Background: Ewing sarcoma (ES) is a developmental bone/soft tissue neoplasia, for which mesenchymal stem cell (MSC) is the putative cell of origin. Although the outcome of ES patients bearing primary tumors has improved significantly,... more
Background: Ewing sarcoma (ES) is a developmental bone/soft tissue neoplasia, for which mesenchymal stem cell (MSC) is the putative cell of origin. Although the outcome of ES patients bearing primary tumors has improved significantly, patients with metastatic ES still have a poor prognosis. The MSC marker endoglin/CD105 (ENG) and the ENG-shedding protease MMP14 have been, respectively, described as cell migration and invasion regulators. In our attempt to better understand the mechanism of ES progression, we are focusing on the role of ENG and MMP14 in ES cells. Methods: In vitro ENG-knockdown (ENG-KD; RM82 and SKNMC cell lines) and ENG-overexpressing (pENG; TC71 cell lines) models were generated by shRNA technology and pDisplay vectors, respectively, and confirmed by qRT-PCR, western blotting and flow cytometry. In vitro MMP14-knockout (MMP14-KO; RM82 and SKNMC) models were generated by CRISPR-Cas9 and confirmed by western blotting and Sanger sequencing. Transwell migration/invasion, adhesion, clonogenicity and proliferation assays were performed in vitro. Actin filament organization was evaluated by immunofluorescence. Extracellular vesicles (EVs) were isolated by ultracentrifugation and density gradient separation. Transcriptomic profiling was performed by Clarion S Human arrays followed by Gene Set Enrichment Analysis. HiPathia tool was used to recode the transcriptomic data into measurements of cell activity by applying mechanistic models. Reverse Phase Protein Array (RPPA) analysis was evaluated on the ENG-KD RM82 model. ENG-targeting immunoprecipitations (IP) from wild type RM82 protein extracts were analyzed by mass spectrometry (MS). Results: The impaired clonogenic potential of ENG-KD RM82 and SKNMC models suggests a role of ENG on the stemness capacity of ES, without affecting the proliferation rate. The reduced migratory and adhesive phenotype of the ENG-KD RM82 model and the transcriptomic profiling of both ENG-KD RM82 and SKNMC cells, suggest a role of ENG in ES cell migration and adhesion. The altered organization of actin filaments in ENG-KD and pENG models could support a role of ENG in actin filament remodeling. The transcriptomic profiling of ENG-KD models and the RPPA results underscore the regulation of FAK signaling by ENG. The IP-MS analysis of ENG partners identifies Integrin-Linked Kinase-Associated serine/threonine Phosphatase (ILKAP) as one of the main interactors of ENG in RM82 cells, suggesting its possible association with the focal adhesion member ILK. ENG is enriched in EVs from different ES cell lines. The transcriptomic profiling of MMP14-KO SKNMC cells highlights the involvement of MMP14 activity in the extracellular matrix organization of ES cells. Conclusion: ENG favors a pro-migratory phenotype in ES cells. These findings help to better understand the molecular landscape of tumoral cells from ES patients, and might have potential clinical implications. Citation Format: Pilar Puerto-Camacho, Ana Teresa Amaral, Juan Diaz-Martin, Alfonso Bolado-Carrancio, Carmen Jordan-Perez, Carmen Salguero-Aranda, Marina Esteban-Medina, Inmaculada Alamo-Alvarez, Joaquin Dopazo, Carmelo Bernabeu, Adam Byron, Valerie G. Brunton, Enrique de Álava. The relevance of endoglin and MMP14 to the metastatic potential of Ewing sarcoma cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6158.
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Research Interests: Immunology, Biology, Cancer Research, Medicine, Signal Transduction, and 10 moreHumans, Female, Clinical Sciences, Public health systems and services research, Transforming Growth Factor Beta, Transforming Growth Factor, fibroblasts, vascular cell adhesion molecule 1, Endoglin, and In Vitro Techniques
Abstract Endoglin is an integral membrane glycoprotein that binds transforming growth factor-β1 (TGF-β1) with high affinity and it is strongly expressed on syncytiotrophoblasts throughout pregnancy. Here, we describe the expression of... more
Abstract Endoglin is an integral membrane glycoprotein that binds transforming growth factor-β1 (TGF-β1) with high affinity and it is strongly expressed on syncytiotrophoblasts throughout pregnancy. Here, we describe the expression of endoglin by the ...
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CD105 (endoglin) is an important component of the transforming growth factor-beta (TGF-beta) receptor complex and is highly expressed in endothelial cells in tissues undergoing angiogenesis such as healing wounds, infarcts and in a wide... more
CD105 (endoglin) is an important component of the transforming growth factor-beta (TGF-beta) receptor complex and is highly expressed in endothelial cells in tissues undergoing angiogenesis such as healing wounds, infarcts and in a wide range of tumours. In an attempt to understand the molecular mechanism by which CD105 exerts its effects on angiogenesis by modulating TGF-beta1 signalling, in this preliminary communication, CD105 transfected rat myoblasts were utilized as an in vitro model. Overexpression of CD105 in these transfectants antagonised TGF-beta1-mediated inhibition of cell proliferation and reduced TGF-beta1-mediated p3TP-Lux (PAI-1 promoter) luciferase activity. It also reduced (CAGA)12-Luc luciferase activity in response to TGF-beta1. The CAGA sequence is specific for Smad3/4 binding, implying that CD105 is involved in inhibition of TGF-beta1/Smad3 signalling. Furthermore, CD105 overexpression reduced serine phosphorylation of Smad3 and inhibited subsequent nuclear translocation of Smad3. CD105 resulted in high phosphorylation of JNK1, which is able to activate c-Jun. c-Jun is known to inhibit Smad3 transcriptional activity on CAGA sites, suggesting that CD105 may also inhibit Smad3 signalling through JNK1.
Research Interests: Biology, Medicine, Angiogenesis, Western blotting, Signal Transduction, and 15 moreHumans, Pubmed, Animals, Phosphorylation, Luciferase, Mitogen Activated Protein Kinase, Anticancer, Rats, Transforming Growth Factor Beta, Transfection, DNA binding proteins, Transforming Growth Factor, Mitogen- Activated Protein Kinases, Endoglin, and Muscle cells
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Endoglin (Eng) is a co-receptor of the transforming growth factor β superfamily playing an important role in endothelial dysfunction. TRC105 (carotuximab) is a monoclonal antibody that blocks Eng and its downstream Smad signaling pathway.... more
Endoglin (Eng) is a co-receptor of the transforming growth factor β superfamily playing an important role in endothelial dysfunction. TRC105 (carotuximab) is a monoclonal antibody that blocks Eng and its downstream Smad signaling pathway. Here we have investigated for the first time the effects of TRC105 treatment on the development of endothelial dysfunction induced by 7-ketocholesterol (7K) or high glucose (HG), focusing on Eng expression, signaling, and function. In the hypercholesterolemia study, human aortic endothelial cells (HAoECs) were treated with TRC105 (300 μg/ml) for 1 h, followed by the addition of 7K (10 μg/ml) for another 12 h. In the hyperglycemia study, HAoECs were exposed to HG (45 mM) for 60 h, followed by the addition of TRC105 for another 12 h, and cells treated with 5mM glucose and 40 mM mannitol served as control. Protein levels, adhesion, and transmigration of monocytes were assessed by flow cytometry, mRNA expression was measured by qRT-PCR. 7K and HG treat...
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Immune electron microscopy is providing information on the three dimensional locations of many functional sites on the ribosome. In particular, localization of the exit site of the newly synthesized protein chain is expected to provide... more
Immune electron microscopy is providing information on the three dimensional locations of many functional sites on the ribosome. In particular, localization of the exit site of the newly synthesized protein chain is expected to provide information on the mechanisms of protein synthesis and on the nature of the ribosome-membrane association. Theoretically the nascent chains of the largest proteins could be visible as they leave the ribosome, but in practice they have not been observed. In order to map their exit site we have made antibodies against one particular enzyme and then applied immune electron microscopy to a polysome fraction enriched for that special protein.In a mutant of Escherichia coli constitutive for the enzyme beta-galac- tosidase, this enzyme accounts for approximately 25% of the total protein (1). Because of the high molecular weight of beta-galactosidase, a fraction of larger polysomes, further enriched in this emzyme, was obtained by differential centrifugation....
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Ascending Thoracic Aortic Aneurysms (ATAA) is a devastating disease, consisting in dilatation of ascending aorta, often resulting in aortic wall dissection. In spite of its heterogeneous aetiology, histologic ATAA features invariably... more
Ascending Thoracic Aortic Aneurysms (ATAA) is a devastating disease, consisting in dilatation of ascending aorta, often resulting in aortic wall dissection. In spite of its heterogeneous aetiology, histologic ATAA features invariably involve marked extracellular matrix (ECM) fragmentation, vascular smooth muscle cell (VSMC) necrosis, cystic degeneration. ATAA seems to arise from a disrupted ECM depostion/degradation homeostasis, abnormally imbalanced towards demolition. Although the complex cellular/molecular ATAA-determining mechanisms are still poorly understood, several studies suggest a key role played by Transforming-Growth–Factor-ß/Bone-Morphogenetic-Protein (TGFß/BMP) superfamily cytokines in ATAA pathogenesis. Many mendelian syndromes including ATAA in their clinical spectrum are caused by mutations in TGFß/BMP-signalling-cascade coding genes, such as TGFBR2, ACVRL5/TGFBR1, MADH3. Furthermore, impaired TGFß/BMP signalling has been described in both syndromic and non-syndromic ATAA samples [2]. Several authors have proposed that ATAA aortic wall cells display an altered TGFß/BMP receptorial reservoir, responsible for impaired TGFß/BMP responsiveness, similarly to pathogenic mechanisms already described for atherosclerosis [3-5] ALK1/ACVRL1 and Endoglin (ENG) are components of the signalling pathway of at least three members of TGFß/BMP superfamily, namely BMP9, TGFß-1 and TGFß-3. Moreover, ALK1/ACVRL1 and ENG genes are mutated in the vascular mendelian syndrome Hereditary Haemorrhagic Telangiectasia (HHT). Despite the well-known pivotal role of the two genes in vascular homeostasis, their involvement in ATAA has never been investigated
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KUMAR, P., WANG, JM and BERNABEAU, C.(1996), EDITORIAL. CD105 AND ANGIOGENESIS. The Journal of Pathology, 178: 363366. doi: 10.1002/(SICI) 1096-9896 (199604) 178: 4< 363:: AID-PATH491> 3.0. CO; 2-8
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Research Interests: Biology, Gene Therapy, Immunohistochemistry, Medicine, Gene expression, and 15 moreBiological Sciences, Humans, Liver, Mice, Animals, Lung, Gene Delivery, Gene Targeting, Gastrointestinal bleeding, Gene Transfer, Cell Adhesion Molecules, Autosomal Dominant, Genetic Therapy, Endoglin, and Medical and Health Sciences
43 O Fecha de publicación de la solicitud: 07.05.2010 ... 43 O Fecha de publicación del folleto de la solicitud: 07.05.2010 ... 71 O Solicitante/s: Consejo Superior de Investigaciones Científicas (CSIC) (Titular al 80 %) c/ Serrano, 117... more
43 O Fecha de publicación de la solicitud: 07.05.2010 ... 43 O Fecha de publicación del folleto de la solicitud: 07.05.2010 ... 71 O Solicitante/s: Consejo Superior de Investigaciones Científicas (CSIC) (Titular al 80 %) c/ Serrano, 117 28006 Madrid, ES Centro de Investigación ...
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Endoglin (CD105) is a cell surface component of the transforming growth factor-β (TGF-β) receptor complex highly expressed by endothelial cells. Mutations in the endoglin gene are responsible for the hereditary hemorrhagic telangiectasia... more
Endoglin (CD105) is a cell surface component of the transforming growth factor-β (TGF-β) receptor complex highly expressed by endothelial cells. Mutations in the endoglin gene are responsible for the hereditary hemorrhagic telangiectasia type 1 (HHT1), also known as Osler-Weber-Rendu syndrome (OMIM 187300). This is an autosomal dominant vascular disorder probably caused by a haploinsufficiency mechanism displaying low levels of the normal protein. To understand the mechanisms underlying the regulated expression of endoglin, a genomic DNA clone containing 3.3 kb of the 5′-flanking sequence of the human endoglin gene has been isolated. The 5′-flanking region of the endoglin gene lacks consensus TATA and CAAT boxes, but contains two GC-rich regions and consensus motifs for Sp1, ets, GATA, AP-2, NFκB, and Mad, as well as TGF-β–, glucocorticoid-, vitamin D-, and estrogen-responsive elements. As determined by primer extension and 5′ RACE experiments, a cluster of transcriptional start sit...
Research Interests: Molecular Biology, Biology, Medicine, Humans, Animals, and 15 moreBlood, Promoter, Vascular endothelium, Gene, Clinical Sciences, Cattle, Molecular cloning, Transforming Growth Factor Beta, Amino Acid Sequence, Base Sequence, Gene Expression Regulation, Molecular Sequence Data, Endoglin, Cardiovascular medicine and haematology, and Paediatrics and reproductive medicine
We have analyzed the cellular and humoral immunity to the mycobacterial 65 KDa heat shock protein (hsp65) in a group of Freund&#39;s Adjuvant-immunized rats with a limited susceptibility to Adjuvant arthritis. According to the... more
We have analyzed the cellular and humoral immunity to the mycobacterial 65 KDa heat shock protein (hsp65) in a group of Freund&#39;s Adjuvant-immunized rats with a limited susceptibility to Adjuvant arthritis. According to the arthritis indices during the period of study (35 days), two different groups of rats could be distinguished; a) autoimmune Adjuvant arthritic rats (AA), and b) Non-arthritic animals (NA), including both rats which did not display any disease symptoms and rats suffering mild transient inflammation. The cellular response to the immunizing agent (Mycobacterium tuberculosis) or the mitogen Concanavalin A was comparable between both groups of rats. However, we detected an impaired cellular response to the individual hsp65 antigen in the animals that did not develop the disease. On the contrary, the level of hsp65-specific antibodies was much higher in NA animals than in AA rats suggesting a protective role for the hsp65 specific antibodies.
Research Interests: Immunology, Autoimmunity, Medicine, Disease susceptibility, Animals, and 15 moreMale, Immunization, Arthritis, Immune system, Antibody, Heat Shock Proteins, Heat Shock Protein, Enzyme Linked Immunosorbent Assay, Antigen, Adjuvant Arthritis, Adjuvant, Antigen presenting cells, Humoral Immunity, Chaperonins, and In Vitro Techniques
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Research Interests: Microbiology, Thermodynamics, Chemistry, Immunology, Kinetics, and 13 moreMedical Microbiology, Biology, Medicine, Protein Structure and Function, Fourier Analysis, Circular Dichroism, Escherichia coli, Protein Secondary Structure, Protein Secondary Structure Prediction, Protein Conformation, Gel electrophoresis, Biochemistry and cell biology, and Sodium Dodecyl Sulfate
Research Interests: Wound Healing, Medicine, Angiogenesis, Humans, Endothelial Cells, and 13 moreFemale, Hereditary Hemorrhagic Telangiectasia (HHT), Haemostasis and Thrombosis, Haploinsufficiency, Clinical Sciences, Aged, Middle Aged, Telangiectasia, Pilot Projects, Hemorrhage, Selective estrogen receptor modulators, Endoglin, and Cardiovascular medicine and haematology
Research Interests: Molecular Biology, Biology, Cell Biology, Medicine, Cell Division, and 15 moreBiological Sciences, Cell line, Humans, TGF Beta, Cell, Phosphorylation, Integrins, Transforming Growth Factor Beta, The cell, Transfection, Base Sequence, Cell Adhesion Molecules, Molecular Sequence Data, Endoglin, and Medical and Health Sciences
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Endoglin is a membrane glycoprotein that acts as a coreceptor for transforming growth factor-β. We and others have previously suggested a function of endoglin as a tumor suppressor in epithelial cancer. Here, we study the expression of... more
Endoglin is a membrane glycoprotein that acts as a coreceptor for transforming growth factor-β. We and others have previously suggested a function of endoglin as a tumor suppressor in epithelial cancer. Here, we study the expression of endoglin during chemical mouse skin carcinogenesis. We find that shedding of membrane endoglin, allowing the secretion of a soluble endoglin form, is a late event associated with progression from squamous to spindle cell carcinomas. Knockdown of endoglin in transformed keratinocytes activates the Smad2/3 signaling pathway resulting in cell growth arrest, delayed tumor latencies, and a squamous to spindle phenotypic conversion. Forced expression of the long endoglin isoform in spindle carcinoma cells blocks transforming growth factor-β1 stimulation of Smad2/3 signaling and prevents tumor formation. In contrast, expression of the short endoglin isoform has no effect on spindle cell growth in vitro or in vivo. Our results show that endoglin behaves as a suppressor of malignancy during the late stages of carcinogenesis. Therefore, disruption of membrane endoglin emerges as a crucial event for progression to spindle cell carcinomas. [Cancer Res 2007;67(21):10268–77]
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CIBER (Center for Biomedical Network Research; Centro de Investigación Biomédica En Red) is a public national consortium created in 2006 under the umbrella of the Spanish National Institute of Health Carlos III (ISCIII). This innovative... more
CIBER (Center for Biomedical Network Research; Centro de Investigación Biomédica En Red) is a public national consortium created in 2006 under the umbrella of the Spanish National Institute of Health Carlos III (ISCIII). This innovative research structure comprises 11 different specific areas dedicated to the main public health priorities in the National Health System. CIBERER, the thematic area of CIBER focused on Rare Diseases currently consists of 75 research groups belonging to universities, research centers and hospitals of the entire country. CIBERER's mission is to be a center prioritizing and favoring collaboration and cooperation between biomedical and clinical research groups, with special emphasis on the aspects of genetic, molecular, biochemical and cellular research of rare diseases. This research is the basis for providing new tools for the diagnosis and therapy of low-prevalence diseases, in line with the International Rare Diseases Research Consortium (IRDiRC) objectives, thus favoring translational research between the scientific environment of the laboratory and the clinical setting of health centers. In this paper, we intend to review CIBERER's 15-year journey and summarize the main results obtained in terms of internationalization, scientific production, contributions towards the discovery of new therapies and novel genes associated to diseases, cooperation with patients' associations and many other topics related to rare disease research. This article is protected by copyright. All rights reserved.
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<b><i>Aims:</i></b> A soluble form of endoglin (sEng) was proposed to participate in the induction of endothelial dysfunction in small blood vessels. Here, we tested the hypothesis that high levels of sEng combined... more
<b><i>Aims:</i></b> A soluble form of endoglin (sEng) was proposed to participate in the induction of endothelial dysfunction in small blood vessels. Here, we tested the hypothesis that high levels of sEng combined with a high-fat diet induce endothelial dysfunction in an atherosclerosis-prone aorta. <b><i>Methods and Results:</i></b> Six-month-old female and male transgenic mice overexpressing human sEng (<i>Sol-Eng</i><sup><i>+</i></sup>) with low (<i>Sol-Eng</i><sup><i>+</i></sup><i>low</i>) or high (<i>Sol-Eng</i><sup><i>+</i></sup><i>high</i>) levels of plasma sEng were fed a high-fat rodent diet containing 1.25% cholesterol and 40% fat for 3 months. The plasma cholesterol and mouse sEng levels did not differ in the <i>Sol-Eng</i><sup><i>+</i></sup><i>high</i> and <i>Sol-Eng</i><sup><i>+</i></sup><i>low</i> mice. The expression of proinflammatory (P-selectin, ICAM-1, pNFκB and COX-2) and oxidative-stress-related markers (HO-1, NOX-1 and NOX-2) in the aortas of <i>Sol-Eng</i><sup><i>+</i></sup><i>high</i> female mice was significantly higher than in <i>Sol-Eng</i><sup><i>+</i></sup><i>low </i>female mice. Endothelium-dependent vasodilatation induced by acetylcholine was preserved better in the <i>Sol-Eng</i><sup><i>+ </i></sup><i>high</i> female mice than in the <i>Sol-Eng</i><sup><i>+</i></sup><i>low </i>female mice. <b><i>Conclusion:</i></b> These results suggest that high concentrations of sEng in plasma in combination with a high-fat diet induce the simultaneous activation of proinflammatory, pro-oxidative and vasoprotective mechanisms in mice aorta and the balance of these biological processes determines whether the final endothelial phenotype is adaptive or maladaptive.