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1.
Mol Med Rep ; 27(1)2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36416346

RESUMO

Carnosine is a vital endogenous dipeptide that has anti­inflammatory, antiaging, anti­crosslinking, antitumor and immune regulatory effects. Numerous cell and animal model studies have proved that carnosine and its compounds promote the proliferation and differentiation of osteoblasts, inhibit osteoclasts and protect chondrocytes. They also regulate the cell cycle of bone progenitor cells and the differentiation of bone marrow mesenchymal stem cells, accelerate fracture healing, delay bone tumor development and ameliorate osteopenia induced by estrogen deficiency or disuse. The correlations between carnosine and activation signal molecules, pluripotent differentiation of bone marrow mesenchymal stem cells and interaction between bone cells are unclear. However, studies have proved that carnosine and its compounds have benefits in preventing and treating specific bone diseases. This makes them potential agents for the treatment of osteoporosis and bone tumors. The present review summarized the existing research on carnosine and its compounds in bone cells and tissue. It focused on the physiological function of carnosine and its compounds in the bone and their effect on bone metabolism­related diseases, thus providing support for developing new strategies for targeted therapy.


Assuntos
Carnosina , Células-Tronco Mesenquimais , Animais , Carnosina/farmacologia , Carnosina/uso terapêutico , Osteoclastos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Osteoblastos , Diferenciação Celular
2.
Cells ; 11(22)2022 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-36429027

RESUMO

Osteoporosis is a systemic skeletal disorder where osteoclasts are prevalent among osteoblasts. Oxidative stress is one of the main causes of osteoporosis, and nuclear factor erythroid-2-related factor 2 (Nrf2) is the master regulator of antioxidant responses. Phytol, a diterpene isolated from Stevia rebaudiana leaves, has many biological effects, including antimicrobial, antioxidant, and anti-inflammatory effects. This study investigated the crosstalk between Nrf2 and osteoclast differentiation in the presence of phytol. Phytol inhibited osteoclast differentiation through TRAP-positive and F-actin formation. The expression of anti-nuclear factor of activated T cells-c1 (NFATc1) and c-Fos was suppressed by phytol, as shown using Western blot and RT-PCR analysis. Phytol inhibited oxidative stress by suppressing reactive oxidant species (ROS) accumulation while recovering antioxidant enzymes, including superoxide dismutase and catalase. Additionally, phytol ameliorated osteoclast-specific differentiation, function, and oxidative stress through Nrf2 regulation by siRNA transfection. In conclusion, these data demonstrate the inhibitory effect of phytol on osteoclast differentiation through Nrf2 regulation, suggesting its potential use in oxidative stress-related osteoporosis and bone diseases.


Assuntos
Fator 2 Relacionado a NF-E2 , Osteoporose , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Fator de Transcrição de Proteínas de Ligação GA/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Osteoclastos/metabolismo , Osteoporose/metabolismo , Estresse Oxidativo , Fitol/metabolismo , Fitol/farmacologia , Animais , Camundongos
3.
Arch Biochem Biophys ; 732: 109464, 2022 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-36368367

RESUMO

Abnormal increases in osteoclast differentiation and activity contribute to excessive bone resorption in inflammatory bone diseases. The specific m6A-binding protein YT521-B homology domain family 1 (YTHDF1) participates in many physiopathological processes by regulating mRNA stability or translation. However, whether YTHDF1 is involved in the regulation of inflammatory osteoclastogenesis remains a mystery. This study revealed that YTHDF1 expression was upregulated during lipopolysaccharide (LPS)-stimulated osteoclast differentiation. Knockdown of Ythdf1 inhibited osteoclast formation, bone resorption and the expression of osteoclast-related genes (Tnfrsf11a, Traf6, Mmp9 and Acp5). Analysis of RNA sequencing data showed that the genes downregulated by Ythdf1 knockdown were closely associated with endoplasmic reticulum (ER) stress and osteoclast differentiation. Western blotting confirmed that Ythdf1 depletion suppressed activation of the ER stress-related PERK, IRE1α and ATF6 signaling pathways. The ER stress activator tunicamycin (Tm) partially rescued the decreased expression of Mmp9 and Acp5 caused by Ythdf1 deficiency. Meanwhile, Ythdf1 depletion inhibited the phosphorylation levels of key proteins in the NF-κB, MAPK and PI3K-AKT signaling pathways and decreased the mRNA stability of Tnfrsf11a, which is the major upstream signaling molecule that mediates the activation of these pathways during osteoclast differentiation. In conclusion, our findings suggest that Ythdf1 knockdown inhibits inflammatory osteoclast differentiation and function by suppressing ER stress signaling pathways. Ythdf1 knockdown also inactivates the signaling pathways involved in osteoclast differentiation by inhibiting Tnfrsf11a mRNA stability. These findings will help shed light on the molecular mechanisms of m6A-mediated epigenetic regulation in inflammatory osteoclastogenesis.


Assuntos
Reabsorção Óssea , NF-kappa B , Humanos , NF-kappa B/metabolismo , Osteogênese , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Endorribonucleases , Estresse do Retículo Endoplasmático , Epigênese Genética , Proteínas Serina-Treonina Quinases , Osteoclastos/metabolismo , Reabsorção Óssea/metabolismo , Transdução de Sinais , Ligante RANK/metabolismo , Diferenciação Celular
4.
Biochem Biophys Res Commun ; 636(Pt 2): 87-96, 2022 Dec 25.
Artigo em Inglês | MEDLINE | ID: mdl-36368159

RESUMO

To verify the osteoclast differentiation ability of MDSCs from mice of different ages and explore the effect of AOPPs on the osteoclast differentiation of bone marrow MDSCs. Bone marrow cells from C57BL/6 (a.k.a C57) mice of different ages were subjected to flow cytometry, and CD11b+Ly6C+Ly6G+ MDSCs were sorted out. After induction of osteoclast differentiation, these cells were subjected to tartrate-resistant acid phosphatase (TRAP) and F-actin. MDSCs from bone marrows of old mice were injected into the tibial medullary cavity of young mice. One week later, the bone marrows were subjected to histological examination, TRAP, and cell count. MDSCs from bone marrows of old mice were sorted for induction of osteoclast differentiation, intervened with reactive oxygen species (ROS) scavenger, inducible nitric oxide synthase (iNOS) inhibitor, and nitric oxide (NO) scavenger, and then subjected to TRAP. 8-weeks-old C57 mice were injected with the same concentrations of either AOPPs or mouse serum albumin (MSA). Four weeks later, MDSCs from bone marrows were sorted and subjected to induction of osteoclast differentiation, followed by IHC staining and TRAP. MDSCs of 8-weeks-old C57 mice were extracted and subjected to in vitro induction of osteoclast differentiation with different concentrations of AOPPs, followed by TRAP training. The number of MDSCs in the bone marrows of old mice was significantly higher than that in young mice. MDSCs from bone marrows of old mice differentiated into large multinucleated TRAP+ osteoclasts, which were significantly different from those in the middle-aged and young mice in terms of cell quantity and morphology. The actin rings formed in the differentiated osteoclasts from MDSCs of bone marrows were densely distributed in the whole field of view, which were significantly denser than those in the middle-aged and young mice. After injection of MDSCs of old mice, the number of TRAP + osteoclasts in the tibial medullary cavity of young mice was significantly increased. NO inhibitor can significantly inhibit the osteoclast differentiation capacity of MDSCs from bone marrows of old mice. In vivo treatment with AOPPs significantly increased the proportion of MDSCs in the bone marrow, which is up to 55.2%. After injection of AOPPs in 8-week-old mice and induction of osteoclast differentiation from the MDSCs, the ratios of CD11b+ and Gr1+ cells were significantly higher than that in the control and MSA groups but was not significantly different from that in the 15-month-old mice. Upon in vitro treatment with different concentrations of AOPPs, the MDSCs did not show any sign of osteoclast differentiation. MDSCs can directly undergo osteoclast differentiation, the capacity of which is stronger in MDSCs of bone marrows of old mice; the NO pathway is a potential mechanism underlying this phenomenon. In vivo but not in vitro AOPPs treatment can induce osteoclast differentiation of MDSCs, indicating there might be other factors in the body that can interact with AOPPs to induce osteoclast differentiation of MDSCs.


Assuntos
Células Supressoras Mieloides , Osteoclastos , Camundongos , Animais , Osteoclastos/metabolismo , Produtos da Oxidação Avançada de Proteínas/metabolismo , Camundongos Endogâmicos C57BL , Diferenciação Celular , Envelhecimento
5.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi ; 36(11): 1428-1433, 2022 Nov 15.
Artigo em Chinês | MEDLINE | ID: mdl-36382463

RESUMO

Objective: To summarize the characteristics of the occurrence and development of osteonecrosis of the femoral head (ONFH), and to review the important regulatory role of immune cells in the progression of ONFH. Methods: The domestic and foreign literature on the immune regulation of ONFH was reviewed, and the relationship between immune cells and the occurrence and development of ONFH was analyzed. Results: The ONFH region has a chronic inflammatory reaction and an imbalance between osteoblast and osteoclast, while innate immune cells such as macrophages, neutrophils, dendritic cells, and immune effector cells such as T cells and B cells are closely related to the maintenance of bone homeostasis. Conclusion: Immunotherapy targeting the immune cells in the ONFH region and the key factors and proteins in their regulatory pathways may be a feasible method to delay the occurrence, development, and even reverse the pathology of ONFH.


Assuntos
Necrose da Cabeça do Fêmur , Células-Tronco Mesenquimais , Osteonecrose , Humanos , Cabeça do Fêmur/patologia , Necrose da Cabeça do Fêmur/etiologia , Necrose da Cabeça do Fêmur/patologia , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Osteoclastos/metabolismo
6.
J Nutr Sci Vitaminol (Tokyo) ; 68(Supplement): S113-S115, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36436988

RESUMO

Bone health is an important medical concern in rapidly aging demographics worldwide. Excessive bone resorption, due to enhanced activity of osteoclasts, is a major underlying cause of bone disorders such as osteoporosis. Inflammation and oxidative stress are key factors contributing to increased osteoclastic activity. Like increased activity of osteoclasts, depletion of osteoblasts also contributes to weakened structural integrity of bone. Considering the epidemiology of bone disorders and aging demographics there is a substantial need for novel bone health therapeutics. IRW (Ile-Arg-Trp), an egg-derived tripeptide, exhibits a spectrum of pharmacological activity. In our recent work, we have shown that IRW inhibits osteoclastogenesis and promotes osteogenesis in the mouse macrophage RAW 264.7 and MC3T3-E1 cells. IRW treatment (25 and 50 µM) significantly inhibited osteoclastogenesis-associated factors [TRAF6 (TNF Receptor Associated Factor 6), Fos Proto-Oncogene (c-Fos), Nuclear Factor of Activated T Cells 1 (NFATc1), and cathepsin K] and upregulated osteogenesis-associated factors [RUNX2 (Runt-related transcription factor 2) and RANKL (Receptor activator of nuclear factor kappa-B ligand)] in the two cell lines. Currently, we are conducting studies to analyze the impact of IRW on Angiotensin II (Ang II)-induced stress in vitro and in vivo. In summary, our recent work presents the ability of IRW to prevent LPS-induced inflammatory bone resorption and activation of osteogenesis activity via multiple signaling pathways.


Assuntos
Reabsorção Óssea , Osteoporose , Camundongos , Animais , Osteoclastos/metabolismo , Osteoporose/prevenção & controle , Osteoporose/metabolismo , Reabsorção Óssea/prevenção & controle , Reabsorção Óssea/metabolismo , Suplementos Nutricionais
7.
Front Endocrinol (Lausanne) ; 13: 939959, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36425467

RESUMO

Background: Ceritinib is used for the treatment of patients with anaplastic lymphoma kinase (ALK)-rearranged non-small cell lung cancer (NSCLC), who are at the risk of developing bone metastasis. During bone metastasis, tumor cells release factors that induce osteoclast formation, resulting in osteolysis. However, the effect of ceritinib on osteoclast formation remains unclear. Methods: Osteoclastogenesis was induced to assess the effect of ceritinib on osteoclast formation and osteoclast-specific gene expression. Western blotting was used to examine the molecular mechanisms underlying the effect of ceritinib on osteoclast differentiation. An in vivo ovariectomized mouse model was established to validate the effect of ceritinib in suppressing osteoclast formation and preventing bone loss. Results: The differentiation of osteoclasts and the expression of osteoclast-specific genes were inhibited upon ceritinib stimulation. Ceritinib suppressed Akt and p65 phosphorylation during the receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastogenesis. The administration of ceritinib to ovariectomized mice ameliorated trabecular bone loss by inhibiting osteoclast formation. Conclusions: Ceritinib is beneficial in preventing bone loss by suppressing osteoclastic Akt and nuclear factor κB (NF-κB) signaling.


Assuntos
Doenças Ósseas Metabólicas , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Camundongos , Animais , Osteoclastos/metabolismo , NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Pulmonares/patologia , Doenças Ósseas Metabólicas/patologia
8.
Biochem Biophys Res Commun ; 636(Pt 1): 178-183, 2022 Dec 25.
Artigo em Inglês | MEDLINE | ID: mdl-36334442

RESUMO

Inhibition of osteoclast differentiation is a promising approach for the treatment of osteoporosis and rheumatoid arthritis. Receptor activator of nuclear factor kappa B (NF-κB) (RANK), which is an essential molecule for osteoclast differentiation, interacts with tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) to transduce downstream signals. Both RANK and TRAF6 have homo-trimeric structures, forming a multivalent interaction between the Pro-X-Glu-X-X-(aromatic/acidic) motif of RANK and the C-terminal domain of TRAF6 (TRAF-C), that markedly increases the binding affinity. Here, we designed a tetravalent peptide, RANK-tet, containing the TRAF-C-binding motif of RANK and found that RANK-tet binds to TRAF-C with high affinity. In contrast, a monomeric form of RANK-tet (RANK-mono) with the same TRAF-C-binding motif did not bind to TRAF-C, clearly indicating the multivalent interaction is strictly required for the high-affinity binding to TRAF-C. RANK-tet did not bind to a series of TRAF-C-mutants with an amino acid substitution in the RANK-binding region, indicating that RANK-tet specifically targets the RANK-binding region of TRAF-C. A cell-permeable form of RANK-tet that has poly-Arg residues at each C-terminal of the TRAF-C-binding motif efficiently inhibited the RANK ligand (RANKL)-induced differentiation of bone marrow cells to osteoclasts. Thus, this compound can be an effective anti-osteoclastogenic agent.


Assuntos
Ligante RANK , Fator 6 Associado a Receptor de TNF , Fator 6 Associado a Receptor de TNF/metabolismo , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Ligante RANK/metabolismo , Osteoclastos/metabolismo , NF-kappa B/metabolismo , Peptídeos/farmacologia , Peptídeos/metabolismo , Diferenciação Celular/fisiologia
9.
Int J Mol Sci ; 23(21)2022 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-36362318

RESUMO

Bone absorption is necessary for the maintenance of bone homeostasis. An osteoclast (OC) is a monocyte-macrophage lineage cell that absorbs bone tissue. Extracellular signal-regulated kinases (ERKs) are known to play important roles in regulating OC growth and differentiation. In this study, we examined specific downstream signal pathways affected by ERK inhibition during OC differentiation. Our results showed that the ERK inhibitors PD98059 and U0126 increased receptor activator of NF-κB ligand (RANKL)-induced OC differentiation in RAW 264.7 cells, implying a negative role in OC differentiation. This is supported by the effect of ERK2-specific small interfering RNA on increasing OC differentiation. In contrast to our findings regarding the RAW 264.7 cells, the ERK inhibitors attenuated the differentiation of bone marrow-derived cells into OCs. The ERK inhibitors significantly increased the phosphorylation of adenosine 5'-monophosphate-activated protein kinase (AMPK) but not the activation of p38 MAPK, Lyn, and mTOR. In addition, while the ERK inhibition increased the expression of the RANKL receptor RANK, it decreased the expression of negative mediators of OC differentiation, such as interferon regulatory factor-8, B-cell lymphoma 6, and interferon-γ. These dichotomous effects of ERK inhibition suggest that while ERKs may play positive roles in bone marrow-derived cells, ERKs may also play negative regulatory roles in RAW 264.7 cells. These data provide important information for drug development utilizing ERK inhibitors in OC-related disease treatment.


Assuntos
Proteínas Quinases Ativadas por AMP , Reabsorção Óssea , Camundongos , Animais , Proteínas Quinases Ativadas por AMP/metabolismo , Células RAW 264.7 , Ligante RANK/farmacologia , Ligante RANK/metabolismo , Osteoclastos/metabolismo , Osteogênese , Diferenciação Celular , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Reabsorção Óssea/metabolismo
10.
Biomed Res Int ; 2022: 8862278, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36330454

RESUMO

Bone remodels via resorption and formation, two phenomena that continuously occur in bone turnover. The RANKL/RANK/OPG pathway is one of the several mechanisms that affect bone turnover. The RANKL/OPG ratio has a substantial role in bone resorption. An imbalance between formation and resorption is related to an increased RANKL/OPG balance. OPG, a member of this system, can bind to RANKL and suppress RANK-RANKL interaction, and subsequently, inhibit further osteoclastogenesis. The serum levels of RANKL and OPG in the bone microenvironment are vital for osteoclasts formation. The RANK/RANKL/OPG system plays a role in the pathogenesis of bone disorders. This system can be considered a new treatment target for bone disorders. Soy isoflavones affect the RANK/RANKL/OPG system through numerous mechanisms. Soy isoflavones decrease RANKL levels and increase OPG levels. Therefore, isoflavones improve bone metabolism and decrease bone resorption. Soy isoflavones decrease serum markers of bone resorption and improve bone metabolism. However, while the available data are promising, the results of several studies reported no change in RANKL and OPG levels with isoflavones supplementation. In this regard, current evidence is insufficient for conclusive approval of the efficacy of isoflavones on RANKL/RANK/OPG and further research, including animal and human studies, are needed to confirm the effect of soy isoflavones on the RANKL/RANK/OPG pathway. This study was a review of available evidence to determine the role of isoflavones in bone hemostasis and the RANK/RANKL/OPG pathway. The identification of the effects of isoflavones on the RANKL/RANK/OPG pathway directs future studies and leads to the development of effective treatment strategies for bone disorders.


Assuntos
Doenças Ósseas , Reabsorção Óssea , Isoflavonas , Animais , Humanos , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Osteoprotegerina/metabolismo , Densidade Óssea , Ligante RANK/metabolismo , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/metabolismo , Osteoclastos/metabolismo , Doenças Ósseas/metabolismo , Isoflavonas/farmacologia
11.
Nat Commun ; 13(1): 6648, 2022 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-36333322

RESUMO

The impact of bone cell activation on bacterially-induced osteolysis remains elusive. Here, we show that matrix-embedded osteocytes stimulated with bacterial pathogen-associated molecular patterns (PAMPs) directly drive bone resorption through an MYD88-regulated signaling pathway. Mice lacking MYD88, primarily in osteocytes, protect against osteolysis caused by calvarial injections of bacterial PAMPs and resist alveolar bone resorption induced by oral Porphyromonas gingivalis (Pg) infection. In contrast, mice with targeted MYD88 restoration in osteocytes exhibit osteolysis with inflammatory cell infiltration. In vitro, bacterial PAMPs induce significantly higher expression of the cytokine RANKL in osteocytes than osteoblasts. Mechanistically, activation of the osteocyte MYD88 pathway up-regulates RANKL by increasing binding of the transcription factors CREB and STAT3 to Rankl enhancers and by suppressing K48-ubiquitination of CREB/CREB binding protein and STAT3. Systemic administration of an MYD88 inhibitor prevents jawbone loss in Pg-driven periodontitis. These findings reveal that osteocytes directly regulate inflammatory osteolysis in bone infection, suggesting that MYD88 and downstream RANKL regulators in osteocytes are therapeutic targets for osteolysis in periodontitis and osteomyelitis.


Assuntos
Perda do Osso Alveolar , Osteólise , Osteomielite , Periodontite , Camundongos , Animais , Osteócitos/metabolismo , Osteólise/induzido quimicamente , Osteólise/complicações , Osteólise/metabolismo , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Padrões Moleculares Associados a Patógenos/metabolismo , Ligante RANK/metabolismo , Porphyromonas gingivalis/metabolismo , Periodontite/metabolismo , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Osteoclastos/metabolismo
12.
Int Immunopharmacol ; 113(Pt A): 109302, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36257255

RESUMO

Osteoporoticfractures become increasingly common in postmenopausal women over age 55 years and men after age 65 years, bringing about substantial bone-associated morbidities, and augmented mortality and health-care costs. Advanced researches have led to a more accurate assessment of osteoporosis (OP) and have broadened the range of therapeutic approaches available to prevent osteoporotic fractures. Single-cell RNA sequencing (scRNA-seq) analysis is an evolutionary method that quantifies the majority of transcripts in individual cells at isoform resolution, paving the way for more detailed analyses of gene regulation in biology and medicine. We have extracted 19,102 cells and 4097 dynamical genes with significant expression changes. Several new subtypes of macrophages and differentially over-expressed genes were discovered in the trajectory of osteoclasts formation. The zinc finger protein 36, C3H type-like 1 (ZFP36L1) and defensin alpha 3 (DEFA3) were identified as novel bone metabolism-related genes. RETN-CAP1 was newly found to be involved in the interaction between osteoclasts and immunocytes, indicating that osteo-immunology microenvironment substantially contributed to the pathology of osteoporosis or osteopenia. In this research, we have performed Single-cell RNA sequencing analysis to display the trajectory of osteoclast formation and reveal the possible gene targets and signaling pathways that probably play an important role in osteoporosis.


Assuntos
Doenças Ósseas Metabólicas , Osteoporose , Masculino , Feminino , Humanos , Pessoa de Meia-Idade , Idoso , Osteoporose/genética , Osteoporose/metabolismo , Osteoclastos/metabolismo , Regulação da Expressão Gênica , Análise de Sequência de RNA
13.
Int J Mol Sci ; 23(19)2022 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-36233351

RESUMO

Erythropoietin (EPO) is a pleiotropic cytokine that classically drives erythropoiesis but can also induce bone loss by decreasing bone formation and increasing resorption. Deletion of the EPO receptor (EPOR) on osteoblasts or B cells partially mitigates the skeletal effects of EPO, thereby implicating a contribution by EPOR on other cell lineages. This study was designed to define the role of monocyte EPOR in EPO-mediated bone loss, by using two mouse lines with conditional deletion of EPOR in the monocytic lineage. Low-dose EPO attenuated the reduction in bone volume (BV/TV) in Cx3cr1Cre EPORf/f female mice (27.05%) compared to controls (39.26%), but the difference was not statistically significant. To validate these findings, we increased the EPO dose in LysMCre model mice, a model more commonly used to target preosteoclasts. There was a significant reduction in both the increase in the proportion of bone marrow preosteoclasts (CD115+) observed following high-dose EPO administration and the resulting bone loss in LysMCre EPORf/f female mice (44.46% reduction in BV/TV) as compared to controls (77.28%), without interference with the erythropoietic activity. Our data suggest that EPOR in the monocytic lineage is at least partially responsible for driving the effect of EPO on bone mass.


Assuntos
Eritropoetina , Receptores da Eritropoetina , Animais , Eritropoetina/metabolismo , Eritropoetina/farmacologia , Feminino , Camundongos , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Receptores da Eritropoetina/genética , Receptores da Eritropoetina/metabolismo , Transdução de Sinais
14.
Nutrients ; 14(19)2022 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-36235607

RESUMO

Trimethylamine-N-oxide (TMAO), an important gut microbiota (GM)-derived metabolite, has been shown to be abnormally increased in osteoporosis. However, the role and underlying mechanism of TMAO in regulating bone loss during osteoporosis have not been fully investigated. In the current study, we found that 100-400 µM TMAO dose-dependently enhanced TRAP-positive osteoclasts, F-actin ring formation, and resorption area on bovine bone slices and up-regulated osteoclast-related gene expression (Calcr, Traf6, Dcstamp, Acp5, C-Fos, and NFATc1). Western blotting validated that TMAO not only activated NF-κB signaling pathway but also stimulated c-Fos and NFATc1 protein expression in a dose-dependent manner. Furthermore, BAY 11-7082, an NF-κB inhibitor, pretreatment markedly suppressed TRAP-positive osteoclast formation and osteoclast-related genes under TMAO treatment. BAY 11-7082 also inhibited p-p65/p65, c-Fos, and NFATc1 protein expression promoted by TMAO. Moreover, TMAO significantly increased ROS production, which was inhibited by N-acetylcysteine (NAC), an ROS antagonist. In addition, we proved that NAC pretreatment could inhibit TMAO-promoted NF-κB activation. NAC also suppressed TRAP-positive osteoclast formation, osteoclast-related gene expression, and protein expression of c-Fos and NFATc1 under TMAO treatment. In vivo studies showed significantly decreased bone mass and increased TRAP-positive osteoclasts in TMAO-treated C57BL/6 mice. Moreover, western-blotting and immunohistochemical staining showed that TMAO administration markedly stimulated NF-κB p65 expression. Additionally, TMAO administration significantly promoted the gene and protein expression of C-Fos and NFATc1. In conclusion, TMAO could promote osteoclast differentiation and induce bone loss in mice by activating the ROS-dependent NF-κB signaling pathway.


Assuntos
Reabsorção Óssea , Osteoporose , Acetilcisteína/metabolismo , Actinas/metabolismo , Animais , Reabsorção Óssea/metabolismo , Bovinos , Diferenciação Celular , Metilaminas , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Nitrilas , Osteoclastos/metabolismo , Osteogênese , Osteoporose/metabolismo , Óxidos/metabolismo , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ligante RANK/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Sulfonas , Fator 6 Associado a Receptor de TNF/metabolismo
15.
Life Sci ; 310: 121073, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36244411

RESUMO

AIM: The low-density lipoprotein receptor (LDLR) plays a crucial role in regulating lipid metabolism. However, whether LDLR deficiency affects bone mass and morphology remains controversial. This study aimed to analyze the bone phenotypes of LDLR knockout (LDLR-/-) mice. MAIN METHODS: Eight-week-old LDLR-/- and wild-type (WT) mice were subjected to microcomputed tomography to detect bone phenotypes. Enzyme-linked immunosorbent assay kits were used to detect the serum estrogen levels and matrix metalloproteinase 9 (MMP-9) levels in tissue homogenates. Von Kossa, toluidine blue, tartrate-resistant acid phosphatase (TRAP) staining, and calcein labeling were performed to explore bone turnover parameters. In vitro, osteoclastogenesis was induced in bone marrow cells from LDLR-/- mice and WT mice in the presence or absence of 17ß-estradiol. The microphotographs and number of osteoclasts were validated using TRAP staining. Relative gene expression during osteoclast differentiation and maturation was determined by quantitative real-time polymerase chain reaction. KEY FINDINGS: LDLR deficiency results in reduced bone mineral density of the tibial cancellous bone, indicating bone loss to some extent in LDLR-/- mice. LDLR deficiency significantly increased the number of osteoclasts, but not osteoblasts. In vitro, bone marrow cells from LDLR-/- mice displayed enhanced osteoclastic potential along with increased expression of TRAP, cathepsin K, nuclear factor of activated T-cells 1 (NFATc1), c-fos, and MMP-9 and inhibited dendritic cell-specific transmembrane protein expression. Moreover, 17ß-estradiol treatment can inhibit osteoclastogenesis in vitro. SIGNIFICANCE: Our data demonstrated that LDLR deficiency promoted osteoclastogenesis by upregulating c-fos and NFATc1 expression, reducing cancellous bone mass in LDLR-/- mice.


Assuntos
Reabsorção Óssea , Ligante RANK , Receptores de LDL , Animais , Camundongos , Densidade Óssea , Reabsorção Óssea/metabolismo , Diferenciação Celular , Estradiol/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/metabolismo , Osteogênese , Ligante RANK/metabolismo , Microtomografia por Raio-X , Receptores de LDL/genética , Camundongos Knockout
16.
Biochem Biophys Res Commun ; 635: 227-235, 2022 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-36283335

RESUMO

TREM2 (Triggering receptor expressed on myeloid cells 2) is the causative gene for Nasu-Hakola disease, which is characterized by multiple bone cysts and leukoencephalopathy. In addition, mutations in this gene have been found to be correlated with the onset of Alzheimer's disease. TREM2 is an immunoreceptor expressed on dendritic cells, microglia, osteoclasts, and macrophages. TREM2 on the cell membrane is shed by some proteases and released as soluble TREM2 (sTREM2). Meanwhile, several TREM2 ligands have been reported, and lipopolysaccharide (LPS) is one of the candidates. Using RNA interference to examine TREM2-mediated LPS response in macrophages, we identified five chemokines whose expression was induced via TREM2. Furthermore, we showed that LPS-induced expression of CXC-motif chemokine ligand (Cxcl10) and Cxcl11 among the five chemokines was mediated in part through sTREM2. These results suggest that sTREM2 has cytokine-like functions in macrophages.


Assuntos
Doença de Alzheimer , Quimiocinas CXC , Glicoproteínas de Membrana , Receptores Imunológicos , Animais , Camundongos , Doença de Alzheimer/metabolismo , Proteínas de Transporte/metabolismo , Quimiocinas CXC/metabolismo , Ligantes , Lipopolissacarídeos/metabolismo , Macrófagos/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Microglia/metabolismo , Osteoclastos/metabolismo , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo
17.
Biochem Biophys Res Commun ; 632: 40-47, 2022 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-36198202

RESUMO

Autophagy is a non-selective action in which cells degrade parts of themselves, reusing degraded cellular components. Among autophagy-related gene (ATG) family members, ATG4 proteins play crucial roles in the microtubule-associated protein 1 light chain 3 (LC3) phosphatidylethanolamine (PE) system which is essential for autophagosome maturation. Although autophagy has been shown to be involved in osteoclastic bone resorption, the role of ATG4/LC3 in bone resorption remains unclear. When mouse bone marrow cells were treated with various concentrations of NSC185058 (NSC), a specific inhibitor of ATG4B, 1 h prior to treatment with receptor activator of NF-κB ligand (RANKL) in the presence of macrophage colony stimulating factor (M-CSF), NSC inhibited osteoclastogenesis in a dose-dependent manner. Addition of NSC in the late stages of osteoclast differentiation suppressed multinucleation and reduced the expression of markers for mature osteoclasts such as Dc-stamp, Mmp9, and Ctsk. NSC also suppressed actin ring formation and pit formation in mature osteoclasts. When a periodontitis model involving eight-week-old male mice in which the right maxillary second molar had been ligated with silk thread was injected with or without NSC, alveolar bone resorption was suppressed by a decrease in the number of osteoclasts in the NSC-treated group. These results suggest that LC3 is important for the maturation of osteoclasts and that LC3 inhibition is a new therapeutic strategy for periodontal disease.


Assuntos
Diferenciação Celular , Osteoclastos , Animais , Masculino , Camundongos , Actinas/metabolismo , Perda do Osso Alveolar , Ligantes , Fator Estimulador de Colônias de Macrófagos/farmacologia , Fator Estimulador de Colônias de Macrófagos/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Proteínas Associadas aos Microtúbulos/antagonistas & inibidores , Proteínas Associadas aos Microtúbulos/metabolismo , Osteoclastos/metabolismo , Fosfatidiletanolaminas/metabolismo , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Seda , Reabsorção Óssea/genética , Reabsorção Óssea/metabolismo , Diferenciação Celular/genética , Proteínas Relacionadas à Autofagia/antagonistas & inibidores , Proteínas Relacionadas à Autofagia/metabolismo , Cisteína Endopeptidases/metabolismo
18.
J Oral Biosci ; 64(4): 410-421, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36241157

RESUMO

OBJECTIVES: Interleukin-6 (IL-6) contributes to the regulation of functions in various tissues and organs. Even though IL-6 has been reported to modulate bone metabolism in previous studies, this finding is controversial. This study aims to evaluate the possible involvement of IL-6 in bone metabolism by examining the histological activity of osteoblasts and osteoclasts in the femora of Il-6 deficient (Il-6-/-) mice. METHODS: Eight-week-old male Il-6-/- mice and their wild-type littermates were fixed with a paraformaldehyde solution, and their femora were extracted for micro-CT analysis, immunohistochemistry, and real-time PCR analysis. RESULTS: Il-6-/- femora showed an increased bone volume/tissue volume (TV) but a reduced bone mineral density compared with the wild-type. Furthermore, the tissue-nonspecific alkaline phosphatase positive area/TV ratio, the expression of Runx2, Osterix, and Rankl, and the number of tartrate-resistant acid phosphatase-positive osteoclasts were all increased in the Il-6-/- mice. A considerable number of unmineralized areas within the bone matrix and abundant sclerostin-reactive osteocytes were observed in Il-6-/- femoral metaphyses but not in the wild-type. Interestingly, the gene expression of Cd206 was elevated in Il-6-/- femora, and many F4/80-positive macrophages/monocytes and CD206-immunoreactive macrophages in the primary trabeculae had migrated closer to the growth plate, where intense RANKL immunoreactivity was detected. These results suggest that, in an IL-6-deficient state, CD206-positive macrophages may differentiate into osteoclasts when in contact with RANKL-reactive osteoblastic cells. CONCLUSION: In a state of IL-6 deficiency, the population and cell activities of osteoblast, osteoclasts, and macrophages seemed to be facilitated, except for the reduced mineralization in bone.


Assuntos
Remodelação Óssea , Interleucina-6 , Camundongos , Masculino , Animais , Interleucina-6/genética , Remodelação Óssea/genética , Osteoclastos/metabolismo , Osteoblastos/metabolismo , Osso e Ossos/diagnóstico por imagem
19.
Biol Pharm Bull ; 45(10): 1426-1431, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36184499

RESUMO

Vacuolar-type ATPase (V-ATPase) shares its structure and rotational catalysis with F-type ATPase (F-ATPase, ATP synthase). However, unlike subunits of F-ATPase, those of V-ATPase have tissue- and/or organelle-specific isoforms. Structural diversity of V-ATPase generated by different combinations of subunit isoforms enables it to play diverse physiological roles in mammalian cells. Among these various roles, this review focuses on the functions of lysosome-specific V-ATPase in bone resorption by osteoclasts. Lysosomes remain in the cytoplasm in most cell types, but in osteoclasts, secretory lysosomes move toward and fuse with the plasma membrane to secrete lysosomal enzymes, which is essential for bone resorption. Through this process, lysosomal V-ATPase harboring the a3 isoform of the a subunit is relocated to the plasma membrane, where it transports protons from the cytosol to the cell exterior to generate the acidic extracellular conditions required for secreted lysosomal enzymes. In addition to this role as a proton pump, we recently found that the lysosomal a3 subunit of V-ATPase is essential for anterograde trafficking of secretory lysosomes. Specifically, a3 interacts with Rab7, a member of the Rab guanosine 5'-triphosphatase (GTPase) family that regulates organelle trafficking, and recruits it to the lysosomal membrane. These findings revealed the multifunctionality of lysosomal V-ATPase in osteoclasts; V-ATPase is responsible not only for the formation of the acidic environment by transporting protons, but also for intracellular trafficking of secretory lysosomes by recruiting organelle trafficking factors. Herein, we summarize the molecular mechanism underlying secretory lysosome trafficking in osteoclasts, and discuss the possible regulatory role of V-ATPase in organelle trafficking.


Assuntos
Reabsorção Óssea , ATPases Vacuolares Próton-Translocadoras , Trifosfato de Adenosina/metabolismo , Animais , Reabsorção Óssea/metabolismo , GTP Fosfo-Hidrolases/metabolismo , Guanosina/metabolismo , Humanos , Lisossomos/metabolismo , Mamíferos/metabolismo , Osteoclastos/metabolismo , Isoformas de Proteínas/metabolismo , Prótons , ATPases Vacuolares Próton-Translocadoras/metabolismo
20.
Int J Mol Sci ; 23(20)2022 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-36293046

RESUMO

Pathogenic variants of the gene Eda cause X-linked hypohidrotic ectodermal dysplasia (XLHED), which is characterized by structural abnormalities or lack of ectodermal appendages. Signs of dysplasia are not restricted to derivatives of the ectodermal layer, but mesodermal abnormalities, such as craniofacial dysmorphism, are also frequently observed, suggesting close reciprocal interactions between the ectoderm and mesoderm; however, a causal link has remained unsubstantiated. We investigated the functional impact of defective ectodysplasin A1 (Eda1) signaling on postnatal bone homeostasis in Eda1-deficient Tabby mice. Interestingly, Eda1 was detected in wild-type mouse calvariae throughout postnatal lifetime. In calvariae, bone-lining Osterix (Osx)+ osteoblasts stained positive for Eda1, and osteoclasts were revealed as Eda receptor (Edar)-positive. Moreover, adult Eda1-deficient calvarial bone showed osteopetrosis-like changes with significantly diminished marrow space, which was maintained during adulthood. Concomitantly with osteopetrosis-like changes, Tabby calvarial bone and Tabby bone marrow-derived osteoclasts had far less osteoclastic activity-associated co-enzymes including cathepsin K, Mmp9, Trap, and Tcirg1 (V-type proton ATPase a3 subunit) compared with wild-type calvariae in vivo or osteoclasts in vitro, indicating that Eda1 deficiency may affect the activity of osteoclasts. Finally, we confirmed that nuclear Nfatc1-positive osteoclasts were strongly diminished during mature osteoclastic differentiation under M-CSF and RANKL in the Tabby model, while Fc-EDA treatment of Tabby-derived osteoclasts significantly increased nuclear translocation of Nfatc1. Furthermore, we identified enhanced Nfatc1 and NF-κB transcriptional activity following Fc-EDA treatment in vitro using luciferase assays. Overall, the results indicate that diminished expressions of osteoclastic activity-associated co-enzymes may lead to disturbed bone homeostasis in Tabby calvariae postnatally.


Assuntos
Displasia Ectodérmica Anidrótica Tipo 1 , Osteopetrose , Camundongos , Animais , Ectodisplasinas/genética , Catepsina K/genética , Fator Estimulador de Colônias de Macrófagos , Metaloproteinase 9 da Matriz , NF-kappa B/metabolismo , Osteopetrose/genética , Osteoclastos/metabolismo , Prótons , Luciferases , Crânio/metabolismo , Adenosina Trifosfatases
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