Photoionization spectroscopy of CH3C3N in the vacuum-ultraviolet range

International audienceUsing vacuum-ultraviolet (VUV) synchrotron radiation, threshold and dissociative photoionization of cyanopropyne (CH3C3N) in the gas phase have been studied from 86 000 cm−1 up to 180 000 cm−1 by recording Threshold-PhotoElectron Spectrum (TPES) and PhotoIon Yield (PIY). Ionization energies of the four lowest electronic states X̃+2E,Ã+2A1,B̃+2E and C̃+ of CH3C3N+ are derived from the TPES with a better accuracy than previously reported. The adiabatic ionization potential of CH3C3N is measured as 86872±20 cm−1. A description of the vibrational structure of these states is proposed leading to the first determination of the vibrational frequencies for most modes. The vibrational assignments of the X̃+ state are supported by density functional theory calculations. In addition, dissociative photoionization spectra have been recorded for several cationic fragments in the range 12–15.5 eV (96 790–125 000 cm−1) and they bring new information on the photophysics of CH3C3N+. Threshold energies for the cationic dissociative channels leading to CH2C3N+, CHC3N+, HC3H+, HCNH+ and CH3+ have been measured for the first time and are compared with quantum chemical calculations

The Gut Microbiota Regulates Intestinal CD4 T Cells Expressing RORγt and Controls Metabolic Disease

SummaryA high-fat diet (HFD) induces metabolic disease and low-grade metabolic inflammation in response to changes in the intestinal microbiota through as-yet-unknown mechanisms. Here, we show that a HFD-derived ileum microbiota is responsible for a decrease in Th17 cells of the lamina propria in axenic colonized mice. The HFD also changed the expression profiles of intestinal antigen-presenting cells and their ability to generate Th17 cells in vitro. Consistent with these data, the metabolic phenotype was mimicked in RORγt-deficient mice, which lack IL17 and IL22 function, and in the adoptive transfer experiment of T cells from RORγt-deficient mice into Rag1-deficient mice. We conclude that the microbiota of the ileum regulates Th17 cell homeostasis in the small intestine and determines the outcome of metabolic disease

Marking Embryonic Stem Cells with a 2A Self-Cleaving Peptide: A NKX2-5 Emerald GFP BAC Reporter

Fluorescent reporters are useful for assaying gene expression in living cells and for identifying and isolating pure cell populations from heterogeneous cultures, including embryonic stem (ES) cells. Multiple fluorophores and genetic selection markers exist; however, a system for creating reporter constructs that preserve the regulatory sequences near a gene's native ATG start site has not been widely available.Here, we describe a series of modular marker plasmids containing independent reporter, bacterial selection, and eukaryotic selection components, compatible with both Gateway recombination and lambda prophage bacterial artificial chromosome (BAC) recombineering techniques. A 2A self-cleaving peptide links the reporter to the native open reading frame. We use an emerald GFP marker cassette to create a human BAC reporter and ES cell reporter line for the early cardiac marker NKX2-5. NKX2-5 expression was detected in differentiating mouse ES cells and ES cell-derived mice.Our results describe a NKX2-5 ES cell reporter line for studying early events in cardiomyocyte formation. The results also demonstrate that our modular marker plasmids could be used for generating reporters from unmodified BACs, potentially as part of an ES cell reporter library

ER-Alpha-cDNA As Part of a Bicistronic Transcript Gives Rise to High Frequency, Long Term, Receptor Expressing Cell Clones

Within the large group of Estrogen Receptor alpha (ERα)-negative breast cancer patients, there is a subgroup carrying the phenotype ERα−, PR−, and Her2−, named accordingly “Triple-Negative” (TN). Using cell lines derived from this TN group, we wished to establish cell clones, in which ERα is ectopically expressed, forming part of a synthetic lethality screening system. Initially, we generated cell transfectants expressing a mono-cistronic ERα transcription unit, adjacent to a separate dominant selectable marker transcription unit. However, the yield of ERα expressing colonies was rather low (5–12.5%), and only about half of these displayed stable ectopic ERα expression over time. Generation and maintenance of such cell clones under minimal exposure to the ERα ligand, did not improve yield or expression stability. Indeed, other groups have also reported grave difficulties in obtaining ectopic expression of ERα in ERα-deficient breast carcinoma cells. We therefore switched to transfecting these cell lines with pERα-IRES, a plasmid vector encoding a bicistronic translation mRNA template: ERα Open Reading Frame (ORF) being upstream followed by a dominant-positive selectable marker (hygroR) ORF, directed for translation from an Internal Ribosome Entry Site (IRES). Through usage of this bicistronic vector linkage system, it was possible to generate a very high yield of ERα expressing cell clones (50–100%). The stability over time of these clones was also somewhat improved, though variations between individual cell clones were evident. Our successful experience with ERα in this system may serve as a paradigm for other genes where ectopic expression meets similar hardships

Tixagevimab–cilgavimab for treatment of patients hospitalised with COVID-19: a randomised, double-blind, phase 3 trial

Background: Tixagevimab–cilgavimab is a neutralising monoclonal antibody combination hypothesised to improve outcomes for patients hospitalised with COVID-19. We aimed to compare tixagevimab–cilgavimab versus placebo, in patients receiving remdesivir and other standard care. Methods: In a randomised, double-blind, phase 3, placebo-controlled trial, adults with symptoms for up to 12 days and hospitalised for COVID-19 at 81 sites in the USA, Europe, Uganda, and Singapore were randomly assigned in a 1:1 ratio to receive intravenous tixagevimab 300 mg–cilgavimab 300 mg or placebo, in addition to remdesivir and other standard care. Patients were excluded if they had acute organ failure including receipt of invasive mechanical ventilation, extracorporeal membrane oxygenation, vasopressor therapy, mechanical circulatory support, or new renal replacement therapy. The study drug was prepared by an unmasked pharmacist; study participants, site study staff, investigators, and clinical providers were masked to study assignment. The primary outcome was time to sustained recovery up to day 90, defined as 14 consecutive days at home after hospital discharge, with co-primary analyses for the full cohort and for participants who were neutralising antibody-negative at baseline. Efficacy and safety analyses were done in the modified intention-to-treat population, defined as participants who received a complete or partial infusion of tixagevimab–cilgavimab or placebo. This study is registered with ClinicalTrials.gov, NCT04501978 and the participant follow-up is ongoing. Findings: From Feb 10 to Sept 30, 2021, 1455 patients were randomly assigned and 1417 in the primary modified intention-to-treat population were infused with tixagevimab–cilgavimab (n=710) or placebo (n=707). The estimated cumulative incidence of sustained recovery was 89% for tixagevimab–cilgavimab and 86% for placebo group participants at day 90 in the full cohort (recovery rate ratio [RRR] 1·08 [95% CI 0·97–1·20]; p=0·21). Results were similar in the seronegative subgroup (RRR 1·14 [0·97–1·34]; p=0·13). Mortality was lower in the tixagevimab–cilgavimab group (61 [9%]) versus placebo group (86 [12%]; hazard ratio [HR] 0·70 [95% CI 0·50–0·97]; p=0·032). The composite safety outcome occurred in 178 (25%) tixagevimab–cilgavimab and 212 (30%) placebo group participants (HR 0·83 [0·68–1·01]; p=0·059). Serious adverse events occurred in 34 (5%) participants in the tixagevimab–cilgavimab group and 38 (5%) in the placebo group. Interpretation: Among patients hospitalised with COVID-19 receiving remdesivir and other standard care, tixagevimab–cilgavimab did not improve the primary outcome of time to sustained recovery but was safe and mortality was lower. Funding: US National Institutes of Health (NIH) and Operation Warp Speed

Efficacy and safety of two neutralising monoclonal antibody therapies, sotrovimab and BRII-196 plus BRII-198, for adults hospitalised with COVID-19 (TICO): a randomised controlled trial

BACKGROUND: We aimed to assess the efficacy and safety of two neutralising monoclonal antibody therapies (sotrovimab [Vir Biotechnology and GlaxoSmithKline] and BRII-196 plus BRII-198 [Brii Biosciences]) for adults admitted to hospital for COVID-19 (hereafter referred to as hospitalised) with COVID-19. METHODS: In this multinational, double-blind, randomised, placebo-controlled, clinical trial (Therapeutics for Inpatients with COVID-19 [TICO]), adults (aged ≥18 years) hospitalised with COVID-19 at 43 hospitals in the USA, Denmark, Switzerland, and Poland were recruited. Patients were eligible if they had laboratory-confirmed SARS-CoV-2 infection and COVID-19 symptoms for up to 12 days. Using a web-based application, participants were randomly assigned (2:1:2:1), stratified by trial site pharmacy, to sotrovimab 500 mg, matching placebo for sotrovimab, BRII-196 1000 mg plus BRII-198 1000 mg, or matching placebo for BRII-196 plus BRII-198, in addition to standard of care. Each study product was administered as a single dose given intravenously over 60 min. The concurrent placebo groups were pooled for analyses. The primary outcome was time to sustained clinical recovery, defined as discharge from the hospital to home and remaining at home for 14 consecutive days, up to day 90 after randomisation. Interim futility analyses were based on two seven-category ordinal outcome scales on day 5 that measured pulmonary status and extrapulmonary complications of COVID-19. The safety outcome was a composite of death, serious adverse events, incident organ failure, and serious coinfection up to day 90 after randomisation. Efficacy and safety outcomes were assessed in the modified intention-to-treat population, defined as all patients randomly assigned to treatment who started the study infusion. This study is registered with ClinicalTrials.gov, NCT04501978. FINDINGS: Between Dec 16, 2020, and March 1, 2021, 546 patients were enrolled and randomly assigned to sotrovimab (n=184), BRII-196 plus BRII-198 (n=183), or placebo (n=179), of whom 536 received part or all of their assigned study drug (sotrovimab n=182, BRII-196 plus BRII-198 n=176, or placebo n=178; median age of 60 years [IQR 50-72], 228 [43%] patients were female and 308 [57%] were male). At this point, enrolment was halted on the basis of the interim futility analysis. At day 5, neither the sotrovimab group nor the BRII-196 plus BRII-198 group had significantly higher odds of more favourable outcomes than the placebo group on either the pulmonary scale (adjusted odds ratio sotrovimab 1·07 [95% CI 0·74-1·56]; BRII-196 plus BRII-198 0·98 [95% CI 0·67-1·43]) or the pulmonary-plus complications scale (sotrovimab 1·08 [0·74-1·58]; BRII-196 plus BRII-198 1·00 [0·68-1·46]). By day 90, sustained clinical recovery was seen in 151 (85%) patients in the placebo group compared with 160 (88%) in the sotrovimab group (adjusted rate ratio 1·12 [95% CI 0·91-1·37]) and 155 (88%) in the BRII-196 plus BRII-198 group (1·08 [0·88-1·32]). The composite safety outcome up to day 90 was met by 48 (27%) patients in the placebo group, 42 (23%) in the sotrovimab group, and 45 (26%) in the BRII-196 plus BRII-198 group. 13 (7%) patients in the placebo group, 14 (8%) in the sotrovimab group, and 15 (9%) in the BRII-196 plus BRII-198 group died up to day 90. INTERPRETATION: Neither sotrovimab nor BRII-196 plus BRII-198 showed efficacy for improving clinical outcomes among adults hospitalised with COVID-19. FUNDING: US National Institutes of Health and Operation Warp Speed

Association Between Early Treatment With Tocilizumab and Mortality Among Critically Ill Patients With COVID-19

Importance: Therapies that improve survival in critically ill patients with coronavirus disease 2019 (COVID-19) are needed. Tocilizumab, a monoclonal antibody against the interleukin 6 receptor, may counteract the inflammatory cytokine release syndrome in patients with severe COVID-19 illness. Objective: To test whether tocilizumab decreases mortality in this population. Design, Setting, and Participants: The data for this study were derived from a multicenter cohort study of 4485 adults with COVID-19 admitted to participating intensive care units (ICUs) at 68 hospitals across the US from March 4 to May 10, 2020. Critically ill adults with COVID-19 were categorized according to whether they received or did not receive tocilizumab in the first 2 days of admission to the ICU. Data were collected retrospectively until June 12, 2020. A Cox regression model with inverse probability weighting was used to adjust for confounding. Exposures: Treatment with tocilizumab in the first 2 days of ICU admission. Main Outcomes and Measures: Time to death, compared via hazard ratios (HRs), and 30-day mortality, compared via risk differences. Results: Among the 3924 patients included in the analysis (2464 male [62.8%]; median age, 62 [interquartile range {IQR}, 52-71] years), 433 (11.0%) received tocilizumab in the first 2 days of ICU admission. Patients treated with tocilizumab were younger (median age, 58 [IQR, 48-65] vs 63 [IQR, 52-72] years) and had a higher prevalence of hypoxemia on ICU admission (205 of 433 [47.3%] vs 1322 of 3491 [37.9%] with mechanical ventilation and a ratio of partial pressure of arterial oxygen to fraction of inspired oxygen of <200 mm Hg) than patients not treated with tocilizumab. After applying inverse probability weighting, baseline and severity-of-illness characteristics were well balanced between groups. A total of 1544 patients (39.3%) died, including 125 (28.9%) treated with tocilizumab and 1419 (40.6%) not treated with tocilizumab. In the primary analysis, during a median follow-up of 27 (IQR, 14-37) days, patients treated with tocilizumab had a lower risk of death compared with those not treated with tocilizumab (HR, 0.71; 95% CI, 0.56-0.92). The estimated 30-day mortality was 27.5% (95% CI, 21.2%-33.8%) in the tocilizumab-treated patients and 37.1% (95% CI, 35.5%-38.7%) in the non-tocilizumab–treated patients (risk difference, 9.6%; 95% CI, 3.1%-16.0%). Conclusions and Relevance: Among critically ill patients with COVID-19 in this cohort study, the risk of in-hospital mortality in this study was lower in patients treated with tocilizumab in the first 2 days of ICU admission compared with patients whose treatment did not include early use of tocilizumab. However, the findings may be susceptible to unmeasured confounding, and further research from randomized clinical trials is needed.The writing committee was supported by grants F32HL149337 (Dr. Admon), K23DK120811 (Dr. Srivastava), R01HL085757 (Dr. Parikh), R01HL144566 and R01DK125786 (Dr. Leaf), K12HL138039 (Dr. Donnelly), K23HL130648 (Dr. Mathews), R37AI102634 (Dr. Hernán), F32DC017342 (Dr. Gupta), K08GM134220 and R03AG060179 (Dr. Shaefi), K23HL143053 (Dr. Semler), and R01HL153384 (Dr. Hayek) from the NIH and grant U-M G024231 from the Frankel Cardiovascular Center COVID-19: Impact Research Ignitor (Dr. Hayek)

Analysis of van-der-Waals vibrations in small (n = 1 to 4) aromatic-(rare gas)

A set of new, or improved, experimental R2P2CI spectra of the various structural isomers of aniline-(argon)n clusters with n = 1 to 4 have been obtained in the region of the Si-So origin band. They exhibit a very large number of sharp VdW excited bands. A simphfied mechanical model has been developped to describe the solvent to chromophore motions in the different isomers. From this restricted harmonic description of the force field, force constants and reduced masses were derived from fitting the spectra of the simple (1|0), (1|1), (2|0), (3|0) clusters, which allowed to make predictions for the VdW frequencies of the more complex (2|1) and (3|1) clusters and thus to propose assignments for die experimentally observed bands