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Although lactococcal phages have been observed in cheese production settings for almost a century, few longitudinal studies have been performed. This 20-year study describes the close monitoring of dairy lactococcal phages in a cheddar cheese factory.

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To assess tocilizumab (TCZ) efficacy associated to standard of care (SOC) compared to SOC alone in severe coronavirus associated disease 2019 (COVID-19) patients. In a matched case-control study from 3 French Hospital COVID-19 Departments, 27 patients with severe COVID-19 treated with TCZ and SOC were matched for baseline epidemiological and clinical features and compared to 27 severe COVID-19 patients treated with SOC alone. Baseline characteristics of the study population were comparable between groups. Eleven patients (20%) died. TCZ was not associated with clinical improvement as compared to SOC regarding oxygen-free status (44% vs 63%) and death (18.5% vs 22%), despite a higher decrease of the C-reactive protein at Day 7 (10.7 vs 52 mg/L; P 

Heterocapsa triquetra (Ht), a non-toxic dinophyte, was selected to investigate the presence of pigments inhibiting melanoma cells growth. We developed an innovative microwave-assisted extraction process (MAE), determined the Ht pigment composition using Reverse Phase-High Performance Liquid Chromatography (HPLC) and high-resolution Ultra Performance Liquid Chromatography-Mass Spectrometry E (UPLC-MSE) analysis and assessed their antiproliferative activity in human invasive melanoma cells (A2058) grown in vitro. MAE allowed a fast and efficient extraction of Ht pigments as compared with conventional processes (soaking, sonication). Seventeen pigments and derivatives were unequivocally identified by UPLC-MSᴱ in the Ht extract, among which two were supposed to be produced during the extraction (hydroxylated chlorophyll a and pheophytin a). Pigment composition was fully coherent with that usually described for the DINO-1 peridiniales order. It included the uncommon carotenoids P457, peridininol, peridinin, pyrrhoxanthin, diadinochrome and diadinoxanthin. Ht pigment ethanol extract induced 18% growth inhibition of A2058 malignant melanoma cell line after a 72 h treatment at 100 μg·mL− 1 and pigment purification confirmed the highly significant antiproliferative activity of chlorophyll c2, peridinin, dinoxanthin and diatoxanthin on this cell line. Our data demonstrate for the first time the effectiveness of microwave irradiation to extract pigments from dinophytes. They also indicate that Heterocapsa triquetra and other dinophytes are interesting sources to purify bioactive pigments with antiproliferative activities in chemoresistant melanoma cells. These results strongly support previous findings about the anticancer activity of phytoplankton pigments and reinforce the possible interest of these molecules as natural cytostatic drugs.

Proteomic profiling of extracellular vesicles derived from adipocytes reveals the presence of key metabolic proteins involved in the development of cardiovascular diseases associated with obesity

Hedgehog (Hh) is a critical regulator of adipogenesis. Extracellular vesicles are natural Hh carriers, as illustrated by activated/apoptotic lymphocytes specifically shedding microparticles (MP) bearing the morphogen (MPHh+). We show that MPHh+ inhibit adipocyte differentiation and orientate mesenchymal stem cells towards a pro-osteogenic program. Despite a Smoothened (Smo)-dependency, MPHh+ anti-adipogenic effects do not activate a canonical Hh signalling pathway in contrast to those elicited either by the Smo agonist SAG or recombinant Sonic Hedgehog. The Smo agonist GSA-10 recapitulates many of the hallmarks of MPHh+ anti-adipogenic effects. The adipogenesis blockade induced by MPHh+ and GSA-10 was abolished by the Smo antagonist LDE225. We further elucidate a Smo/Lkb1/Ampk axis as the non-canonical Hh pathway used by MPHh+ and GSA-10 to inhibit adipocyte differentiation. Our results highlight for the first time the ability of Hh-enriched MP to signal via a non-canonical pathway opening new perspectives to modulate fat development.

Resistance to fluconazole antifungal is an ongoing impediment to a successful treatment of Candida albicans infections. One of the most prevalent mechanisms leading to azole resistance is genetic alterations of the 14α-demethylase, the target of azole antifungals, through point mutations. Site-directed mutagenesis and molecular modeling of 14α-demethylase rationalize biological data about the role of protein substitutions in the azole treatment failure. In this work, we investigated the role of N136Y substitution by site-directed mutagenesis into Pichia pastoris guided by structural analysis. Single amino acid substitutions were created by site-directed mutagenesis into P. pastoris with C. albicans ERG11 gene as template. In vitro susceptibility of P. pastoris transformants expressing wild-type and mutants to azole compounds was determined by CLSI M27-A2 and spot agar methods. The fluconazole effect on ergosterol biosynthesis was analyzed by gas chromatography-mass spectrometry. By microdilution and spot tests, N136Y transformants showed a reduced in vitro susceptibility to fluconazole compared to wild-type controls. As expected, ergosterol/lanosterol ratios were higher in N136Y transformants compared to the wild-type controls after treatment with fluconazole. Molecular modeling suggests that residue Asn136 located within the first mutation hot spot, could play a role during heme and azole binding. These results provide new insights into the structural basis for 14α-demethylase-azole interaction and could guide the design of novel azole antifungals.

International audience; Extracellular vesicles (EVs), including microvesicles (MVs) and exosomes, are small vesicles secreted from a wide variety of cells. Whereas MVs are particles released by the outward budding of the plasma membrane, exosomes are derived from endocytic compartments. Secretion of EVs can be enhanced by specific stimuli, and increased plasma circulating levels of EVs have been correlated with pathophysiological situations. MVs, already present in the blood of healthy individuals, are considerably elevated in several cardiovascular diseases associated with inflammation, suggesting that they can mediate deleterious effects such as endothelial dysfunction or thrombosis. Nonetheless, very recent studies also demonstrate that MVs may act as biological information vectors transferring proteins or genetic material to maintain cell homeostasis, favor cell repair, or even promote angiogenesis. Additionally, exosomes have also been shown to have pro-angiogenic and cardio-protective properties. These beneficial effects, therefore, reveal the potential therapeutical use of EVs in the field of cardiovascular medicine and regenerative therapy. In this review, we will provide an update of cellular processes modulated by EVs of specific interest in the treatment of cardiovascular pathologies. A special focus will be made on the morphogen sonic hedgehog (Shh) associated with EVs (EVsShh+), which have been shown to mediate many pro-angiogenic effects. In addition to offer a potential source of cardiovascular markers, therapeutical potential of EVs reveal exciting opportunities to deliver specific agents by non-immunogenic means to cardiovascular system.

Objectif Le morphogene Sonic Hedgehog (Shh) diffuse dans l’organisme sous forme libre ou associee a des vesicules extracellulaires. Nos travaux montrent que certains types de MVs vehiculant specifiquement Shh peuvent corriger la dysfonction endotheliale souvent associee aux pathologies cardiovasculaires. Shh a aussi ete identifie comme un facteur anti-adipogenique dans des etudes utilisant la proteine recombinante non-lipidee (RecShh). L’objectif est de caracteriser l’effet de Shh associe aux MVs sur la differenciation adipocytaire. Materiels et methodes Le modele cellulaire 3T3-L1 est utilisee pour etudier la differenciation adipocytaire. Les preadipocytes sont incubes en presence de RecShh ou de MVs issues d’une lignee lymphocytaire T (T-CEM) portant ou non Shh (MVsShh + et MVsShh-). Resultats Le traitement de 3T3-L1 par RecShh ou par des MVsShh + inhibe la differenciation adipocytaire et induit une diminution des facteurs cles de l’adipogenese (C/EBP et PPARγ). RecShh ou les MVsShh + ne sont plus capables d’inhiber la differentiation adipocytaire sur des 3T3-L1 deficientes pour le recepteur Smoothened. Cependant, les acteurs en aval de Smoothened conduisant a l’inhibition de l’adipogenese different pour RecShh ou les MVsShh +. RecShh induit une translocation de Smoothened dans le cil primaire et l’expression des facteurs Gli, et son effet inhibiteur est prevenu par la cyclopamine. L’effet des MVsShh + est quant a lui independant de la translocation de Smoothened, de l’induction des Gli et insensible a la cyclopamine. En revanche, les MVsShh + induisent une activation de l’AMPK qui pourrait sous-tendre a l’effet inhibiteur observe. Conclusion Shh inhibe la differentiation adipocytaire par des mecanismes differents sous sa forme recombinante ou portee par les MVs. De maniere originale, les MVsShh + mettent en jeu l’activation d’une voie hedgehog non-canonique. Ces travaux revelent une nouvelle voie de signalisation specifique des MVsShh + qui pourrait participer a la regulation de la masse adipeuse en modulant le pool de preadipocytes. Declaration d’interet Les auteurs declarent ne pas avoir d’interet direct ou indirect (financier ou en nature) avec un organisme prive, industriel ou commercial en relation avec le sujet presente.

International audience; Context:Focusing on mitochondrial function and thyroid tumorigenesis, we used an integrative approach to identify relevant biomarkers for borderline thyroid lesions.Design:Using cDNA and microRNA (miRNA) microarrays and quantitative RT-PCR analysis (qPCR), we explored samples of various types of thyroid tumors including 25 benign follicular adenomas represented by macrofollicular variants of thyroid adenomas, 38 oncocytic variants of follicular thyroid tumors, 19 papillary thyroid carcinomas, and 10 tumors of uncertain malignant potential, together with 53 normal thyroid tissue samples.Results:Our transcriptomic analysis, which highlighted discrepancies between controls and tumor tissues, as well as between various tumor types, led to the identification of 13 genes, allowing discrimination between the thyroid adenomas, oncocytic variants of follicular thyroid tumors, and papillary thyroid carcinomas, whereas the tumors of uncertain malignant potential were found to overlap these classes. Five of these genes (TP53, HOXA9, RUNX1, MYD88, and CITED1), with a differential expression confirmed by qPCR analysis, are implicated in tumorigenesis, 4 in mitochondrial metabolism (MRPL14, MRPS2, MRPS28, and COX6A1), and 2 in thyroid metabolic pathways (CaMKIINalpha and TPO). The global miRNA analysis revealed 62 differential miRNAs, the expression level for 10 of these being confirmed by qPCR. The differential expression of the miRNAs was in accordance with the modulation of gene expression and the ontologies revealed by our transcriptomic analysis.Conclusions:These findings reinforce the classification of follicular thyroid tumors established by the World Health Organization, and our technique offers a novel molecular approach to refine the classification of thyroid tumors of uncertain malignant potential.

Zeaxanthin, an abundant carotenoid present in fruits, vegetables and algae was reported to exert antiproliferative activity and induce apoptosis in human uveal melanoma cells. It also inhibited uveal melanoma tumor growth and cell migration in nude mice xenograft models. Here we report that zeaxanthin purified from the rhodophyte Porphyridium purpureum (Bory) K.M.Drew & R.Ross, Porphyridiaceae, promotes apoptosis in the A2058 human melanoma cell line expressing the oncogenic BRAF V600E mutation. Zeaxanthin 40 μM (IC50) induced chromatin condensation, nuclear blebbing, hypodiploidy, accumulation of cells in sub-G1 phase, DNA internucleosomal fragmentation and activation of caspase-3. Western blot analysis revealed that zeaxanthin induced up-regulation of the pro-apoptotic factors Bim and Bid and inhibition of NF-κB transactivation. Additionally, zeaxanthin sensitized A2058 melanoma cells in vitro to the cytotoxic activity of vemurafenib, a BRAF inhibitor widely used for the clinical management of melanoma, suggesting its potential interest as dietary adjuvant increasing melanoma cells sensitivity to chemotherapy. Keywords: Cancer, Carotenoid, Melanoma, Microalgae, Phytoplankton, Zeaxanthin