RICERCA DI MUTAZIONI NEL CANALE PACEMAKER HCN4 IN PAZIENTI AFFETTI DA DISTURBI DEL RITMO CARDIACO

Cardiovascular diseases are widespread and they are a major cause of death in the western part of the world. Therefore, identifying their causes precisely with the aim of establishing a preventive and therapeutic strategy has become subject of research also for their social and economic impact. Many studies have now clearly established that the genetic characteristics of a person can be a background predisposing to the onset of various diseases including cardiac arrhythmias. For example, various mutations in genes that code for ion channels are known and their accessory proteins that lead to the development of heart diseases such as the long QT syndrome and bradycardia. My laboratory of the PhD has shown a correlation between a specific mutation in the hHCN4 (Hyperpolarization-activate Cyclic Nucleotide-gated channel) and sinus bradycardia. The screening is a part of research of monogenic diseases and in particular the focus is on genes directly involved in the genesis of the rhythm (HCN channels) and on interacting proteins (CAV3, MiRP1, FLNA). In my thesis I performed a genetic screening in order to identify polymorphisms and mutations in the gene encoding for the hHCN4 pacemaker channel. The HCN4 protein channel is an isoform of the HCN family responsible of the If current. This channel is more expressed at the level of the sinus node, an anatomical region of the heart in which the source of the heart rhythm is located. SAN (sinus atrial node) cells are able to generate action potentials in an autonomous way that spread through the conduction system allowing the contraction of the heart. The current If plays a key role in the slow diastolic depolarization phase, where it leads the membrane potential to threshold values for a new action potential. The heart rate is determined by the duration of the diastolic depolarization phase: the principal current responsible is the If current. The genetic analysis was performed on the gene coding for the channel hHCN4 in patients with cardiac arrhythmias (sinus bradycardia, sinus tachycardia and atrial fibrillation) in order to identify genetic mutations causally related to the pathological phenotype. Among the various polymorphisms we found, the attention was focused on the R524Q mutation in HCN4, identified in patients of the same family with inappropriate sinus tachycardia (IST): it is a non-conservative missense mutation in heterozygosis (located on exon 4, position 1571G> A mRNA, codon CGG in CAG). R524 is located in the initial part of the C-linker, an important region which connects the last transmembrane domain (S6) to CNBD and which has a function in modulating channel gating. To test whether the R524Q polymorphism affects biophysical properties of the HCN4 pacemaker channel and consequently determines sinus tachycardia in patients carrying the mutation, we conducted electrophysiological experiments in heterologous cells (HEK 293) transfected with plasmids expressing the gene respectively hHCN4 wild type, mutated homozygous and heterozygous (hHCN4 coexpression of plasmids with wild-type and mutated hHCN4 homozygous). In some families that have inheritance of the disease, it was not possible to identify a mutation in HCN4. In collaboration with a specialized center that employs NGS (New Generation Sequencing) technology, the screening has been extended to other genes involved in heart disease. We selected two families: the first with bradycardia and atrial fibrillation, and the second with atrial fibrillation and tachycardia. The first experiments on probands have led to the identification of mutations that will be interesting to investigate in the future. In a vitro system we characterized the I57T mutation found in MiRP1, identified in the second family

A distributed approach for parameter estimation in Systems Biology models

Due to the lack of experimental measurements, biological variability and experimental errors, the value of many parameters of the systems biology Mathematical models is yet unknown or uncertain. A possible computational solution is the parameter estimation, that is the identification of the parameter values that determine the best model fitting respect to experimental data. We have developed an environment to distribute each run of the parameter estimation algorithm on a different computational resource. The key feature of the implementation is a relational database that allows the user to swap the candidate solutions among the working nodes during the computations. The comparison of the distributed implementation with the parallel one showed that the presented approach enables a faster and better parameter estimation of systems biology models

Applying the theory of real options to the optimal timing of timber harvests

Se propone el enfoque de opciones reales como herramienta económico-financiera para la toma de decisiones estratégicas en el sector forestal. En términos de instrumentos financieros, consideraremos en particular una opción exótica conocida como barrier option del tipo knock-in. Suponemos que la proyección del precio de venta promedio de los subproductos sigue un proceso estocástico del tipo Geométrico Browniano, mientras que la producción se determina mediante simulación de un turno forestal. La decisión de talar la masa forestal surge de comparar en cada periodo, el valor de flujo de fondos en cada nodo (FFij(t)) de una rejilla binomial con el valor esperado en el próximo año (X t+1 x e−rΔ t) ). En un ejemplo analizado el criterio tradicional del VAN indica que el mayor valor actual se produce en el instante t = 0 (año 10), mientras que el enfoque de opciones reales arroja que el máximo valor de ejercicio se da en el periodo t = 8 (año 18).We present here a real options approach as a tool for strategic decision-making in the forestry sector. We consider, in particular, an exotic option known as a knock-in barrier option. We use this approach to determine the optimal timing of harvests. The optimal time is determined by comparing at each period, the cash flow corresponding to each node (FFij(t)) in a binomial lattice with the expected value in the next year (X t+1 x e−rΔ t). The traditional NPV indicates, in an hypothetical context analyzed in the paper, that the largest present value is obtained at the tenth year of standing. The real options approach championed here, instead, indicates that the highest value is at the eighteenth year of standing.Fil: Milanesi, Gastón. Universidad Nacional del SurFil: Woitschach, Guillermo B. M.. Universidad Nacional del SurFil: Broz, Diego R.. Universidad Nacional del Su

Etringita y ceolitas formadas en hormigones afectados por la reacción álcali- sílice

En el presente trabajo se estudiaron hormigones deteriorados por efecto de la reacción álcali-sílice (RAS) con el propósito de identificar los diferentes productos de reacción presentes. Las muestras de hormigón investigadas fueron obtenidas a partir de testigos extraídos de una estructura de hormigón en servicio y de probetas moldeadas en laboratorio. Los estudios realizados mediante difractometría de rayos X (DRX) y microscopía electrónica de barrido (SEM) permitieron identificar principalmente dos productos de naturaleza cristalina: clinoptilolita y etringita. Se mencionan las principales características de cada uno de ellos y se comentan los probables mecanismos de formación en base a los conocimientos disponibles

An expansive dolostone from Argentina

Alkali-carbonate reaction is a known harmful reaction that affects concrete durability and constitutes an issue of much controversy between researchers and concrete technologists. Since 1991, the authors have been working in this field to evaluate the possibility of the occurrence of this reaction in Argentina. The studies have shown the existence of a fine-grained dolostone that reacts deleteriously with concrete alkalis following a mechanism similar to ACR. This dolostone has two distinctive characteristics. First of all, the rock shows a clear dedolomitization reaction producing calcite and brucite (no siliceous gel was observed in thin sections). Secondly, the use of known mineral admixtures (fly ash, silica fume, ground granulated blast-furnace slag, natural pozzolan) or lithium compounds does not mitigate concrete expansion. This paper summarizes the results obtained with this dolomitic rock using current characterization techniques (ASTM C 586, ASTM C 227, NBRI, concrete prism expansion test, and polarization microscopy)

Los procesos de deterioro en el hormigón y su relación con la formación de ettringita diferida

La detección de ettringita en hormigones afectados por algún tipo de deterioro prematuro, ha sido motivo de preocupación por numerosos investigadores en lo últimos años. Se han definido dos líneas principales de enfoque del tema. Una la considera como factor principal de la destrucción y la otra asume que su aparición es posterior al hecho precursor. Con la finalidad de contribuir a esclarecer este fenómeno, se proyectó una investigación tendiente a cubrir los aspectos antes citados. En este trabajo se informan las conclusiones obtenidas hasta la fecha, de estudios realizados sobre hormigones que han sido afectados por diferentes mecanismos destructivos, como congelamiento y deshielo, humedecimiento y secado, prefisuración y RAS. Se realizaron observaciones microscópicas, DRX, SEM, EDAX, estabilidad dimensional, porosidad y resistencia mecánica. La formación de ettringita en la mayoría de los casos estuvo vinculada a la existencia de circulación de líquido en la masa del hormigón pero sin definir claramente su participación en los procesos de destrucción

Traceability of the PGI product "Vitellone Bianco dell'Appennino Centrale" by SNP markers

AbstractTraceability of meat has become a key aspect of food-quality assurance and a priority for EU countries, to meet consumer demand for comprehensive and integrated food safety policies. In this context, the traceability of animals and animal products at the breed level might play a key role as it would enable the certification of regional products linked to particular breeds. Technologies based on DNA analysis have the potential to achieve this goal bypassing the large scale and systematic biological sampling necessary for individual fingerprinting.The objective of this work was to test the power of a SNP panel to trace PGI "Vitellone Bianco dell'Appennino Centrale", a product linked to three breeds: Chianina, Marchigiana and Romagnola.A total of 180 unrelated animals belonging to PGI-allowed breeds (Chianina n=22; Romagnola n=22; and Marchigiana n=22) and PGI-not allowed breeds (Piemontese n=22, Maremmana n=24, Italian Red Pied n=24, Italian Brown n=22, Italian Friesian n=22) were sampled and genoty..

Identification of polymorphism in the SCL24A5 gene of cattle

The SLC24A5 (Solute Carrier family 24, member 5) gene is implicated in skin pigmentation in zebrafish and humans as it regulates the morphogenesis of melanosomes, specialized lysosomes involved in melanin deposit. In humans, the ancestral allele predominates in African and East Asian populations, while the allelic variant is nearly fixed in European populations and correlates with lighter pigmentation. Considering the role of melanin in the protecting of DNA from ultraviolet radiation, the lack of information in cattle and the importance of polymorphisms associated with pigmentation phenotypes, we investigated the SLC24A5 gene in cattle with light and dark skin pigmentation. To identify SNPs (Single Nucleotide Polymorphisms) in this gene and their association to dark skin pigmentation in cattle, each of the nine SLC24A5 exons, three introns (1, 3 and 8) and a portion of intron 5, were sequenced in a set of sixteen animals belonging to four Italian cattle breeds, two African zebu breeds and two African sanga breeds. The region spanning exons 3 and 4 was sequenced in fifteen animals belonging to seven additional breeds. A total of sixteen SNPs were identified: eleven positioned in introns (six in intron 1, one in intron 5 and four in intron 8) and five in exons (one in exon 1, two in exon 6 and two in exon 7). Three SNPs (located in exons 1, 6 and 7) were non synonymous, determining Pro19Leu, Ala238Val, and Met341Ile amino acid changes, respectively. All the SNPs identified were polymorphic between Bos taurus, Bos indicus and Sanga, while none of them resulted associated with the studied phenotype and discriminated the three breeds (Chianina, Mucubal and Goudali) characterized by dark pigmented skin from the others