Role of platelet C3G in vesicle trafficking and spreading. Involvement in hemostasis and platelet-mediated inflammation

[ES] En este trabajo presentamos evidencias de un papel de C3G en la modulación de la extensión plaquetaria a través de la regulación de la formación de lamellipodia por su participación en la vía Rac1/WAVE2/Arp2/3 de forma independiente de su función GEF. Además, C3G regularía la secreción de gránulos α modulando la formación de los complejos trans-SNARE y VARP/VAMP-7/Arp2/3, así como la activación de Ral, de forma dependiente de Rap1. Por último, presentamos resultados preliminares, tanto in vitro como in vivo sobre una nueva función de la proteína C3G en la respuesta inflamatoria mediada por plaquetas, a través de la regulación de la formación de PLA, con NE y linfocitos B, y la liberación de factores inflamatorios. [EN] In this work we present evidence of a role for C3G in the modulation of platelet spreading through the regulation of lamellipodia formation via its participation in the Rac1/WAVE2/Arp2/3 pathway, which is independent of its GEF function. In addition, C3G would regulate the secretion of α-granules by modulating the formation of the trans-SNARE and the VARP/VAMP-7/Arp2/3 complexes, as well as the activation of Ral, in a Rap1-dependent manner. Finally, we present preliminary results, both in vitro and in vivo, on a novel role of the C3G protein in the platelet-mediated inflammatory response, through the regulation of PLA formation, with NE and B lymphocytes, and the release of inflammatory factor

Aplicación del polimedia en el ámbito educativo

Polimedia is a room created for benefit of all. Actualy, we can say that a tool with that exists innovating and productive we can record a information, to reproduce it on a presentation to power point, like complementary explanation to the information that is offered. The basic function is to include a information, the vision and oral communication of the author and to offer the possibility, to visualize it so many times as it is necessary. The University of Huelva, has believed in this project, believes in the opportunity to offer an innovating resource and the benefits that contribute are numerous for teaching.Polimedia es una sala creada para beneficio de todos. Actualmente, podemos decir que existe una herramienta innovadora y productiva con la que podemos grabar una información y reproducirla sobre una presentación power point como explicación complementaria a la información que se ofrece. La función básica es incluir a la información textual, la visión y comunicación oral del autor, además de ofrecer la posibilidad de visualizarla tantas veces como sea necesario. La Universidad de Huelva ha creído en este proyecto, pues cree en la oportunidad de ofrecer un recurso innovador y que los beneficios que aporta son cuantiosos para la docencia

Use of social media in secondary school libraries as channels for disseminating their information: The case of Extremadura

Usando como muestra los centros públicos de educación secundaria de la Comunidad de Extremadura en España, el objetivo de este trabajo consiste en descubrir cómo la biblioteca escolar realiza la difusión de información a través de medios sociales, cuáles son los más utilizados para ello y qué contenidos son los más difundidos, así como establecer una clasificación de centros educativos atendiendo al uso que sus bibliotecas hacen de los medios, para lo que se crea una ponderación contrastada de medios y de usos. Los resultados de la muestra señalan que: el 60,61% de las bibliotecas de secundaria participa en algún medio social (con un promedio de 3,66); Twitter, Facebook, Google+ y Pinterest resultan los medios más dinámicos para llevar a cabo la difusión de información; los blogs se asientan como la herramienta más importante para realizar esta labor; y la ‘Población’ y el ‘Producto Interior Bruto per cápita’ del municipio en el que se ubica el centro escolar cuya biblioteca se analiza, no influyen en la posición que éste obtiene en la clasificación de medios sociales. Se concluye con la propuesta de una serie de pautas para una planificación futura sobre la presencia de las bibliotecas escolares de secundaria en medios sociales.The purpose of this study is to find out how school libraries disseminate information through social media, using as an example the public secondary schools of Extremadura, Spain. It looks at which tools are the most used and what contents are the most widespread, as well as setting up a ranking of schools regarding the use that their libraries make of social media. For this a proven weighting of media and uses was created. The results of the sample indicate that 60.61 % of secondary school libraries use some social media (the average is 3.66); Twitter, Facebook, Google+ and Pinterest are the most dynamic media for disseminating information; blogs are the most important tool for this task. ‘Population’ and ‘Per capita gross domestic product’ of the town in which the school under study is located do not affect the position attained by the school in the ranking of social media. To conclude, a set of guidelines is proposed for planning the visibility of secondary school libraries in the social media.peerReviewe

La formación universitaria en entornos virtuales en la Universidad de Huelva

El ámbito de la docencia universitaria debe estar alerta y a la vez, ser pionera en los nuevos cambios que estamos viviendo. Por ello, es imprescindible la adaptación de los sistemas docentes, tanto en metodología como en materias impartidas, si queremos seguir siendo una institución de referencia en el avance cultural y social. Un claro ejemplo, de que se está llevando a cabo estos cambios en las universidades españolas, concretamente en Andalucía, es el caso de la Universidad de Huelva. Que incorpora por primera vez, la modalidad de enseñanza virtual, en una de sus Facultades, con el propósito de incrementar su campo de acción. Esta modalidad, permite al estudiante desarrollar sus estudios a través de una plataforma de teleformación, contando además con clases de apoyo presenciales con periodicidad mensual.University teaching must be alert to the new changes that we are living and at the same time, be pioneering. For that reason, the adaptation of educational systems, in methodology as matters is essential if we want to continue being on institution of reference in this cultural and social advance. A clear example of these changes being carried out in Spanish universities, mainly in Andalusia, is in the case of the University of Huelva. For the first time it incorporates the method of virtual education in one of its faculties in order to increase its field of study. This method allows students to develop their studies through education learning as well trough monthly tutorials

C3G, through its GEF activity, induces megakaryocytic differentiation and proplatelet formation

[Background]: Megakaryopoiesis allows platelet formation, which is necessary for coagulation, also playing an important role in different pathologies. However, this process remains to be fully characterized. C3G, an activator of Rap1 GTPases, is involved in platelet activation and regulates several differentiation processes. [Methods]: We evaluated C3G function in megakaryopoiesis using transgenic mouse models where C3G and C3GΔCat (mutant lacking the GEF domain) transgenes are expressed exclusively in megakaryocytes and platelets. In addition, we used different clones of K562, HEL and DAMI cell lines with overexpression or silencing of C3G or GATA-1. [Results]: We found that C3G participates in the differentiation of immature hematopoietic cells to megakaryocytes. Accordingly, bone marrow cells from transgenic C3G, but not those from transgenic C3GΔCat mice, showed increased expression of the differentiation markers CD41 and CD61, upon thrombopoietin treatment. Furthermore, C3G overexpression increased the number of CD41+ megakaryocytes with high DNA content. These results are supported by data obtained in the different models of megakaryocytic cell lines. In addition, it was uncovered GATA-1 as a positive regulator of C3G expression. Moreover, C3G transgenic megakaryocytes from fresh bone marrow explants showed increased migration from the osteoblastic to the vascular niche and an enhanced ability to form proplatelets. Although the transgenic expression of C3G in platelets did not alter basal platelet counts, it did increase slightly those induced by TPO injection in vivo. Moreover, platelet C3G induced adipogenesis in the bone marrow under pathological conditions. [Conclusions]: All these data indicate that C3G plays a significant role in different steps of megakaryopoiesis, acting through a mechanism dependent on its GEF activity.This work was supported by grants from the Spanish Ministry of Economy and Competitiveness [SAF2013–48210-C2–1-R and SAF2016–76588-C2–2-R to CG, SAF2013–48210-C2–2-R and SAF2016–76588-C2–1-R to AP], and by two grants from the Council of Education of Junta de Castilla y León, Spain [SA157A12–1 and SA017U16 to CG]. All funding was cosponsored by the European FEDER Program

A novel nonsense variant in TPM4 caused dominant macrothrombocytopenia, mild bleeding tendency and disrupted cytoskeleton remodeling

[Background]: Rare inherited thrombocytopenias are caused by alterations in genes involved in megakaryopoiesis, thrombopoiesis and/or platelet release. Diagnosis is challenging due to poor specificity of platelet laboratory assays, large numbers of culprit genes, and difficult assessment of the pathogenicity of novel variants. [Objectives]: To characterize the clinical and laboratory phenotype, and identifying the underlying molecular alteration, in a pedigree with thrombocytopenia of uncertain etiology. [Patients/Methods]: Index case was enrolled in our Spanish multicentric project of inherited platelet disorders due to lifelong thrombocytopenia and bleeding. Bleeding score was recorded by ISTH‐BAT. Laboratory phenotyping consisted of blood cells count, blood film, platelet aggregation and flow cytometric analysis. Genotyping was made by whole‐exome sequencing (WES). Cytoskeleton proteins were analyzed in resting/spreading platelets by immunofluorescence and immunoblotting. [Results]: Five family members displayed lifelong mild thrombocytopenia with a high number of enlarged platelets in blood film, and mild bleeding tendency. Patient's platelets showed normal aggregation and granule secretion response to several agonists. WES revealed a novel nonsense variant (c.322C>T; p.Gln108*) in TPM4 (NM_003290.3), the gene encoding for tropomyosin‐4 (TPM4). This variant led to impairment of platelet spreading capacity after stimulation with TRAP‐6 and CRP, delocalization of TPM4 in activated platelets, and significantly reduced TPM4 levels in platelet lysates. Moreover, the index case displayed up‐regulation of TPM2 and TPM3 mRNA levels. [Conclusions]: This study identifies a novel TPM4 nonsense variant segregating with macrothrombocytopenia and impaired platelet cytoskeletal remodeling and spreading. These findings support the relevant role of TPM4 in thrombopoiesis and further expand our knowledge of TPM4‐related thrombocytopenia.This work was partially supported by grants from Instituto de Salud Carlos III (ISCIII) and Feder (PI17/01966, PI20/00926), Gerencia Regional de Salud (GRS2061A/19, GRS2135/A/2020, GRS2314/A/2021), Fundación Mutua Madrileña (FMM, AP172142019) and Sociedad Española de Trombosis y Hemostasia (SETHFETH; Premio López Borrasca 2019 and Ayuda a Grupos de Trabajo en Patología Hemorrágica 2020 and 2021).Peer reviewe

PiRNA-associated proteins and retrotransposons are differentially expressed in murine testis and ovary of aryl hydrocarbon receptor deficient mice

Previous studies suggested that the aryl hydrocarbon receptor (AhR) contributes to mice reproduction and fertility. However, the mechanisms involved remain mostly unknown. Retrotransposon silencing by Piwi-interacting RNAs (piRNAs) is essential for germ cell maturation and, remarkably, AhR has been identified as a regulator of murine B1-SINE retrotransposons. Here, using littermate AhR+/+ and AhR-/- mice, we report that AhR regulates the general course of spermatogenesis and oogenesis by a mechanism likely to be associated with piRNA-associated proteins, piRNAs and retrotransposons. piRNA-associated proteins MVH and Miwi are upregulated in leptotene to pachytene spermatocytes with a more precocious timing in AhR-/- than in AhR+/+ testes. piRNAs and transcripts from B1-SINE, LINE-1 and IAP retrotransposons increased at these meiotic stages in AhR-null testes. Moreover, B1-SINE transcripts colocalize with MVH and Miwi in leptonema and pachynema spermatocytes. Unexpectedly, AhR-/- males have increased sperm counts, higher sperm functionality and enhanced fertility than AhR+/+ mice. In contrast, piRNA-associated proteins and B1-SINE and IAP-derived transcripts are reduced in adult AhR-/- ovaries. Accordingly, AhR-null female mice have lower numbers of follicles when compared with AhR+/+ mice. Thus, AhR deficiency differentially affects testis and ovary development possibly by a process involving piRNA-associated proteins, piRNAs and transposable elements.Trabajo financiado por: Ministerio de Ciencia e Innovación. Ayuda BFU2011-22678 para Pedro María Fernández Salguero Ministerio de Economía y Competitividad. Ayuda SAF2014-51813-R para Pedro María Fernández Salguero Junta de Extremadura. Ayuda GR15008, para Pedro María Fernández Salguero Red Temática de Investigación Cooperativa en Cáncer (RTICC), Instituto Carlos III y Ministerio de Economía y Competitividad (RD12/0036/0032). Trabajos de laboratorio de Pedro María Fernández Salguero Ministerio de Economía y Competitividad. Ayuda AGL2013-43211-R, para Fernando Juan Peña Vega Red Temática de Investigación Cooperativa en Cáncer (RTICC), Instituto Carlos III. Ayuda para Eva María Rico Leo Ministerio de Economía y Competitividad. Ayuda para Francisco Javier González Rico Junta de Extremadura. Ayuda para Eva María Barrasa Ardila Ministerio de Educación, Cultura y Deportes. Beca de Formación de Personal de Investigación, para Nuria Moreno Marín Ministerio de Educación, Cultura y Deportes. Beca FPU13/03991, de Formación de Profesorado Universitario, para Patricia Martín Muñoz Ministerio de Economía y Competitividad. Beca Juan de la Cierva IJCI-2014-21671, para Cristina Ortega Ferrusola Ministerio de Economía y Competitividad, Beca BFU2014-59307-R, para Alberto M. Pendás y Elena Llano Cuadro MEIONet, Junta de Castilla y León y el Programa de Fondeos FEDER, de la Unión EuropeapeerReviewe

Characterization of the platelet phenotype caused by a germline RUNX1 Variant in a CRISPR/Cas9-generated murine model

RUNX1-related disorder (RUNX1-RD) is caused by germline variants affecting the RUNX1 gene. This rare, heterogeneous disorder has no specific clinical or laboratory phenotype, making genetic diagnosis necessary. Although international recommendations have been established to classify the pathogenicity of variants, identifying the causative alteration remains a challenge in RUNX1-RD. Murine models may be useful not only for definitively settling the controversy about the pathogenicity of certain RUNX1 variants, but also for elucidating the mechanisms of molecular pathogenesis. Therefore, we developed a knock-in murine model, using the CRISPR/Cas9 system, carrying the RUNX1 p.Leu43Ser variant (mimicking human p.Leu56Ser) to study its pathogenic potential and mechanisms of platelet dysfunction. A total number of 75 mice were generated; 25 per genotype (RUNX1WT/WT, RUNX1WT/L43S, and RUNX1L43S/L43S). Platelet phenotype was assessed by flow cytometry and confocal microscopy. On average, RUNX1L43S/L43S and RUNX1WT/L43S mice had a significantly longer tail-bleeding time than RUNX1WT/WT mice, indicating the variant's involvement in hemostasis. However, only homozygous mice displayed mild thrombocytopenia. RUNX1L43S/L43S and RUNX1WT/L43S displayed impaired agonist-induced spreading and α-granule release, with no differences in δ-granule secretion. Levels of integrin αIIbβ3 activation, fibrinogen binding, and aggregation were significantly lower in platelets from RUNX1L43S/L43S and RUNX1WT/L43S using phorbol 12-myristate 13-acetate (PMA), adenosine diphosphate (ADP), and high thrombin doses. Lower levels of PKC phosphorylation in RUNX1L43S/L43S and RUNX1WT/L43S suggested that the PKC-signaling pathway was impaired. Overall, we demonstrated the deleterious effect of the RUNX1 p.Leu56Ser variant in mice via the impairment of integrin αIIbβ3 activation, aggregation, α-granule secretion, and platelet spreading, mimicking the phenotype associated with RUNX1 variants in the clinical setting.This work was partially supported by grants from Instituto de Salud Carlos III (ISCIII) and Feder (PI17/01311, PI17/01966, and CB15/00055), Fundación Séneca (19873/GERM/15), Gerencia Regional de Salud (GRS 2061A/19 and 1647/A/17), Fundación Mutua Madrileña (FMM, AP172142019), and Sociedad Española de Trombosis y Hemostasia (SETH-FETH; Premio López Borrasca 2019 and Ayuda a Grupos de Trabajo en Patología Hemorrágica 2019). The authors' research on IPDs is conducted in accordance with the aims of the Functional and Molecular Characterization of Patients with Inherited Platelet Disorders Project, which is supported by the Hemorrhagic Diathesis Working Group of the Spanish Society of Thrombosis and Haemostasis. A.M.-Q., C.F.-I., and L.H.-C. were supported by predoctoral grants from the Junta de Castilla y León, Spain. E.V. was supported by the predoctoral grant from the University of Salamanca, Spain. IG-T and RB were supported by "Contratos postdoctorales Programa II) from the University of Salamanca, Spain

BNIP3 Is Involved in Muscle Fiber Atrophy in Late-Onset Pompe Disease Patients

Late-onset Pompe disease (LOPD) is a rare genetic disorder produced by mutations in the GAA gene and is characterized by progressive muscle weakness. LOPD muscle biopsies show accumulation of glycogen along with the autophagic vacuoles associated with atrophic muscle fibers. The expression of molecules related to muscle fiber atrophy in muscle biopsies of LOPD patients was studied using immunofluorescence and real-time PCR. BCL2 and adenovirus E1B 19-kDa interacting protein 3 (BNIP3), a well-known atrogene, was identified as a potential mediator of muscle fiber atrophy in LOPD muscle biopsies. Vacuolated fibers in LOPD patient muscle biopsies were smaller than nonvacuolated fibers and expressed BNIP3. The current data suggested that BNIP3 expression is regulated by inhibition of the AKT-mammalian target of rapamycin pathway, leading to phosphorylation of Unc-51 like autophagy activating kinase 1 (ULK1) at Ser317 by AMP-activated protein kinase. Myoblasts and myotubes obtained from LOPD patients and age-matched controls were studied to confirm these results using different molecular techniques. Myotubes derived from LOPD patients were likewise smaller and expressed BNIP3. Conclusively, transfection of BNIP3 into control myotubes led to myotube atrophy. These findings suggest a cascade that starts with the inhibition of the AKT-mammalian target of rapamycin pathway and activation of BNIP3 expression, leading to progressive muscle fiber atrophy. These results open the door to potential new treatments targeting BNIP3 to reduce its deleterious effects on muscle fiber atrophy in Pompe disease.Peer reviewe

Soy Niña

Este libro pretende contribuir al reencuentro de la educación con esas finalidades que verdaderamente importan a una niña o un niño: ser feliz, jugar, vivir juntos y (no) aprender. Para ello hemos puesto el arte, nuestras experiencias y el saber acumulado al servicio del disfrute, el cuestionamiento, el análisis crítico y la construcción común de un presente deseable. Un texto colaborativo coordinado por Ignacio Calderón Almendros y realizado por alumnado de Educación y Cambio Social en el Grado en Educación Infantil de la Universidad de Málaga