Regulation of the Na+/K+-ATPase Ena1 Expression by Calcineurin/Crz1 under High pH Stress: A Quantitative Study

[EN] Regulated expression of the Ena1 Na+-ATPase is a crucial event for adaptation to high salt and/or alkaline pH stress in the budding yeast Saccharomyces cerevisiae. ENA1 expression is under the control of diverse signaling pathways, including that mediated by the calcium-regulatable protein phosphatase calcineurin and its downstream transcription factor Crz1. We present here a quantitative study of the expression of Ena1 in response to alkalinization of the environment and we analyze the contribution of Crz1 to this response. Experimental data and mathematical models substantiate the existence of two stress-responsive Crz1-binding sites in the ENA1 promoter and estimate that the contribution of Crz1 to the early response of the ENA1 promoter is about 60%. The models suggest the existence of a second input with similar kinetics, which would be likely mediated by high pH-induced activation of the Snf1 kinase.This work was supported by grants BFU2011-30197-C3-01, BFU2014-54591-C2-1-P and EUI2009-04147 (SysMo2) to JA. (Ministry of Industry and Competitivity, Spain, and Fondo Europeo de Desarrollo Regional [FEDER]). JA is the recipient of an Ajut 2014SGR-4 award (Generalitat de Catalunya). DC was recipient of a predoctoral fellowship from the Spanish Ministry of Education. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Petrezsélyová, S.; López-Malo, M.; Canadell, D.; Roque, A.; Serra-Cardona, A.; Marques Romero, MC.; Vilaprinyó, E.... (2016). Regulation of the Na+/K+-ATPase Ena1 Expression by Calcineurin/Crz1 under High pH Stress: A Quantitative Study. PLoS ONE. 11(6):e0158424-e0158424. https://doi.org/10.1371/journal.pone.0158424Se0158424e015842411

The evolution of the ventilatory ratio is a prognostic factor in mechanically ventilated COVID-19 ARDS patients

Background: Mortality due to COVID-19 is high, especially in patients requiring mechanical ventilation. The purpose of the study is to investigate associations between mortality and variables measured during the first three days of mechanical ventilation in patients with COVID-19 intubated at ICU admission. Methods: Multicenter, observational, cohort study includes consecutive patients with COVID-19 admitted to 44 Spanish ICUs between February 25 and July 31, 2020, who required intubation at ICU admission and mechanical ventilation for more than three days. We collected demographic and clinical data prior to admission; information about clinical evolution at days 1 and 3 of mechanical ventilation; and outcomes. Results: Of the 2,095 patients with COVID-19 admitted to the ICU, 1,118 (53.3%) were intubated at day 1 and remained under mechanical ventilation at day three. From days 1 to 3, PaO2/FiO2 increased from 115.6 [80.0-171.2] to 180.0 [135.4-227.9] mmHg and the ventilatory ratio from 1.73 [1.33-2.25] to 1.96 [1.61-2.40]. In-hospital mortality was 38.7%. A higher increase between ICU admission and day 3 in the ventilatory ratio (OR 1.04 [CI 1.01-1.07], p = 0.030) and creatinine levels (OR 1.05 [CI 1.01-1.09], p = 0.005) and a lower increase in platelet counts (OR 0.96 [CI 0.93-1.00], p = 0.037) were independently associated with a higher risk of death. No association between mortality and the PaO2/FiO2 variation was observed (OR 0.99 [CI 0.95 to 1.02], p = 0.47). Conclusions: Higher ventilatory ratio and its increase at day 3 is associated with mortality in patients with COVID-19 receiving mechanical ventilation at ICU admission. No association was found in the PaO2/FiO2 variation

Impact of the SARS-CoV-2 (COVID19) pandemic on the morbidity and mortality of high risk patients undergoing surgery: a non-inferiority retrospective observational study

Background: During the COVID-19 crisis it was necessary to generate a specific care network and reconvert operating rooms to attend emergency and high-acuity patients undergoing complex surgery. The aim of this study is to classify postoperative complications and mortality and to assess the impact that the COVID-19 pandemic may have had on the results. Methods: this is a non-inferiority retrospective observational study. Two different groups of surgical patients were created: Pre-pandemic COVID and Pandemic COVID. Severity of illness was rated according to the Diagnosis-related Groups (DRG) score. Comparisons were made between groups and between DRG severity score-matched samples. Non-inferiority was set at up to 10 % difference for grade III to V complications according to the Clavien-Dindo classification, and up to 2 % difference in mortality. Results: A total of 1649 patients in the PreCOVID group and 763 patients in the COVID group were analysed; 371 patients were matched for DRG severity score 3-4 (236 preCOVID and 135 COVID). No differences were found in relation to re-operation (22.5 % vs. 21.5 %) or late admission to critical care unit (5.1 % vs. 4.5 %). Clavien grade III to V complications occurred in 107 patients (45.3 %) in the PreCOVID group and in 56 patients (41.5 %) in the COVID group, and mortality was 12.7 % and 12.6 %, respectively. During the pandemic, 3 % of patients tested positive for Covid-19 on PCR: 12 patients undergoing elective surgery and 11 emergency surgery; there were 5 deaths, 3 of which were due to respiratory failure following Covid-19-induced pneumonia. Conclusions: Although this study has some limitations, it has shown the non-inferiority of surgical outcomes during the COVID pandemic, and indicates that resuming elective surgery is safe

Differential expression of long non-coding RNAs are related to proliferation and histological diversity in follicular lymphomas

Long non‐coding RNA s (lncRNA s) comprise a family of non‐coding transcripts that are emerging as relevant gene expression regulators of different processes, including tumour development. To determine the possible contribution of lncRNA to the pathogenesis of follicular lymphoma (FL ) we performed RNA ‐sequencing at high depth sequencing in primary FL samples ranging from grade 1‐3A to aggressive grade 3B variants using unpurified (n = 16) and purified (n = 12) tumour cell suspensions from nodal samples. FL grade 3B had a significantly higher number of differentially expressed lncRNA s (dif‐lncRNA s) with potential target coding genes related to cell cycle regulation. Nine out of the 18 selected dif‐lncRNA s were validated by quantitative real time polymerase chain reaction in an independent series (n = 43) of FL . RP 4‐694A7.2 was identified as the top deregulated lncRNA potentially involved in cell proliferation. RP 4‐694A7.2 silencing in the WSU ‐FSCCL FL cell line reduced cell proliferation due to a block in the G1/S phase. The relationship between RP 4‐694A7.2 and proliferation was confirmed in primary samples as its expression levels positively related to the Ki‐67 proliferation index. In summary, lncRNA s are differentially expressed across the clinico‐biological spectrum of FL and a subset of them, related to cell cycle, may participate in cell proliferation regulation in these tumours.Fil: Roisman, Alejandro. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; España. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Castellano, Giancarlo. Universidad de Barcelona; EspañaFil: Navarro, Alba. Centro de Investigación Biomédica en Red de Cáncer; España. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; EspañaFil: Gonzalez Farre, Blanca. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; España. Universidad de Barcelona; EspañaFil: Pérez Galan, Patricia. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; EspañaFil: Esteve Codina, Anna. Universitat Pompeu Fabra; España. Barcelona Institute Of Science And Technology.; EspañaFil: Dabad, Marc. Barcelona Institute Of Science And Technology.; España. Universitat Pompeu Fabra; EspañaFil: Heath, Simon. Universitat Pompeu Fabra; España. Barcelona Institute Of Science And Technology.; EspañaFil: Gut, Marta. Universitat Pompeu Fabra; España. Barcelona Institute Of Science And Technology.; EspañaFil: Bosio, Mattia. Barcelona Supercomputing Center - Centro Nacional de Supercomputacion; EspañaFil: Bellot, Pau. Barcelona Institute Of Science And Technology. Institut Català D'investigació Química.; EspañaFil: Salembier, Philippe. Universidad Politécnica de Catalunya; EspañaFil: Oliveras, Albert. Universidad Politécnica de Catalunya; EspañaFil: Slavutsky, Irma Rosa. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Magnano, Andrea Laura. Hospital Clínico de Barcelona; España. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Horn, Heike. Eberhard Karls Universität Tübingen.; AlemaniaFil: Rosenwald, Andreas. University of Würzburg; AlemaniaFil: Ott, German. Department of Clinical Pathology; AlemaniaFil: Aymerich, Marta. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; EspañaFil: López Guillermo, Armando. Hospital Clínico de Barcelona; EspañaFil: Jares, Pedro. Universidad de Barcelona; España. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; EspañaFil: Martín Subero, José I.. Centro de Investigación Biomédica en Red de Cáncer; España. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; EspañaFil: Campo, Elías. Centro de Investigación Biomédica en Red de Cáncer; España. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; España. Hospital Clínico de Barcelona; EspañaFil: Hernández, Luis. Lymphoid Neoplasm Programme, Institut d'Investigacions Biomèdiques August Pi i Sunyer; España. Centro de Investigación Biomédica en Red de Cáncer; Españ

Guidelines for the use and interpretation of assays for monitoring autophagy

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Guidelines for the use and interpretation of assays for monitoring autophagy

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field