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1.
J Biol Chem ; 280(4): 2498-502, 2005 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-15545262

RESUMO

High bone mass diseases are caused both by activating mutations in the Wnt pathway and by loss of SOST, a bone morphogenetic protein (BMP) antagonist, leading to the activation of BMP signaling. Given the phenotypic similarity between mutations that activate these signaling pathways, it seems likely that BMPs and Wnts operate in parallel or represent components of the same pathway, modulating osteoblast differentiation. In this study, we show that in C3H10T1/2 cells, Wnt-3A and BMP-6 proteins were inducers of osteoblast differentiation, as measured by alkaline phosphatase (ALP) induction. Surprisingly, sclerostin, noggin, and human BMP receptor 1A (BMPR1A)-FC fusion proteins blocked Wnt-3A-induced ALP as well as BMP-6-induced ALP activity. Dkk-1, a Wnt inhibitor, blocked Wnt-induced ALP activity but not BMP-induced ALP activity. Early Wnt-3A signaling as measured by beta-catenin accumulation was not affected by the BMP antagonists but was blocked by Dkk-1. Wnt-3A induced the appearance of BMP-4 mRNA 12 h prior to that of ALP in C3H10T1/2 cells. We propose that sclerostin and other BMP antagonists do not block Wnt signaling directly. Sclerostin blocks Wnt-induced ALP activity by blocking the activity of BMP proteins produced by Wnt treatment. The expression of BMP proteins in this autocrine loop is essential for Wnt-3A-induced osteoblast differentiation.


Assuntos
Proteínas Morfogenéticas Ósseas/farmacologia , Proteínas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Fosfatase Alcalina/metabolismo , Animais , Proteína Morfogenética Óssea 6 , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular , Proteínas do Citoesqueleto/metabolismo , Relação Dose-Resposta a Droga , Marcadores Genéticos , Glicoproteínas , Peptídeos e Proteínas de Sinalização Intercelular , Camundongos , Camundongos Endogâmicos C3H , Modelos Biológicos , Mutação , Osteoblastos/metabolismo , Fenótipo , Proteínas Recombinantes de Fusão/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Fatores de Tempo , Transativadores/metabolismo , Proteínas Wnt , Proteína Wnt3 , Proteína Wnt3A , beta Catenina
2.
Bone ; 35(4): 828-35, 2004 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-15454089

RESUMO

A null mutation in the SOST gene is associated with sclerosteosis, an inherited disorder characterized by a high bone mass phenotype. The protein product of the SOST gene, sclerostin, is a bone morphogenetic protein (BMP) antagonist that decreases osteoblast activity and reduces the differentiation of osteoprogenitors. We sought to delineate the mechanism by which sclerostin modulated osteoblastic function by examining the effects of the protein on differentiating cultures of human mesenchymal stem cells (hMSC). Sclerostin significantly decreased alkaline phosphatase (ALP) activity and the proliferation of hMSC cells. In addition, hMSC cells treated with sclerostin displayed a marked increase in caspase activity. Elevated levels of fragmented histone-associated DNA in these cells were detected by ELISA and by TUNEL staining. Other BMP antagonists including noggin, Chordin, Gremlin, and Twisted gastrulation did not affect caspase activity. The sclerostin-mediated increase in caspase activity was blocked by caspase-1 and caspase-3 inhibitors. Sclerostin-induced changes in ALP activity and the survival of hMSC cells were partially restored by BMP-6, suggesting the involvement of additional growth factors. These findings show that sclerostin selectively controls the apoptosis of bone cells. The ability of sclerostin to interact with important growth factors such as BMPs likely serves as the basis by which it modulates the survival of osteoblasts. By making these growth factors unavailable for cell function, sclerostin promotes the apoptosis of bone cells, providing a novel level of control in the regulation of bone formation.


Assuntos
Apoptose , Proteínas Morfogenéticas Ósseas/metabolismo , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteogênese , Proteínas Adaptadoras de Transdução de Sinal , Fosfatase Alcalina/metabolismo , Apoptose/efeitos dos fármacos , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Proteínas Morfogenéticas Ósseas/genética , Inibidores de Caspase , Caspases/metabolismo , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Regulação da Expressão Gênica , Marcadores Genéticos/genética , Humanos , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Osteogênese/genética
3.
J Biol Chem ; 279(35): 36293-8, 2004 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-15199066

RESUMO

Noggin and sclerostin are bone morphogenetic protein (BMP) antagonists that modulate mitogenic activity through sequestering BMPs. Little is known of the interactions among this class of proteins. We show that recombinant sclerostin and noggin bound to each other with high affinity (K(D) = 2.92 nm). This observation has been extended to naturally expressed noggin and sclerostin from the rat osteosarcoma cell line, ROS 17/2.8, supporting a role for the complex in natural systems. The noggin-sclerostin complex was competitive with BMP binding and mutually attenuated the activity of each BMP antagonist. Collectively, the data demonstrate a novel and exquisite paradigm for the regulation of BMP activity through direct neutralization of the BMP and activation by co-localized BMP antagonist expression. The pleiotrophic nature of noggin and sclerostin represents a novel mechanism for the fine-tuning of BMP activity in bone homeostasis.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Ligação Competitiva , Western Blotting , Proteína Morfogenética Óssea 6 , Osso e Ossos/metabolismo , Proteínas de Transporte , Linhagem Celular , Linhagem Celular Tumoral , Meios de Cultivo Condicionados/farmacologia , Proteínas de Ligação a DNA/metabolismo , Relação Dose-Resposta Imunológica , Ensaio de Imunoadsorção Enzimática , Marcadores Genéticos , Glicoproteínas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Cinética , Camundongos , Camundongos Endogâmicos C3H , Osteossarcoma/metabolismo , Testes de Precipitina , Ligação Proteica , Ratos , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Proteínas Smad , Fatores de Tempo , Transativadores/metabolismo
4.
EMBO J ; 22(23): 6267-76, 2003 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-14633986

RESUMO

There is an unmet medical need for anabolic treatments to restore lost bone. Human genetic bone disorders provide insight into bone regulatory processes. Sclerosteosis is a disease typified by high bone mass due to the loss of SOST expression. Sclerostin, the SOST gene protein product, competed with the type I and type II bone morphogenetic protein (BMP) receptors for binding to BMPs, decreased BMP signaling and suppressed mineralization of osteoblastic cells. SOST expression was detected in cultured osteoblasts and in mineralizing areas of the skeleton, but not in osteoclasts. Strong expression in osteocytes suggested that sclerostin expressed by these central regulatory cells mediates bone homeostasis. Transgenic mice overexpressing SOST exhibited low bone mass and decreased bone strength as the result of a significant reduction in osteoblast activity and subsequently, bone formation. Modulation of this osteocyte-derived negative signal is therapeutically relevant for disorders associated with bone loss.


Assuntos
Desenvolvimento Ósseo/fisiologia , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Proteínas Morfogenéticas Ósseas/fisiologia , Marcadores Genéticos/fisiologia , Osteócitos/fisiologia , Proteínas Adaptadoras de Transdução de Sinal , Fosfatase Alcalina/metabolismo , Animais , Sequência de Bases , Densidade Óssea , Doenças Ósseas Metabólicas/genética , Proteína Morfogenética Óssea 6 , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas Morfogenéticas Ósseas/farmacologia , Linhagem Celular , Clonagem Molecular , Primers do DNA , Marcadores Genéticos/genética , Glicoproteínas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Cinética , Mesoderma/efeitos dos fármacos , Mesoderma/fisiologia , Camundongos , Camundongos Endogâmicos C3H , Camundongos Transgênicos , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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