Mass Spectrometry-based Absolute Quantification of 20S Proteasome Status for Controlled Ex-vivo Expansion of Human Adipose-derived Mesenchymal Stromal/Stem Cells

International audienceIn Brief 20S proteasomes are very heterogeneous protein complexes involved in many cellular processes. In the present study, we combined an MRM-based assay with the production and purification of entire SILAC labelled pro-teasome to monitor absolute quantities of the different 20S proteasome subtypes in various human cells and tissues. This method applied to adipocyte-derived stem cells (ADSCs) amplified under various conditions highlights an increased expression of immunoproteasome when this type of cell is primed with IFN␥ or amplified in a 20% O 2 environment. Graphical Abstract Highlights • Design of an MRM assay to determine the absolute quantity and stoichiometry of ubiquitous and tissue-specific human 20S proteasome subtypes. • Use of purified isotopically labelled 20S proteasome as internal standard for accurate quantification. • Variation in the expression of immunoproteasome in adipocyte-derived stem cells (ADSCs) grown under different O 2 levels might be causal for change in cells differentiation capacity. • The status of 20S proteasome during ADSCs expansion might constitute an additional relevant quality control parameter to contribute to predict, among other quality markers, their therapeutic capacity

Admission criteria and management of critical care patients in a pandemic context: position of the Ethics Commission of the French Intensive Care Society, update of April 2021.

Intensive care unit professionals have experience in critical care and its proportionality, collegial decision-making, withholding or withdrawal of treatment deemed futile, and communication with patients' relatives. These elements rely on ethical values from which we must not deviate in a pandemic situation. The recommendations made by the Ethics Commission of the French Intensive Care Society reflect an approach of responsibility and solidarity towards our citizens regarding the potential impact of a pandemic on critical care resources in France, with the fundamental requirement of respect for human dignity and equal access to health care for all

Frequency, prognostic impact, and subtype association of 8p12, 8q24, 11q13, 12p13, 17q12, and 20q13 amplifications in breast cancers

BACKGROUND: Oncogene amplification and overexpression occur in tumor cells. Amplification status may provide diagnostic and prognostic information and may lead to new treatment strategies. Chromosomal regions 8p12, 8q24, 11q13, 17q12 and 20q13 are recurrently amplified in breast cancers. METHODS: To assess the frequencies and clinical impact of amplifications, we analyzed 547 invasive breast tumors organized in a tissue microarray (TMA) by fluorescence in situ hybridization (FISH) and calculated correlations with histoclinical features and prognosis. BAC probes were designed for: (i) two 8p12 subregions centered on RAB11FIP1 and FGFR1 loci, respectively; (ii) 11q13 region centered on CCND1; (iii) 12p13 region spanning NOL1; and (iv) three 20q13 subregions centered on MYBL2, ZNF217 and AURKA, respectively. Regions 8q24 and 17q12 were analyzed with MYC and ERBB2 commercial probes, respectively. RESULTS: We observed amplification of 8p12 (amplified at RAB11FIP1 and/or FGFR1) in 22.8%, 8q24 in 6.1%, 11q13 in 19.6%, 12p13 in 4.1%, 17q12 in 9.9%, 20q13(Z )(amplified at ZNF217 only) in 9.9%, and 20q13(Co )(co-amplification of two or three 20q13 loci) in 8.5% of cases. The 8q24, 12p13, and 17q12 amplifications were correlated with high grade. The most frequent single amplifications were 8p12 (9.8%), 8q24 (3.3%) and 12p13 (3.3%), 20q13(Z )and 20q13(Co )(1.6%) regions. The 17q12 and 11q13 regions were never found amplified alone. The most frequent co-amplification was 8p12/11q13. Amplifications of 8p12 and 17q12 were associated with poor outcome. Amplification of 12p13 was associated with basal molecular subtype. CONCLUSION: Our results establish the frequencies, prognostic impacts and subtype associations of various amplifications and co-amplifications in breast cancers

DMTs and Covid-19 severity in MS: a pooled analysis from Italy and France

We evaluated the effect of DMTs on Covid-19 severity in patients with MS, with a pooled-analysis of two large cohorts from Italy and France. The association of baseline characteristics and DMTs with Covid-19 severity was assessed by multivariate ordinal-logistic models and pooled by a fixed-effect meta-analysis. 1066 patients with MS from Italy and 721 from France were included. In the multivariate model, anti-CD20 therapies were significantly associated (OR = 2.05, 95%CI = 1.39–3.02, p < 0.001) with Covid-19 severity, whereas interferon indicated a decreased risk (OR = 0.42, 95%CI = 0.18–0.99, p = 0.047). This pooled-analysis confirms an increased risk of severe Covid-19 in patients on anti-CD20 therapies and supports the protective role of interferon

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/)

Role(s) of CcpA and HPr, two actors of the carbon catabolite repression of the pathogen Streptococcus agalactiae

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Determination of the Regulon of CcpA, a Pleiotropic Regulator, in the Pathogen Streptococcus agalactiae

National audienc

Cholinergic Differentiation of Neural Progenitors in Adult Mouse Motor Facial Nucleus

International audienceEnvironmental cues are critical determinants of the fate of neural progenitors (NPs) upon transplantation into the central nervous system. In the present study, we assessed the differentiation potential of NPs implanted in a cholinergic environment of the adult mouse brain. Neurospheres containing NPs issued from fetal ganglionic eminences of transgenic mice expressing the green fluorescent protein (GFP) were transplanted either inside or outside the mouse cholinergic facial motor nucleus. In some mice, a pre-degenerated nerve releasing trophic factors was grafted into this nucleus to favor NP survival and improve axonal growth into the graft. The fate of NPs was analyzed 6 to 9 days or 2 months post-transplantation by immunofluorescence under confocal microscopy. Transplanted NPs were observed both inside and outside the facial nucleus after 6 to 9 days, but almost exclusively inside after 2 months regardless of the presence of a pre-degenerated nerve. NPs expressed markers of undifferentiated cells, astrocytes, oligodendrocytes, neurons, or cholinergic cells. The cholinergic phenotype of NPs engrafted inside the facial nucleus increased with time and the presence of a pre-degenerated nerve. Large GFP cholinergic somata and abundant long cholinergic GFP axons projecting into the nerve graft were also observed. Our results show that NPs, isolated from fetal mouse brain and transplanted into the non-neurogenic environment of the adult mouse facial nucleus, differentiate into cholinergic cells capable to project axons. This environment and the nerve graft favored NP differentiation into cholinergic neuron

Involvement of the pleiotropic regulator CcpA in the physiology and stress response of the pathogen Streptococcus agalactiae

Session : BactériologieInternational audienceStreptococcus agalactiae is a leading cause of infections in neonates. This opportunistic pathogen colonizes the vagina where it has to cope with acid pH and hydrogen peroxide produced by lactobacilli. Thus, in the host, this bacterium has the necessity to set up numerous adaptation mechanisms of which the pleiotropic regulators are part. Notably, several studies have shown links between regulation of carbon metabolism and mechanisms of adaptation and virulence. This is the case of the transcriptional regulator CcpA (catabolite control protein A) that has previously been demonstrated as a major regulator involved in carbon catabolite repression in Gram-positive bacteria. This pleiotropic regulator regulates genes involved in carbon metabolism but also in other numerous mechanisms. By a transcriptomic analysis, we highlighted here that CcpA regulates 13.5% of the genome. We confirmed by electrophoretic mobility shift assays (EMSAs) that the DNA binding sites called cis-acting catabolite responsive element (cre) determined for other streptococci was effective in S. agalactiae. We reported here that CcpA is of capital importance for survival in acid and oxidative stresses, and is implicated in macrophages survival by regulating several genes putatively or already described as involved in stress response. Among them, we focused our study on SAK_1689 that codes for a putative UspA protein. We first demonstrated that SAK_1689, highly repressed by CcpA, is overexpressed in oxidative stress. We showed that the overexpression in the ∆ccpAmutant under oxidative stress was harmful for the bacterium

La responsabilité civile de l'industriel pollueur

Her Vio Anne, Lecorvaisier Stéphanie, Le Dro Maryvonne, Lemoine Emmanuelle, Le Roux Valérie, Pain Ghislaine, Jugault Joseph. La responsabilité civile de l'industriel pollueur. In: Revue juridique de l'Ouest, N° Spécial 1995. La mer. pp. 169-201