Estudio del proceso de bruñido de superficies complejas

Millorar l’acabat superficial d’una peça plana o cilíndrica, pot fer-se fàcilment mitjançant un procés de rectificat, tanmateix, millorar la qualitat superficial d’una superfície de configuració complexa (com motlles o matrius) és un problema difícil de tractar. Per aquest motiu, es recorre al procés conegut com a “brunyit”. El brunyit és una operació tecnològica que s’aplica després del mecanitzat i que deforma plàsticament les irregularitats superficials de la peça mitjançant l’ús d’una eina que aplica força sobre la zona de treball. En aquest projecte, es descriu el procés de brunyit d’una superfície complexa amb una eina de puntes intercanviables que té com a element que brunyeix una punta esfèrica de carbur de Tungstè (o carbur de Wolframi), a diferència del brunyit convencional que es realitza mitjançant l’ús d’una esfera que rota sobre si mateixa. L’objectiu principal d’aquest treball és definir els paràmetres òptims de funcionament de l’eina de treball amb la finalitat de determinar quins permeten obtenir millors acabats, arribant a valors de rugositat similars als obtinguts en un procés de polit manual. Per a aconseguir-ho, s’han realitzat dos experiments basats en el brunyit de diverses mostres sotmeses a diferents combinacions de paràmetres: força, velocitat de rotació, passada lateral, etc. El material de la peça sobre la qual s’han fet els assajos és un bloc d’acer 1045 (o C45). Un cop analitzades les mostres, no s’aconsegueixen determinar els paràmetres òptims de funcionament de l’eina. No obstant això, es determinen paràmetres recomanables que milloren dràsticament l’acabat superficial de la peça quan es treballa amb mostres d’acer 1045. Pel que fa al brunyit de superfícies complexes, es determina que treballant amb velocitats de rotació elevades (n > 600 rpm) i amb forces inferiors a 100 N, s’obtenen rugositats Ra per sota de 0,6µm, independentment de si han estat brunyides en una fresadora de 3 o 5 eixos. En canvi, no es recomana l’ús d’aquesta eina quan es requereixi millorar l’acabat de superfícies planes, ja que s’obtenen valors de rugositat per sobre de 0,8 µm, arribant a superar 1 µm.Mejorar el acabado superficial de una pieza plana o cilíndrica puede hacerse fácilmente mediante un proceso de rectificado, sin embargo, mejorar la calidad superficial de una superficie de configuración geométrica compleja (como moldes y matrices) es un problema difícil de tratar. Por ello, se recurre al proceso conocido con el nombre de “bruñido”. El bruñido es una operación tecnológica que se aplica después del mecanizado y que deforma plásticamente las irregularidades superficiales de la pieza mediante el uso de una herramienta que aplica fuerza sobre la zona de trabajo. En este proyecto, se describe el proceso de bruñido de una superficie compleja con una herramienta de puntas intercambiables que tiene como elemento que bruñe una punta esférica de carburo de Tungsteno (o carburo de Wolframio), a diferencia del bruñido convencional que se realiza mediante el uso de una esfera que rota sobre sí misma. El objetivo principal de este trabajo es definir los parámetros óptimos de funcionamiento de la herramienta de trabajo con el fin de determinar cuáles permiten obtener mejores acabados, llegando a valores de rugosidad similares a los obtenidos en un proceso de pulido manual. Para ello, se han realizado dos experimentos basados en el bruñido de varias muestras sometidas a distinta combinación de parámetros: fuerza, velocidad de rotación de la herramienta, paso lateral, etc. El material de la pieza sobre la que se han realizado los ensayos es un bloque de acero 1045 (o C45). Tras el análisis de los distintos ensayos, no se logran determinar los parámetros óptimos de funcionamiento de la herramienta. Sin embargo, se obtienen parámetros recomendables que conllevan una mejoría drástica en el acabado de la pieza cuando se trabaja con muestras de acero 1045. Por lo que refiere al bruñido de superficies complejas, se determina que trabajando a velocidades de rotación elevadas (n > 600 rpm) y con fuerzas aplicadas inferiores a 100 N, se obtienen rugosidades Ra por debajo de 0,6 µm, independientemente de si han sido bruñidas en una fresadora de 3 o de 5 ejes. En cambio, no se recomienda el uso de esta herramienta cuando se quiera mejorar el acabado de superficies planas, ya que se obtienen valores de rugosidad por encima de 0,8 µm, llegando a superar 1 µm.The finishing of a flat or cylindrical part can be easily done by a grinding process, however, to improve the surface quality of a complex surface (such as molds and dies) is a difficult problem to deal with. Therefore, the process known as “burnishing” is used. Burnishing is a technological operation that is applied after machining process and that plastically deforms surface irregularities by using a tool that applies force on the selected area. In this project, the process of burnishing a complex surface with an interchangeable tip tool is described. The tool has as a burnishing element a spherical tip of Tungsten carbide, unlike conventional burnishing which is done by using a sphere that rotates on itself. The main objective of this project is to define the optimal operating parameters of the tool in order to determine which allow to obtain better finishes, reaching roughness values similar to those obtained in a manual polishing process. Therefore, two experiments have been carried out based on the burnishing of several samples subjected to different combinations of parameters like: force, tool rotation speed, step, etc. The material that has been used for the experiments is 1045 (or C45) Steel. After the analysis of the different experiments, it has been not possible to determine the optimal operating parameters of the tool. However, some recommended parameters that lead to a drastic improvement in the finish of the part when working with 1045 Steel samples are obtained. Regarding the burnishing of complex surfaces, it is determined that working at high rotation speeds (n > 600 rpm) and with applied forces lower than 100 N, roughness Ra < 0,6 µm is obtained, regardless of whether they have been done on a 3 o 5-axis milling machine. On the other hand, the use of this tool is not recommended if you want to improve the finish of a flat surface, since roughness values above 0,8 µm are obtained, reaching more than 1 µm in some cases

Two unconventional Metallothioneins in the apple snail Pomacea Bridgesii have lost their metal specificity during adaptation to freshwater habitats

Metallothioneins (MTs) are a diverse group of proteins responsible for the control of metal homeostasis and detoxification. To investigate the impact that environmental conditions might have had on the metal-binding abilities of these proteins, we have characterized the MTs from the apple snail Pomacea bridgesii, a gastropod species belonging to the class of Caenogastropoda with an amphibious lifestyle facing diverse situations of metal bioavailability. P. bridgesii has two structurally divergent MTs, named PbrMT1 and PbrMT2, that are longer than other gastropod MTs due to the presence of extra sequence motifs and metal-binding domains. We have characterized the Zn(II), Cd(II), and Cu(I) binding abilities of these two MTs after their heterologous expression in E. coli. Our results have revealed that despite their structural differences, both MTs share an unspecific metal-binding character, and a great ability to cope with elevated amounts of different metal ions. Our analyses have also revealed slight divergences in their metal-binding features: PbrMT1 shows a more pronounced Zn(II)-thionein character than PbrMT2, while the latter has a stronger Cu(I)-thionein character. The characterization of these two unconventional PbrMTs supports the loss of the metal-binding specificity during the evolution of the MTs of the Ampullariid family, and further suggests an evolutionary link of this loss with the adaptation of these gastropod lineages to metal-poor freshwater habitats

Modular Evolution and Population Variability of Oikopleura dioica Metallothioneins

Chordate Oikopleura dioica probably is the fastest evolving metazoan reported so far, and thereby, a suitable system in which to explore the limits of evolutionary processes. For this reason, and in order to gain new insights on the evolution of protein modularity, we have investigated the organization, function and evolution of multi-modular metallothionein (MT) proteins in O. dioica. MTs are a heterogeneous group of modular proteins defined by their cysteine (C)-rich domains, which confer the capacity of coordinating different transition metal ions. O. dioica has two MTs, a bi-modular OdiMT1 consisting of two domains (t-12C and 12C), and a multi-modular OdiMT2 with six t-12C/12C repeats. By means of mass spectrometry and spectroscopy of metal-protein complexes, we have shown that the 12C domain is able to autonomously bind four divalent metal ions, although the t-12C/12C pair -as it is found in OdiMT1- is the optimized unit for divalent metal binding. We have also shown a direct relationship between the number of the t-12C/12C repeats and the metal-binding capacity of the MTs, which means a stepwise mode of functional and structural evolution for OdiMT2. Finally, after analyzing four different O. dioica populations worldwide distributed, we have detected several OdiMT2 variants with changes in their number of t-12C/12C domain repeats. This finding reveals that the number of repeats fluctuates between current O. dioica populations, which provides a new perspective on the evolution of domain repeat proteins

Metal-Specificity Divergence between Metallothioneins of Nerita peloronta (Neritimorpha, Gastropoda) Sets the Starting Point for a Novel Chemical MT Classification Proposal

Metallothioneins' (MTs) biological function has been a matter of debate since their discovery. The importance to categorize these cysteine-rich proteins with high coordinating capacity into a specific group led to numerous classification proposals. We proposed a classification based on their metal-binding abilities, gradually sorting them from those with high selectivity towards Zn/Cd to those that are Cu-specific. However, the study of the NpeMT1 and NpeMT2isoforms of Nerita peloronta, has put a new perspective on this classification. N. peloronta has been chosen as a representative mollusk to elucidate the metal-binding abilities of Neritimorpha MTs, an order without any MTs characterized recently. Both isoforms have been recombinantly synthesized in cultures supplemented with ZnII, CdII, or CuII, and the purified metal-MT complexes have been thoroughly characterized by spectroscopic and spectrometric methods, leading to results that confirmed that Neritimorpha share Cd-selective MTs with Caenogastropoda and Heterobranchia, solving a so far unresolved question. NpeMTs show high coordinating preferences towards divalent metal ions, although one of them (NpeMT1) shares features with the so-called genuine Zn-thioneins, while the other (NpeMT2) exhibits a higher preference for Cd. The dissimilarities between the two isoforms let a window open to a new proposal of chemical MT classification

The Modular architecture of metallothioneins facilitates domain rearrangements and contributes to their evolvability in metal-accumulating mollusks.

Protein domains are independent structural and functional modules that can rearrange to create new proteins. While the evolution of multidomain proteins through the shuffling of different preexisting domains has been well documented, the evolution of domain repeat proteins and the origin of new domains are less understood. Metallothioneins (MTs) provide a good case study considering that they consist of metal-binding domain repeats, some of them with a likely de novo origin. In mollusks, for instance, most MTs are bidomain proteins that arose by lineage-specific rearrangements between six putative domains: α, β1, β2, β3, γ and δ. Some domains have been characterized in bivalves and gastropods, but nothing is known about the MTs and their domains of other Mollusca classes. To fill this gap, we investigated the metal-binding features of NpoMT1 of Nautilus pompilius (Cephalopoda class) and FcaMT1 of Falcidens caudatus (Caudofoveata class). Interestingly, whereas NpoMT1 consists of α and β1 domains and has a prototypical Cd2+ preference, FcaMT1 has a singular preference for Zn2+ ions and a distinct domain composition, including a new Caudofoveata-specific δ domain. Overall, our results suggest that the modular architecture of MTs has contributed to MT evolution during mollusk diversification, and exemplify how modularity increases MT evolvability

The Modular Architecture of Metallothioneins Facilitates Domain Rearrangements and Contributes to Their Evolvability in Metal-Accumulating Mollusks

Protein domains are independent structural and functional modules that can rearrange to create new proteins. While the evolution of multidomain proteins through the shuffling of different preexisting domains has been well documented, the evolution of domain repeat proteins and the origin of new domains are less understood. Metallothioneins (MTs) provide a good case study considering that they consist of metal-binding domain repeats, some of them with a likely de novo origin. In mollusks, for instance, most MTs are bidomain proteins that arose by lineage-specific rearrangements between six putative domains: α, β1, β2, β3, γ and δ. Some domains have been characterized in bivalves and gastropods, but nothing is known about the MTs and their domains of other Mollusca classes. To fill this gap, we investigated the metal-binding features of NpoMT1 of Nautilus pompilius (Cephalopoda class) and FcaMT1 of Falcidens caudatus (Caudofoveata class). Interestingly, whereas NpoMT1 consists of α and β1 domains and has a prototypical Cd preference, FcaMT1 has a singular preference for Zn ions and a distinct domain composition, including a new Caudofoveata-specific δ domain. Overall, our results suggest that the modular architecture of MTs has contributed to MT evolution during mollusk diversification, and exemplify how modularity increases MT evolvability

Reduced heart rate variability predicts fatigue severity in individuals with chronic fatigue syndrome/myalgic encephalomyelitis

Heart rate variability (HRV) is an objective, non-invasive tool to assessing autonomic dysfunction in chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME). People with CFS/ME tend to have lower HRV; however, in the literature there are only a few previous studies (most of them inconclusive) on their association with illness-related complaints. To address this issue, we assessed the value of different diurnal HRV parameters as potential biomarker in CFS/ME and also investigated the relationship between these HRV indices and self-reported symptoms in individuals with CFS/ME. In this case-control study, 45 female patients who met the 1994 CDC/Fukuda definition for CFS/ME and 25 age- and gender-matched healthy controls underwent HRV recording-resting state tests. The intervals between consecutive heartbeats (RR) were continuously recorded over three 5-min periods. Time- and frequency-domain analyses were applied to estimate HRV variables. Demographic and clinical features, and self-reported symptom measures were also recorded. CFS/ME patients showed significantly higher scores in all symptom questionnaires (p < 0.001), decreased RR intervals (p < 0.01), and decreased HRV time- and frequency-domain parameters (p < 0.005), except for the LF/HF ratio than in the healthy controls. Overall, the correlation analysis reached significant associations between the questionnaires scores and HRV time- and frequency-domain measurements (p < 0.05). Furthermore, separate linear regression analyses showed significant relationships between self-reported fatigue symptoms and mean RR (p = 0.005), RMSSD (p = 0.0268) and HFnu indices (p = 0.0067) in CFS/ME patients, but not in healthy controls. Our findings suggest that ANS dysfunction presenting as increased sympathetic hyperactivity may contribute to fatigue severity in individuals with ME/CFS. Further studies comparing short- and long-term HRV recording and self-reported outcome measures with previous studies in larger CFS/ME cohorts are urgently warranted

Modularity in protein evolution: modular organization and de novo domain evolution in mollusc metallothionein

Metallothioneins (MTs) are proteins devoted to the control of metal homeostasis and detoxification, and therefore, MTs have been crucial for the adaptation of the living beings to variable situations of metal bioavailability. The evolution of MTs is, however, not yet fully understood, and to provide new insights into it, we have investigated the MTs in the diverse classes of Mollusks. We have shown that most molluskan MTs are bimodular proteins that combine six domains α, β1, β2, β3, γ, and δ in a lineage-specific manner. We have functionally characterized the Neritimorpha β3β1 and the Patellogastropoda γβ1 MTs, demonstrating the metal-binding capacity of the new γ domain. Our results have revealed a modular organization of mollusk MT, whose evolution has been impacted by duplication, loss, and de novo emergence of domains. MTs represent a paradigmatic example of modular evolution probably driven by the structural and functional requirements of metal binding

Understanding the formation of heartwood in Larch using synchrotron infrared imaging combined with multivariate analysis and atomic force microscope infrared spectroscopy

Formation of extractive-rich heartwood is a process in live trees that make them and the wood obtained from them more resistant to fungal degradation. Despite the importance of this natural mechanism, little is known about the deposition pathways and cellular level distribution of extractives. Here we follow heartwood formation in Larix gmelinii var. Japonica by use of synchrotron infrared images analyzed by the unmixing method Multivariate Curve Resolution - Alternating Least Squares (MCR-ALS). A subset of the specimens was also analyzed using atomic force microscopy infrared spectroscopy. The main spectral changes observed in the transition zone when going from sapwood to heartwood was a decrease in the intensity of a peak at approximately 1660 cm-1 and an increase in a peak at approximately 1640 cm-1. There are several possible interpretations of this observation. One possibility that is supported by the MCR-ALS unmixing is that heartwood formation in larch is a type II or Juglans-type of heartwood formation, where phenolic precursors to extractives accumulate in the sapwood rays. They are then oxidized and/or condensed in the transition zone and spread to the neighboring cells in the heartwood

The epidermal growth factor receptor (EGRF) in lung cancer

Altres ajuts: Work in Dr Rosell's laboratory is partially supported by a grant from Fundació La CaixaIn the last decade, important advances have been made in understanding of cancer biology, particularly non-small-cell lung cancer (NSCLC) with the discovery of oncogenic drivers of the disease. The epidermal growth factor receptor (EGFR) gene and its pathways was the first oncogenic driver discovered to be mutated and treatable in lung cancer. Treatment with EGFR tyrosine kinase inhibitors (TKIs) is the standard of care for molecularly selected EGFR -mutant patients, while its role in unselected lung cancer patients is nowadays controversial. This review will provide an overview of the EGFR pathway and options for its treatment of lung cancer