Ras, hSpry2 y PGA₁: efectos en la nducción de la vía ERK/Elk1 y en la vía de NF-ĸB

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Medicina. Departamento Bioquímica. Fecha de lectura: 19 de Diciembre del 200

Sprouty2 and Spred1-2 Proteins Inhibit the Activation of the ERK Pathway Elicited by Cyclopentenone Prostanoids

Sprouty and Spred proteins have been widely implicated in the negative regulation of the fibroblast growth factor receptor-extracellular regulated kinase (ERK) pathway. In considering the functional role of these proteins, we explored their effects on ERK activation induced by cyclopentenone prostanoids, which bind to and activate Ras proteins. We therefore found that ectopic overexpression in HeLa cells of human Sprouty2, or human Spred1 or 2, inhibits ERK1/2 and Elk-1 activation triggered by the cyclopentenone prostanoids PGA1 and 15d-PGJ2. Furthermore, we found that in HT cells that do not express Sprouty2 due to hypermethylation of its gene-promoter, PGA1-provoked ERK activation was more intense and sustained compared to other hematopoietic cell lines with unaltered Sprouty2 expression. Cyclopentenone prostanoids did not induce Sprouty2 tyrosine phosphorylation, in agreement with its incapability to activate tyrosine-kinase receptors. However, Sprouty2 Y55F, which acts as a defective mutant upon tyrosine-kinase receptor stimulation, did not inhibit cyclopentenone prostanoids-elicited ERK pathway activation. In addition, Sprouty2 did not affect the Ras-GTP levels promoted by cyclopentenone prostanoids. These results unveil both common and differential features in the activation of Ras-dependent pathways by cyclopentenone prostanoids and growth factors. Moreover, they provide the first evidence that Sprouty and Spred proteins are negative regulators of the ERK/Elk-1 pathway activation induced not only by growth-factors, but also by reactive lipidic mediators

Treatment with tocilizumab or corticosteroids for COVID-19 patients with hyperinflammatory state: a multicentre cohort study (SAM-COVID-19)

Objectives: The objective of this study was to estimate the association between tocilizumab or corticosteroids and the risk of intubation or death in patients with coronavirus disease 19 (COVID-19) with a hyperinflammatory state according to clinical and laboratory parameters. Methods: A cohort study was performed in 60 Spanish hospitals including 778 patients with COVID-19 and clinical and laboratory data indicative of a hyperinflammatory state. Treatment was mainly with tocilizumab, an intermediate-high dose of corticosteroids (IHDC), a pulse dose of corticosteroids (PDC), combination therapy, or no treatment. Primary outcome was intubation or death; follow-up was 21 days. Propensity score-adjusted estimations using Cox regression (logistic regression if needed) were calculated. Propensity scores were used as confounders, matching variables and for the inverse probability of treatment weights (IPTWs). Results: In all, 88, 117, 78 and 151 patients treated with tocilizumab, IHDC, PDC, and combination therapy, respectively, were compared with 344 untreated patients. The primary endpoint occurred in 10 (11.4%), 27 (23.1%), 12 (15.4%), 40 (25.6%) and 69 (21.1%), respectively. The IPTW-based hazard ratios (odds ratio for combination therapy) for the primary endpoint were 0.32 (95%CI 0.22-0.47; p < 0.001) for tocilizumab, 0.82 (0.71-1.30; p 0.82) for IHDC, 0.61 (0.43-0.86; p 0.006) for PDC, and 1.17 (0.86-1.58; p 0.30) for combination therapy. Other applications of the propensity score provided similar results, but were not significant for PDC. Tocilizumab was also associated with lower hazard of death alone in IPTW analysis (0.07; 0.02-0.17; p < 0.001). Conclusions: Tocilizumab might be useful in COVID-19 patients with a hyperinflammatory state and should be prioritized for randomized trials in this situatio

A first update on mapping the human genetic architecture of COVID-19

peer reviewe

Modification and activation of Ras proteins by electrophilic prostanoids with different structure are site-selective

Cyclopentenone prostanoids (cyP) arise as important modulators of inflammation and cell proliferation. Although their physiological significance has not been fully elucidated, their potent biological effects have spurred their study as leads for the development of therapeutic agents. A key determinant of cyP action is their ability to bind to thiol groups in proteins or in glutathione through Michael addition. Even though several protein targets for cyP addition have been identified, little is known about the structural determinants from the protein or the cyP that drive this modification. The results herein presented provide the first evidence that cyP with different structures target distinct thiol sites in a protein molecule, namely, H-Ras. Whereas 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) and Δ12-PGJ2 preferentially target the C-terminal region containing cysteines 181 and 184, PGA1 and 8-iso-PGA1 bind mainly to cysteine 118, located in the GTP-binding motif. The biological counterparts of this specificity are the site-selective modification and activation of H-Ras in cells and the differential interaction of cyP with H, N, and K-Ras proteins. Cysteine 184 is unique to H-Ras, whereas cysteine 118 is present in the three Ras homologues. Consistent with this, PGA1 binds to and activates H-, N-, and K-Ras, thus differing from the preferential interaction of 15d-PGJ2 with H-Ras. These results put forward the possibility of influencing the selectivity of cyP-protein addition by modifying cyP structure. Furthermore, they may open new avenues for the development of cyP-based drugs. © 2007 American Chemical Society.Peer Reviewe

On the relationship between N content, textural properties and catalytic performance for the oxygen reduction reaction of N/CNT

Non-precious metal catalysts for the oxygen reduction reaction (ORR) based upon the incorporation of different amounts of N into the CC network of multiwalled carbon nanotubes (CNTs) have been prepared by using CNTs and urea as the carbon and nitrogen sources, respectively. First, and with the aim of generating different levels of defects in the carbon network, the CNTs have been subjected to ballmilling during different periods of time between 0 and 150 h. Then, urea was mixed with the treated CNTs, subjected to further ballmilling and pyrolized at 800 °C. The number of defects, and as a consequence, the amount of N incorporated into the CNTs, increases with the duration of the ballmilling time. Moreover, the structure of the CNTs obtained after longer ballmilling times collapses leading to a carbon material with a high degree of microporisity. The performance of the N/CNT for the ORR, in terms of both the onset potential and mass current activity, increases with the amount of N actually incorporated into the CNT. Moreover, the H2O2 formation during ORR varies with the morphology of the catalyst. Thus, the formation of H2O2 is favored with the electrocatalysts in which the CNT structure is preserved, whereas the total reduction of O2 to H2O is favored for the electrocatalysts in which micropores are formed.This project was funded by the Deanship of Scientific Research (DSR), King Abdulaziz University, Jeddah, under grant number (D-006-432). The authors, therefore, acknowledge with thanks DSR technical and financial support. Economic support from Project 201080E116 from the CSIC is also acknowledged.Peer Reviewe

On the relationship between the amount of N and ORR performance of N/Carbon nanotube electrodes

Trabajo presentado en la European Hydrogen Energy Conference - EHEC, celebrada en Sevilla (España) del 12 al 14 de mayo de 2014.Peer Reviewe

Endothelial nitric oxide synthase regulates N-Ras activation on the Golgi complex of antigen-stimulated T cells

Ras/ERK signaling plays an important role in T cell activation and development. We recently reported that endothelial nitric oxide synthase (eNOS)-derived NO regulates T cell receptor (TCR)- dependent ERK activation by a cGMP-independent mechanism. Here, we explore the mechanisms through which eNOS exerts this regulation. We have found that eNOS-derived NO positively regulates Ras/ERK activation in T cells stimulated with antigen on antigenpresenting cells (APCs). Intracellular activation of N-, H-, and K-Ras was monitored with fluorescent probes in T cells stably transfected with eNOS-GFP or its G2A point mutant, which is defective in activity and cellular localization. Using this system, we demonstrate that eNOS selectively activates N-Ras but not K-Ras on the Golgi complex of T cells engaged with APC, even though Ras isoforms are activated in response to NO from donors. We further show that activation of N-Ras involves eNOS-dependent S-nitrosylation on Cys118, suggesting that upon TCR engagement, eNOS-derived NO directly activates N-Ras on the Golgi. Moreover, wild-type but not C118S N-Ras increased TCR-dependent apoptosis, suggesting that S-nitrosylation of Cys118 contributes to activation-induced T cell death. Our data define a signaling mechanism for the regulation of the Ras/ERK pathway based on the eNOS-dependent differential activation of N-Ras and K-Ras at specific cell compartments.Fondo de InvestigacionesInstituto de Salud Carlos IIIFundación ‘‘la Caixa"Centro Nacional de Investigaciones CardiovascularesBancajaComunidad Autonoma de MadridDepto. de Bioquímica y Biología MolecularFac. de FarmaciaTRUEpu

Effect of transition metal (M: Fe, Co or Mn) for the oxygen reduction reaction with non-precious metal catalysts in acid medium

The effect of the metal for the oxygen reduction reaction (ORR) in acid medium with non-precious metal catalysts has been investigated. A series of non-precious metal catalysts with typical formulation M/N/C with M being Mn, Co or Fe have been prepared by incorporating N onto an active carbon matrix by means of thermal treatments under inert atmospheres. The N-containing active carbons were further treated with the M-containing precursors based upon Mn, Co or Fe phthalocyanines and thermally treated under inert atmosphere. The performance for the ORR in acid medium of all of the catalysts has been evaluated by means of electrochemical techniques. The activity, both in terms of onset potential for the ORR and maximum current density at representative potentials between 900 and 700 mV follows the trend Fe > Co > Mn. In addition, the performance of the Fe-based catalysts obtained during the different stages of the catalyst preparation has been also evaluated. The catalysts obtained after the pyrolysis step are the only ones showing measurable rates for the ORR. Although the amount of N and Fe incorporated onto the carbon matrix decreases the pyrolysis treatment, this treatment leads to the formation of the real active sites for the ORR irrespectively of the nature of the transition metal.This project was funded by the Deanship of Scientific Research (DSR), King Abdulaziz University, Jeddah, under grant number (D-006-432). The authors, therefore, acknowledge with thanks DSR technical and financial support. Economic support from projects ENE2010-15381 from the Spanish Ministry of Science and Innovation and Project 201080E116 from the CSIC is also acknowledged.Peer Reviewe