Proteomics of Stored Red Blood Cell Membrane and Storage-Induced Microvesicles Reveals the Association of Flotillin-2 With Band 3 Complexes

The storage of erythrocyte concentrates (ECs) induces lesions that notably affect metabolism, protein activity, deformability of red blood cells (RBCs), as well as the release of oxygen. Band 3 is one of the proteins affected during the ex vivo aging of RBCs. This membrane protein is an anion transporter, an anchor site for the cytoskeleton and other membrane proteins as well as a binding site for glycolytic enzymes and bears blood group antigens. In the present study, band 3 complexes were isolated from RBCs stored for 7 and 42 days in average (n = 3), as well as from microvesicles (n = 3). After extraction of membrane proteins with a deoxycholate containing buffer, band 3 complexes were co-immunoprecipitated on magnetic beads coated with two anti-band 3 antibodies. Both total membrane protein extracts and eluates (containing band 3 complexes) were separated on SDS-PAGE and analyzed by bottom-up proteomics. It revealed that three proteins were present or absent in band 3 complexes stemming from long-stored or short-stored ECs, respectively, whereas the membrane protein contents remained equivalent. These potential markers for storage-induced RBC aging are adenylosuccinate lyase (ADSL), alpha-adducin and flotillin-2, and were further analyzed using western blots. ADSL abundance tended to increase during storage in both total membrane protein and band 3 complexes, whereas alpha-adducin mainly tended to stay onto the membrane extract. Interestingly, flotillin-2 was equivalently present in total membrane proteins whereas it clearly co-immunoprecipitated with band 3 complexes during storage (1.6-fold-change, p = 0.0024). Moreover, flotillin-2 was enriched (almost threefold) in RBCs compared to microvesicles (MVs) (p < 0.001) and the amount found in MVs was associated to band 3 complexes. Different types of band 3 complexes are known to exist in RBCs and further studies will be required to better understand involvement of this protein in microvesiculation during the storage of RBC

Biomarker Analysis of Stored Blood Products: Emphasis on Pre-Analytical Issues

Millions of blood products are transfused every year; many lives are thus directly concerned by transfusion. The three main labile blood products used in transfusion are erythrocyte concentrates, platelet concentrates and fresh frozen plasma. Each of these products has to be stored according to its particular components. However, during storage, modifications or degradation of those components may occur, and are known as storage lesions. Thus, biomarker discovery of in vivo blood aging as well as in vitro labile blood products storage lesions is of high interest for the transfusion medicine community. Pre-analytical issues are of major importance in analyzing the various blood products during storage conditions as well as according to various protocols that are currently used in blood banks for their preparations. This paper will review key elements that have to be taken into account in the context of proteomic-based biomarker discovery applied to blood banking

Propofol Directly Increases Tau Phosphorylation

In Alzheimer's disease (AD) and other tauopathies, the microtubule-associated protein tau can undergo aberrant hyperphosphorylation potentially leading to the development of neurofibrillary pathology. Anesthetics have been previously shown to induce tau hyperphosphorylation through a mechanism involving hypothermia-induced inhibition of protein phosphatase 2A (PP2A) activity. However, the effects of propofol, a common clinically used intravenous anesthetic, on tau phosphorylation under normothermic conditions are unknown. We investigated the effects of a general anesthetic dose of propofol on levels of phosphorylated tau in the mouse hippocampus and cortex under normothermic conditions. Thirty min following the administration of propofol 250 mg/kg i.p., significant increases in tau phosphorylation were observed at the AT8, CP13, and PHF-1 phosphoepitopes in the hippocampus, as well as at AT8, PHF-1, MC6, pS262, and pS422 epitopes in the cortex. However, we did not detect somatodendritic relocalization of tau. In both brain regions, tau hyperphosphorylation persisted at the AT8 epitope 2 h following propofol, although the sedative effects of the drug were no longer evident at this time point. By 6 h following propofol, levels of phosphorylated tau at AT8 returned to control levels. An initial decrease in the activity and expression of PP2A were observed, suggesting that PP2A inhibition is at least partly responsible for the hyperphosphorylation of tau at multiple sites following 30 min of propofol exposure. We also examined tau phosphorylation in SH-SY5Y cells transfected to overexpress human tau. A 1 h exposure to a clinically relevant concentration of propofol in vitro was also associated with tau hyperphosphorylation. These findings suggest that propofol increases tau phosphorylation both in vivo and in vitro under normothermic conditions, and further studies are warranted to determine the impact of this anesthetic on the acceleration of neurofibrillary pathology

Dimethyl Sulfoxide Induces Both Direct and Indirect Tau Hyperphosphorylation

Dimethyl sulfoxide (DMSO) is widely used as a solvent or vehicle for biological studies, and for treatment of specific disorders, including traumatic brain injury and several forms of amyloidosis. As Alzheimer’s disease (AD) brains are characterized by deposits of β-amyloid peptides, it has been suggested that DMSO could be used as a treatment for this devastating disease. AD brains are also characterized by aggregates of hyperphosphorylated tau protein, but the effect of DMSO on tau phosphorylation is unknown. We thus investigated the impact of DMSO on tau phosphorylation in vitro and in vivo. One hour following intraperitoneal administration of 1 or 2 ml/kg DMSO in mice, no change was observed in tau phosphorylation. However, at 4 ml/kg, tau was hyperphosphorylated at AT8 (Ser202/Thr205), PHF-1 (Ser396/Ser404) and AT180 (Thr231) epitopes. At this dose, we also noticed that the animals were hypothermic. When the mice were maintained normothermic, the effect of 4 ml/kg DMSO on tau hyperphosphorylation was prevented. On the other hand, in SH-SY5Y cells, 0.1% DMSO induced tau hyperphosphorylation at AT8 and AT180 phosphoepitopes in normothermic conditions. Globally, these findings demonstrate that DMSO can induce tau hyperphosphorylation indirectly via hypothermia in vivo, and directly in vitro. These data should caution researchers working with DMSO as it can induce artifactual results both in vivo and in vitro

HIV-2 interaction with cell coreceptors: amino acids within the V1/V2 region of viral envelope are determinant for CCR8, CCR5 and CXCR4 usage

© 2014 Santos-Costa et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Background: Human immunodeficiency virus 1 and 2 (HIV-1 and HIV-2) use cellular receptors in distinct ways. Besides a more promiscuous usage of coreceptors by HIV-2 and a more frequent detection of CD4-independent HIV-2 isolates, we have previously identified two HIV-2 isolates (HIV-2MIC97 and HIV-2MJC97) that do not use the two major HIV coreceptors: CCR5 and CXCR4. All these features suggest that in HIV-2 the Env glycoprotein subunits may have a different structural organization enabling distinct - although probably less efficient - interactions with cellular receptors. Results: By infectivity assays using GHOST cell line expressing CD4 and CCR8 and blocking experiments using CCR8-specific ligand, I-309, we show that efficient replication of HIV-2MIC97 and HIV-2MJC97 requires the presence of CCR8 at plasma cell membrane. Additionally, we disclosed the determinants of chemokine receptor usage at the molecular level, and deciphered the amino acids involved in the usage of CCR8 (R8 phenotype) and in the switch from CCR8 to CCR5 or to CCR5/CXCR4 usage (R5 or R5X4 phenotype). The data obtained from site-directed mutagenesis clearly indicates that the main genetic determinants of coreceptor tropism are located within the V1/V2 region of Env surface glycoprotein of these two viruses. Conclusions: We conclude that a viral population able to use CCR8 and unable to infect CCR5 or CXCR4-positive cells, may exist in some HIV-2 infected individuals during an undefined time period, in the course of the asymptomatic stage of infection. This suggests that in vivo alternate molecules might contribute to HIV infection of natural target cells, at least under certain circumstances. Furthermore we provide direct and unequivocal evidence that the usage of CCR8 and the switch from R8 to R5 or R5X4 phenotype is determined by amino acids located in the base and tip of V1 and V2 loops of HIV-2 Env surface glycoprotein.This work was supported by grants from: Fundação para a Ciência e Tecnologia (FCT; PPCDT/SAU-IMI/55726/2004); Fundação para a Ciência e Tecnologia and Ministério da Saúde de Portugal (VIH/SAU/0006/2011); and from Gilead Sciences Portugal (Programa Gilead Génese).info:eu-repo/semantics/publishedVersio

COVID-19 symptoms at hospital admission vary with age and sex: results from the ISARIC prospective multinational observational study

Background: The ISARIC prospective multinational observational study is the largest cohort of hospitalized patients with COVID-19. We present relationships of age, sex, and nationality to presenting symptoms. Methods: International, prospective observational study of 60 109 hospitalized symptomatic patients with laboratory-confirmed COVID-19 recruited from 43 countries between 30 January and 3 August 2020. Logistic regression was performed to evaluate relationships of age and sex to published COVID-19 case definitions and the most commonly reported symptoms. Results: ‘Typical’ symptoms of fever (69%), cough (68%) and shortness of breath (66%) were the most commonly reported. 92% of patients experienced at least one of these. Prevalence of typical symptoms was greatest in 30- to 60-year-olds (respectively 80, 79, 69%; at least one 95%). They were reported less frequently in children (≤ 18 years: 69, 48, 23; 85%), older adults (≥ 70 years: 61, 62, 65; 90%), and women (66, 66, 64; 90%; vs. men 71, 70, 67; 93%, each P < 0.001). The most common atypical presentations under 60 years of age were nausea and vomiting and abdominal pain, and over 60 years was confusion. Regression models showed significant differences in symptoms with sex, age and country. Interpretation: This international collaboration has allowed us to report reliable symptom data from the largest cohort of patients admitted to hospital with COVID-19. Adults over 60 and children admitted to hospital with COVID-19 are less likely to present with typical symptoms. Nausea and vomiting are common atypical presentations under 30 years. Confusion is a frequent atypical presentation of COVID-19 in adults over 60 years. Women are less likely to experience typical symptoms than men

Le parodonte sain de l'adulte (le connaître et le maintenir en vue de la réalisation d'une couronne sur dent naturelle)

BORDEAUX2-BU Sci.Homme/Odontol. (330632102) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Basinga: A cell‐by‐cell GIS toolbox for computing basin average scaling factors, cosmogenic production rates and denudation rates

International audienceThe calculation of denudation rates from the measured cosmogenic nuclide concentrations in river sediments requires assumptions and approximations. Several different approaches and numerical tools are available in the literature. A widely used analytical approach represents the muogenic production with one or two exponentials, assumes the attenuation length of muons to be constant and also neglects temporal variations in the Earth's magnetic field. The denudation rates are then calculated directly and analytically from the measured concentrations. A second numerical and iterative approach was more recently proposed and considers a more rigorous muogenic production law based on pre-calculated variable attenuation length of muons and accounts for temporal changes of the magnetic field. It also assumes a specific distribution of denudation rates throughout the basin and uses an iterative approach to calculate the basin average denudation rates. We tested the two approaches across several natural basins and found that both approaches provide similar denudation results. Hence, assuming exponential muogenic production and constant attenuation length of muons in the rock has little impact on the derived denudation rates. Therefore, unless a priori known distributions of denudation rates are to be tested, there does not appear to be any particular gain from using the second iterative method which is computationally less effective. Based on these findings, we developed and describe here Basinga, a new ArcGIS® and QGIS toolbox which computes the basin average scaling factors, cosmogenic production rates and denudation rates for several tens of drainage basins together. Basinga follows either the Lal/Stone or the Lifton/Sato/Dunai scaling schemes and includes several optional tools for correcting for topographic shielding, ice cover and lithology. We have also developed an original method for correcting the cosmogenic production rates for past variations in the Earth's magnetic field