In vitro and in vivo development of mice morulae after storage in non-frozen conditions

Background Interchange of genetically modified (GM) mice between laboratories using embryos provides several advantages. Not only is transport stress avoided, but also the health status of the recipient colony is not compromised. Embryos do not need to be shipped in frozen stage, which requires expensive packaging in addition to a certain degree of expertise in order to freeze and thaw them correctly. The aim of this study was to examine different storage conditions and their effect on embryo viability in order to establish the feasibility of practical, non-frozen conditions for embryo shipment. Methods Mouse morulae developed in vivo (collected from donors 2.5d post coitum) or in vitro (zygotes cultured until morulae stage) were stored, combining two different media (KSOMeq or KSOM-H) and temperatures (4 degrees C, 15 degrees C and 37 degrees C) throughout 24 or 48 hours. After storage in vitro viability was assessed determining percentage of development to blastocyst and total cell number. In vivo viability was determined based on the number of implantations and living fetuses after embryo transfer of stored embryos. The storage effect at the molecular level was assessed by studying a gene pool involved in early development by quantitative RT-PCR. Results In vivo-produced morulae stored for 24 hours did not show differences in development up to the blastocyst stage, regardless of the storage type. Even though a decrease in the total cell number in vivo was observed, embryo development after embryo transfer was not affected. All 24 hour storage conditions tested provided a similar number of implantations and fetuses at day 14 of pregnancy. Morulae obtained from in vitro embryo culture collected at the 1-cell stage showed a decreased ability to develop to blastocyst after 24 hours of storage at 15degrees C both in KSOMeq and KSOM-H. Concomitantly, a significant decrease of embryo implantation rates after transfer to recipients was also found. In order to further characterize the effect of non-frozen storage combining a molecular approach with the ordinary in vitro culture evaluation, embryos collected at the morula stage were submitted to the same storage conditions described throughout 48 hours. In vitro culture of those embryos showed a significant decrease in their developmental rate to blastocyst in both KSOMeq and KSOM-H at 15degrees C, which also affected the total number of cells. Gene transcription studies confirmed significant alterations in retrotransposons (Erv4 and Iap) after 48 h of storage at 15degrees C. Conclusions Our results show that both KSOMeq and KSOM-H can be equally used, and that several temperature conditions allow good survival rates in vitro and in vivo. Some of these storage conditions can substitute freezing in order to maintain embryo viability for 24–48 hours, providing a reliable and less demanding technical alternative for embryo interchanges.This work was performed as part of, and financed by, Project AGL2004-00332. JDH received a Ph D. grant from the INIA (Ministry of Science and Innovation). We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its unit of Information Resources for Research (URICI).Peer Reviewe

Estudio experimental de procesos de calentamiento y enfriamiento. Resultados e implicaciones didácticas

A detailed experimental study on different processes of heating and cooling allows to deduce that temperature variation, in these processes, in the course of time is not always the same, but it depends, to a great extent, on how the process is carried out. So to confuse the T-Q relations (always lineal) wirth T-t relations (dependent on the process) is a mistake that is necessary to avoid in our classes

Cre recombinase microinjection for single-cell tracing and localised gene targeting.

Tracing and manipulating cells in embryos are essential to understand development. Lipophilic dye microinjections, viral transfection and iontophoresis have been key to map the origin of the progenitor cells that form the different organs in the post-implantation mouse embryo. These techniques require advanced manipulation skills and only iontophoresis, a demanding approach of limited efficiency, has been used for single-cell labelling. Here, we perform lineage tracing and local gene ablation using cell-permeant Cre recombinase (TAT-Cre) microinjection. First, we map the fate of undifferentiated progenitors to the different heart chambers. Then, we achieve single-cell recombination by titrating the dose of TAT-Cre, which allows clonal analysis of nascent mesoderm progenitors. Finally, injecting TAT-Cre to Mycnflox/flox embryos in the primitive heart tube revealed that Mycn plays a cell-autonomous role in maintaining cardiomyocyte proliferation. This tool will help researchers identify the cell progenitors and gene networks involved in organ development, helping to understand the origin of congenital defects.This research was supported by grant PGC2018-096486-B-I00 from the Spanish Ministerio de Ciencia e Innovación and grant H2020-MSCA-ITN-2016-722427 from the EU Horizon 2020 program to M.T. M.S. was supported by a “la Caixa” Foundation PhD fellowship (LCF/BQ/DE18/11670014) and The Company of Biologists travelling fellowship (DEVTF181145). O.H.O. is supported by the Ministerio de Ciencia e Innovación (grant RTI2018-097617-J-I00). J.N.D. received funding from grant 1380918 from the European Regional Development Fund Andalucıa 2014-2020 ́ Operating Program. Open access funding provided by Centro Nacional de Investigaciones Cardiovasculares. Deposited in PMC for immediate release.S

Scaling up climate finance in the context of Covid-19: A science-based call for financial decision-makers

The sooner we act, the lower the risks of climate change and the higher the synergies between climate action and other societal benefits. That is a clear conclusion from the IPCC Special Report on the impacts of global warming of 1.5°C above pre-industrial levels. Financing a rapid transition to achieve the Paris Agreement goals requires significantly more investment and investment in a different set of low emission, climate resilient assets. The Covid-19 crisis increases the imperative to scale up climate action before these goals are out of reach. In particular, it is critical to increase the ability of developing countries to realize their climate ambitions in the context of the pandemic without increasing their debt burden.This new report, Scaling up climate finance in the context of Covid-19, aims to help financial decision-makers to align finance with sustainable development, accelerating the transition to a net-zero, climate resilient economy, based on the latest scientific findings and policy developments. It proposes four sets of actions to support developing countries in achieving this transition.This report aims to help financial decision-makers to align finance with sustainable development, accelerating the transition to a net-zero, climate resilient economy, based on the latest scientific findings and policy developments. It proposes four interventions to achieve this objective in the context of Covid-19

PRI COMINTRISQ – PRI Des communautés internationales et des risques

Marie-Angèle Hermitte, Dominique Pestre, Francis Chateauraynaud, Jean-Charles Hourcade, directeurs d’étudesGiorgio Blundo, maître de conférencesSoraya Boudia, professeur à l’Université Strasbourg-I/Louis-PasteurRafael Encinas de Munagorri, professeur à l’Université de NantesJean-Michel Fourniau, directeur de recherche à l’IFSTTARChristelle Gramaglia, ingénieure de recherche au Cemagref Des communautés internationales et des risques. Science, justice et politique dans les crises contemporaines..

Using mutagenesis and structural biology to map the binding site for the plasmodium falciparum merozoite Protein PfRh4 on the human immune adherence receptor

To survive and replicate within the human host, malaria parasites must invade erythrocytes. Invasion can be mediated by the P. falciparum reticulocyte-binding homologue protein 4 (PfRh4) on the merozoite surface interacting with complement receptor type 1 (CR1, CD35) on the erythrocyte membrane. The PfRh4 attachment site lies within the three N-terminal complement control protein modules (CCPs 1–3) of CR1, which intriguingly also accommodate binding and regulatory sites for the key complement activation-specific proteolytic products, C3b and C4b. One of these regulatory activities is decay-accelerating activity. Although PfRh4 does not impact C3b/C4b binding, it does inhibit this convertase disassociating capability. Here, we have employed ELISA, co-immunoprecipitation, and surface plasmon resonance to demonstrate that CCP 1 contains all the critical residues for PfRh4 interaction. We fine mapped by homologous substitution mutagenesis the PfRh4-binding site on CCP 1 and visualized it with a solution structure of CCPs 1–3 derived by NMR and small angle x-ray scattering. We cross-validated these results by creating an artificial PfRh4-binding site through substitution of putative PfRh4-interacting residues from CCP 1 into their homologous positions within CCP 8; strikingly, this engineered binding site had an ∼30-fold higher affinity for PfRh4 than the native one in CCP 1. These experiments define a candidate site on CR1 by which P. falciparum merozoites gain access to human erythrocytes in a non-sialic acid-dependent pathway of merozoite invasion

Early Spanish meteorological records (1780-1850)

This article summarizes recent efforts on early instrumental data recovery in Spain conducted under the Salvà-Sinobas project. We have retrieved and digitized more than 100 000 meteorological observations prior to 1850 in Spain. This data set contains measurements of air temperature, atmospheric pressure, wind direction and state of the atmosphere in 16 places located in Iberia and the Balearic Islands. Most of the observations are made on a daily basis. However, monthly and annual information has also been retrieved. The time coverage of the series is not homogeneous, with the earliest records starting in Seville in 1780. Prior to this work only two series were available in Spain (i.e. Cádiz and Barcelona), so this data set represents a great advance in the early data availability for Spain. Due to the lack of metadata in most of the series, their interpretation must be made with caution