La vitrification en une seule étape d’embryons murins définit de nouveaux standards de cryopréservation

La cryopréservation d’embryons est un des outils les plus efficaces, économiques et utiles au niveau éthique pour préserver indéfiniment la génétique des animaux de laboratoire. Les bénéfices liés à son utilisation sont nombreux, et incluent la réduction des coûts associés à la pérennisation de lignées, la limitation de l’apparition et de la dissémination de mutations non voulues, la facilité et la sécurité pour les transferts internationaux, et la réduction du nombre d’animaux à élever en captivité. La vitrification a été démontrée comme étant plus efficace que la congélation lente en procréation médicalement assistée humaine, où elle constitue maintenant la méthode de référence. Il en va de même pour la cryopréservation des embryons de souris, où la vitrification préserve mieux l’intégrité de la chromatine, réduit la pénétration intracytoplasmique d’agents cryoprotecteurs potentiellement toxiques, et in fine permet une meilleure survie et un meilleur développement après réchauffement que la congélation lente. Cependant, pour être efficaces, les méthodes actuelles de vitrification nécessitent plusieurs étapes d’exposition des embryons à des solutions spécifiques avant le refroidissement, mais aussi au cours de leur réchauffement ultérieur. Cette relative complexité peut s’avérer difficile à gérer de manière optimale quand un grand nombre d’embryons doivent être traités au cours d’une même session. Nous avons développé et breveté une technologie de vitrification d’embryons en une étape (« one-step ») qui est aussi efficace que les meilleures méthodes de vitrification multi-étapes. De plus, nos milieux sont chimiquement définis (sans sérum ou autre composant biologique non défini), et des supports permettant la vitrification aseptique peuvent être utilisés sans perte de rendement. Notre technologie de vitrification one-step d’embryons de rongeurs répond ainsi aux problèmes d’ergonomie liés aux méthodes classiques de vitrification, et fournit ainsi aux scientifiques une solution efficace, biologiquement sûre et facile à utiliser pour la cryopréservation d’embryons de rongeurs. Elle rend ainsi l’efficacité de la vitrification plus aisément applicable aux rongeurs de laboratoire, contribuant ainsi à la réduction du nombre d’animaux nécessaires à la pérennisation et à la diffusion de lignées ou de colonies utiles.Vitricell: développement et valorisation de kits de vitrification de cellules en conditions aseptiques et chimiquement définie

Single nanoparticle tracking of N-methyl-d-aspartate receptors in cultured and intact brain tissue

This work was supported by the Centre National de la Recherche Scientifique (CNRS), Agence Nationale de la Recherche, Conseil Régional d’Aquitaine, and Marie Curie Individual Fellowship No. 326442.Recent developments in single-molecule imaging have revealed many biological mechanisms, providing high spatial and temporal resolution maps of molecular events. In neurobiology, these techniques unveiled that plasma membrane neurotransmitter receptors and transporters laterally diffuse at the surface of cultured brain cells. The photostability of bright nanoprobes, such as quantum dots (QDs), has given access to neurotransmitter receptor tracking over long periods of time with a high spatial resolution. However, our knowledge has been restricted to cultured systems, i.e., neurons and organotypic slices, therefore lacking several aspects of the intact brain rheology and connectivity. Here, we used QDs to track single glutamatergic N-methyl-d-aspartate receptors (NMDAR) in acute brain slices. By delivering functionalized nanoparticles in vivo through intraventricular injections to rats expressing genetically engineered-tagged NMDAR, we successfully tracked the receptors in native brain tissue. Comparing NMDAR tracking to different classical brain preparations (acute brain slices, cultured organotypic brain slices, and cultured neurons) revealed that the surface diffusion properties shared several features and are also influenced by the nature of the extracellular environment. Together, we describe the experimental procedures to track plasma membrane NMDAR in dissociated and native brain tissue, paving the way for investigations aiming at characterizing receptor diffusion biophysics in intact tissue and exploring the physiopathological roles of receptor surface dynamics.Publisher PDFPeer reviewe

Rare coding variants in PLCG2, ABI3, and TREM2 implicate microglial-mediated innate immunity in Alzheimer's disease

We identified rare coding variants associated with Alzheimer’s disease (AD) in a 3-stage case-control study of 85,133 subjects. In stage 1, 34,174 samples were genotyped using a whole-exome microarray. In stage 2, we tested associated variants (P<1×10-4) in 35,962 independent samples using de novo genotyping and imputed genotypes. In stage 3, an additional 14,997 samples were used to test the most significant stage 2 associations (P<5×10-8) using imputed genotypes. We observed 3 novel genome-wide significant (GWS) AD associated non-synonymous variants; a protective variant in PLCG2 (rs72824905/p.P522R, P=5.38×10-10, OR=0.68, MAFcases=0.0059, MAFcontrols=0.0093), a risk variant in ABI3 (rs616338/p.S209F, P=4.56×10-10, OR=1.43, MAFcases=0.011, MAFcontrols=0.008), and a novel GWS variant in TREM2 (rs143332484/p.R62H, P=1.55×10-14, OR=1.67, MAFcases=0.0143, MAFcontrols=0.0089), a known AD susceptibility gene. These protein-coding changes are in genes highly expressed in microglia and highlight an immune-related protein-protein interaction network enriched for previously identified AD risk genes. These genetic findings provide additional evidence that the microglia-mediated innate immune response contributes directly to AD development

The James Webb Space Telescope Mission

Twenty-six years ago a small committee report, building on earlier studies, expounded a compelling and poetic vision for the future of astronomy, calling for an infrared-optimized space telescope with an aperture of at least $4m$. With the support of their governments in the US, Europe, and Canada, 20,000 people realized that vision as the $6.5m$ James Webb Space Telescope. A generation of astronomers will celebrate their accomplishments for the life of the mission, potentially as long as 20 years, and beyond. This report and the scientific discoveries that follow are extended thank-you notes to the 20,000 team members. The telescope is working perfectly, with much better image quality than expected. In this and accompanying papers, we give a brief history, describe the observatory, outline its objectives and current observing program, and discuss the inventions and people who made it possible. We cite detailed reports on the design and the measured performance on orbit.Comment: Accepted by PASP for the special issue on The James Webb Space Telescope Overview, 29 pages, 4 figure

Extracorporeal Membrane Oxygenation for Severe Acute Respiratory Distress Syndrome associated with COVID-19: An Emulated Target Trial Analysis.

RATIONALE: Whether COVID patients may benefit from extracorporeal membrane oxygenation (ECMO) compared with conventional invasive mechanical ventilation (IMV) remains unknown. OBJECTIVES: To estimate the effect of ECMO on 90-Day mortality vs IMV only Methods: Among 4,244 critically ill adult patients with COVID-19 included in a multicenter cohort study, we emulated a target trial comparing the treatment strategies of initiating ECMO vs. no ECMO within 7 days of IMV in patients with severe acute respiratory distress syndrome (PaO2/FiO2 <80 or PaCO2 ≥60 mmHg). We controlled for confounding using a multivariable Cox model based on predefined variables. MAIN RESULTS: 1,235 patients met the full eligibility criteria for the emulated trial, among whom 164 patients initiated ECMO. The ECMO strategy had a higher survival probability at Day-7 from the onset of eligibility criteria (87% vs 83%, risk difference: 4%, 95% CI 0;9%) which decreased during follow-up (survival at Day-90: 63% vs 65%, risk difference: -2%, 95% CI -10;5%). However, ECMO was associated with higher survival when performed in high-volume ECMO centers or in regions where a specific ECMO network organization was set up to handle high demand, and when initiated within the first 4 days of MV and in profoundly hypoxemic patients. CONCLUSIONS: In an emulated trial based on a nationwide COVID-19 cohort, we found differential survival over time of an ECMO compared with a no-ECMO strategy. However, ECMO was consistently associated with better outcomes when performed in high-volume centers and in regions with ECMO capacities specifically organized to handle high demand. This article is open access and distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives License 4.0 (http://creativecommons.org/licenses/by-nc-nd/4.0/)

Cadasil (étude de quatre cas dijonnais et revue de la littérature en 2005)

DIJON-BU Médecine Pharmacie (212312103) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

One-step vitrification of murine embryos redefines cryopreservation standards

Cryopreservation of embryos is amongst the most powerful and efficient tools for indefinitely preserving the genetics of laboratory animals. The ensuing benefits are numerous and include reduction of costs associated to strain perpetuation, limitation of mutations occurrence and spreading, ease and safety of transnational shipping, and reduction of live animal husbandry. It has been demonstrated that vitrification is more efficient than slow freezing in human assisted reproduction, where it stands now as the gold standard. This is equally true for murine embryos, where vitrification has been shown to better preserve chromatin integrity, induce lower intracellular ingress of cryoprotectants and ultimately yield better embryo survival and development than slow freezing. Beside these benefits, current vitrification procedures require multiple pre-cooling and post-warming exposure steps to dedicated solutions to reach maximum effectiveness, which appears difficult to deal with when many embryos must be cryopreserved in one single session. We have developed and patented a unique one-step embryo vitrification procedure which is as efficient as the best multi-step vitrification methods. Moreover, our media are chemically defined (no serum nor undefined biological component), and aseptic vitrification carriers can be used without any yield loss. Our one-step vitrification kits and media for rodents (VitriMice™, VitriCell) address the poor ergonomy issues of classical vitrification, providing scientists with efficient, biologically safe and user-friendly solutions for embryo cryopreservation. Consequently, our one-step vitrification technology improves efficiency and applicability of cryopreservation for laboratory rodents, thereby contributing to the reduction of the number of live animals required to perpetuate and spread useful strains and colonies.Vitricell: développement et valorisation de kits de vitrification de cellules en conditions aseptiques et chimiquement définie

ONE-STEP VITRIFICATION OF MURINE EMBRYOS CHALLENGES CURRENT PARADIGMS OF CRYOBIOLOGY

peer reviewedCryopreservation of embryos is amongst the most powerful tools for preserving the genetics of laboratory animals. Vitrification is widely recognized as more efficient than slow freezing e.g. in human assisted reproduction technologies and for preserving murine embryos. Unfortunately, current vitrification procedures require multiple pre-cooling and post-warming exposure steps to dedicated solutions to reach maximum effectiveness, which appears difficult to deal with when many embryos are cryopreserved in one single session. To help solving this issue, we have developed a one-step embryo vitrification procedure. Briefly, embryos are exposed to a unique –chemically defined- vitrification solution before plunging in the liquid nitrogen. Subsequent warming is performed by immersing the vitrified embryos directly into the culture medium. Murine embryos at the zygote, two cells and morula stages have undergone our one-step procedure either under aseptic or non-aseptic conditions. After warming, direct in vitro survival, development to the blastocyst stage and in vivo development to birth were recorded as endpoints. Short exposure times to the vitrification solution (less than 60 seconds) before cooling and direct warming into culture medium led to results equivalent or better than after classical vitrification. Longer exposure times to the vitrification solution (between 90 and 150 seconds) decreased efficiency. These results demonstrate that intracellular vitrification after our one-step procedure occurs by combined effects of fast cooling (or warming) and cell dehydration, with minimal, if any, ingress of cryoprotectants. The absence of deleterious effect of warming directly into the culture medium and the relatively low sensitivity to thermal inertia (aseptic vs non-aseptic) of the carrier is a confirmation thereof. Consequently, beyond bringing a methodological simplification without any loss of efficiency, our patented one-step vitrification procedure dramatically lowers if not suppresses intracellular concentration of cryoprotectants and associated toxicity, thereby challenging some commonly accepted concepts of cryobiology.Vitricell: développement et valorisation de kits de vitrification de cellules en conditions aseptiques et chimiquement définie

Stagnation pondérale chez le nourrisson allaité (conduite à tenir. A propos de 7 observations)

STRASBOURG-Medecine (674822101) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF