A novel biocompatible polymer derived from D-mannitol used as a vector in the field of genetic engineering of eukaryotic cells

The design and preparation of new vectors to transport genetic material and increase the transfection efficiency continue being an important research line. Here, a novel biocompatible sugar-based polymer derived from Dmannitol has been synthesized to be used as a gene material nanocarrier in human (gene transfection) and microalga cells (transformation process). Its low toxicity allows its use in processes with both medical and industrial applications. A multidisciplinary study about the formation of polymer/p-DNA polyplexes has been carried out using techniques such as gel electrophoresis, zeta potential, dynamic light scattering, atomic force microscopy, and circular dichroism spectroscopy. The nucleic acids used were the eukaryotic expression plasmid pEGFP-C1 and the microalgal expression plasmid Phyco69, which showed different behaviors. The importance of DNA supercoiling in both transfection and transformation processes was demonstrated. Better results were obtained in microalga cells nuclear transformation than in human cells gene transfection. This was related to the plasmid’s conformational changes, in particular to their superhelical structure. It is noteworthy that the same nanocarrier has been used with eukaryotic cells from both human and microalga.This work was supported by the Consejería de Conocimiento, Innovación y Universidades de la Junta de Andalucía (FQM-206, FQM-274, FQM-135 and P20–01234); VI Plan Propio Universidad de Sevilla (PP2019/00000748), and the European Union (Feder Funds)

Impairment of autophagy in scrapie-infected transgenic mice at the clinical stage

Autophagy appears to play a role in the etiology and progress of misfolded protein disorders. Although this process is dysregulated in prion diseases, it is unknown whether this impairment is a cause or a consequence of prion neuropathology. The study of autophagy during the progress of the disease could elucidate its role. For this purpose, we have investigated its regulation at different stages of the disease in Tg338 mice, a transgenic murine model that overexpresses the highly susceptible ovine VRQ prion protein allele. Mice were intracerebrally inoculated with mouse adapted classical scrapie and euthanized at the preclinical and clinical stages of the disease. Regulation of autophagy was investigated analyzing the distribution of LC3-B and p62 proteins by immunohistochemistry. Moreover, the expression of genes involved in autophagy regulation was quantified by real-time PCR. LC3-B and p62 proteins were downregulated and upregulated, respectively, in the central nervous system of infected mice with clinical signs of scrapie. Accumulation of p62 correlated with scrapierelated lesions, suggesting an impairment of autophagy in highly prion-affected areas. In addition, Gas5 (growth arrestspecific 5), Atg5 (autophagy-related 5), and Fbxw7 (F-box and WD repeat domain containing 7) transcripts were downregulated in mesencephalon and cervical spinal cord of the same group of animals. The impairment of autophagic machinery seems to be part of the pathological process of scrapie, but only during the late stage of prion infection. Similarities between Tg338 mice and the natural ovine disease make them a reliable in vivo model to study prion infection and autophagy side by side

Dysregulation of autophagy in the central nervous system of sheep naturally infected with classical scrapie

Autophagy is a dynamic cellular mechanism involved in protein and organelle turnover through lysosomal degradation. Autophagy regulation modulates the pathologies associated with many neurodegenerative diseases. Using sheep naturally infected with scrapie as a natural animal model of prion diseases, we investigated the regulation of autophagy in the central nervous system (CNS) during the clinical phase of the disease. We present a gene expression and protein distribution analysis of different autophagy-related markers and investigate their relationship with prion-associated lesions in several areas of the CNS. Gene expression of autophagy markers ATG5 and ATG9 was downregulated in some areas of scrapie brains. In contrast, ATG5 protein accumulates in medulla oblongata and positively correlates with prion deposition and scrapie-related lesions. The accumulation of this protein and p62, a marker of autophagy impairment, suggests that autophagy is decreased in the late phases of the disease. However, the increment of LC3 proteins and the mild expression of p62 in basal ganglia and cerebellum, primarily in Purkinje cells, suggests that autophagy machinery is still intact in less affected areas. We hypothesize that specific cell populations of the CNS may display neuroprotective mechanisms against prion-induced toxicity through the induction of PrPSc clearance by autophagy

Diffuse illuminance availability on horizontal and vertical surfaces at Madrid, Spain

The overall objective of this work is to provide diffuse illuminance availability at Madrid (Spain) through a statistical analysis of illuminance values corresponding to a long-term data series. The illuminance values are obtained from irradiance measurements by means of different empirical models for luminous efficacy. The values of diffuse illuminance on a horizontal and on vertical surfaces facing the four cardinal points are estimated and the different aspects related to daylight availability in an area with specific climatic conditions are analyzed. The experimental data consist of global and diffuse irradiance measurements on a horizontal surface provided by the National Meteorological Agency in Spain (AEMET) for Madrid. These data consist of hourly values measured in the period of 1980–2005. The statistical results derived correspond to a daylight typical year for the five surfaces considered. This information will be useful to building experts to estimate natural illumination availability when daylighting techniques are applied in building design with the main aim of electric energy savings

Cerebrospinal fluid and plasma small extracellular vesicles and mirnas as biomarkers for prion diseases

Diagnosis of transmissible spongiform encephalopathies (TSEs), or prion diseases, is based on the detection of proteinase K (PK)-resistant PrPSc in post-mortem tissues as indication of infection and disease. Since PrPSc detection is not considered a reliable method for in vivo diagnosis in most TSEs, it is of crucial importance to identify an alternative source of biomarkers to provide useful alternatives for current diagnostic methodology. Ovine scrapie is the prototype of TSEs and has been known for a long time. Using this natural model of TSE, we investigated the presence of PrPSc in exosomes derived from plasma and cerebrospinal fluid (CSF) by protein misfolding cyclic amplification (PMCA) and the levels of candidate microRNAs (miRNAs) by quantitative PCR (qPCR). Significant scrapie-associated increase was found for miR-21-5p in plasma-derived but not in CSF-derived exosomes. However, miR-342-3p, miR-146a-5p, miR-128-3p and miR-21-5p displayed higher levels in total CSF from scrapie-infected sheep. The analysis of overexpressed miRNAs in this biofluid, together with plasma exosomal miR-21-5p, could help in scrapie diagnosis once the presence of the disease is suspected. In addition, we found the presence of PrPSc in most CSF-derived exosomes from clinically affected sheep, which may facilitate in vivo diagnosis of prion diseases, at least during the clinical stage

Finanzas sostenibles: Entre todos y para todos

Proyecto en el que participan dos universidades: Universidad Complutense de Madrid y Universidad Rey Juan Carlos, y un Instituto de Enseñanza Secundario: Gerardo DiegoDepto. de Economía Aplicada, Pública y PolíticaFac. de Ciencias Económicas y EmpresarialesFALSEUniversidad Complutense de Madridsubmitte

Identification of clinical variants beyond the exome in inborn errors of metabolism

Inborn errors of metabolism (IEM) constitute a huge group of rare diseases affecting 1 in every 1000 newborns. Next-generation sequencing has transformed the diagnosis of IEM, leading to its proposed use as a second-tier technology for confirming cases detected by clinical/biochemical studies or newborn screening. The diagnosis rate is, however, still not 100%. This paper reports the use of a personalized multi-omics (metabolomic, genomic and transcriptomic) pipeline plus functional genomics to aid in the genetic diagnosis of six unsolved cases, with a clinical and/or biochemical diagnosis of galactosemia, mucopolysaccharidosis type I (MPS I), maple syrup urine disease (MSUD), hyperphenylalaninemia (HPA), citrullinemia, or urea cycle deficiency. Eight novel variants in six genes were identified: six (four of them deep intronic) located in GALE, IDUA, PTS, ASS1 and OTC, all affecting the splicing process, and two located in the promoters of IDUA and PTS, thus affecting these genes’ expression. All the new variants were subjected to functional analysis to verify their pathogenic effects. This work underscores how the combination of different omics technologies and functional analysis can solve elusive cases in clinical practic

Materialidad pictórica. Estudio y percepción de la pintura tardogótica en la actualidad

Estudio con carácter científico e interdisciplinar de los estudios físicos, químicos y visuales de los aspectos materiales e históricos de las pinturas tardogóticas en Castilla, y su difusión a través de las redes sociales de forma visual y didáctica

MicroRNA Alterations in a Tg501 Mouse Model of Prion Disease

MicroRNAs (miRNAs) may contribute to the development and pathology of many neurodegenerative diseases, including prion diseases. They are also promising biomarker candidates due to their stability in body fluids. We investigated miRNA alterations in a Tg501 mouse model of prion diseases that expresses a transgene encoding the goat prion protein (PRNP). Tg501 mice intracranially inoculated with mouse-adapted goat scrapie were compared with age-matched, mock inoculated controls in preclinical and clinical stages. Small RNA sequencing from the cervical spinal cord indicated that miR-223-3p, miR-151-3p, and miR-144-5p were dysregulated in scrapie-inoculated animals before the onset of symptoms. In clinical-stage animals, 23 significant miRNA alterations were found. These miRNAs were predicted to modify the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways including prion disease, extracellular matrix interactions, glutaminergic synapse, axon guidance, and transforming growth factor-beta signaling. MicroRNAs miR-146a-5p (up in cervical spinal cord) and miR-342-3p (down in cervical spinal cord, cerebellum and plasma), both indicated in neurodegenerative diseases earlier, were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Minimal changes observed before the disease onset suggests that most miRNA alterations observed here are driven by advanced prion-associated pathology, possibly limiting their use as diagnostic markers. However, the results encourage further mechanistic studies on miRNA-regulated pathways involved in these neurodegenerative conditions

BAMBI and CHGA in Prion Diseases: Neuropathological Assessment and Potential Role as Disease Biomarkers

Prion diseases affect both animals and humans. Research in the natural animal model of the disease could help in the understanding of neuropathological mechanisms and in the development of biomarkers for human pathologies. For this purpose, we studied the expression of 10 genes involved in prion propagation in vitro in the central nervous system of scrapie-infected sheep. Dysregulated genes (BAMBI and CHGA) were further analysed in a transgenic murine model (Tg338) of scrapie, and their protein distribution was determined using immunohistochemistry and Western blot. Their potential as biomarkers was finally assessed using enzyme-linked immunosorbent assay (ELISA) in cerebrospinal fluid (CSF) of scrapie sheep and Creutzfeldt-Jakob disease (CJD) patients. Protein BAMBI was upregulated in highly affected brain areas and CHGA was overexpressed along the brain in both models. Moreover, BAMBI and CHGA immunostaining scores strongly correlated with spongiosis and microgliosis in mice. Finally, levels of BAMBI were significantly higher in the CSF of clinical sheep and CJD patients. In addition to their potential as biomarkers, our work confirms the role of BAMBI and CHGA in prion neuropathology in vivo, but besides prion replication, they seem to be involved in the characteristic neuroinflammatory response associated to prion infection