , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/patología , Ratones Endogámicos C3H , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Inhibidor NF-kappaB alfa/metabolismo , Fosforilación , Factor de Transcripción SOX9/genética , Factor de Transcripción SOX9/metabolismo , Proteína Sequestosoma-1/genética , Proteína Sequestosoma-1/metabolismo , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología , Factores de TiempoRESUMEN
Osteoporosis is a metabolic disease characterized by osteopenia and bone microstructural deterioration. Osteoclasts are the primary effector cells that degrade bone matrix and their abnormal function leads to the development of osteoporosis. Reactive oxygen species (ROS) accumulation during cellular metabolism promotes osteoclast proliferation and differentiation, therefore, playing an important role in osteoporosis. Cistanche deserticola polysaccharide (CDP) possesses antitumor, anti-inflammatory, and antioxidant activity. However, the impact of CDP on osteoclasts is unclear. In this study, tartrate-resistant acid phosphatase staining, immunofluorescence, reverse transcription-polymerase chain reaction, and western blot analysis were utilized to demonstrate that CDP inhibited osteoclastogenesis and hydroxyapatite resorption. In addition, CDP also inhibited the expression of osteoclast maker genes including Ctsk, Mmp9, and Acp5 and had no effect on receptor activator of nuclear factor κB (RANK) expression. Mechanistic analyses revealed that CDP increases the expression of antioxidant enzymes to attenuate RANKL-mediated ROS production in osteoclasts and inhibits nuclear factor of activated T cells and mitogen-activated protein kinase activation. These results suggest that CDP may represent a candidate drug for the treatment of osteoporosis caused by excessive osteoclast activity.
Asunto(s)
Resorción Ósea/tratamiento farmacológico , Cistanche/química , Osteoporosis/tratamiento farmacológico , Polisacáridos/farmacología , Ligando RANK/genética , Resorción Ósea/genética , Resorción Ósea/patología , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Proteína Quinasa 1 Activada por Mitógenos/genética , Osteoclastos/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Osteoporosis/genética , Osteoporosis/patología , Polisacáridos/química , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Linfocitos T/efectos de los fármacosRESUMEN
The extracellular matrix (ECM) contains rich biological cues for cell recruitment, proliferationm, and even differentiation. The osteoinductive potential of scaffolds could be enhanced through human bone marrow mesenchymal stem cell (hBMSC) directly depositing ECM on surface of scaffolds. However, the role and mechanism of human umbilical cord mesenchymal stem cells (hUCMSC)-secreted ECM in bone formation remain unknown. We tested the osteoinductive properties of a hUCMSC-secreted ECM construct (hUCMSC-ECM) in a large femur defect of a severe combined immunodeficiency (SCID) mouse model. The hUCMSC-ECM improved the colonization of endogenous MSCs and bone regeneration, similar to the hUCMSC-seeded scaffold and superior to the scaffold substrate. Besides, the hUCMSC-ECM enhanced the promigratory molecular expressions of the homing cells, including CCR2 and TßRI. Furthermore, the hUCMSC-ECM increased the number of migrated MSCs by nearly 3.3 ± 0.1-fold, relative to the scaffold substrate. As the most abundant cytokine deposited in the hUCMSC-ECM, insulin-like growth factor binding protein 3 (IGFBP3) promoted hBMSC migration in the TßRI/II- and CCR2-dependent mechanisms. The hUCMSC-ECM integrating shRNA-mediated silencing of Igfbp3 that down-regulated IGFBP3 expression by approximately 60%, reduced the number of migrated hBMSCs by 47%. In vivo, the hUCMSC-ECM recruited 10-fold more endogenous MSCs to initiate bone formation compared to the scaffold substrate. The knock-down of Igfbp3 in the hUCMSC-ECM inhibited nearly 60% of MSC homing and bone regeneration capacity. This research demonstrates that IGFBP3 is an important MSC homing molecule and the therapeutic potential of hUCMSC-ECM in bone regeneration is enhanced by improving MSC homing in an IGFBP3-dependent mechanism.