Interoperability Issues Between Learning Object Repositories and Metadata Harvesters

In this paper we describe an open learning object repository on Statistics based on DSpace which contains true learning objects, that is, exercises, equations, data sets, etc. This repository is part of a large project intended to promote the use of learning object repositories as part of the learning process in virtual learning environments. This involves the creation of a new user interface that provides users with additional services such as resource rating, commenting and so. Both aspects make traditional metadata schemes such as Dublin Core to be inadequate, as there are resources with no title or author, for instance, as those fields are not used by learners to browse and search for learning resources in the repository. Therefore, exporting OAI-PMH compliant records using OAI-DC is not possible, thus limiting the visibility of the learning objects in the repository outside the institution. We propose an architecture based on ontologies and the use of extended metadata records for both storing and refactoring such descriptions

Interoperability Issues Between Learning Object Repositories and Metadata Harvesters

In this paper we describe an open learning object repository on Statistics based on DSpace which contains true learning objects, that is, exercises, equations, data sets, etc. This repository is part of a large project intended to promote the use of learning object repositories as part of the learning process in virtual learning environments. This involves the creation of a new user interface that provides users with additional services such as resource rating, commenting and so. Both aspects make traditional metadata schemes such as Dublin Core to be inadequate, as there are resources with no title or author, for instance, as those fields are not used by learners to browse and search for learning resources in the repository. Therefore, exporting OAI-PMH compliant records using OAI-DC is not possible, thus limiting the visibility of the learning objects in the repository outside the institution. We propose an architecture based on ontologies and the use of extended metadata records for both storing and refactoring such descriptions

Epigenetic alterations in Fanconi Anaemia: Role in pathophysiology and therapeutic potential

Fanconi anaemia (FA) is an inherited disorder characterized by chromosomal instability. The phenotype is variable, which raises the possibility that it may be affected by other factors, such as epigenetic modifications. These play an important role in oncogenesis and may be pharmacologically manipulated. Our aim was to explore whether the epigenetic profiles in FA differ from non-FA individuals and whether these could be manipulated to alter the disease phenotype. We compared expression of epigenetic genes and DNA methylation profile of tumour suppressor genes between FA and normal samples. FA samples exhibited decreased expression levels of genes involved in epigenetic regulation and hypomethylation in the promoter regions of tumour suppressor genes. Treatment of FA cells with histone deacetylase inhibitor Vorinostat increased the expression of DNM3T beta and reduced the levels of CIITA and HDAC9, PAK1, USP16, all involved in different aspects of epigenetic and immune regulation. Given the ability of Vorinostat to modulate epigenetic genes in FA patients, we investigated its functional effects on the FA phenotype. This was assessed by incubating FA cells with Vorinostat and quantifying chromosomal breaks induced by DNA cross-linking agents. Treatment of FA cells with Vorinostat resulted in a significant reduction of aberrant cells (81\% on average). Our results suggest that epigenetic mechanisms may play a role in oncogenesis in FA. Epigenetic agents may be helpful in improving the phenotype of FA patients, potentially reducing tumour incidence in this population.publishersversionpublishe

Development of a mouse model for spontaneous oral squamous cell carcinoma in Fanconi anemia

Altres ajuts: European Regional Development Fund (FEDER); the European Union; the Spanish Fundacion Anemia de Fanconi and Fanconi Anemia Research Fund USA; Comunidad de Madrid (ref PEJ-2019-TL_BMD-12905).Fanconi anemia (FA) patients frequently develop oral squamous cell carcinoma (OSCC). This cancer in FA patients is diagnosed within the first 3-4 decades of life, very often preceded by lesions that suffer a malignant transformation. In addition, they respond poorly to current treatments due to toxicity or multiple recurrences. Translational research on new chemopreventive agents and therapeutic strategies has been unsuccessful partly due to scarcity of disease models or failure to fully reproduce the disease. Here we report that Fanca gene knockout mice (Fanca ) frequently display pre-malignant lesions in the oral cavity. Moreover, when these animals were crossed with animals having conditional deletion of Trp53 gene in oral mucosa (K14cre;Trp53), they spontaneously developed OSCC with high penetrance and a median latency of less than ten months. Tumors were well differentiated and expressed markers of squamous differentiation, such as keratins K5 and K10. In conclusion, Fanca and Trp53 genes cooperate to suppress oral cancer in mice, and Fanca;K14cre;Trp53 mice constitute the first animal model of spontaneous OSCC in FA

Upregulation of NKG2D ligands impairs hematopoietic stem cell function in Fanconi anemia

Altres ajuts: Fondo Europeo de Desarrollo Regional (FEDER); Next Generation EU; EUROFANCOLEN); Comunidad de Madrid (AvanCell, B2017/BMD-3692); ICREA-Academia program.Fanconi anemia (FA) is the most prevalent inherited bone marrow failure (BMF) syndrome. Nevertheless, the pathophysiological mechanisms of BMF in FA have not been fully elucidated. Since FA cells are defective in DNA repair, we hypothesized that FA hematopoietic stem and progenitor cells (HSPCs) might express DNA damage-associated stress molecules such as natural killer group 2 member D ligands (NKG2D-Ls). These ligands could then interact with the activating NKG2D receptor expressed in cytotoxic NK or CD8+ T cells, which may result in progressive HSPC depletion. Our results indeed demonstrated upregulated levels of NKG2D-Ls in cultured FA fibroblasts and T cells, and these levels were further exacerbated by mitomycin C or formaldehyde. Notably, a high proportion of BM CD34+ HSPCs from patients with FA also expressed increased levels of NKG2D-Ls, which correlated inversely with the percentage of CD34+ cells in BM. Remarkably, the reduced clonogenic potential characteristic of FA HSPCs was improved by blocking NKG2D-NKG2D-L interactions. Moreover, the in vivo blockage of these interactions in a BMF FA mouse model ameliorated the anemia in these animals. Our study demonstrates the involvement of NKG2D-NKG2D-L interactions in FA HSPC functionality, suggesting an unexpected role of the immune system in the progressive BMF that is characteristic of FA

Analysis of the transcriptional activity of endogenous NFAT5 in primary cells using transgenic NFAT-luciferase reporter mice

<p>Abstract</p> <p>Background</p> <p>The transcription factor NFAT5/TonEBP regulates the response of mammalian cells to hypertonicity. However, little is known about the physiopathologic tonicity thresholds that trigger its transcriptional activity in primary cells. Wilkins et al. recently developed a transgenic mouse carrying a luciferase reporter (9xNFAT-Luc) driven by a cluster of NFAT sites, that was activated by calcineurin-dependent NFATc proteins. Since the NFAT site of this reporter was very similar to an optimal NFAT5 site, we tested whether this reporter could detect the activation of NFAT5 in transgenic cells.</p> <p>Results</p> <p>The 9xNFAT-Luc reporter was activated by hypertonicity in an NFAT5-dependent manner in different types of non-transformed transgenic cells: lymphocytes, macrophages and fibroblasts. Activation of this reporter by the phorbol ester PMA plus ionomycin was independent of NFAT5 and mediated by NFATc proteins. Transcriptional activation of NFAT5 in T lymphocytes was detected at hypertonic conditions of 360–380 mOsm/kg (isotonic conditions being 300 mOsm/kg) and strongly induced at 400 mOsm/kg. Such levels have been recorded in plasma in patients with osmoregulatory disorders and in mice deficient in aquaporins and vasopressin receptor. The hypertonicity threshold required to activate NFAT5 was higher in bone marrow-derived macrophages (430 mOsm/kg) and embryonic fibroblasts (480 mOsm/kg). Activation of the 9xNFAT-Luc reporter by hypertonicity in lymphocytes was insensitive to the ERK inhibitor PD98059, partially inhibited by the PI3-kinase inhibitor wortmannin (0.5 μM) and the PKA inhibitor H89, and substantially downregulated by p38 inhibitors (SB203580 and SB202190) and by inhibition of PI3-kinase-related kinases with 25 μM LY294002. Sensitivity of the reporter to FK506 varied among cell types and was greater in primary T cells than in fibroblasts and macrophages.</p> <p>Conclusion</p> <p>Our results indicate that NFAT5 is a sensitive responder to pathologic increases in extracellular tonicity in T lymphocytes. Activation of NFAT5 by hypertonicity in lymphocytes was mediated by a combination of signaling pathways that differed from those required in other cell types. We propose that the 9xNFAT-Luc transgenic mouse model might be useful to study the physiopathological regulation of both NFAT5 and NFATc factors in primary cells.</p

Epigenetic Alterations in Fanconi Anaemia: Role in Pathophysiology and Therapeutic Potential.

Fanconi anaemia (FA) is an inherited disorder characterized by chromosomal instability. The phenotype is variable, which raises the possibility that it may be affected by other factors, such as epigenetic modifications. These play an important role in oncogenesis and may be pharmacologically manipulated. Our aim was to explore whether the epigenetic profiles in FA differ from non-FA individuals and whether these could be manipulated to alter the disease phenotype. We compared expression of epigenetic genes and DNA methylation profile of tumour suppressor genes between FA and normal samples. FA samples exhibited decreased expression levels of genes involved in epigenetic regulation and hypomethylation in the promoter regions of tumour suppressor genes. Treatment of FA cells with histone deacetylase inhibitor Vorinostat increased the expression of DNM3Tβ and reduced the levels of CIITA and HDAC9, PAK1, USP16, all involved in different aspects of epigenetic and immune regulation. Given the ability of Vorinostat to modulate epigenetic genes in FA patients, we investigated its functional effects on the FA phenotype. This was assessed by incubating FA cells with Vorinostat and quantifying chromosomal breaks induced by DNA cross-linking agents. Treatment of FA cells with Vorinostat resulted in a significant reduction of aberrant cells (81% on average). Our results suggest that epigenetic mechanisms may play a role in oncogenesis in FA. Epigenetic agents may be helpful in improving the phenotype of FA patients, potentially reducing tumour incidence in this population

The ALFA project: A research platform to identify early pathophysiological features of Alzheimer's disease

Introduction: The preclinical phase of Alzheimer's disease (AD) is optimal for identifying early pathophysiological events and developing prevention programs, which are shared aims of the ALFA project, including the ALFA registry and parent cohort and the nested ALFA+ cohort study. Methods: The ALFA parent cohort baseline visit included full cognitive evaluation, lifestyle habits questionnaires, DNA extraction, and MRI. The nested ALFA+ study adds wet and imaging biomarkers for deeper phenotyping. Results: A total of 2743 participants aged 45 to 74 years were included in the ALFA parent cohort. We show that this cohort, mostly composed of cognitively normal offspring of AD patients, is enriched for AD genetic risk factors. Discussion: The ALFA project represents a valuable infrastructure that will leverage with different studies and trials to prevent AD. The longitudinal ALFA+ cohort will serve to untangle the natural history of the disease and to model the preclinical stages to develop successful trials

DNA methylation pattern on tumour suppressor genes in peripheral blood mononuclear cells of FA patients.

<p>PBMC from FA patients and healthy donor were isolated as in A, DNA was isolated and methylation profile was identified in 2 FA samples and 2 healthy donors samples using EpiTect Methyl II Complete PCR array (SABiosciences). Samples CTR1 and CTR2 represent control and FA1 and FA2 represent FA patients. Each row represents a tumour-supressor genes and each column represent a single DNA sample. The methylation degree are represented by the level of intensity of the square, red representing greater than 10% promoter hypermethylation, and green representing less than 10% promoter methylation (unmethylated) alleles for the tumour-suppressor gene.</p

Number of breaks per cell in cultured lymphocytes from FA patients.

<p>The effect of the Vorinostat was calculated by the percentage of reduction of the number of breaks per cell in the Vorinostat treatments relatively to Spontaneous breaks and DEB.</p><p>Number of breaks per cell in cultured lymphocytes from FA patients.</p