Aquaporins can be involved in the swelling caused by shiga toxin type 2 on hgec and hk-2 cells

Hemolytic uremic syndrome related to Shiga toxin?producing Escherichiacoli (STEC-HUS) is the principal etiology of acute kidney injuryin children in Argentina.Previously, we demonstrated that Shiga toxin type 2 (Stx2) damageshuman glomerular endothelial cells (HGEC) and HK-2 human proximaltubular epithelial cell line by inducing swelling and detachment.In this work, we analyzed cell volume changes of HGEC and HK-2exposed or not to Stx2 or a hypoosmotic (HYPO) medium, in thepresence or not of aquaporins (AQPs) inhibitors (mercuric chloride:HgCl2 and tetraethylammonium: TEA), or an inhibitor of Stx2 receptor(Gb3) synthesis, Eliglustat (EG). For controls, an isosmotic (ISO)medium was used.Cells were grown on 12 well plates and pretreated for 30 minuteswith HgCl2 (10 μM) or TEA (100 μM) or pretreated during 24 h withEG (10 μM). Then, HGEC and HK-2 were incubated with Stx2 (50μM) for an additional 40 minutes. Cell volume was analyzed by lightmicroscopy and measuring cell area by using Image J software.After Stx2 and HYPO medium treatments, a significant increase inthe cell volume of HGEC (Stx2: 42%; HYPO: 36%, n= 3, p<0.05) andHK-2 (Stx2: 70%; HYPO: 55%, n= 3, p<0.05) was detected respectto ISO medium. However, when HGEC and HK-2 were pretreatedwith HgCl2 or TEA a significant swelling prevention was obtained forHGEC (Stx2+HgCl2:100%; Stx2+TEA:86%; HYPO+HgCl2: 42.5%;HYPO+TEA: 83%, n=3, p<0.05) and HK-2 (Stx2+HgCl2: 90%; Stx-2+TEA: 85%; HYPO+HgCl2: 55%; HYPO+TEA: 75%, n=3, p<0.05).In addition, EG also was able to prevent HK-2 swelling in 87 % (n=1)with respect to Stx2 treatment.Results show that AQPs may be involved in the water movement insideHGEC and HK-2 induced by Stx2, since HgCl2 and TEA avoidedthis effect. Furthermore, binding of Stx2 to Gb3 could be the initialstep for the development of cellular mechanisms that possibly triggerthe entry of solutes into the cells and the consequent osmoticgradient responsible for the hypotonic effect.Fil: Gomez, Fernando Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Repetti, Julieta. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Romero, Romina. Hospital Nacional Profesor Alejandro Posadas.; ArgentinaFil: Sacerdoti, Flavia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Ibarra, Cristina Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Amaral, María Marta. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaLXVI Reunión Anual de la Sociedad Argentina de Investigación Clínica: LXIX Reunión Anual de la Sociedad Argentina de Inmunología; LIII Reunión Anual de la Asociación Argentina de Farmacología Experimental y XI Reunión Anual de la Asociación Argentina de NanomedicinasArgentinaSociedad Argentina de Investigación ClínicaSociedad Argentina de InmunologíaAsociación Argentina de Farmacología ExperimentalAsociación Argentina de Nanomedicina

Familial hypercholesterolaemia in children and adolescents from 48 countries: a cross-sectional study

Background: Approximately 450 000 children are born with familial hypercholesterolaemia worldwide every year, yet only 2·1% of adults with familial hypercholesterolaemia were diagnosed before age 18 years via current diagnostic approaches, which are derived from observations in adults. We aimed to characterise children and adolescents with heterozygous familial hypercholesterolaemia (HeFH) and understand current approaches to the identification and management of familial hypercholesterolaemia to inform future public health strategies. Methods: For this cross-sectional study, we assessed children and adolescents younger than 18 years with a clinical or genetic diagnosis of HeFH at the time of entry into the Familial Hypercholesterolaemia Studies Collaboration (FHSC) registry between Oct 1, 2015, and Jan 31, 2021. Data in the registry were collected from 55 regional or national registries in 48 countries. Diagnoses relying on self-reported history of familial hypercholesterolaemia and suspected secondary hypercholesterolaemia were excluded from the registry; people with untreated LDL cholesterol (LDL-C) of at least 13·0 mmol/L were excluded from this study. Data were assessed overall and by WHO region, World Bank country income status, age, diagnostic criteria, and index-case status. The main outcome of this study was to assess current identification and management of children and adolescents with familial hypercholesterolaemia. Findings: Of 63 093 individuals in the FHSC registry, 11 848 (18·8%) were children or adolescents younger than 18 years with HeFH and were included in this study; 5756 (50·2%) of 11 476 included individuals were female and 5720 (49·8%) were male. Sex data were missing for 372 (3·1%) of 11 848 individuals. Median age at registry entry was 9·6 years (IQR 5·8-13·2). 10 099 (89·9%) of 11 235 included individuals had a final genetically confirmed diagnosis of familial hypercholesterolaemia and 1136 (10·1%) had a clinical diagnosis. Genetically confirmed diagnosis data or clinical diagnosis data were missing for 613 (5·2%) of 11 848 individuals. Genetic diagnosis was more common in children and adolescents from high-income countries (9427 [92·4%] of 10 202) than in children and adolescents from non-high-income countries (199 [48·0%] of 415). 3414 (31·6%) of 10 804 children or adolescents were index cases. Familial-hypercholesterolaemia-related physical signs, cardiovascular risk factors, and cardiovascular disease were uncommon, but were more common in non-high-income countries. 7557 (72·4%) of 10 428 included children or adolescents were not taking lipid-lowering medication (LLM) and had a median LDL-C of 5·00 mmol/L (IQR 4·05-6·08). Compared with genetic diagnosis, the use of unadapted clinical criteria intended for use in adults and reliant on more extreme phenotypes could result in 50-75% of children and adolescents with familial hypercholesterolaemia not being identified. Interpretation: Clinical characteristics observed in adults with familial hypercholesterolaemia are uncommon in children and adolescents with familial hypercholesterolaemia, hence detection in this age group relies on measurement of LDL-C and genetic confirmation. Where genetic testing is unavailable, increased availability and use of LDL-C measurements in the first few years of life could help reduce the current gap between prevalence and detection, enabling increased use of combination LLM to reach recommended LDL-C targets early in life

Aquaporins can be involved in the swell- ing caused by shiga toxin type 2 on hgec and hk-2 cells

Hemolytic uremic syndrome related to Shiga toxin?producing Escherichia coli (STEC-HUS) is the principal etiology of acute kidney in-jury in children in Argentina.Previously, we demonstrated that Shiga toxin type 2 (Stx2) damageshuman glomerular endothelial cells (HGEC) and HK-2 human proxi-mal tubular epithelial cell line by inducing swelling and detachment.In this work, we analyzed cell volume changes of HGEC and HK-2exposed or not to Stx2 or a hypoosmotic (HYPO) medium, in thepresence or not of aquaporins (AQPs) inhibitors (mercuric chloride:HgCl2 and tetraethylammonium: TEA), or an inhibitor of Stx2 receptor(Gb3) synthesis, Eliglustat(EG).For controls,an isosmotic(ISO)medium was used. Cells were grown on 12 well plates and pretreated for 30 minuteswith HgCl2 (10 μM) or TEA (100 μM) or pretreated during 24 h withEG (10 μM). Then, HGEC and HK-2 were incubated with Stx2 (50μM) for an additional 40 minutes. Cell volume was analyzed by lightmicroscopy and measuring cell area by using Image J software.After Stx2 and HYPO medium treatments, a signi cant increase inthe cell volume of HGEC (Stx2: 42%; HYPO: 36%, n= 3, p<0.05) andHK-2 (Stx2: 70%; HYPO: 55%, n= 3, p<0.05) was detected respectto ISO medium. However, when HGEC and HK-2 were pretreatedwith HgCl2 or TEA a signi cant swelling prevention was obtained forHGEC (Stx2+HgCl2:100%; Stx2+TEA:86%; HYPO+HgCl2: 42.5%;HYPO+TEA: 83%, n=3, p<0.05) and HK-2 (Stx2+HgCl2: 90%; Stx-2+TEA: 85%; HYPO+HgCl2: 55%; HYPO+TEA: 75%, n=3, p<0.05).In addition, EG also was able to prevent HK-2 swelling in 87 % (n=1)with respect to Stx2 treatment. Results show that AQPs may be involved in the water movement in-side HGEC and HK-2 induced by Stx2, since HgCl2 and TEA avoid-ed this effect. Furthermore, binding of Stx2 to Gb3 could be the initialstep for the development of cellular mechanisms that possibly trig-ger the entry of solutes into the cells and the consequent osmoticgradient responsible for the hypotonic effect.Fil: Gomez, Fernando Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Repetti, Julieta. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Romero, Romina. Hospital Nacional Profesor Alejandro Posadas.; ArgentinaFil: Sacerdoti, Flavia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Ibarra, Cristina Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Amaral, María Marta. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaLXVI Reunión Anual de la Sociedad Argentina de Investigación Clínica; LXIX Reunión Anual de la Sociedad Argentina de Inmunología; LIII Reunión Anual de la Asociación Argentina de Farmacología Experimental y XI Reunión Anual de la Asociación Argentina de NanomedicinasArgentinaSociedad Argentina de Investigación ClínicaSociedad Argentina de InmunologíaAsociación Argentina de Farmacología ExperimentalAsociación Argentina de Nanomedicina

Sublethal exposure to imidacloprid in commercial Apis mellifera colonies in early spring: performance of honey bees and insecticide transference between in-hive products

Honey bees have an important role in ecosystems as pollinators. However, in recent years, bee populations have declined, with habitat destruction, pesticide use, and climate change contributing to the decline. One of the most important risk factors is the use of neurotoxic pesticides, such as neonicotinoids. The aim of this work was to study the effects of imidacloprid in commercial Apis mellifera L. colonies artificially fed with syrups spiked with this insecticide and its possible transfer among in-hive products such as honey and larvae. For this purpose, 30 colonies were placed in the same apiary; once a week for 7 weeks in early spring, each colony was fed with 0.5 L of syrup with the following doses of imidacloprid: 0, 15, 30, 120, and 240 µg kg−1. The colony strength was evaluated by monitoring: the number of adult bees and brood combs, queenlessness, unhealthy colonies (by detection of Nosema spp. spores and European foulbrood), as well as pollen and honey storage. Worker bees, larvae, honey, and beeswax were sampled to evaluate imidacloprid transfer within the hive. Trends in the persistence of the compound showed that up to 60% of the parent (not metabolized) was stored in honey, and the absence of residues in the larvae suggests that they were not exposed. Another result showed a certain impact in the honey reserves and honey yield with a reduction of this resource in the colonies exposed to imidacloprid.EEA RafaelaFil: Michlig, Melina Paola. Universidad Nacional del Litoral. Facultad de Ingeniería Química. Programa de Investigación y Análisis de Residuos y Contaminantes Químicos (PRINARC); ArgentinaFil: Michlig, Melina Paola. Consejo Nacional de Investigaciones Científicas Técnicas; ArgentinaFil: Pacini, Adriana Cecilia. Instituto Nacional de Tecnología Agropecuaria. Estación Experimental Agropecuaria Rafaela. Instituto de Investigación de la Cadena Láctea (IDICAL); ArgentinaFil: Pacini, Adriana Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigación de la Cadena Láctea (IDICAL); ArgentinaFil: Merke, Julieta. Instituto Nacional de Tecnología Agropecuaria. Estación Experimental Agropecuaria Rafaela. Instituto de Investigación de la Cadena Láctea (IDICAL); ArgentinaFil: Merke, Julieta. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigación de la Cadena Láctea (IDICAL); ArgentinaFil: Orellano, Emanuel. Instituto Nacional de Tecnología Agropecuaria. Estación Experimental Agropecuaria Rafaela. Instituto de Investigación de la Cadena Láctea (IDICAL); ArgentinaFil: Orellano, Emanuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigación de la Cadena Láctea (IDICAL); ArgentinaFil: Brasca, Melina. Universidad Nacional del Litoral. Facultad de Ingeniería Química. Programa de Investigación y Análisis de Residuos y Contaminantes Químicos (PRINARC); ArgentinaFil: Brasca, Melina. Consejo Nacional de Investigaciones Científicas Técnicas; ArgentinaFil: Repetti, María Rosa. Universidad Nacional del Litoral. Facultad de Ingeniería Química. Programa de Investigación y Análisis de Residuos y Contaminantes Químicos (PRINARC); Argentin

Cómo la cultura digital (TICs) afecta el desarrollo cognitivo y los paradigmas de aprendizaje

A pesar de que un gran número de educadores se han volcado hacia lo digital y se han capacitado en el uso de herramientas digitales, no existe un cambio real en los paradigmas de enseñanza y aprendizaje. Se han adoptado nuevas herramientas, pero también se han adaptado a las estructuras vigentes. Si bien se ha producido un cambio drástico y nos encontramos inmersos en una suerte de revolución digital, no se ha logrado aun una transformación real en los paradigmas de enseñanza y aprendizaje. Sin embargo, no escapa a nadie el hecho de que las formas de adquirir conocimiento se han desplazado de las fronteras tradicionales y han ocupado otros espacios (desde el hogar a lo recreativo).¿Cómo arribar, entonces, a una solución? ¿Cómo podría lograrse una transformación real? ¿Cómo propiciar el cambio para la redefinición de roles y competencias? En parte, a través de la capacitación y el desarrollo docente, para que los educadores se desplacen desde su zona de confort hacia un cambio de paradigma que conlleve la adopción de nuevos roles y competencias, nuevos escenarios y ecologías.Las teorías del desarrollo cognitivo brindarán también la información contextual necesaria para internalizar los cambios de paradigma y analizarán el impacto de las TICs en dicho desarrollo.Despite the fact that many education leaders have gone digital and are trained and coached into how to use digital tools, there hasn’t been an actual change in their learning paradigms. New tools have been adopted and adapted as well, to the old structure. Though the change has been drastic and we are currently immersed in a sort of digital revolution, no real transformation has been achieved yet. Transference of skills, tasks and activities has taken place, but there has been no real transformation in their learning paradigms. Yet everybody is aware that the sources for acquiring knowledge have escaped the traditional boundaries and have invaded other spaces (ranging from home to leisure places).How can this be solved? How can the transformation become real? How can change be fostered so that roles and competencies are redefined? Partially through teacher training and development;this will ‘push’ educational leaders out of their comfort zone into changing the paradigm by adoptingnew roles and competencies and embracing new scenarios and ecologies.Cognitive development theories will provide, as well, the background information needed tointernalize this change in paradigms and will analyze the impact ICTs have in this development

Determination of imidacloprid in beehive samples by UHPLC-MS/MS

Imidacloprid is a systemic insecticide belonging to the neonicotinoid family. It was the first neonicotinoid introduced in the mid-1990s, and since then, its use has grown rapidly to control pests in a variety of agricultural crops. Several studies have shown that neonicotinoids translocate to the nectar and pollen of treated plants, which represents a potential risk to pollinators. Therefore, an open-field feeding study was carried out. For this purpose, 30 beehives of Apis mellifera L. were installed in the same apiary. All colonies were in similar health and population conditions when assays were started. For seven weeks, colonies were fed with sucrose syrup with different concentrations of imidacloprid: 15, 30, 120 and 240 μg kg −1 . Thus, the assays were divided into four treatments and a witness (Control) with no added imidacloprid. To check the hives’ exposure to imidacloprid and evaluate its distribution, sampling of adult worker bees and larvae was performed before, during and after the whole feeding period (7 weeks). Furthermore, in the 15th week, honey and beeswax (honeycomb) samples were collected from the brood chamber and honey super of all hives. Analytical methodologies for sample preparation based on the QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) procedure were optimized and validated. After soaking the bees and honey samples and the extraction using acetonitrile with MgSO 4 and NaCl salts, a dispersive solid-phase extraction (d-SPE) step with MgSO 4 , PSA and C18 was applied. Melted beeswax was subjected to an acetonitrile extraction, followed by freeze-out and d-SPE with PSA and C18. Extracts were evaluated in a UHPLC-MS/MS system. LOQ (μg kg −1 ) values were 0.25, 0.50 and 1 for honey, bees and beeswax, respectively. Satisfactory recovery performance was achieved with relative standard deviation ≤20%. Residue concentrations of imidacloprid in samples showed correlation with the doses supplied, indicating exposure of the beehives to the insecticide. Honey stored approximately 60% of the loaded imidacloprid through syrup feeding.Fil: Michlig, Melina Paola. Universidad Nacional del Litoral; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Merke, Julieta. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Santa Fe. Estación Experimental Agropecuaria Rafaela; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Pacini, Adriana Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires; ArgentinaFil: Orellano, Ramiro Emanuel. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires; ArgentinaFil: Beldomenico, Horacio Ramon. Universidad Nacional del Litoral. Facultad de Ingeniería Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Repetti, María Rosa. Universidad Nacional del Litoral. Facultad de Ingeniería Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Comparative assessment of individual and mixture chronic toxicity of glyphosate and glufosinate ammonium on amphibian tadpoles: A multibiomarker approach

The aim of the present study was to assess the ecotoxicity of glyphosate and glufosinate ammonium mixtures on amphibian tadpoles and the potential impact of mixture in aquatic ecosystems health. The bonding properties of the mixture based on computational chemistry and an experimental bioassay on morphology, DNA damage and biochemical biomarkers on tadpoles of the common toad Rhinella arenarum were studied. The results of the density functional theory analysis showed trends of the pesticides clustering to form exothermic mixtures, suggesting the likelihood of hot-spots of pesticides in real aquatic systems. In addition, biological effects of individual pesticides and the mixture were studied on tadpoles over 45 days-chronic bioassay. The bioassay consisted of four treatments: a negative control (CO), 2.5 mg L-1 of a glyphosate-based herbicide (GBH), 2.5 mg L-1 of a glufosinate ammonium-based herbicide (GABH) and their 50:50 (% v/v) mixture (GBH-GABH). Morphological abnormality rates were significantly higher in all herbicide treatments with respect to CO at 48 h of exposure. Abdominal edema was the most frequent type of abnormality recorded at 48 h, 10 and 45 days of exposure. DNA damage was recorded in all herbicides treatments. Thyroxin increased only in GABH treatment. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) significantly increased in GBH treatment, indicating a GBH-neurotoxic effect. Glutathione S-transferase decreased in GABH and GBH-GABH treatments, while catalase decreased in individual GBH and GABH treatments. Overall, teratogenicity, DNA damage, hormonal disruption (T4), and oxidative stress were greater in GABH-treated tadpoles than GBH-treated tadpoles. This study also highlights the robust chemical interaction between the active ingredients of both herbicides, which is reflected on antagonisms in most of analyzed biomarkers, as well as potentiation and additivity in others. Based on our results, the GABH had a higher toxicity than GBH for amphibian tadpoles.Fil: Cuzziol Boccioni, Ana Paula. Universidad Nacional del Litoral. Facultad de Humanidades y Ciencias. Laboratorio de Ecotoxicologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Lener, German. Universidad Nacional de Córdoba. Facultad de Cs.químicas. Departamento de Química Teórica y Computacional; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Físico-química de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Instituto de Investigaciones en Físico-química de Córdoba; ArgentinaFil: Peluso, Julieta. Universidad Nacional de San Martín. Instituto de Investigación e Ingeniería Ambiental. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación e Ingeniería Ambiental; ArgentinaFil: Peltzer, Paola. Universidad Nacional del Litoral. Facultad de Humanidades y Ciencias. Laboratorio de Ecotoxicologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Attademo, Andres Maximiliano. Universidad Nacional del Litoral. Facultad de Humanidades y Ciencias. Laboratorio de Ecotoxicologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Aronzon, Carolina Mariel. Universidad Nacional de San Martín. Instituto de Investigación e Ingeniería Ambiental. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación e Ingeniería Ambiental; ArgentinaFil: Simoniello, Maria Fernanda. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Cátedra de Toxicología y Bioquímica Legal; ArgentinaFil: Demonte, Luisina Delma. Universidad Nacional del Litoral. Facultad de Ingeniería Química. Programa de Investigación y Análisis de Residuos y Contaminantes Químicos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Repetti, María Rosa. Universidad Nacional del Litoral. Facultad de Ingeniería Química. Programa de Investigación y Análisis de Residuos y Contaminantes Químicos; ArgentinaFil: Lajmanovich, Rafael Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Humanidades y Ciencias. Laboratorio de Ecotoxicologia; Argentin

Natural tannin extracts supplementation for COVID-19 patients (TanCOVID): a structured summary of a study protocol for a randomized controlled trial.

This research aims to study the efficacy of tannins co-supplementation on disease duration, severity and clinical symptoms, microbiota composition and inflammatory mediators in SARS-CoV2 patients. This is a prospective, double-blind, randomized, placebo-controlled, parallel-group trial to evaluate the efficacy of the administration of the dietary supplement ARBOX, a molecular blend of quebracho and chestnut tannins extract and Vit B12, in patients affected by COVID-19. 18 years of age or older, admitted to Hospital de Clinicas Jose de San Martin, Buenos Aires University (Argentina), meeting the definition of "COVID-19 confirmed case" ( https://www.argentina.gob.ar/salud/coronavirus-COVID-19/definicion-de-caso ). Inclusion Criteria Participants are eligible to be included in the study if the following criteria apply: 1. Any gender 2. ≥18 years old 3. Informed consent for participation in the study 4. Virological diagnosis of SARS-CoV-2 infection (real-time PCR) Exclusion Criteria Participants are excluded from the study if any of the following criteria apply: 1. Pregnant and lactating patients 2. Patients who cannot take oral therapy (with severe cognitive decline, assisted ventilation, or impaired consciousness) 3. Hypersensitivity to polyphenols 4. Patients already in ICU or requiring mechanical ventilation 5. Patients already enrolled in other clinical trials 6. Decline of consent INTERVENTION AND COMPARATOR: Experimental: TREATED ARM Participants will receive a supply of 28 -- 390 mg ARBOX capsules for 14 days. Patients will be supplemented with 2 capsules of ARBOX per day. Placebo Comparator: CONTROL ARM Participants will receive placebo supply for 14 days. The placebo will be administered with the identical dose as described for the test product. All trial participants will receive standard therapy, which includes: Antipyretics or Lopinavir / Ritonavir, Azithromycin and Hydroxychloroquine, as appropriate (treatment currently recommended by the department of Infectious Diseases of the Hospital de Clínicas that could undergo to modifications). In addition, if necessary: supplemental O2, non-invasive ventilation, antibiotic therapy. Primary Outcome Measures: Time to hospital discharge, defined as the time from first dose of ARBOX to hospital discharge [ Time Frame: Throughout the Study (Day 0 to Day 28) ] Secondary Outcome Measures: 28-day all-cause mortality [ Time Frame: Throughout the Study (Day 0 to Day 28) ]-proportion Invasive ventilation on day 28 [ Time Frame: Throughout the Study (Day 0 to Day 28) ]-proportion Level of inflammation parameters and cytokines [ Time Frame: day 1-14 ] -mean difference Difference in fecal intestinal microbiota composition and intestinal permeability [ Time Frame: day 1-14 ] Negativization of COVID-PCR at day 14 [ Time Frame: day 14 ]-proportion RANDOMIZATION: Potential study participants were screened for eligibility 24 hours prior to study randomization. Patients were randomly assigned via computer-generated random numbering (1:1) to receive standard treatment coupled with tannin or standard treatment plus placebo (control group). Study personnel and participants are blinded to the treatment allocation, as both ARBOX and placebo were packed in identical containers. Thus, all the used capsules had identical appearance. Considering an alpha error of 5%, a power of 80% a sample size of 70 patients per branch was estimated. 140 patients in total. The protocol version is number V2, dated May 23, 2020. The first patient, first visit was on June 12, 2020; the recruitment end date was October 6, 2020. The protocol was not submitted earlier because the enrollment of some patients took place after the closure of the recruitment on the clinicaltrials platform. In fact, due to the epidemiological conditions, due to the decrease of the cases in Argentina during the summer period, the recruitment stopped t before reaching the number of 140 patients (as indicated in the webpage). However, since there was a new increase in cases, the enrolment was resumed in order to reach the number of patients initially planned in the protocol. The final participant was recruited on February 14, 2021. ClinicalTrials.gov, number: NCT04403646 , registered on May 27th, 2020. The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol

A cohort-based study of host gene expression: tumor suppressor and innate immune/inflammatory pathways associated with the HIV reservoir size

&lt;p&gt;The major barrier to an HIV cure is the HIV reservoir: latently-infected cells that persist despite effective antiretroviral therapy (ART). Most prior studies of host genetic predictors of HIV control have focused on "elite controllers," rare individuals able to control virus in the absence of ART. However, there have been few genetic studies among ART-suppressed non-controllers, who make up the majority of people living with HIV (PLWH). We performed host RNA sequencing and HIV reservoir quantification (total DNA [tDNA], unspliced RNA [usRNA], intact DNA) from peripheral CD4+ T cells from 191 HIV+ ART-suppressed non-controllers. After adjusting for nadir CD4+ count, timing of ART initiation, and genetic ancestry, we identified two host genes for which higher expression was significantly associated with smaller total DNA viral reservoir size, &lt;em&gt;P3H3&lt;/em&gt; and &lt;em&gt;NBL1&lt;/em&gt;, both known tumor suppressor genes. We then identified 17 host genes for which lower expression was associated with higher residual transcription (HIV usRNA). These included novel associations with membrane channel (&lt;em&gt;KCNJ2&lt;/em&gt;, &lt;em&gt;GJB2&lt;/em&gt;), inflammasome (&lt;em&gt;IL1A, CSF3, TNFAIP5, TNFAIP6, TNFAIP9, CXCL3, CXCL10&lt;/em&gt;), and innate immunity (TLR7) genes (FDR-adjusted q&lt;0.05). Gene set enrichment analyses further identified significant associations of HIV usRNA with TLR4/microbial translocation (q=0.006), IL-1/NRLP3 inflammasome (q=0.008), and IL-10 (q=0.037) signaling. Protein validation assays using ELISA and multiplex cytokine assays supported these observed inverse host gene correlations, with P3H3, IL-10, and TNF-a protein associations achieving statistical significance (p&lt;0.05). Of note, plasma IL-10 was also significantly inversely associated with HIV DNA (p=0.016). HIV intact DNA was not associated with differential host gene expression, although this may have been due to a large number of undetectable values in our study. Further data are needed to validate these findings, including functional genomic studies, larger cohorts including underrepresented PLWH in research, and those including dedicated assays to measure the replication-competent HIV reservoir.&lt;/p&gt;&lt;p&gt;Funding provided by: National Institute of Allergy and Infectious Diseases&lt;br&gt;Crossref Funder Registry ID: https://ror.org/043z4tv69&lt;br&gt;Award Number: K23GM112526&lt;/p&gt;&lt;p&gt;Funding provided by: National Institute of Allergy and Infectious Diseases&lt;br&gt;Crossref Funder Registry ID: https://ror.org/043z4tv69&lt;br&gt;Award Number: U19 AI096109&lt;/p&gt;&lt;p&gt;Funding provided by: National Institute of Allergy and Infectious Diseases&lt;br&gt;Crossref Funder Registry ID: https://ror.org/043z4tv69&lt;br&gt;Award Number: UM1 AI126623&lt;/p&gt;&lt;p&gt;Funding provided by: National Center for Advancing Translational Sciences&lt;br&gt;Crossref Funder Registry ID: https://ror.org/04pw6fb54&lt;br&gt;Award Number: KL2TR002317&lt;/p&gt;&lt;p&gt;Funding provided by: American Foundation for AIDS Research&lt;br&gt;Crossref Funder Registry ID: https://ror.org/02jesmk44&lt;br&gt;Award Number: 108072-50-RGRL&lt;/p&gt;&lt;p&gt;Funding provided by: Bill &amp; Melinda Gates Foundation&lt;br&gt;Crossref Funder Registry ID: https://ror.org/0456r8d26&lt;br&gt;Award Number: INV-008500&lt;/p&gt;&lt;p&gt;Funding provided by: National Institute of Allergy and Infectious Diseases&lt;br&gt;Crossref Funder Registry ID: https://ror.org/043z4tv69&lt;br&gt;Award Number: &lt;/p&gt;&lt;p&gt;HIV+ ART-suppressed non-controllers from the UCSF SCOPE and Options HIV+ cohorts were included in the study.&lt;/p&gt; &lt;p&gt;Cryopreserved PBMCs were enriched for CD4+ T cells (StemCell, Vancouver, Canada), and RNA was extracted from CD4+ T cells using the AllPrep Universal Kit (Qiagen, Hilden, Germany) with one aliquot set aside for HIV reservoir quantification and a second aliquot for host RNA sequencing. Host RNA sequencing was analyzed using the HTStream pre-processing pipeline (s4hts.github.io/htstream/) was used for removing PCR duplicates, adapters, N characters, PolyA/T sequences, Phix contaminants, and poor-quality sequences (with quality score &lt;20 with sliding window of 10 base pairs). The quality of raw reads was assessed using FastQC. All samples had a per base quality score and sequence quality score &gt;30. RNA-seq reads were then mapped to the human genome (GRCh38) with a corresponding annotation file from the GENCODE project. Alignment and gene quantification were performed using the STAR alignment tool and its quantification protocol. Gene expression was converted to counts per million (CPM). The mean-variance trend was estimated to assign observational weights based on predicted variance on log&lt;sub&gt;2&lt;/sub&gt;-counts per million (log&lt;sub&gt;2&lt;/sub&gt;-CPM) using the Limma-Voom pipeline.&lt;/p&gt; &lt;p&gt;For validation of intracellularly expressed or membrane-associated encoded proteins, we performed ELISA from peripheral CD4-enriched T cells. CD4+ T cells were isolated from PBMC by negative selection, using the EasySep Human CD4+ T Cell Isolation Kit (StemCell Technologies, Vancouver, BC, Canada), following manufacturer's guidelines. The Muse Human CD4 T Cell kit (Luminex, Austin, TX) in combination with the Guava Muse Cell Analyzer was used to determine the concentration and percentages of CD4+ T cells after the isolation. To generate cellular lysates, purified CD4+ T cells were subjected to three cycles of freezing/thawing, using a dry ice (frozen CO&lt;sub&gt;2&lt;/sub&gt;) /absolute ethanol mixture and a 37˚C water bath. Complete lysis was verified by trypan blue staining and microscopic analysis. Lysates were spun down at 1,500 g for 10 min at 4˚C (to remove cellular debris) and the supernatants diluted 1:5 with PBS and kept at -80˚C until the time of protein quantification. Total protein concentration of the CD4+ T cell lysates was determined using the Pierce®BCA assay (Thermo Fisher Scientific). The mean value obtained was 0.94 mg/ml (range: 0.66 mg/ml – 1.18 mg/ml). Chemiluminescence or absorbance was read on a SpectraMax® iD5 multi-mode plate reader (Molecular Devices, San Jose, CA) and reported in relative light units (RLU). A standard curve was constructed by plotting the log mean RLU reading for each standard on the y-axis against the log of known concentrations on the x-axis using the SoftMax Pro 7.1 software (Molecular Devices, San Jose, CA). Data were normalized by total protein concentration to accurately reflect the total population of cells (live and dead). Briefly, a 1:5 dilution factor (based on supernatant dilution with PBS at the time of CD4+ T cell isolation) was used to calculate the concentration in the lysate before the dilution. Each protein marker was then quantified using the marker-specific ELISA (again, taking into account the 1:5 dilution performed before cryopreservation of the lysates). Data normalization was performed by dividing the concentration of each protein in the final lysate (e.g., for P3H3 in pg/ml) by the total protein concentration (mg/ml).&lt;/p&gt; &lt;p&gt;For validation of the several inflammatory pathway genes identified in association with HIV usRNA, most of the encoded proteins were secreted proteins, and thus, we performed high-sensitivity multiplex plasma cytokine quantification (Meso Scale Diagnostics). Plasma levels of IP-10 (the encoded protein for &lt;em&gt;CXCL10&lt;/em&gt;), G-CSF (&lt;em&gt;GCSF&lt;/em&gt;) and pentraxin 3 (&lt;em&gt;TNFAIP5&lt;/em&gt;) were quantified using the electrochemiluminescence-based 3-plex mesoscale discovery (MSD) platform (U-Plex mesoscale discovery, Rockville, MA); IL-10 (&lt;em&gt;IL10&lt;/em&gt;), IL-1β (&lt;em&gt;IL1B&lt;/em&gt;) and TNF-α (&lt;em&gt;TNFA&lt;/em&gt;) were measured in a separate 3-plex S-plex Proinflammatory panel kit, and IL‑1α (&lt;em&gt;IL1A&lt;/em&gt;) was quantified by a V-plex kit. In all these assays, undiluted samples were run in duplicate following manufacturer's instructions, and protein concentrations were determined using MSD Discovery Workbench (version 4.0.13) analysis software. The light intensities from the samples were interpolated using a four-parameter logistic fit (FourPL) to a standard curve of electrochemiluminescence generated from eight calibrators of know concentrations. The lower limit of detection for each marker can be found on the manufacturer's website (MesoScale Diagnostics, &lt;a href="https://www.mesoscale.com/~/media/files/handout/assaylist.pdf"&gt;https://www.mesoscale.com/~/media/files/handout/assaylist.pdf&lt;/a&gt;).&lt;/p&gt