RESUMEN
Background: Periprosthetic osteolysis is a prevalent complication following total ankle arthroplasty (TAA), implicating various cytokines in osteoclastogenesis as pivotal in this process. This study aimed to evaluate the relationship between osteolysis and the concentrations of osteoclastogenesis-related cytokines in synovial fluid and investigate its clinical value following TAA. Methods: Synovial fluid samples from 23 ankles that underwent revision surgery for osteolysis following TAA were analyzed as the osteolysis group. As a control group, we included synovial fluid samples obtained from 23 ankles during primary TAA for osteoarthritis. The receptor activator of nuclear factor-κB ligand (RANKL)/osteoprotegerin (OPG) ratio in these samples was quantified using sandwich enzyme-linked immunosorbent assay techniques, and a bead-based multiplex immunoassay facilitated the detection of specific osteoclastogenesis-related cytokines. Results: RANKL levels averaged 487.9 pg/mL in 14 of 23 patients in the osteolysis group, with no detection in the control group's synovial fluid. Conversely, a significant reduction in OPG levels was observed in the osteolysis group (p = 0.002), resulting in a markedly higher mean RANKL/OPG ratio (0.23) relative to controls (p = 0.020). Moreover, the osteolysis group had increased concentrations of various osteoclastogenesis-related cytokines (tumor necrosis factor-α, interleukin [IL]-1ß, IL-6, IL-8, IP-10, and monocyte chemotactic protein-1) in the synovial fluid relative to the control group. Conclusions: Our results demonstrated that periprosthetic osteolysis was associated with osteoclastogenesis activation through an elevated RANKL/OPG ratio following TAA. We assume that RANKL and other osteoclastogenesis-related cytokines in the synovial fluid have clinical value as a potential marker for the development and progression of osteolysis following TAA.
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Artroplastia de Reemplazo de Tobillo , Biomarcadores , Osteólisis , Osteoprotegerina , Ligando RANK , Líquido Sinovial , Humanos , Líquido Sinovial/metabolismo , Líquido Sinovial/química , Osteólisis/metabolismo , Osteólisis/etiología , Masculino , Femenino , Ligando RANK/metabolismo , Anciano , Persona de Mediana Edad , Artroplastia de Reemplazo de Tobillo/efectos adversos , Osteoprotegerina/metabolismo , Osteoprotegerina/análisis , Biomarcadores/metabolismo , Biomarcadores/análisis , Anciano de 80 o más Años , Citocinas/metabolismo , Citocinas/análisis , ReoperaciónRESUMEN
OBJECTIVE AND DESIGN: This observational study investigated the regulatory mechanism of Pim-1 in inflammatory signaling pathways. MATERIALS: THP-1, RAW 264.7, BV2, and Jurkat human T cell lines were used. TREATMENT: None. METHODS: Lipopolysaccharide (LPS) was used to induce inflammation, followed by PIM1 knockdown. Western blot, immunoprecipitation, immunofluorescence, and RT-PCR assays were used to assess the effect of PIM1 knockdown on LPS-induced inflammation. RESULTS: PIM1 knockdown in macrophage-like THP-1 cells suppressed LPS-induced upregulation of pro-inflammatory cytokines, inducible nitric oxide synthase, cyclooxygenase-2, phosphorylated Janus kinase, signal transducer and activator of transcription 3, extracellular signal-regulated kinase, c-Jun N-terminal kinase, p38, and nuclear factor kappa B p65 (NF-κB p65). It also suppressed upregulation of inhibitor of NF-κB kinase α/ß and enhanced the nuclear translocation of NF-κB p65. Moreover, it inhibited the upregulation of Nod-like receptor family pyrin domain-containing 3 (NLRP3) and cleavage of caspase-1 induced by co-treatment of LPS with adenosine triphosphate. Additionally, p-transforming growth factor-ß-activated kinase 1 (TAK1) interacted with Pim-1. All three members of Pim kinases (Pim-1, Pim-2, and Pim-3) were required for LPS-mediated inflammation in macrophages; however, unlike Pim-1 and Pim-3, Pim-2 functioned as a negative regulator of T cell activity. CONCLUSIONS: Pim-1 interacts with TAK1 in LPS-induced inflammatory responses and is involved in MAPK/NF-κB/NLRP3 signaling pathways. Additionally, considering the negative regulatory role of Pim-2 in T cells, further in-depth studies on their respective functions are needed.
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Several members of the transforming growth factor beta (TGF-ß) superfamily regulate the proliferation, differentiation, and function of bone-forming osteoblasts and bone-resorbing osteoclasts. However, it is still unknown whether Nodal, a member of the TGF-ß superfamily, serves a function in bone cells. In this study, we found that Nodal did not have any function in osteoblasts but instead negatively regulated osteoclast differentiation. Nodal inhibited RANKL-induced osteoclast differentiation by downregulating the expression of pro-osteoclastogenic genes, including c-fos, Nfatc1, and Blimp1, and upregulating the expression of antiosteoclastogenic genes, including Bcl6 and Irf8. Nodal activated STAT1 in osteoclast precursor cells, and STAT1 downregulation significantly reduced the inhibitory effect of Nodal on osteoclast differentiation. These findings indicate that Nodal activates STAT1 to downregulate or upregulate the expression of pro-osteoclastogenic or antiosteoclastogenic genes, respectively, leading to the inhibition of osteoclast differentiation. Moreover, the inhibitory effect of Nodal on osteoclast differentiation contributed to the reduction of RANKL-induced bone loss in vivo.
Asunto(s)
Diferenciación Celular , Proteína Nodal , Osteoclastos , Factor de Transcripción STAT1 , Animales , Ratones , Resorción Ósea/metabolismo , Resorción Ósea/genética , Resorción Ósea/patología , Factores Reguladores del Interferón/metabolismo , Factores Reguladores del Interferón/genética , Factores de Transcripción NFATC/metabolismo , Factores de Transcripción NFATC/genética , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteogénesis/genética , Fosforilación , Factor 1 de Unión al Dominio 1 de Regulación Positiva/metabolismo , Factor 1 de Unión al Dominio 1 de Regulación Positiva/genética , Proteínas Proto-Oncogénicas c-bcl-6/metabolismo , Proteínas Proto-Oncogénicas c-bcl-6/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-fos/genética , Ligando RANK/metabolismo , Transducción de Señal , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT1/genética , Masculino , Ratones Endogámicos ICR , Proteína Nodal/genética , Proteína Nodal/metabolismo , Proteína Nodal/farmacologíaRESUMEN
Human monocyte chemoattractant protein-1 (MCP-1) in mice has two orthologs, MCP-1 and MCP-5. MCP-1, which is highly expressed in osteoclasts rather than in osteoclast precursor cells, is an important factor in osteoclast differentiation. However, the roles of MCP-5 in osteoclasts are completely unknown. In this study, contrary to MCP-1, MCP-5 was downregulated during receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclast differentiation and was considered an inhibitory factor in osteoclast differentiation. The inhibitory role of MCP-5 in osteoclast differentiation was closely related to the increase in Ccr5 expression and the inhibition of IκB degradation by RANKL. Transgenic mice expressing MCP-5 controlled by Mx-1 promoter exhibited an increased bone mass because of a decrease in osteoclasts. This result strongly supported that MCP-5 negatively regulated osteoclast differentiation. MCP-5 also prevented severe bone loss caused by RANKL.
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Diferenciación Celular , Glicoproteínas de Membrana , Proteínas Quimioatrayentes de Monocitos , Osteoclastos , Animales , Humanos , Masculino , Ratones , Células Cultivadas , Glicoproteínas de Membrana/metabolismo , Ratones Endogámicos ICR , Proteínas Quimioatrayentes de Monocitos/genética , Proteínas Quimioatrayentes de Monocitos/metabolismo , Proteínas Quimioatrayentes de Monocitos/farmacología , FN-kappa B/metabolismo , Osteoclastos/citología , Osteoclastos/metabolismo , Ligando RANK/farmacología , Ligando RANK/metabolismo , Receptor Activador del Factor Nuclear kappa-B/metabolismo , Regulación hacia ArribaRESUMEN
Sestrins are stress-responsive proteins with antioxidant properties. They participate in cellular redox balance and protect against oxidative damage. This study investigated the effects of Sestrin2 (Sesn2) on osteoclast differentiation and function. Overexpressing Sesn2 in osteoclast precursor cells significantly inhibited receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclastogenesis. This was assessed as reduced expression of various osteoclast markers, including c-Fos, nuclear factor of activated T cells 1 (NFATc1), osteoclast-associated receptor, tartrate-resistant acid phosphatase, and cathepsin K. Conversely, downregulation of Sesn2 produced the opposite effect. Mechanistically, Sesn2 overexpression enhanced AMPK activation and the nuclear translocation of nuclear factor erythroid-derived factor 2-related factor 2 (Nrf2), promoting antioxidant enzymes. Moreover, azithromycin (Azm) induced Sesn2 expression, which suppressed RANKL-induced osteoclast differentiation. Specifically, Azm treatment reduced RANKL-induced production of reactive oxygen species in osteoclasts. Furthermore, intraperitoneal administration of Azm ameliorated RANKL-induced bone loss by reducing osteoclast activity in mice. Taken together, our results suggested that Azm-induced Sesn2 act as a negative regulator of RANKL-induced osteoclast differentiation through the AMPK/NFATc1 signaling pathway. Concisely, targeting Sesn2 can be a potential pharmacological intervention in osteoporosis.
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Osteogénesis , Ligando RANK , Animales , Ratones , Osteogénesis/genética , Especies Reactivas de Oxígeno/metabolismo , Ligando RANK/genética , Ligando RANK/farmacología , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Antioxidantes/farmacología , Osteoclastos/metabolismo , FN-kappa B/metabolismo , Factores de Transcripción NFATC/genética , Factores de Transcripción NFATC/metabolismo , Diferenciación CelularRESUMEN
Ginsenosides, among the most active components of ginseng, exhibit several therapeutic effects against cancer, diabetes, and other metabolic diseases. However, the molecular mechanism underlying the anti-osteoporotic activity of ginsenoside Rg2, a major ginsenoside, has not been clearly elucidated. This study aimed to determine the effects of ginsenoside Rg2 on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation. Results indicate that ginsenoside Rg2 inhibits RANKLinduced osteoclast differentiation of bone marrow macrophages (BMMs) without cytotoxicity. Pretreatment with ginsenoside Rg2 significantly reduced the RANKL-induced gene expression of c-fos and nuclear factor of activated T-cells (Nfatc1), as well as osteoclast-specific markers tartrate-resistant acid phosphatase (TRAP, Acp5) and osteoclast-associated receptor (Oscar). Moreover, RANKL-induced phosphorylation of mitogen-activated protein kinases (MAPKs) was decreased by ginsenoside Rg2 in BMM. Therefore, we suggest that ginsenoside Rg2 suppresses RANKLinduced osteoclast differentiation through the regulation of MAPK signaling-mediated osteoclast markers and could be developed as a therapeutic drug for the prevention and treatment of osteoporosis. [BMB Reports 2023; 56(10): 551-556].
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Resorción Ósea , Ginsenósidos , Humanos , Osteogénesis , Ginsenósidos/farmacología , Ginsenósidos/metabolismo , Osteoclastos/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , FN-kappa B/metabolismo , Factores de Transcripción NFATC/metabolismo , Ligando RANK/farmacología , Ligando RANK/metabolismo , Diferenciación Celular , Resorción Ósea/metabolismoRESUMEN
There has been increasing interest in adjunctive use of anti-inflammatory drugs to control periodontitis. This study was performed to examine the effects of pirfenidone (PFD) on alveolar bone loss in ligature-induced periodontitis in mice and identify the relevant mechanisms. Experimental periodontitis was established by ligating the unilateral maxillary second molar for 7 days in mice (n = 8 per group), and PFD was administered daily via intraperitoneal injection. The micro-computed tomography and histology analyses were performed to determine changes in the alveolar bone following the PFD administration. For in vitro analysis, bone marrow macrophages (BMMs) were isolated from mice and cultured with PFD in the presence of RANKL or LPS. The effectiveness of PFD on osteoclastogenesis, inflammatory cytokine expression, and NF-κB activation was determined with RT-PCR, Western blot, and immunofluorescence analyses. PFD treatment significantly inhibited the ligature-induced alveolar bone loss, with decreases in TRAP-positive osteoclasts and expression of inflammatory cytokines in mice. In cultured BMM cells, PFD also inhibited RANKL-induced osteoclast differentiation and LPS-induced proinflammatory cytokine (IL-1ß, IL-6, TNF-a) expression via suppressing the NF-κB signal pathway. These results suggest that PFD can suppress periodontitis progression by inhibiting osteoclastogenesis and inflammatory cytokine production via inhibiting the NF-κB signal pathway, and it may be a promising candidate for controlling periodontitis.
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Pérdida de Hueso Alveolar , Periodontitis , Ratones , Animales , FN-kappa B/metabolismo , Pérdida de Hueso Alveolar/tratamiento farmacológico , Pérdida de Hueso Alveolar/etiología , Pérdida de Hueso Alveolar/metabolismo , Microtomografía por Rayos X , Lipopolisacáridos/farmacología , Transducción de Señal , Osteoclastos/metabolismo , Periodontitis/tratamiento farmacológico , Periodontitis/etiología , Periodontitis/metabolismo , Citocinas/metabolismo , Ligando RANK/metabolismoRESUMEN
CrkII, a member of the adaptor protein family, is known to participate in bone homeostasis via the regulation of osteoclasts and osteoblasts. Therefore, silencing CrkII would beneficially impact the bone microenvironment. In this study, CrkII siRNA encapsulated by a bone-targeting peptide (AspSerSer)6-liposome was evaluated for its therapeutic applications using a receptor activator of nuclear factor kappa-B ligand (RANKL)-induced bone loss model. (AspSerSer)6-liposome-siCrkII maintained its gene-silencing ability in both osteoclasts and osteoblasts in vitro and significantly reduced osteoclast formation while increasing osteoblast differentiation in vitro. Fluorescence image analyses showed that the (AspSerSer)6-liposome-siCrkII was present largely in bone, where it remained present for up to 24 hours and was cleared by 48 hours, even when systemically administrated. Importantly, microcomputed-tomography revealed that bone loss induced by RANKL administration was recovered by systemic administration of (AspSerSer)6-liposome-siCrkII. Collectively, the findings of this study suggest that (AspSerSer)6-liposome-siCrkII is a promising therapeutic strategy for the development of treatments for bone diseases, as it overcomes the adverse effects derived from ubiquitous expression via bone-specific delivery of siRNA.
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Enfermedades Óseas , Resorción Ósea , Humanos , Osteogénesis , ARN Interferente Pequeño/metabolismo , Resorción Ósea/tratamiento farmacológico , Resorción Ósea/genética , Resorción Ósea/metabolismo , Liposomas/metabolismo , Osteoclastos , Osteoblastos , Enfermedades Óseas/metabolismo , Diferenciación CelularRESUMEN
BACKGROUND: Osteolysis is one of the most common problems that occurs after total hip and knee arthroplasty and has recently become a significant problem after total ankle arthroplasty (TAA). In this study, we investigated the role of LIM homeobox transcription factor 1-ß (Lmx1b) in osteoclast differentiation. By evaluating the expression profiles associated with osteolysis following TAA treatment, Lmx1b was found to be differentially expressed in patients with osteolysis after TAA. METHODS: To identify the important genes associated with osteolysis after TAA, RNA sequencing was performed by analyzing 8 patient samples: 5 primary TAA samples (control group) and 3 TAA samples revised for flexion instability (osteolysis group). By analyzing the differentially expressed genes and gene ontologies, Lmx1b expression was found to be upregulated in the osteolysis group compared to that in the control group. Focusing on the role of Lmx1b in bone cells, Lmx1b was overexpressed by a retrovirus in osteoclast precursor cells. The cultured cells were stained with tartrate-resistant acid phosphatase, and the expression of osteoclast-related genes was analyzed using real-time polymerase chain reaction. RESULTS: Lmx1b overexpression in osteoclast precursors suppresses osteoclast formation and resorptive activity. The expression of osteoclast marker genes was significantly reduced during osteoclast differentiation by Lmx1b overexpression. Furthermore, Lmx1b is associated with nuclear factor of activated T cells 1 (NFATc1) and inhibited NFATc1 translocation into the nucleus. CONCLUSIONS: These results provide novel insights into the anti-bone resorptive effect of Lmx1b on osteolysis after TAA and may lead to the development of effective preventative and therapeutic strategies for peri-implant osteolysis.
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Neurogenin 1 (Ngn1) belongs to the basic helix-loop-helix (bHLH) transcription factor family and plays important roles in specifying neuronal differentiation. The present study aimed to determine whether forced Ngn1 expression contributes to bone homeostasis. Ngn1 inhibited the p300/CREB-binding protein-associated factor (PCAF)-induced acetylation of nuclear factor of activated T cells 1 (NFATc1) and runt-related transcription factor 2 (Runx2) through binding to PCAF, which led to the inhibition of osteoclast and osteoblast differentiation, respectively. In addition, Ngn1 overexpression inhibited the TNF-α- and IL-17A-mediated enhancement of osteoclast differentiation and IL-17A-induced osteoblast differentiation. These findings indicate that Ngn1 can serve as a novel therapeutic agent for treating ankylosing spondylitis with abnormally increased bone formation and resorption.
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Osteoclastos , Osteogénesis , Diferenciación Celular , Interleucina-17/metabolismo , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteogénesis/genéticaRESUMEN
The LIM-homeodomain transcription factor Lmx1b plays a key role in body pattern formation during development. Although Lmx1b is essential for the normal development of multiple tissues, its regulatory mechanism in bone cells remains unclear. Here, we demonstrated that Lmx1b negatively regulates bone morphogenic protein 2 (BMP2)-induced osteoblast differentiation. Overexpressed Lmx1b in the osteoblast precursor cells inhibited alkaline phosphatase (ALP) activity and nodule formation, as well as the expression of osteoblast maker genes, including runt-related transcription factor 2 (Runx2), alkaline phosphatase (Alpl), bone sialoprotein (Ibsp), and osteocalcin (Bglap). Conversely, the knockdown of Lmx1b in the osteoblast precursors enhanced the osteoblast differentiation and function. Lmx1b physically interacted with and repressed the transcriptional activity of Runx2 by reducing the recruitment of Runx2 to the promoter region of its target genes. In vivo analysis of BMP2-induced ectopic bone formation revealed that the knockdown of Lmx1b promoted osteogenic differentiation and bone regeneration. Our data demonstrate that Lmx1b negatively regulates osteoblast differentiation and function through regulation of Runx2 and provides a molecular basis for therapeutic targets for bone diseases.
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Subunidad alfa 1 del Factor de Unión al Sitio Principal , Factores de Transcripción , Fosfatasa Alcalina/genética , Fosfatasa Alcalina/metabolismo , Proteína Morfogenética Ósea 2/genética , Proteína Morfogenética Ósea 2/metabolismo , Diferenciación Celular/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Osteoblastos/metabolismo , Osteogénesis/genética , Factores de Transcripción/metabolismoRESUMEN
Activating transcription factor 3 (ATF3) has been identified as a negative regulator of osteoblast differentiation in in vitro study. However, it was not associated with osteoblast differentiation in in vivo study. To provide an understanding of the discrepancy between the in vivo and in vitro findings regarding the function of ATF3 in osteoblasts, we investigated the unidentified roles of ATF3 in osteoblast biology. ATF3 enhanced osteoprotegerin (OPG) production, not only in osteoblast precursor cells, but also during osteoblast differentiation and osteoblastic adipocyte differentiation. In addition, ATF3 increased nodule formation in immature osteoblasts and decreased osteoblast-dependent osteoclast formation, as well as the transdifferentiation of osteoblasts to adipocytes. However, all these effects were reversed by the OPG neutralizing antibody. Taken together, these results suggest that ATF3 contributes to bone homeostasis by regulating the differentiation of various cell types in the bone microenvironment, including osteoblasts, osteoclasts, and adipocytes via inducing OPG production.
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Osteoclastos , Osteoprotegerina , Factor de Transcripción Activador 3/genética , Factor de Transcripción Activador 3/metabolismo , Adipocitos/metabolismo , Diferenciación Celular , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteogénesis , Osteoprotegerina/genética , Osteoprotegerina/metabolismo , Ligando RANK/metabolismoRESUMEN
Vascular calcification increases morbidity and mortality in patients with cardiovascular and renal diseases. Previously, we reported that histone deacetylase 1 prevents vascular calcification, whereas its E3 ligase, mouse double minute 2 homolog (MDM2), induces vascular calcification. In the present study, we identified the upstream regulator of MDM2. By utilizing cellular models and transgenic mice, we confirmed that E3 ligase activity is required for vascular calcification. By promoter analysis, we found that both msh homeobox 1 (Msx1) and msh homeobox 2 (Msx2) bound to the MDM2 promoter region, which resulted in transcriptional activation of MDM2. The expression levels of both Msx1 and Msx2 were increased in mouse models of vascular calcification and in calcified human coronary arteries. Msx1 and Msx2 potentiated vascular calcification in cellular and mouse models in an MDM2-dependent manner. Our results establish a novel role for MSX1/MSX2 in the transcriptional activation of MDM2 and the resultant increase in MDM2 E3 ligase activity during vascular calcification.
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Proteínas de Homeodominio/metabolismo , Factor de Transcripción MSX1/metabolismo , Proteínas Proto-Oncogénicas c-mdm2/genética , Ubiquitina-Proteína Ligasas/genética , Calcificación Vascular/etiología , Calcificación Vascular/metabolismo , Animales , Biomarcadores , Calcio/metabolismo , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Masculino , Ratones , Ratones Noqueados , Modelos Biológicos , Mutación , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Elementos de Respuesta , Ubiquitina-Proteína Ligasas/metabolismo , Calcificación Vascular/patologíaRESUMEN
Interferon regulatory factors (IRFs) play roles in various biological processes including cytokine signaling, cell growth regulation and hematopoietic development. Although it has been reported that several IRFs are involved in bone metabolism, the role of IRF2 in bone cells has not been elucidated. Here, we investigated the involvement of IRF2 in RANKL-induced osteoclast differentiation. IRF2 overexpression in osteoclast precursor cells enhanced osteoclast differentiation by regulating the expression of NFATc1, a master regulator of osteoclastogenesis. Conversely, IRF2 knockdown inhibited osteoclast differentiation and decreased the NFATc1 expression. Moreover, IRF2 increased the translocation of NF-κB subunit p65 to the nucleus in response to RANKL and subsequently induced the expression of NFATc1. IRF2 plays an important role in RANKL-induced osteoclast differentiation by regulating NF-κB/NFATc1 signaling pathway. Taken together, we demonstrated the molecular mechanism of IRF2 in osteoclast differentiation, and provide a molecular basis for potential therapeutic targets for the treatment of bone diseases characterized by excessive bone resorption. [BMB Reports 2021; 54(9): 482-487].
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Diferenciación Celular/efectos de los fármacos , Factor 2 Regulador del Interferón/metabolismo , Osteogénesis/efectos de los fármacos , Ligando RANK/farmacología , Transducción de Señal/efectos de los fármacos , Animales , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Núcleo Celular/metabolismo , Factor 2 Regulador del Interferón/antagonistas & inhibidores , Factor 2 Regulador del Interferón/genética , Masculino , Ratones , Ratones Endogámicos ICR , Factores de Transcripción NFATC/metabolismo , Osteoclastos/citología , Osteoclastos/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Factor de Transcripción ReIA/metabolismoRESUMEN
BACKGROUND: Multiple members of the transforming growth factor-ß (TGF-ß) superfamily have well-established roles in bone homeostasis. Anti-Müllerian hormone (AMH) is a member of TGF-ß superfamily of glycoproteins that is responsible for the regression of fetal Müllerian ducts and the transcription inhibition of gonadal steroidogenic enzymes. However, the involvement of AMH in bone remodeling is unknown. Therefore, we investigated whether AMH has an effect on bone cells as other TGF-ß superfamily members do. METHODS: To identify the roles of AMH in bone cells, we administered AMH during osteoblast and osteoclast differentiation, cultured the cells, and then stained the cultured cells with Alizarin red and tartrate-resistant acid phosphatase, respectively. We analyzed the expression of osteoblast- or osteoclast-related genes using real-time polymerase chain reaction and western blot. RESULTS: AMH does not affect bone morphogenetic protein 2-mediated osteoblast differentiation but inhibits receptor activator of nuclear factor-κB (NF-κB) ligand-induced osteoclast differentiation. The inhibitory effect of AMH on osteoclast differentiation is mediated by IκB-NF-κB signaling. CONCLUSIONS: AMH negatively regulates osteoclast differentiation without affecting osteoblast differentiation.
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Carcinoma de Pulmón de Células no Pequeñas/genética , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Mutación/genética , Miosinas/genética , Acrilamidas/uso terapéutico , Afatinib/uso terapéutico , Compuestos de Anilina/uso terapéutico , Animales , Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Modelos Animales de Enfermedad , Gefitinib/uso terapéutico , Técnicas In Vitro , Neoplasias Pulmonares/tratamiento farmacológico , RatonesRESUMEN
Coupled signaling between bone-forming osteoblasts and bone-resorbing osteoclasts is crucial to the maintenance of bone homeostasis. We previously reported that v-crk avian sarcoma virus CT10 oncogene homolog-like (CrkL), which belongs to the Crk family of adaptors, inhibits bone morphogenetic protein 2 (BMP2)-mediated osteoblast differentiation, while enhancing receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclast differentiation. In this study, we investigated whether CrkL can also regulate the coupling signals between osteoblasts and osteoclasts, facilitating bone homeostasis. Osteoblastic CrkL strongly decreased RANKL expression through its inhibition of runt-related transcription factor 2 (Runx2) transcription. Reduction in RANKL expression by CrkL in osteoblasts resulted in the inhibition of not only osteoblast-dependent osteoclast differentiation but also osteoclast-dependent osteoblast differentiation, suggesting that CrkL participates in the coupling signals between osteoblasts and osteoclasts via its regulation of RANKL expression. Therefore, CrkL bifunctionally regulates osteoclast differentiation through both a direct and indirect mechanism while it inhibits osteoblast differentiation through its blockade of both BMP2 and RANKL reverse signaling pathways. Collectively, these data suggest that CrkL is involved in bone homeostasis, where it helps to regulate the complex interactions of the osteoblasts, osteoclasts, and their coupling signals.
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Proteínas Adaptadoras Transductoras de Señales/fisiología , Remodelación Ósea/genética , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Animales Recién Nacidos , Diferenciación Celular/genética , Células Cultivadas , Células HEK293 , Humanos , Ratones , Ratones Endogámicos ICR , Osteoblastos/fisiología , Osteoclastos/fisiología , Osteogénesis/genéticaRESUMEN
STAT5 is a transcription factor that is activated by various cytokines, hormones, and growth factors. Activated STAT5 is then translocated to the nucleus and regulates the transcription of target genes, affecting several biological processes. Several studies have investigated the role of STAT5 in adipogenesis, but unfortunately, its role in adipogenesis remains controversial. In the present study, we generated adipocyte-specific Stat5 conditional knockout (cKO) (Stat5fl/fl;Apn-cre) mice to investigate the role of STAT5 in the adipogenesis of bone marrow mesenchymal stem cells (BMSCs). BMSC adipogenesis was significantly inhibited upon overexpression of constitutively active STAT5A, while it was enhanced in the absence of Stat5 in vitro. In vivo adipose staining and histological analyses revealed increased adipose volume in the bone marrow of Stat5 cKO mice. ATF3 is the target of STAT5 during STAT5-mediated inhibition of adipogenesis, and its transcription is regulated by the binding of STAT5 to the Atf3 promoter. ATF3 overexpression was sufficient to suppress the enhanced adipogenesis of Stat5-deficient adipocytes, and Atf3 silencing abolished the STAT5-mediated inhibition of adipogenesis. Stat5 cKO mice exhibited reduced bone volume due to an increase in the osteoclast number, and coculture of bone marrow-derived macrophages with Stat5 cKO adipocytes resulted in enhanced osteoclastogenesis, suggesting that an increase in the adipocyte number may contribute to bone loss. In summary, this study shows that STAT5 is a negative regulator of BMSC adipogenesis and contributes to bone homeostasis via direct and indirect regulation of osteoclast differentiation; therefore, it may be a leading target for the treatment of both obesity and bone loss-related diseases.
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Adipocitos/metabolismo , Huesos/metabolismo , Diferenciación Celular , Regulación de la Expresión Génica , Homeostasis , Células Madre Mesenquimatosas/metabolismo , Factor de Transcripción STAT5/metabolismo , Adipocitos/citología , Adipogénesis/genética , Animales , Huesos/diagnóstico por imagen , Huesos/patología , Células Cultivadas , Modelos Animales de Enfermedad , Humanos , Células Madre Mesenquimatosas/citología , Ratones , Ratones Noqueados , Ratones Transgénicos , Osteogénesis/genética , Unión Proteica , Factor de Transcripción STAT5/genética , Transducción de SeñalRESUMEN
Selenoproteins containing selenium in the form of selenocysteine are critical for bone remodeling. However, their underlying mechanism of action is not fully understood. Herein, we report the identification of selenoprotein W (SELENOW) through large-scale mRNA profiling of receptor activator of nuclear factor (NF)-κΒ ligand (RANKL)-induced osteoclast differentiation, as a protein that is downregulated via RANKL/RANK/tumour necrosis factor receptor-associated factor 6/p38 signaling. RNA-sequencing analysis revealed that SELENOW regulates osteoclastogenic genes. SELENOW overexpression enhances osteoclastogenesis in vitro via nuclear translocation of NF-κB and nuclear factor of activated T-cells cytoplasmic 1 mediated by 14-3-3γ, whereas its deficiency suppresses osteoclast formation. SELENOW-deficient and SELENOW-overexpressing mice exhibit high bone mass phenotype and osteoporosis, respectively. Ectopic SELENOW expression stimulates cell-cell fusion critical for osteoclast maturation as well as bone resorption. Thus, RANKL-dependent repression of SELENOW regulates osteoclast differentiation and blocks osteoporosis caused by overactive osteoclasts. These findings demonstrate a biological link between selenium and bone metabolism.