SARS-CoV-2 transmission via apical syncytia release from primary bronchial epithelia and infectivity restriction in children epithelia

The beta-coronavirus SARS-CoV-2 is at the origin of a persistent worldwide pandemic. SARS-CoV-2 infections initiate in the bronchi of the upper respiratory tract and are able to disseminate to the lower respiratory tract eventually causing acute severe respiratory syndrome with a high degree of mortality in the elderly. Here we use reconstituted primary bronchial epithelia from adult and children donors to follow the infection dynamic following infection with SARS-CoV-2. We show that in bronchial epithelia derived from adult donors, infections initiate in multi-ciliated cells. Then, infection rapidly spread within 24-48h throughout the whole epithelia. Within 3-4 days, large apical syncytia form between multi-ciliated cells and basal cells, which dissipate into the apical lumen. We show that these syncytia are a significant source of the released infectious dose. In stark contrast to these findings, bronchial epithelia reconstituted from children donors are intrinsically more resistant to virus infection and show active restriction of virus spread. This restriction is paired with accelerated release of IFN compared to adult donors. Taken together our findings reveal apical syncytia formation as an underappreciated source of infectious virus for either local dissemination or release into the environment. Furthermore, we provide direct evidence that children bronchial epithelia are more resistant to infection with SARS-CoV-2 providing experimental support for epidemiological observations that SARS-CoV-2 cases’ fatality is linked to age. Significance Statement Bronchial epithelia are the primary target for SARS-CoV-2 infections. Our work uses reconstituted bronchial epithelia from adults and children. We show that infection of adult epithelia with SARS-CoV-2 is rapid and results in the synchronized release of large clusters of infected cells and syncytia into the apical lumen contributing to the released infectious virus dose. Infection of children derived bronchial epithelia revealed an intrinsic resistance to infection and virus spread, probably as a result of a faster onset of interferon secretion. Thus, our data provide direct evidence for the epidemiological observation that children are less susceptible to SARS-CoV-2

Gold nanoparticle trophic transfer from natural biofilm to grazer fish

Nanoparticle (NP) trophic transfer is reported as an important exposure route for organisms in aquatic ecosystems. This study focused on gold nanoparticle (AuNP, PEG-coated, 10 nm diameter) transfer using an experimental benthic food chain which included two trophic levels: natural river biofilm and grazer fish Hypostomus plecostomus. AuNP biofilm accumulation was assessed via water AuNP concentrations and total biofilm mass. An extended range of six AuNP concentrations in water (0, 0.0048, 0.048, 0.48, 4.8, and 48 mg Au L−1) was set. A dose-dependent relation between gold concentrations in water and natural river biofilm was observed after a 48-h exposure. This pointed out the high propensity of natural biofilms to accumulate gold. Additionally, total biofilm mass appeared to influence AuNP accumulation at the highest exposure levels. This first step enables the set-up of the transfer experiment in which grazer fish were exposed for 21 days to natural biofilms, previously contaminated by low AuNP concentrations in water (NP0.1: 0.48 and NP1: 4.8 mg Au L−1). Gold was quantified in eight fish organs, and histology was observed. Gold was transferred from biofilms to fish; bioaccumulation was organ- and exposure level-dependent. Interestingly, the brain showed significant gold accumulation at the highest exposure level (NP1). Histological observations indicated distinct inflammatory responses in fish liver, spleen, and muscle. The overall results suggest the potential hazards of subchronic nanoparticle exposure in aquatic organisms.Approches à différentes échelles pour caractériser les interactions cellulaires, le transfert trophique et les impacts toxiques de nanoparticules métalliques chez les organismes aquatiquesInitiative d'excellence de l'Université de BordeauxDéveloppment d'une infrastructure française distribuée coordonné

Role of G(i/o)-Src kinase-PI3K/Akt pathway and caveolin-1 in β₂-adrenoceptor coupling to endothelial NO synthase in mouse pulmonary artery.

Activation of the β₂-adrenoceptor (β₂-AR) elicits an endothelial nitric oxide synthase (eNOS)-dependent relaxation in mouse pulmonary artery, which, contrary to the muscarinic receptor-dependent relaxation, is preserved in hypoxic pulmonary arterial hypertension. We therefore characterized the signaling pathways underlying the β₂-AR-mediated eNOS activation, with special focus on G(i/o) proteins, protein kinases and caveolae. Functional studies (for evaluation of vasorelaxant response), Western blotting (for assessment of eNOS and caveolin-1 phosphorylation) and transmission electron microscopy (for visualization of caveolae) were conducted in pulmonary arteries from wild-type or caveolin-1 knockout mice. In wild-type isolated arteries, relaxation to the selective β₂-AR agonist procaterol was reduced by inhibitors of G(i/o) proteins (pertussis toxin, PTX), phosphatidylinositol 3-kinase (PI3K; wortmannin or LY 294002), Akt (Akt inhibitor X) and Src-kinase (PP2) and by cholesterol depletion (using methyl-β-cyclodextrin). Procaterol induced eNOS phosphorylation at Ser(1177), which was prevented by PTX, PP2 or Akt inhibitor. Procaterol also promoted caveolin-1 phosphorylation at Tyr(14), which was decreased by PTX or PP2. Caveolin-1 gene deletion resulted in endothelial caveolae disruption in mouse pulmonary artery and in potentiation of procaterol-induced relaxation. Unlike procaterol, acetylcholine-induced relaxation was unaffected by PTX, methyl-β-cyclodextrin or caveolin-1 gene deletion. To conclude, the mouse pulmonary endothelial β₂-AR is coupled to a G(i/o)-Src kinase-PI3K/Akt pathway to promote eNOS phosphorylation at Ser(1177) leading to a NO-dependent vasorelaxation. Caveolin-1 exerts a negative control on this response that is abrogated by its phosphorylation at Tyr(14), through a G(i/o)-Src kinase pathway. Since pulmonary β₂-AR- and muscarinic receptor-mediated relaxations differentiate in their respective signaling pathways leading to eNOS activation and sensitivities during hypoxia-induced pulmonary arterial hypertension, mechanisms underlying eNOS activation might be key determinants of pulmonary endothelial dysfunction

Monocore vs multicore magnetic iron oxide nanoparticles: uptake by glioblastoma cells and efficiency for magnetic hyperthermia

PEGylated magnetic iron oxide nanoparticles (IONPs) were synthesised with the aim to provide proof of concept results of remote cancer cell killing by magnetic fluid hyperthermia. The IONPs were produced by the polyol synthetic route also called “forced hydrolysis pathway” yielding highly superparamagnetic, readily-dispersible, and biocompatible IONPs. As shown previously, adjusting parameters of the reaction led to either monocore or multicore IONPs, with on-demand morphology and magnetic properties. Polyethylene glycol (PEG) was grafted onto the nanoparticles in a single final step, using a phosphonic acid-terminated PEG synthesised separately, a strategy named “convergent”. The magnetic properties of the IONPs were preserved in physiological media thanks to this biocompatible shell. The interaction of the PEGylated IONPs with a glioblastoma cell line was studied, from the stability of IONPs in appropriate cell culture medium to the remotely magnetically triggered cell death. Cellular internalisation of the IONPs was studied, along with their fate after application of an alternating magnetic field (AMF). This investigation highlights the superior efficiency of multicore (nanoflowers) vs monocore (nanospheres) IONPs for magnetic hyperthermia, leading to 80 % cancer cells death in medically translatable conditions.Magnéto-Chimiothérapie : Modélisation de la Délivrance Induite par Champ Magnétique Radiofréquence d'Anticancéreux par des Nano-Vésicules Polymères et Suivi par IRM d'un Modèle de GlioblastomeDéveloppment d'une infrastructure française distribuée coordonnéeMultifunctional Nanoparticles for Magnetic Hyperthermia and Indirect Radiation Therap

Characterization of a Cell-Assembled extracellular Matrix and the effect of the devitalization process

We have previously shown that the Cell-Assembled extracellular Matrix (CAM) synthesized by normal, human, skin fibroblasts in vitro can be assembled in a completely biological vascular graft that was successfully tested in the clinic. The goal of this study was to perform a detailed analysis of the composition and the organization of this truly bio-material. In addition, we investigated whether the devitalization process (dehydration) used to store the CAM, and thus, make the material available "off-the-shelf," could negatively affect its organization and mechanical properties. We demonstrated that neither the thickness nor the mechanical strength of CAM sheets were significantly changed by the dehydration/freezing/rehydration cycle. The identification of over 50 extracellular matrix proteins highlighted the complex composition of the CAM. Histology showed intense collagen and glycosaminoglycan staining throughout the CAM sheet. The distribution of collagen I, collagen VI, thrombospondin-1, fibronectin-1, fibrillin-1, biglycan, decorin, lumican and versican showed various patterns that were not affected by the devitalization process. Transmission electron microscopy analysis revealed that the remarkably dense collagen network was oriented in the plane of the sheet and that neither fibril density nor diameter was changed by devitalization. Second harmonic generation microscopy revealed an intricate, multi-scale, native-like collagen fiber orientation. In conclusion, this bio-material displayed many tissue-like properties that could support normal cell-ECM interactions and allow implantation without triggering degradative responses from the host's innate immune system. This is consistent with its success in vivo. In addition, the CAM can be devitalized without affecting its mechanical or unique biological architecture. STATEMENT OF SIGNIFICANCE: The extracellular matrix (ECM) defines biological function and mechanical properties of tissues and organs. A number of promising tissue engineering approaches have used processed ECM from cadaver/animal tissues or cell-assembled ECM in vitro combined with scaffolds. We have shown the clinical potential of a scaffold-free approach based on an entirely biological material produced by human cells in culture without chemical processing. Here, we perform a comprehensive analysis of the properties of what can truly be called a bio-material. We also demonstrate that this material can be stored dried without losing its remarkable biological architecture

Broadband forward light scattering by architectural design of core–shell silicon particles

A goal in the field of nanoscale optics is the fabrication of nanostructures with strong directional light scattering at visible frequencies. Here, we demonstrate the synthesis of Mie-resonant coreshell particles with overlapping electric and magnetic dipole resonances in the visible spectrum. The core consists of silicon surrounded by a lower index silicon oxynitride (SiOxNy) shell with an adjustable thickness. Optical spectroscopies coupled to Mie theory calculations give the first experimental evidence that the relative position and intensity of the magnetic and electric dipole resonances are tuned by changing the core-shell architecture. Specifically, coating a high-index particle with a low-index shell coalesces the dipoles, while maintaining a high scattering efficiency, thus generating broadband forward scattering. This synthetic strategy opens a route towards metamaterial fabrication with unprecedented control over visible light manipulation.Advanced Materials by DesignInitiative d'excellence de l'Université de BordeauxDéveloppment d'une infrastructure française distribuée coordonné

Microwave-assisted and metal-induced crystallization: a rapid and low temperature combination

Here, we present a new crystallization process which, by combining microwaves and metal-induced devitrification, reduces both the time and the temperature of crystallization compared to other known methods. Titania crystallization initiates at a temperature as low as 125 °C within a few minutes of microwave radiation. Several cations induce this low-temperature crystallization, namely, Mn2+, Co2+, Ni2+, Al3+, Cu2+ and Zn2+. The crystallization mechanism is probed with electron microscopy, elemental mapping, single-particle inductively coupled plasma mass spectrometry, X-ray photoelectron spectroscopy, Auger electron spectroscopy, and scanning Auger mapping. These techniques show that the metal ion migration through the vitreous titania under microwave radiation occurs prior to crystallization. The crystalline particles are suspended in solution at the end of the treatment, avoiding particle aggregation and sintering. The crystalline suspensions are thus ready for processing into a material or employment in any other application. This combination of microwaves and metal-induced crystallization is applied here to TiO2, but we are investigating its application to other materials as an ecofriendly crystallization method.Développment d'une infrastructure française distribuée coordonnéeInitiative d'excellence de l'Université de Bordeau

Multicenter Pilot Study to Assess a Biphasic Calcium Phosphate Implant for Functional and Aesthetic Septorhinoplasty

Importance: A validated biomaterial would have several medical advantages in septorhinoplasties requiring a large-volume graft such as avoiding donor site morbidity, making ambulatory surgery possible, and reducing surgical costs. Objective: To assess the safety and efficacy of a ceramic to treat saddle and crooked noses. The main endpoint was the biocompatibility of the implant. The secondary endpoint was its functional and aesthetic efficacy. Design, Setting, and Participants: The nasal septum (NASEPT) study is a pilot multicenter noncomparative prospective phase IIa clinical trial. The biomaterial tested was a biphasic calcium phosphate implant composed of 75% hydroxyapatite and 25% beta tri calcium phosphate. This versatile material can be used to replace septal skeleton when it is absent or nonusable. We included 25 patients with a multifractured osseous and cartilaginous framework after several traumas or surgeries. The implant placement technique was identical to an extracorporeal septoplasty through the external approach. Main Outcomes and Measures: The primary endpoint was the occurrence of expected adverse and severe adverse events. The secondary endpoints were clinical functional and aesthetic results and histological microscopic modifications. Results: Any extrusion, infection, pain, and epistaxis were observed. All implants were placed in a sagittal, straight, and solid position without extralobular depression. Comparisons between pre- and postoperative symptoms showed that nasal comfort (p < 10(-4)) and quality of life (p < 10(-4)) were dramatically improved in all patients. The nasolabial angle (p = 0.047) and the columellar projection (p = 0.024) were improved after surgery. Histological data showed little submucosal inflammation at 6 months with well-differentiated epithelium. The mean follow-up was 23 months: three patients underwent revision surgery for functional or aesthetic details and four implants were removed (16%) owing to a foreign body reaction between 17 and 74 months. Conclusion and Relevance: The NASEPT implant meets functional and aesthetic requirements in complex septorhinoplasties but its long-term biocompatibility needs to be improved. It could potentially avoid donor site morbidity

Cellular and molecular mechanisms of NiONPs toxicity on eel hepatocytes HEPA-E1: An illustration of the impact of Ni release from mining activity in New Caledonia

International audienceAnthropic activities such as open pit mining, amplify the natural erosion of metals contained in the soils, particularly in New Caledonia, leading to atmospheric emission of nickel oxide nanoparticles (NiONPs). These particles are produced during extraction end up in aquatic ecosystems through deposition or leaching in the rivers. Despite alarming freshwater Ni concentrations, only few studies have focused on the cellular and molecular mechanisms of NiONPs toxicity on aquatic organisms and particularly on eels. Those fish are known to be sensitive to metal contamination, especially their liver, which is a key organ for lipid metabolism, detoxification and reproduction. The objective of this study was to assess in vitro the cytotoxic effects of NiONPs on Anguilla japonica hepatocytes, HEPA-E1. HEPA-E1 were exposed to NiONPs (0.5-5 μg/cm2) for 4 or 24 h. Several endpoints were studied: (i) viability, (ii) ROS production, SOD activity and selected anti-oxidant genes expression, (iii) inflammation, (iv) calcium signalling, (v) mitochondrial function and (vi) apoptosis. The results evidenced that NiONPs induce a decrease of cell viability and an increase in oxidative stress with a significant superoxide anion production. An increase of mitochondrial calcium concentration and a decrease of mitochondrial membrane potential were observed, leading to apoptosis. These results underline the potential toxic impact of NiONPs on eels living in mining areas. Therefore, eel exposure to NiONPs can affect their migration and reproduction in New Caledonia