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1.
J Pathol ; 255(3): 330-342, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34357595

RESUMO

Chondrocytes in mice developing osteoarthritis (OA) exhibit an aberrant response to the secreted cytokine transforming growth factor (TGF)-ß, consisting in a potentiation of intracellular signaling downstream of the transmembrane type I receptor kinase activin receptor-like kinase (ALK)1 against canonical TGF-ß receptor ALK5-mediated signaling. Unfortunately, the underlying mechanisms remain elusive. In order to identify novel druggable targets for OA, we aimed to investigate novel molecules regulating the ALK1/ALK5 balance in OA chondrocytes. We performed gene expression analysis of TGF-ß signaling modulators in joints from three different mouse models of OA and found an upregulated expression of the TGF-ß co-receptor Cripto (Tdgf1), which was validated in murine and human cartilage OA samples at the protein level. In vitro and ex vivo, elevated expression of Cripto favors the hypertrophic differentiation of chondrocytes, eventually contributing to tissue calcification. Furthermore, we found that Cripto participates in a TGF-ß-ALK1-Cripto receptor complex in the plasma membrane, thereby inducing catabolic SMAD1/5 signaling in chondrocytes. In conclusion, we demonstrate that Cripto is expressed in OA and plays a functional role promoting chondrocyte hypertrophy, thereby becoming a novel potential therapeutic target in OA, for which there is no efficient cure or validated biomarker. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.


Assuntos
Condrócitos/patologia , Proteínas Ligadas por GPI/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Neoplasias/metabolismo , Osteoartrite/patologia , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Fator de Crescimento Epidérmico/metabolismo , Humanos , Hipertrofia/patologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Transdução de Sinais/fisiologia
2.
Mol Cell ; 51(5): 559-72, 2013 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-23973329

RESUMO

TGF-ß signaling is a therapeutic target in advanced cancers. We identified tumor necrosis factor receptor-associated factor 4 (TRAF4) as a key component mediating pro-oncogenic TGF-ß-induced SMAD and non-SMAD signaling. Upon TGF-ß stimulation, TRAF4 is recruited to the active TGF-ß receptor complex, where it antagonizes E3 ligase SMURF2 and facilitates the recruitment of deubiquitinase USP15 to the TGF-ß type I receptor (TßRI). Both processes contribute to TßRI stabilization on the plasma membrane and thereby enhance TGF-ß signaling. In addition, the TGF-ß receptor-TRAF4 interaction triggers Lys 63-linked TRAF4 polyubiquitylation and subsequent activation of the TGF-ß-activated kinase (TAK)1. TRAF4 is required for efficient TGF-ß-induced migration, epithelial-to-mesenchymal transition, and breast cancer metastasis. Elevated TRAF4 expression correlated with increased levels of phosphorylated SMAD2 and phosphorylated TAK1 as well as poor prognosis among breast cancer patients. Our results demonstrate that TRAF4 can regulate the TGF-ß pathway and is a key determinant in breast cancer pathogenesis.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator 4 Associado a Receptor de TNF/genética , Fator 4 Associado a Receptor de TNF/metabolismo , Animais , Linhagem Celular Tumoral , Movimento Celular , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , MAP Quinase Quinase Quinases/metabolismo , Camundongos , Fosforilação , Poliubiquitina/metabolismo , Prognóstico , Proteínas Serina-Treonina Quinases/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Transdução de Sinais , Proteína Smad2/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Ubiquitina-Proteína Ligases/metabolismo
3.
J Pathol ; 247(1): 9-20, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30246251

RESUMO

Bone morphogenetic proteins (BMPs) are secreted cytokines that were initially discovered on the basis of their ability to induce bone. Several decades of research have now established that these proteins function in a large variety of physiopathological processes. There are about 15 BMP family members, which signal via three transmembrane type II receptors and four transmembrane type I receptors. Mechanistically, BMP binding leads to phosphorylation of the type I receptor by the type II receptor. This activated heteromeric complex triggers intracellular signaling that is initiated by phosphorylation of receptor-regulated SMAD1, 5, and 8 (also termed R-SMADs). Activated R-SMADs form heteromeric complexes with SMAD4, which engage in specific transcriptional responses. There is convergence along the signaling pathway and, besides the canonical SMAD pathway, BMP-receptor activation can also induce non-SMAD signaling. Each step in the pathway is fine-tuned by positive and negative regulation and crosstalk with other signaling pathways. For example, ligand bioavailability for the receptor can be regulated by ligand-binding proteins that sequester the ligand from interacting with receptors. Accessory co-receptors, also known as BMP type III receptors, lack intrinsic enzymatic activity but enhance BMP signaling by presenting ligands to receptors. In this review, we discuss the role of BMP receptor signaling and how corruption of this pathway contributes to cardiovascular and musculoskeletal diseases and cancer. We describe pharmacological tools to interrogate the function of BMP receptor signaling in specific biological processes and focus on how these agents can be used as drugs to inhibit or activate the function of the receptor, thereby normalizing dysregulated BMP signaling. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.


Assuntos
Receptores de Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas Morfogenéticas Ósseas/metabolismo , Doenças Cardiovasculares/metabolismo , Doenças Musculoesqueléticas/metabolismo , Neoplasias/metabolismo , Transdução de Sinais , Animais , Receptores de Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/genética , Doenças Cardiovasculares/genética , Doenças Cardiovasculares/patologia , Doenças Cardiovasculares/fisiopatologia , Humanos , Ligantes , Doenças Musculoesqueléticas/genética , Doenças Musculoesqueléticas/patologia , Doenças Musculoesqueléticas/fisiopatologia , Neoplasias/genética , Neoplasias/patologia , Neoplasias/fisiopatologia , Fosforilação , Proteínas Smad Reguladas por Receptor/metabolismo
4.
J Pathol ; 247(3): 333-346, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30430573

RESUMO

Endothelial-to-mesenchymal transition (EndMT) has been unveiled as a common cause for a multitude of human pathologies, including cancer and cardiovascular disease. Vascular calcification is a risk factor for ischemic vascular disorders and slowing calcification may reduce mortality in affected patients. The absence of early biomarkers hampers the identification of patients at risk. EndMT and vascular calcification are induced upon cooperation between distinct stimuli, including inflammatory cytokines and transforming growth factor beta (TGF-ß) family members. However, how these signaling pathways interplay to promote cell differentiation and eventually vascular calcification is not well understood. Using in vitro and ex vivo analysis in animal models and patient-derived tissues, we have identified that the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1ß) induce EndMT in human primary aortic endothelial cells, thereby sensitizing them for BMP-9-induced osteogenic differentiation. Downregulation of the BMP type II receptor BMPR2 is a key event in this process. Rather than compromising BMP canonical signal transduction, loss of BMPR2 results in decreased JNK signaling in ECs, thus enhancing BMP-9-induced mineralization. Altogether, our results point at the BMPR2-JNK signaling axis as a key pathway regulating inflammation-induced EndMT and contributing to calcification. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.


Assuntos
Receptores de Proteínas Morfogenéticas Ósseas Tipo II/fisiologia , Transição Epitelial-Mesenquimal/fisiologia , Calcificação Vascular/fisiopatologia , Animais , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Células Endoteliais/fisiologia , Endotélio Vascular/patologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Mediadores da Inflamação/farmacologia , Interleucina-1beta/farmacologia , Camundongos Endogâmicos C3H , Osteogênese/efeitos dos fármacos , Osteogênese/fisiologia , Transdução de Sinais/fisiologia , Fator de Necrose Tumoral alfa/farmacologia , Calcificação Vascular/patologia
5.
Dev Dyn ; 247(3): 492-508, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-28891150

RESUMO

The process named endothelial-to-mesenchymal transition (EndMT) was observed for the first time during the development of the chicken embryo several decades ago. Of interest, accumulating evidence suggests that EndMT plays a critical role in the onset and progression of multiple postnatal cardiovascular diseases. EndMT is controlled by a set of developmental signaling pathways, very similar to the process of epithelial-to-mesenchymal transition, which determine the activity of several EndMT transcriptional effectors. Once activated, these EndMT effectors regulate the expression of endothelial- and mesenchymal-specific genes, in part by interacting with specific motifs in promoter regions, eventually leading to the down-regulation of endothelial-specific features and acquisition of a fibroblast-like phenotype. Important technical advances in lineage tracing methods combined with experimental mouse models demonstrated the pathophysiological importance of EndMT for human diseases. In this review, we discuss the major signal transduction pathways involved in the activation and regulation of the EndMT program. Furthermore, we will review the latest discoveries on EndMT, focusing on cardiovascular diseases, and in particular on its role in vascular calcification, pulmonary arterial hypertension, and organ fibrosis. Developmental Dynamics 247:492-508, 2018. © 2017 Wiley Periodicals, Inc.


Assuntos
Doenças Cardiovasculares/fisiopatologia , Células Endoteliais/patologia , Células-Tronco Mesenquimais/patologia , Transdução de Sinais/fisiologia , Animais , Calcinose/etiologia , Fibrose/etiologia , Humanos
6.
Ren Fail ; 39(1): 629-642, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28805484

RESUMO

Uncontrolled activation of transforming growth factor beta (TGF-ß) family members is hypothesized to participate in type 2 diabetes (T2D) dependent diabetic nephropathy (DN). We evaluated and compared downstream activation of the Smad2-signaling pathway in kidney samples from T2D patients to kidneys from the T2D model of leptin receptor deficient db/db mouse. Furthermore, expression of TGF-ß family members was evaluated to elucidate molecular mechanisms in the mouse model. Kidney samples from patients with advanced stages of DN showed elevated pSmad2 staining whereas db/db mouse kidneys surprisingly showed a decrease in pSmad2 in the tubular compartment. Structurally, kidney tissue showed dilated tubules and expanded glomeruli, but no clear fibrotic pattern was found in the diabetic mice. Selective TGF-ß family members were up-regulated at the mRNA level. Antagonists of bone morphogenetic protein (BMP) ligands, such as Gremlin1, USAG1 and Sclerostin, were strongly up-regulated suggesting a dampening effect on BMP pathways. Together, these results indicate a lack of translation from T2D patient kidneys to the db/db model with regards to Smad signaling pathway. It is plausible that a strong up-regulation of BMP antagonizing factors account for the lack of Smad1/5/8 activation, in spite of increased expression of several BMP members.


Assuntos
Diabetes Mellitus Tipo 2/complicações , Nefropatias Diabéticas/patologia , Glomérulos Renais/patologia , Túbulos Renais/patologia , Proteína Smad2/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Proteínas Morfogenéticas Ósseas/metabolismo , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patologia , Nefropatias Diabéticas/etiologia , Modelos Animais de Doenças , Feminino , Fibrose , Glicoproteínas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Fosforilação , RNA Mensageiro/metabolismo , Receptores para Leptina/genética , Transdução de Sinais , Fator de Crescimento Transformador beta1 , Regulação para Cima
7.
JBMR Plus ; 3(11): e10230, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31768489

RESUMO

Fibrodysplasia ossificans progressiva (FOP) is an extremely rare congenital form of heterotopic ossification (HO), caused by heterozygous mutations in the activin A type I receptor (ACVR1), that encodes the bone morphogenetic protein (BMP) type I receptor ALK2. These mutations enable ALK2 to induce downstream signaling in response to activins, thereby turning them into bone-inducing agents. To date, there is no cure for FOP. The further development of FOP patient-derived models may contribute to the discovery of novel biomarkers and therapeutic approaches. Nevertheless, this has traditionally been a challenge, as biopsy sampling often triggers HO. We have characterized peripheral blood-derived endothelial colony-forming cells (ECFCs) from three independent FOP donors as a new model for FOP. FOP ECFCs are prone to undergo endothelial-to-mesenchymal transition and exhibit increased ALK2 downstream signaling and subsequent osteogenic differentiation upon stimulation with activin A. Moreover, we have identified a new class of small molecule macrocycles with potential activity against ALK2 kinase. Finally, using FOP ECFCs, we have selected OD36 and OD52 as potent inhibitors with excellent kinase selectivity profiles that potently antagonize mutant ALK2 signaling and osteogenic differentiation. We expect that these results will contribute to the development of novel ALK2 clinical candidates for the treatment of FOP. © 2019 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

8.
Arthritis Res Ther ; 19(1): 112, 2017 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-28569204

RESUMO

BACKGROUND: Dysregulated transforming growth factor ß (TGFß) signaling is implicated in osteoarthritis development, making normalizing TGFß signaling a possible therapy. Theoretically, this can be achieved with small molecule inhibitors specifically targeting the various TGFß receptors and downstream mediators. In this study we explore in primary chondrocytes the use of small molecule inhibitors to target TGFß-induced pSmad1/5/9-, pSmad2/3- and TGFß-activated kinase 1 (TAK1)-dependent signaling. METHOD: Primary bovine chondrocytes and explants were isolated from metacarpophalangeal joints. To modulate TGFß signaling the activin receptor-like kinase (ALK)1/2/3/6 inhibitor LDN-193189, the ALK4/5/7 inhibitor SB-505124 and the TAK1 inhibitor (5Z)-7-Oxozeaenol were used. pSmad1/5 and pSmad2 were measured using western blot analysis and TGFß1-induced gene expression was measured using quantitative real time PCR (qPCR). RESULTS: In chondrocytes, TGFß1 strongly induced both pSmad1/5 and pSmad2. Remarkably, LDN-193189 did not inhibit TGFß-induced pSmad1/5. In contrast, SB-505124 did inhibit both TGFß-induced Smad2 and Smad1/5 phosphorylation. Furthermore, (5Z)-7-Oxozeaenol also profoundly inhibited TGFß-induced pSmad2 and pSmad1/5. Importantly, both SB-505124 and (5Z)-7-Oxozeaenol did not significantly inhibit constitutively active ALK1, making an off-target effect unlikely. Additionally, LDN-193189 was able to potently inhibit BMP2/7/9-induced pSmad1/5, showing its functionality. On gene expression, LDN-193189 did not affect TGFß1-induced regulation, whereas both SB-505124 and (5Z)-7-Oxozeaenol did. Similar results were obtained in cartilage explants, although pSmad1/5 was not strongly induced by addition of TGFß1. CONCLUSION: Our data suggest that ALK5 kinase activity plays a central role in both TGFß-induced Smad1/5 and Smad2/3 phosphorylation, making it difficult to separate both pathways with the use of currently available small molecule inhibitors. Furthermore, our data regarding (5Z)-7-Oxozeaenol suggest that TAK1 facilitates Smad-dependent signaling.


Assuntos
Condrócitos/metabolismo , Inibidores Enzimáticos/farmacologia , MAP Quinase Quinase Quinases/metabolismo , Osteoartrite/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Proteínas Smad/metabolismo , Animais , Benzodioxóis/farmacologia , Bovinos , Condrócitos/efeitos dos fármacos , Imidazóis/farmacologia , Fosforilação , Pirazóis/farmacologia , Piridinas/farmacologia , Pirimidinas/farmacologia , Receptor do Fator de Crescimento Transformador beta Tipo I , Fator de Crescimento Transformador beta1/metabolismo , Zearalenona/análogos & derivados , Zearalenona/farmacologia
9.
Cytokine Growth Factor Rev ; 27: 65-79, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26823333

RESUMO

The vascular system is critical for developmental growth, tissue homeostasis and repair but also for tumor development. Bone morphogenetic protein (BMP) signaling has recently emerged as a fundamental pathway of the endothelium by regulating cardiovascular and lymphatic development and by being causative for several vascular dysfunctions. Two vascular disorders have been directly linked to impaired BMP signaling: pulmonary arterial hypertension and hereditary hemorrhagic telangiectasia. Endothelial BMP signaling critically depends on the cellular context, which includes among others vascular heterogeneity, exposure to flow, and the intertwining with other signaling cascades (Notch, WNT, Hippo and hypoxia). The purpose of this review is to highlight the most recent findings illustrating the clear need for reconsidering the role of BMPs in vascular biology.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Endotélio Linfático/metabolismo , Endotélio Vascular/metabolismo , Transdução de Sinais , Telangiectasia Hemorrágica Hereditária/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/genética , Humanos , Hipertensão/genética , Hipertensão/metabolismo , Telangiectasia Hemorrágica Hereditária/genética
10.
Expert Opin Investig Drugs ; 22(11): 1371-83, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24053899

RESUMO

INTRODUCTION: Formation of blood vessels from pre-existing ones, also termed angiogenesis, is of crucial importance for the outgrowth of tumours beyond 1 - 2 mm³. Therefore, anti-angiogenic therapies, mainly focussing on inhibition of vascular endothelial growth factor (VEGF) are used in clinical therapy. However, although initially reducing tumour size, therapy resistance occurs frequently and new targets are needed. A possible target is activin receptor-like kinase (ALK)-1, a transforming growth factor (TGF)-ß type-I receptor, which binds bone morphogenetic protein (BMP)-9 and -10 with high affinity and has an important role in regulating angiogenesis. AREAS COVERED: Several approaches to interfere with ALK1 signalling have been developed, that is, ALK1 neutralising antibodies and a soluble ALK1 extracellular domain/Fc fusion protein (ALK1-Fc), acting as a ligand trap. In this review, we discuss the involvement of ALK1 in angiogenesis, in a variety of diseases and the current status of the development of ALK1 inhibitors for cancer therapy. EXPERT OPINION: Based on current, mainly preclinical studies on inhibition of ALK1 signalling by ligand traps and neutralising antibodies, targeting ALK1 seems very promising. Both ALK1-Fc and neutralising antibodies strongly inhibit angiogenesis in vitro and in vivo. The results from the first Phase I clinical trials are to be reported soon and multiple Phase II studies are ongoing.


Assuntos
Receptores de Activinas Tipo II/antagonistas & inibidores , Inibidores da Angiogênese/uso terapêutico , Neoplasias/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Receptores de Activinas Tipo II/metabolismo , Animais , Humanos , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia
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