Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 51
Filtrar
Mais filtros

Bases de dados
Tipo de documento
Intervalo de ano de publicação
1.
Physiol Rev ; 102(1): 379-410, 2022 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-34337974

RESUMO

Osteocytes, former osteoblasts encapsulated by mineralized bone matrix, are far from being passive and metabolically inactive bone cells. Instead, osteocytes are multifunctional and dynamic cells capable of integrating hormonal and mechanical signals and transmitting them to effector cells in bone and in distant tissues. Osteocytes are a major source of molecules that regulate bone homeostasis by integrating both mechanical cues and hormonal signals that coordinate the differentiation and function of osteoclasts and osteoblasts. Osteocyte function is altered in both rare and common bone diseases, suggesting that osteocyte dysfunction is directly involved in the pathophysiology of several disorders affecting the skeleton. Advances in osteocyte biology initiated the development of novel therapeutics interfering with osteocyte-secreted molecules. Moreover, osteocytes are targets and key distributors of biological signals mediating the beneficial effects of several bone therapeutics used in the clinic. Here we review the most recent discoveries in osteocyte biology demonstrating that osteocytes regulate bone homeostasis and bone marrow fat via paracrine signaling, influence body composition and energy metabolism via endocrine signaling, and contribute to the damaging effects of diabetes mellitus and hematologic and metastatic cancers in the skeleton.


Assuntos
Remodelação Óssea/fisiologia , Osteoclastos/citologia , Osteócitos/citologia , Osteogênese/fisiologia , Animais , Reabsorção Óssea/metabolismo , Diferenciação Celular/fisiologia , Humanos
2.
Brief Bioinform ; 23(6)2022 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-36198068

RESUMO

Extrachromosomal circular DNA (eccDNA) of chromosomal origin is found in many eukaryotic species and cell types, including cancer, where eccDNAs with oncogenes drive tumorigenesis. Most studies of eccDNA employ short-read sequencing for their identification. However, short-read sequencing cannot resolve the complexity of genomic repeats, which can lead to missing eccDNA products. Long-read sequencing technologies provide an alternative to constructing complete eccDNA maps. We present a software suite, Construction-based Rolling-circle-amplification for eccDNA Sequence Identification and Location (CReSIL), to identify and characterize eccDNA from long-read sequences. CReSIL's performance in identifying eccDNA, with a minimum F1 score of 0.98, is superior to the other bioinformatic tools based on simulated data. CReSIL provides many useful features for genomic annotation, which can be used to infer eccDNA function and Circos visualization for eccDNA architecture investigation. We demonstrated CReSIL's capability in several long-read sequencing datasets, including datasets enriched for eccDNA and whole genome datasets from cells containing large eccDNA products. In conclusion, the CReSIL suite software is a versatile tool for investigating complex and simple eccDNA in eukaryotic cells.


Assuntos
DNA Circular , Genoma , DNA Circular/genética , DNA/genética , Células Eucarióticas
3.
Haematologica ; 2024 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-38385272

RESUMO

Multiple myeloma (MM) remains incurable due to disease relapse and drug resistance. Notch signals from the tumor microenvironment (TME) confer chemoresistance, but the cellular and molecular mechanisms are not entirely understood. Using clinical and transcriptomic datasets, we found that NOTCH3 is upregulated in CD138+ cells from newly diagnosed MM (NDMM) patients compared to healthy individuals and increased in progression/relapsed MM (PRMM) patients. Further, NDMM patients with high NOTCH3 expression exhibited worse responses to Bortezomib (BOR)-based therapies. Cells of the TME, including osteocytes, upregulated NOTCH3 in MM cells and protected them from apoptosis induced by BOR. NOTCH3 activation (NOTCH3OE) in MM cells decreased BOR anti-MM efficacy and its ability to improve survival in in vivo myeloma models. Molecular analyses revealed that NDMM and PRMM patients with high NOTCH3 exhibit CXCL12 upregulation. TME cells upregulated CXCL12 and activated the CXCR4 pathway in MM cells in a NOTCH3-dependent manner. Moreover, genetic or pharmacologic inhibition of CXCL12 in NOTCH3OE MM cells restored sensitivity to BOR regimes in vitro and in human bones bearing NOTCH3OE MM tumors cultured ex vivo. Our clinical and preclinical data unravel a novel NOTCH3-CXCL12 pro-survival signaling axis in the TME and suggest that osteocytes transmit chemoresistance signals to MM cells.

4.
Curr Osteoporos Rep ; 22(2): 266-272, 2024 04.
Artigo em Inglês | MEDLINE | ID: mdl-38457001

RESUMO

PURPOSE OF REVIEW: To describe the contributions of osteocytes to the lesions in Paget's disease, which are characterized by locally overactive bone resorption and formation. RECENT FINDINGS: Osteocytes, the most abundant cells in bone, are altered in Paget's disease lesions, displaying increased size, decreased canalicular length, incomplete differentiation, and less sclerostin expression compared to controls in both patients and mouse models. Pagetic lesions show increased senescent osteocytes that express RANK ligand, which drives osteoclastic bone resorption. Abnormal osteoclasts in Paget's disease secrete abundant IGF1, which enhances osteocyte senescence, contributing to lesion formation. Recent data suggest that osteocytes contribute to lesion formation in Paget's disease by responding to high local IGF1 released from abnormal osteoclasts. Here we describe the characteristics of osteocytes in Paget's disease and their role in bone lesion formation based on recent results with mouse models and supported by patient data.


Assuntos
Osteíte Deformante , Osteoclastos , Osteócitos , Osteíte Deformante/metabolismo , Osteíte Deformante/patologia , Osteócitos/metabolismo , Osteócitos/patologia , Humanos , Animais , Osteoclastos/metabolismo , Ligante RANK/metabolismo , Reabsorção Óssea/metabolismo , Camundongos , Fator de Crescimento Insulin-Like I/metabolismo , Modelos Animais de Doenças , Senescência Celular
5.
Haematologica ; 2023 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-37981834

RESUMO

Multiple myeloma (MM) is a malignancy of plasma cells whose antibody secretion creates proteotoxic stress relieved by the N-end rule pathway, a proteolytic system that degrades Narginylated proteins in the proteasome. When the proteasome is inhibited, protein cargo is alternatively targeted for autophagic degradation by binding to the ZZ-domain of p62/sequestosome-1. Here, we demonstrate that XRK3F2, a selective ligand for the ZZ-domain, dramatically improved two major responses to the proteasome inhibitor bortezomib by increasing: 1) killing of human MM cells by stimulating both bortezomib mediated apoptosis and necroptosis, a process regulated by p62; and 2) preservation of bone mass by stimulating osteoblasts differentiation and inhibiting osteoclastic bone destruction. Co-administration of bortezomib and XRK3F2 inhibited both branches of the bimodal N-end rule pathway exhibited synergistic anti-MM effects on MM cell lines and CD138+ cells from MM patients, and prevented stromal-mediated MM cell survival. In mice with established human MM, coadministration of bortezomib and XRK3F2 decreased tumor burden and prevented the progression of MM-induced osteolytic disease by inducing new bone formation more effectively than either single agent alone. The results suggest that p62-ZZ ligands enhance the anti-MM efficacy of proteasome inhibitors and can reduce MM morbidity and mortality by improving bone health.

6.
FASEB J ; 36(3): e22196, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35137455

RESUMO

Parathyroid hormone (PTH) signaling downstream of the PTH 1 receptor (Pth1r) results in both bone anabolic and catabolic actions by mechanisms not yet fully understood. In this study, we show that Pth1r signaling upregulates the expression of several components of the Notch pathway and that Notch signals contribute to the catabolic actions of PTH in bone. We found that constitutive genetic activation of PTH receptor signaling in osteocytes (caPth1rOt ) or treatment with PTH daily increased the expression of several Notch ligands/receptors in bone. In contrast, sustained elevation of endogenous PTH did not change Notch components expression. Deletion of the PTH receptor or sclerostin overexpression in osteocytes abolished Notch increases by PTH. Further, deleting the canonical Notch transcription factor Rbpjk in osteocytes decreased bone mass and increased resorption and Rankl expression in caPth1rOt mice. Moreover, pharmacological bone-targeted Notch inhibition potentiated the bone mass gain induced by intermittent PTH by reducing bone resorption and preserving bone formation. Thus, Notch activation lies downstream of anabolic signaling driven by PTH actions in osteocytes, and Notch pharmacological inhibition maximizes the bone anabolic effects of PTH.


Assuntos
Reabsorção Óssea/metabolismo , Osteogênese , Hormônio Paratireóideo/metabolismo , Receptores Notch/metabolismo , Animais , Reabsorção Óssea/genética , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Osteócitos/metabolismo , Receptor Tipo 1 de Hormônio Paratireóideo/metabolismo , Receptores Notch/genética , Transdução de Sinais
7.
Int J Mol Sci ; 24(15)2023 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-37569774

RESUMO

The evidence sustaining the regenerative properties of mesenchymal stem cells' (MSCs) secretome has prompted a paradigm change, where MSCs have shifted from being considered direct contributors to tissue regeneration toward being seen as cell factories for producing biotech medicines. We have previously designed a method to prime MSCs towards osteogenic differentiation by silencing the Wnt/ß-Catenin inhibitor Sfpr1. This approach produces a significant increase in bone formation in osteoporotic mice. In this current work, we set to investigate the contribution of the secretome from the MSCs where Sfrp1 has been silenced, to the positive effect seen on bone regeneration in vivo. The conditioned media (CM) of the murine MSCs line C3H10T1/2, where Sfrp1 has been transiently silenced (CM-Sfrp1), was found to induce, in vitro, an increase in the osteogenic differentiation of this same cell line, as well as a decrease of the expression of the Wnt inhibitor Dkk1 in murine osteocytes ex vivo. A reduction in the RANKL/OPG ratio was also detected ex vivo, suggesting a negative effect of CM-Sfrp1 on osteoclastogenesis. Moreover, this CM significantly increases the mineralization of human primary MSCs isolated from osteoportotic patients in vitro. Proteomic analysis identified enrichment of proteins involved in osteogenesis within the soluble and vesicular fractions of this secretome. Altogether, we demonstrate the pro-osteogenic potential of the secretome of MSCs primmed in this fashion, suggesting that this is a valid approach to enhance the osteo-regenerative properties of MSCs' secretome.


Assuntos
Osteogênese , Proteômica , Humanos , Animais , Camundongos , Osteogênese/genética , Secretoma , Peptídeos e Proteínas de Sinalização Intracelular/farmacologia , Diferenciação Celular/genética
8.
Curr Osteoporos Rep ; 19(3): 247-255, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33818732

RESUMO

PURPOSE OF REVIEW: The goal of this manuscript is to review the current knowledge on the role of osteocytes in cancer in the bone, discuss the potential of osteocytes as a therapeutic target, and propose future research needed to understand the crosstalk between cancer cells and osteocytes in the tumor niche. RECENT FINDINGS: Numerous studies have established that cancer cells manipulate osteocytes to facilitate invasion and tumor progression in bone. Moreover, cancer cells dysregulate osteocyte function to disrupt physiological bone remodeling, leading to the development of bone disease. Targeting osteocytes and their derived factors has proven to effectively interfere with the progression of cancer in the bone and the associated bone disease. Osteocytes communicate with cancer cells and are also part of the vicious cycle of cancer in the bone. Additional studies investigating the role of osteocytes on metastases to the bone and the development of drug resistance are needed.


Assuntos
Doenças Ósseas/patologia , Osteócitos/patologia , Animais , Remodelação Óssea , Progressão da Doença , Humanos , Invasividade Neoplásica/patologia , Transdução de Sinais
9.
FASEB J ; 32(5): 2878-2890, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29401593

RESUMO

Parathyroid hormone (PTH) affects the skeleton by acting on osteocytes (Ots) in bone through yet unclear mechanisms. We report that matrix metalloproteinase 14 (MMP14) expression/activity are increased in bones from mice with genetic constitutive activation (ca) of the PTH receptor 1 (PTH1R) in Ots (caPTH1ROt) and in bones from mice exposed to elevated PTH levels but not in mice lacking [conditional knockout (cKO)] the PTH1R in Ots (cKOPTH1ROt). Furthermore, PTH upregulates MMP14 in human bone cultures and in Ot-enriched bones from floxed control mice but not from cKOPTH1ROt mice. MMP14 activity increases soluble receptor activator of NF-κΒ ligand production, which in turn, stimulates osteoclast differentiation and resorption. Pharmacologic inhibition of MMP14 activity reduced the high bone remodeling exhibited by caPTH1ROt mice or induced by chronic PTH elevation and decreased bone resorption but allowed full stimulation of bone formation induced by PTH injections, thereby potentiating bone gain. Thus, MMP14 is a new member of the intricate gene network activated in Ots by PTH1R signaling that can be targeted to adjust the skeletal responses to PTH in favor of bone preservation.-Delgado-Calle, J., Hancock, B., Likine, E. F., Sato, A. Y., McAndrews, K., Sanudo, C., Bruzzaniti, A., Riancho, J. A., Tonra, J. R., Bellido, T. MMP14 is a novel target of PTH signaling in osteocytes that controls resorption by regulating soluble RANKL production.


Assuntos
Reabsorção Óssea/metabolismo , Metaloproteinase 14 da Matriz/metabolismo , Osteócitos/metabolismo , Hormônio Paratireóideo/metabolismo , Ligante RANK/biossíntese , Transdução de Sinais/fisiologia , Animais , Reabsorção Óssea/genética , Células Cultivadas , Redes Reguladoras de Genes/fisiologia , Metaloproteinase 14 da Matriz/genética , Camundongos , Camundongos Knockout , Osteoclastos/citologia , Osteoclastos/metabolismo , Osteócitos/citologia , Osteogênese/fisiologia , Hormônio Paratireóideo/genética , Ligante RANK/genética , Receptor Tipo 1 de Hormônio Paratireóideo/genética , Receptor Tipo 1 de Hormônio Paratireóideo/metabolismo
10.
Genome Res ; 25(1): 27-40, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25271306

RESUMO

In differentiated cells, aging is associated with hypermethylation of DNA regions enriched in repressive histone post-translational modifications. However, the chromatin marks associated with changes in DNA methylation in adult stem cells during lifetime are still largely unknown. Here, DNA methylation profiling of mesenchymal stem cells (MSCs) obtained from individuals aged 2 to 92 yr identified 18,735 hypermethylated and 45,407 hypomethylated CpG sites associated with aging. As in differentiated cells, hypermethylated sequences were enriched in chromatin repressive marks. Most importantly, hypomethylated CpG sites were strongly enriched in the active chromatin mark H3K4me1 in stem and differentiated cells, suggesting this is a cell type-independent chromatin signature of DNA hypomethylation during aging. Analysis of scedasticity showed that interindividual variability of DNA methylation increased during aging in MSCs and differentiated cells, providing a new avenue for the identification of DNA methylation changes over time. DNA methylation profiling of genetically identical individuals showed that both the tendency of DNA methylation changes and scedasticity depended on nongenetic as well as genetic factors. Our results indicate that the dynamics of DNA methylation during aging depend on a complex mixture of factors that include the DNA sequence, cell type, and chromatin context involved and that, depending on the locus, the changes can be modulated by genetic and/or external factors.


Assuntos
Envelhecimento/genética , Metilação de DNA , DNA/genética , Células-Tronco/citologia , Adolescente , Idoso , Idoso de 80 Anos ou mais , Diferenciação Celular , Células Cultivadas , Criança , Pré-Escolar , Cromatina/genética , Epigênese Genética , Histonas/genética , Humanos , Análise em Microsséries , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Processamento de Proteína Pós-Traducional , Análise de Sequência de DNA , Gêmeos Monozigóticos , Adulto Jovem
11.
Proc Natl Acad Sci U S A ; 112(5): E478-86, 2015 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-25605937

RESUMO

Osteocytes, >90% of the cells in bone, lie embedded within the mineralized matrix and coordinate osteoclast and osteoblast activity on bone surfaces by mechanisms still unclear. Bone anabolic stimuli activate Wnt signaling, and human mutations of components along this pathway underscore its crucial role in bone accrual and maintenance. However, the cell responsible for orchestrating Wnt anabolic actions has remained elusive. We show herein that activation of canonical Wnt signaling exclusively in osteocytes [dominant active (da)ßcat(Ot) mice] induces bone anabolism and triggers Notch signaling without affecting survival. These features contrast with those of mice expressing the same daß-catenin in osteoblasts, which exhibit decreased resorption and perinatal death from leukemia. daßcat(Ot) mice exhibit increased bone mineral density in the axial and appendicular skeleton, and marked increase in bone volume in cancellous/trabecular and cortical compartments compared with littermate controls. daßcat(Ot) mice display increased resorption and formation markers, high number of osteoclasts and osteoblasts in cancellous and cortical bone, increased bone matrix production, and markedly elevated periosteal bone formation rate. Wnt and Notch signaling target genes, osteoblast and osteocyte markers, and proosteoclastogenic and antiosteoclastogenic cytokines are elevated in bones of daßcat(Ot) mice. Further, the increase in RANKL depends on Sost/sclerostin. Thus, activation of osteocytic ß-catenin signaling increases both osteoclasts and osteoblasts, leading to bone gain, and is sufficient to activate the Notch pathway. These findings demonstrate disparate outcomes of ß-catenin activation in osteocytes versus osteoblasts and identify osteocytes as central target cells of the anabolic actions of canonical Wnt/ß-catenin signaling in bone.


Assuntos
Osso e Ossos/metabolismo , Osteócitos/fisiologia , Via de Sinalização Wnt , beta Catenina/metabolismo , Animais , Densidade Óssea , Camundongos , Camundongos Transgênicos
13.
J Bone Miner Metab ; 35(2): 150-160, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27038990

RESUMO

Different model systems using osteoblastic cell lines have been developed to help understand the process of bone formation. Here, we report the establishment of two human osteoblastic cell lines obtained from primary cultures upon transduction of immortalizing genes. The resulting cell lines had no major differences to their parental lines in their gene expression profiles. Similar to primary osteoblastic cells, osteocalcin transcription increased following 1,25-dihydroxyvitamin D3 treatment and the immortalized cells formed a mineralized matrix, as detected by Alizarin Red staining. Moreover, these human cell lines responded by upregulating ALPL gene expression after treatment with the demethylating agent 5-aza-2'-deoxycytidine (AzadC), as shown before for primary osteoblasts. We further demonstrate that these cell lines can differentiate in vivo, using a hydroxyapatite/tricalcium phosphate composite as a scaffold, to produce bone matrix. More importantly, we show that these cells respond to demethylating treatment, as shown by the increase in SOST mRNA levels, the gene encoding sclerostin, upon treatment of the recipient mice with AzadC. This also confirms, in vivo, the role of DNA methylation in the regulation of SOST expression previously shown in vitro. Altogether our results show that these immortalized cell lines constitute a particularly useful model system to obtain further insight into bone homeostasis, and particularly into the epigenetic mechanisms regulating sclerostin production.


Assuntos
Linhagem Celular , Epigênese Genética , Osteoblastos/citologia , Osteogênese , Proteínas Adaptadoras de Transdução de Sinal , Animais , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Calcificação Fisiológica , Calcitriol/farmacologia , Diferenciação Celular , Metilação de DNA , Decitabina , Marcadores Genéticos/genética , Humanos , Camundongos , Camundongos Nus , Osteocalcina/genética , Osteocalcina/metabolismo , Alicerces Teciduais
14.
Curr Osteoporos Rep ; 15(6): 532-541, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28956252

RESUMO

PURPOSE OF REVIEW: This review provides a summary of the current knowledge on Sost/sclerostin in cancers targeting the bone, discusses novel observations regarding its potential as a therapeutic approach to treat cancer-induced bone loss, and proposes future research needed to fully understand the potential of therapeutic approaches that modulate sclerostin function. RECENT FINDINGS: Accumulating evidence shows that sclerostin expression is dysregulated in a number of cancers that target the bone. Further, new findings demonstrate that pharmacological inhibition of sclerostin in preclinical models of multiple myeloma results in a robust prevention of bone loss and preservation of bone strength, without apparent effects on tumor growth. These data raise the possibility of targeting sclerostin for the treatment of cancer patients with bone metastasis. Sclerostin is emerging as a valuable target to prevent the bone destruction that accompanies the growth of cancer cells in the bone. Further studies will focus on combining anti-sclerostin therapy with tumor-targeted agents to achieve both beneficial skeletal outcomes and inhibition of tumor progression.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Neoplasias Ósseas/metabolismo , Mieloma Múltiplo/metabolismo , Neoplasias/patologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/secundário , Marcadores Genéticos , Humanos , Terapia de Alvo Molecular , Mieloma Múltiplo/tratamento farmacológico
15.
J Transl Med ; 14(1): 207, 2016 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-27393146

RESUMO

BACKGROUND: Age-associated changes in genomic DNA methylation have been primarily attributed to 5-methylcytosine (5mC). However, the recent discovery of 5-hydroxymethylcytosine (5hmC) suggests that this epigenetic mark might also play a role in the process. METHODS: Here, we analyzed the genome-wide profile of 5hmc in mesenchymal stem cells (MSCs) obtained from bone-marrow donors, aged 2-89 years. RESULTS: We identified 10,685 frequently hydroxymethylated CpG sites in MSCs that were, as in other cell types, significantly associated with low density CpG regions, introns, the histone posttranslational modification H3k4me1 and enhancers. Study of the age-associated changes to 5hmC identified 785 hyper- and 846 hypo-hydroxymethylated CpG sites in the MSCs obtained from older individuals. CONCLUSIONS: DNA hyper-hydroxymethylation in the advanced-age group was associated with loss of 5mC, which suggests that, at specific CpG sites, this epigenetic modification might play a role in DNA methylation changes during lifetime. Since bone-marrow MSCs have many clinical applications, and the fact that the epigenomic alterations in this cell type associated with aging identified in this study could have associated functional effects, the age of donors should be taken into account in clinical settings.


Assuntos
5-Metilcitosina/análogos & derivados , Envelhecimento/genética , Células da Medula Óssea/citologia , Metilação de DNA/genética , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , 5-Metilcitosina/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Cromatina/metabolismo , Ilhas de CpG/genética , Genoma Humano , Genômica , Humanos , Pessoa de Meia-Idade , Adulto Jovem
16.
Calcif Tissue Int ; 96(1): 30-7, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25432767

RESUMO

Osteoporosis causes important morbidity among elderly individuals. Fragility fractures, and especially hip fractures, have a particularly negative impact on the patients' quality of life. The role of epigenetic mechanisms in the pathogenesis of many disorders is increasingly recognized, yet little is known about their role in non-malignant bone disorders such as osteoporosis. The aim of this study was to explore the expression of miRNAs in patients with osteoporotic hip fractures. Trabecular bone samples were obtained from the femoral heads of patients undergoing replacement surgery for osteoporotic hip fractures and non-fracture controls with hip osteoarthritis. Levels of 760 miRNA were analyzed by real-time PCR. Thirteen miRNAs showed nominally significant (p < 0.05) differences between both groups. Six miRNAs (miR-187, miR-193a-3p, miR-214, miR518f, miR-636, and miR-210) were selected for the replication stage. These miRNAs were individually analyzed in a larger group of 38 bone samples. At this stage, we confirmed statistically significant differences across groups for mir-187 and miR-518f. The median relative expression levels of miR-187 were 5.3-fold higher in the non-fracture group (p = 0.002). On the contrary, miR-518f was preferentially expressed in bones from osteoporotic patients (8.6-fold higher in fractures; p = 0.046). In this first hypothesis-free study of the bone microRNome we found two miRNAs, miR-187, and miR-518f, differentially regulated in osteoporotic bone. Further studies are needed to elucidate the mechanisms involved in the association of these miRNAs with fractures.


Assuntos
Osso e Ossos/metabolismo , Fraturas do Quadril/metabolismo , MicroRNAs/metabolismo , Osteoporose/metabolismo , Fraturas por Osteoporose/metabolismo , Idoso , Idoso de 80 Anos ou mais , Densidade Óssea/fisiologia , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Qualidade de Vida
17.
Calcif Tissue Int ; 94(4): 414-22, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24322886

RESUMO

Mechanical stimulation reduces sclerostin expression in rodents. However, few data are available about the effect of physical stimuli in human systems. Recently we observed that the demethylating agent AzadC induces SOST expression in bone cells. This allowed us in this study to explore the effect of mechanical loading on SOST expression by subjecting AzadC-treated human bone cells to pulsating fluid flow (PFF). PFF significantly decreased the AzadC-induced expression of SOST. This effect persisted for at least 24 h, and in fact SOST expression was lower at 24 h after PFF treatment than at 1 h after PFF treatment (PFF/static ratio 0.47 ± 0.04 vs. 0.63 ± 0.03 respectively, p = 0.03). The PFF-induced decrease in SOST expression was not due to a change in the methylation profile of the SOST promoter. However, PFF stimulated nitric oxide (NO) synthesis, which appeared essential for the PFF effect on SOST expression. In fact, the NO synthase inhibitor 1400 W prevented the effect of PFF on SOST expression. Moreover, the NO-donor SNAP decreased SOST mRNA in bone organ cultures. The conditioned medium (CM) of cells subjected to PFF induced a 38 ± 4 % decrease in SOST expression (p = 0.03) in static cultures and diminished the transcriptional activity of reporter vectors with the cloned SOST promoter (Static-CM: 1.47 ± 0.10 vs. PFF-CM: 0.78 ± 0.09, p = 0.02). This is consistent with a PFF-induced secretion of factors that modulate SOST. Our results suggest that NO and other soluble factors are involved in the inhibition of SOST expression by PFF.


Assuntos
Azacitidina/análogos & derivados , Proteínas Morfogenéticas Ósseas/metabolismo , Regulação para Baixo , Óxido Nítrico/química , Estresse Mecânico , Proteínas Adaptadoras de Transdução de Sinal , Azacitidina/química , Osso e Ossos/metabolismo , Linhagem Celular , Meios de Cultivo Condicionados/química , Metilação de DNA , Decitabina , Marcadores Genéticos , Humanos , Osteoblastos/efeitos dos fármacos , Fluxo Pulsátil , Transdução de Sinais , Transcrição Gênica , Transfecção
18.
Calcif Tissue Int ; 95(2): 194-9, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24913258

RESUMO

Sclerostin, the product of the SOST gene, is a key regulator of bone homeostasis. Sclerostin interferes with the Wnt signalling pathway and, therefore, has a negative effect on bone formation. Although the importance of sclerostin in bone homeostasis is well established, many aspects of its biology are still unknown. Due to its restricted pattern of expression, in vitro studies of SOST gene regulation are technically challenging. Furthermore, a more profound investigation of the molecular mechanism controlling sclerostin expression has been hampered by the lack of a good human in vitro model. Here, we describe two cell lines derived from the human osteosarcoma cell line SaOS-2 that produce elevated levels of sclerostin. Analysis of the super-producer cell lines showed that sclerostin levels were still reduced in response to parathyroid hormone treatment or in response to mechanical loading, indicating that these regulatory mechanisms were not affected in the presented cell lines. In addition, we did not find differences between the promoter or ECR5 sequences of our clones and the SaOS-2 parental line. However, the methylation of the proximal CpG island located at the SOST promoter was lower in the super-producer clones, in agreement with a higher level of SOST transcription. Although the underlying biological causes of the elevated levels of sclerostin production in this cell line are not yet clear, we believe that it could be an extremely useful tool to study the molecular mechanisms driving sclerostin expression in humans.


Assuntos
Proteínas Morfogenéticas Ósseas/biossíntese , Linhagem Celular Tumoral/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Marcadores Genéticos , Humanos , Immunoblotting , Análise de Sequência com Séries de Oligonucleotídeos , Osteogênese/fisiologia , Osteossarcoma , Transcriptoma
19.
Arthritis Rheum ; 65(1): 197-205, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23124911

RESUMO

OBJECTIVE: To determine genome-wide methylation profiles of bone from patients with hip osteoarthritis (OA) and those with osteoporotic (OP) hip fractures. METHODS: Trabecular bone pieces were obtained from the central part of the femoral head of 27 patients with hip fractures and 26 patients with hip OA. DNA was isolated, and methylation was explored with Illumina methylation arrays. RNA was extracted, pooled, and deep-sequenced to obtain the whole transcriptome. Differentially methylated regions were identified, and connections between genes with differentially methylated regions were explored by pathway and text-mining analyses. RESULTS: After quality control, methylation of 23,367 CpG sites (13,463 genes) was analyzed. There was a genome-wide inverse relationship between methylation and gene expression in both patient groups. Comparison of OP and OA bones revealed 241 CpG sites, located in 228 genes, with significant differences in methylation (false discovery rate<0.05). Of them, 217 were less methylated in OP than in OA. The absolute methylation differences were >5% in 128 CpG sites and >10% in 45 CpG sites. The differentially methylated genes were enriched for association with bone traits in the genome-wide association study catalog. Pathway analysis and text-mining analysis with Gene Relationships Across Implicated Loci software revealed enrichment in genes participating in glycoprotein metabolism or cell differentiation, and particularly in the homeobox superfamily of transcription factors. CONCLUSION: Genome-wide methylation profiling of bone samples revealed differentially methylated regions in OP and OA. These regions were enriched in genes associated with cell differentiation and skeletal embryogenesis, such as those in the homeobox superfamily, suggesting the existence of a developmental component in the predisposition to these disorders.


Assuntos
Osso e Ossos/metabolismo , Metilação de DNA , Osteoartrite do Quadril/genética , Osteoporose/genética , Fraturas por Osteoporose/genética , Idoso , Idoso de 80 Anos ou mais , Feminino , Perfilação da Expressão Gênica , Estudo de Associação Genômica Ampla , Humanos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Osteoartrite do Quadril/metabolismo , Osteoporose/metabolismo , Fraturas por Osteoporose/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de DNA
20.
Res Sq ; 2024 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-38558984

RESUMO

Breast cancer bone metastases increase fracture risk and are a major cause of morbidity and mortality among women. Upon colonization by tumor cells, the bone microenvironment undergoes profound reprogramming to support cancer progression that disrupts the balance between osteoclasts and osteoblasts, leading to bone lesions. Whether such reprogramming affects matrix-embedded osteocytes remains poorly understood. Here, we demonstrate that osteocytes in breast cancer bone metastasis develop premature senescence and a distinctive senescence-associated secretory phenotype (SASP) that favors bone destruction. Single-cell RNA sequencing identified osteocytes from mice with breast cancer bone metastasis enriched in senescence and SASP markers and pro-osteoclastogenic genes. Using multiplex in situ hybridization and AI-assisted analysis, we detected osteocytes with senescence-associated distension of satellites, telomere dysfunction, and p16Ink4a expression in mice and patients with breast cancer bone metastasis. In vitro and ex vivo organ cultures showed that breast cancer cells promote osteocyte senescence and enhance their osteoclastogenic potential. Clearance of senescent cells with senolytics suppressed bone resorption and preserved bone mass in mice with breast cancer bone metastasis. These results demonstrate that osteocytes undergo pathological reprogramming by breast cancer cells and identify osteocyte senescence as an initiating event triggering bone destruction in breast cancer metastases.

SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA