234 research outputs found
Estudio de los mecanismos moleculares de estrés oxidativo, disfunción mitocondrial, estrés de retículo endoplasmático y autofagia en leucocitos de pacientes con diabetes tipo 2
Introducción:
La diabetes mellitus tipo 2 (DM2) se presenta como un cuadro clínico que incluye diversas alteraciones en el metabolismo, incluyendo hiperglicemia, resistencia a insulina, dislipidemia e inflamación crónica de bajo grado. Supone un gran problema a nivel mundial dada su creciente incidencia y las complicaciones deletéreas que conlleva, principalmente en el sistema cardiovascular. El principal factor determinante de DM2 es la obesidad. A medida que aumenta el índice de masa corporal se desarrolla resistencia a la insulina lo que finalmente originará una pérdida del control homeostático de la glucosa.
Entre las principales complicaciones vasculares de la diabetes están por una parte la enfermedad cerebrovascular, la enfermedad arterial periférica y la cardiopatía isquémica, que representan el grupo de complicaciones macrovasculares y suman el 52% de las muertes en pacientes diabéticos. Por otra parte, la nefropatía, la retinopatía y la neuropatía diabética constituyen el conjunto de complicaciones microvasculares, dentro de las cuales la nefropatía diabética es la más común afectando aproximadamente al 40% de los pacientes. El control glicémico es clave para prevenir la aparición de estas complicaciones en los pacientes con DM2, ya que el principal desencadenante de daño vascular es la hiperglicemia.
La inflamación sistémica es una de las características más representativa de la DM2, caracterizada por unos elevados niveles sistémicos de citoquinas proinflamatorias y una activación crónica del sistema inmune. A nivel local en las paredes vasculares, el proceso inflamatorio contribuye al inicio de la aterosclerosis debido a que convergen la disfunción endotelial, la activación de los leucocitos y el aumento de señales atrayentes (citoquinas y moléculas de adhesión) que favorecen la migración de los leucocitos hacia la pared endotelial. En este contexto es interesante el estudio de la fisiopatología de la DM2 en leucocitos de sangre periférica, ya que estas células participan tanto en la defensa inmune como en el proceso de aterosclerosis e influyen en la aparición de problemas cardiovasculares.
De entre los mecanismos subyacentes a la DM2 que contribuyen al desarrollo de complicaciones vasculares el estrés oxidativo juega un rol principal, así como la disfunción mitocondrial. De hecho, se ha especulado mucho acerca de las rutas de señalización que conectan la situación de hiperglicemia con el incremento de especies reactivas de oxígeno (ERO) y su papel en la inducción de enfermedad cardiovascular. Otros procesos a parte del estrés oxidativo y la disfunción mitocondrial han sido relacionados con la DM2 en los últimos años, como el estrés de retículo endoplasmático (RE) y la autofagia. Sin embargo, el grado de conocimiento acerca del papel que juegan éstos en la DM2 es bastante menor. Además se precisan más estudios acerca de la relación de estos procesos con el proceso inflamatorio y de adhesión de los leucocitos a las paredes vasculares, estableciendo así su posible influencia en la aparición de complicaciones vasculares.
Objetivos:
En la presente Tesis Doctoral nos planteamos evaluar el papel de la disfunción mitocondrial en la activación de los leucocitos en la DM2, así como su relación con el incremento de interacciones leucocito-endotelio y la correlación de estos factores con el desarrollo de cardiopatía isquémica silente (CIS). Por otra parte, quisimos estudiar el papel de la mieloperoxidasa (MPO) en la generación de ERO en la DM2 y evaluar la posible asociación de la MPO con la función endotelial y el desarrollo de nefropatía diabética. También nos propusimos evaluar la influencia del control glicémico de la DM2 sobre la inducción de estrés oxidativo y estrés de RE y el papel que juega el estrés de RE en la inducción de interacciones leucocito-endotelio. Por último nos planteamos determinar si el proceso de autofagia está alterado en leucocitos de pacientes con DM2, así como su posible relación con el estrés oxidativo y el estrés de RE.
Metodología:
Se reclutaron pacientes con DM2 y sujetos control y se determinaron parámetros antropométricos (peso, talla, IMC, índice cintura/cadera) y bioquímicos (en muestras sanguíneas). Se realizó el diagnóstico de la CIS mediante electrocardiograma y estudio coronariográfico seguido en los pacientes con CIS por SPECT (Single Photon Emission Computed Tomography) con 123-I-MIBG (Metayodobencilguanidina) mediante gammacámara. La nefropatía diabética se confirmó en los pacientes que presentaban un ratio albúmina:creatinina en orina mayor a 30mg/g. Para la determinación de MPO, citoquinas y moléculas de adhesión se conservaron alícuotas de suero a -80ºC. La concentración de MPO se determinó mediante un inmunoensayo tipo ELISA, mientras que las citoquinas y las moléculas de adhesión se determinaron mediante la tecnología X-MAP de LUMINEX. La evaluación del estrés oxidativo, la disfunción mitocondrial, el estrés de RE y la autofagia se hizo a partir de leucocitos aislados de sangre periférica mediante gradiente de Ficoll. Los leucocitos se incubaron con los siguientes fluorocromos para la valoración del estado redox: DCFH-DA, indicativo de la producción total de ERO; Mitosox Red, indicativo de la producción superóxido mitocondrial; TMRM, indicativo del potencial de membrana mitocondrial; NAO, indicativo de la masa mitocondrial y Hoescht, indicativo de la morfología nuclear. La fluorescencia emitida por los leucocitos se cuantificó bajo un microscopio de fluorescencia (IX81; Olympus) acoplado al software de citometría estática “ScanR”. La respiración mitocondrial se evaluó mediante un electrodo de O2 tipo Clark. Las interacciones leucocito-endotelio se evaluaron mediante una cámara de flujo paralelo. En este sistema se perfundió una suspensión de leucocitos sobre células endoteliales humanas cultivadas en monocapa (aisladas a partir de cordón umbilical mediante un tratamiento con colagenasa) en condiciones que simulan las del flujo sanguíneo. La interacción de los leucocitos con la monocapa endotelial se observó a través de un microscopio invertido de contraste de fases (Nikon Eclipse TE2000-S) y las imágenes se grabaron con una cámara acoplada (Sony Exware HAD). Se estimaron los procesos de rodamiento y adhesión firme leucocitaria. Se extrajo RNA y proteínas totales de los leucocitos para evaluar la expresión de los distintos marcadores de estrés de RE y de autofagia mediante Real Time PCR y Western Blotting. El análisis estadístico se realizó mediante los programas Graphpad Prism y SPSS.
Resultados y conclusiones:
Comparamos pacientes con DM2 de al menos 2 años de duración con sujetos control ajustados por edad y sexo. Los pacientes con DM2 presentaron un aumento en el IMC, la circunferencia de cintura y la presión arterial. Además mostraron mayores niveles de glucosa y de insulina en ayunas y por tanto un incremento en el índice HOMA-IR, así como un aumento de la HbA1c. Se observó a su vez un aumento de triglicéridos y unos niveles de colesterol-HDL reducidos en los pacientes diabéticos. Los marcadores de inflamación PCR, IL6 y TNFα mostraron niveles incrementados en el suero de los pacientes con DM2 además de un aumento significativo de las moléculas E-selectina y VCAM-1. Estas observaciones sugieren que existe un desequilibrio metabólico en los pacientes con DM2, que junto con la obesidad abdominal que presentan favorece la aparición de una inflamación sistémica de bajo grado y de origen metabólico.
En cuanto a los leucocitos, como consecuencia de la situación de inflamación crónica en los pacientes con DM2, manifestaron una menor velocidad de rodamiento sobre el endotelio, un mayor flujo de rodamiento y más adhesión. Cuando estas interacciones están incrementadas se favorece la disfunción endotelial y el daño vascular que precede a múltiples enfermedades cardiovasculares. Uno de los mecanismos que podría estar favoreciendo este fenotipo protrombótico en los leucocitos es el estrés oxidativo y la disfunción mitocondrial, y de hecho lo pudimos evidenciar a partir de nuestras observaciones. En los pacientes diabéticos, los leucocitos presentaron una excesiva producción de radicales libres junto con una disminución de las defensas antioxidantes, una menor masa mitocondrial, una reducción en el potencial de membrana y una menor tasa de respiración mitocondrial. Estas características se manifestaron mucho más aumentadas en los pacientes que además presentaban CIS, y a su vez la producción de ERO mitocondriales mostró una fuerte correlación con el aumento en las interacciones leucocito-endotelio pudiendo por tanto indicar que la disfunción mitocondrial sería un factor determinante en la inducción de complicaciones vasculares como la CIS.
Por otra parte observamos unos niveles aumentados de MPO en suero de pacientes con DM2, que fueron mayores en los pacientes que sufrían nefropatía, habiendo además una correlación positiva entre la concentración de MPO y el ratio albúmina/creatinina. Los pacientes con nefropatía mostraron una mayor frecuencia de contacto entre leucocitos y endotelio, además de un mayor incremento en los niveles de TNFα y E-selectina. Estos resultados plantean que la MPO participaría en la generación excesiva de ERO en leucocitos de pacientes con DM2 y podría tener un papel importante en el desarrollo de nefropatía mediado por la disfunción endotelial.
En cuanto a la inducción de estrés de RE en los pacientes con DM2, vimos que los niveles de GRP78 se aumentaron en los leucocitos de pacientes con DM2 de manera general. Sin embargo, el control glicémico de los pacientes parecía estar influenciando el patrón de expresión de los marcadores de estrés de RE, siendo adaptativo en los pacientes con un buen control de los niveles de glucosa, y proapoptótico en los pacientes mal controlados. También en este grupo de pacientes con mal control de la glicemia se mostraron mayores niveles de PCR, TNFα y E-selectina, así como un mayor flujo de rodamiento sobre el endotelio. La relación entre el estrés de RE y el proceso de interacción de los leucocitos con el endotelio se pudo intuir al observar que los niveles de GRP78 y sXBP1 se correlacionaban negativavente con la velocidad de rodamiento y los de CHOP exhibían una correlación positiva con el grado de adhesión leucocitaria, sugiriendo que bajo un buen control de la glicemia durante la DM2 los leucocitos tienden a frenar sobre las paredes endoteliales sin adherirse, en cambio cuando no hay un buen control de los niveles de glucosa la adhesión leucocitaria se acentúa.
Por último, evaluamos el papel de la autofagia en la DM2, determinando los niveles de LC3-II y Beclina 1 en leucocitos. Observamos que en los pacientes con DM2 estos marcadores de autofagia estaban aumentados junto con un patrón de condensación de la cromatina acentuado indicativo de muerte celular. Estos mismos leucocitos mostraban signos de disfunción mitocondrial, con una menor masa mitocondrial y una mayor producción de ERO, así como signos de estrés de RE, con un aumento en los marcadores GRP78, p-eIF2α y ATF6. Además, los niveles de Beclina 1 se correlacionan positivamente con la producción de ERO y también Beclina 1 y LC3-II se correlacionan positivamente con los distintos marcadores de estrés de RE estudiados. Estas observaciones sugieren que la DM2 altera la función autofágica en los leucocitos, y que esta función está regulada de igual forma que el estrés oxidativo y el estrés de RE durante la DM2.
En conjunto, estas evidencias ponen de manifiesto que los mecanismos moleculares de disfunción mitocondrial, estrés oxidativo, estrés de RE y autofagia ocurren en leucocitos de pacientes con DM2 y potencian la interacción de estas células con el endotelio, pudiendo ser determinantes en la inducción de complicaciones en el sistema cardiovascular.Introduction:
Type 2 diabetes (T2D) is a metabolic disorder characterized by hyperglycemia, insulin resistance, dyslipidemia and chronic low-grade inflammation. T2D is becoming a major public health issue due to its prevalence, which is increasing worldwide, and deleterious associated complications in the cardiovascular system. Obesity is the main determinant in the development of T2D. As body mass index (BMI) increases, insulin resistance develops and leads to an imbalance in glucose homeostasis.
Among the main vascular complications of T2D, cerebrovascular disease, peripheral artery disease and myocardial ischemia are macrovascular complications and represent 52% of deaths in T2D population. On the other hand, nephropathy, retinopathy and diabetic neuropathy are microvascular complications of the disease, with nephropathy being the most common, as it develops in approximately 40% of diabetic patients. Adequate glycemic control is the key to preventing these complications, as hyperglycemia is the main inducer of vascular damage.
Systemic inflammation is one of the main features of T2D and is characterized by enhanced circulatory levels of proinflammatory cytokines and chronic activation of the immune system. In the vascular wall, this inflammatory process contributes to the onset of atherosclerosis, as endothelial dysfunction, leukocyte activation and an increase in cytokines and adhesion molecules converge and promote leukocyte migration to the endothelial wall. In this context, it is interesting to study the physiopathology of T2D in peripheral blood cells such as leukocytes, since they participate in immunity and in the development of atherosclerosis, which can promote cardiovascular disease.
Oxidative stress and mitochondrial dysfunction play a main role in the development of vascular complications in T2D. In fact, a growing number of studies have described different signalling pathways that connect the hyperglycaemic situation with the induction of cardiovascular diseases, in which oxidative stress is the major contributor.
Other mechanisms besides oxidative stress and mitochondrial dysfunction have been related with T2D in recent years, such as endoplasmic reticulum (ER) stress and autophagy. However, less is known about how these processes participate in the physiopathology of T2D. Moreover, further research is needed to assess the relationship between the inflammatory/adhesive process in the vascular walls and intracellular mechanisms, such as oxidative and ER stress or autophagy in leukocytes, that can eventually lead to vascular complications.
Objectives:
In the current Doctoral Thesis we have aimed to evaluate the role of mitochondrial dysfunction in the activation of leukocytes in T2D, as well as its relationship with the interactions between leukocytes and endothelial cells, and the potential correlation among these factors and the onset of silent myocardial ischemia (SMI).
On the other hand, our goal was to assess the potential participation of myeloperoxidase (MPO) in reactive oxygen species (ROS) generation in patients with T2D, and to evaluate is potential relationship with endothelial function and the development of diabetic nephropathy.
We also decided to assess in T2D patients with different extent of glycemic control, the induction of oxidative stress and ER stress and the role of the latter in the promotion of leukocyte-endothelial interactions.
Finally, we aimed to determine whether the process of autophagy is altered in the leukocytes of T2D patients, as well as exploring its connection with oxidative and ER stress.
Methods:
T2D patients and healthy subjects were recruited and anthropometric parameters (weight, height, BMI, waist and hip circumferences, blood pressure) and biochemical assessments (from blood samples) were determined. SMI diagnosis was confirmed by electrocardiogram and a coronary angiography test and SPECT (Single Photon Emission Computed Tomography). Diabetic nephropathy was confirmed when a urinary albumin excretion over 30 mg/g creatinine was detected in T2D patients. To assess the concentration of MPO, cytokines and adhesion molecules, serum aliquots were stored at -80ºC after collection. MPO concentration was determined by an ELISA immunoassay, whereas cytokines and adhesion molecules were studied by X-MAP technology in LUMINEX equipment. Oxidative stress, mitochondrial dysfunction, ER stress and autophagy were evaluated following isolation of peripheral blood leukocytes by Ficoll gradient. Leukocytes were incubated with the following fluorescent probes to evaluate redox balance: DCFH-DA, a marker of total ROS production; Mitosox Red, an indicator of mitochondrial ROS production; TMRM, a marker of mitochondrial membrane potential; NAO, an indicator of mitochondrial mass; and Hoescht, a nuclear morphology probe. Fluorescence was quantified using a fluorescence microscope (IX81; Olympus) coupled to the static cytometry software “ScanR”. The mitochondrial respiration rate was assessed by a Clark-type oxygen electrode. Leukocyte-endothelial interactions were estimated using a parallel flow chamber in which leukocytes were drawn across an endothelial cell monolayer (isolated from human umbilical cords) at a controlled flow rate similar to the in vivo vascular system. Recordings of this process were obtained using a video camera (Sony Exware HAD) coupled to a phase contrast microscope (Nikon Eclipse TE2000-S) and finally, the processes of leukocyte rolling and adhesion were estimated. To assess the levels of ER stress and autophagy markers, the RNA and proteins from leukocytes were isolated and Real-Time PCR and Western Blotting were performed, respectively. Statistical analyses were performed by means of Graphpad Prism and SPSS software.
Results and conclusions:
We compared T2D patients (with at least 2 years of disease duration) with age and sex matched control subjects. T2D patients showed increases in BMI, waist circumference and blood pressure. Furthermore, they displayed increased fasting glucose and insulin levels and, therefore, an enhanced HOMA-IR index, as well as elevated HbA1c levels. A rise in triglycerides and reduced HDL content was also observed in T2D patients. The inflammatory parameters PCR, IL6 and TNFα were increased in the serum of T2D patients, as were the adhesion molecules E-selectin and VCAM-1. These results suggest there is an imbalanced metabolism in T2D patients, which, together with abdominal obesity, promotes a low-grade inflammatory state of metabolic origin.
As a consequence of the chronic inflammatory condition in T2D, leukocytes display diminished rolling velocity and enhanced rolling flux and adhesion over the endothelium. When these properties are increased, endothelial dysfunction is promoted and leads to the vascular damage that precedes cardiovascular diseases. One of the mechanisms that promotes this prothrombotic state in leukocytes may be oxidative stress and mitochondrial dysfunction, as our results confirm. Leukocytes from T2D patients showed enhanced ROS production, impaired antioxidant response, less mitochondrial mass, reduced membrane potential and a reduction in the oxygen consumption rate when compared to those from the control group. These features were even more pronounced in patients with SMI. At the same time, mitochondrial ROS production was strongly correlated with the increase in leukocyte-endothelial interactions, suggesting that mitochondrial dysfunction is a determinant factor in the induction of SMI.
On the other hand, we observed a higher MPO concentration in serum from T2D patients, which was even higher in those with diabetic nephropathy. Furthermore, MPO levels correlated positively with the albumin/creatinine ratio. The interaction between leukocytes and the endothelium was more frequent in diabetic patients with nephropathy, and was accompanied by a rise in TNFα and E-selectin levels. These findings indicate that MPO contributes to an enhanced ROS generation in the leukocytes of T2D patients, which could have an important role in the development of diabetic nephropathy mediated by endothelial dysfunction.
Regarding the potential activation of ER stress in T2D, we observed a rise in GRP78 levels in general. However, glycemic control seemed to influence the pattern of expression of several ER stress markers, and appeared to follow an adaptive pattern in well-controlled patients and a proapoptotic pattern in poorly controlled T2D patients. Moreover, the group of poorly controlled patients displayed enhanced levels of inflammatory markers and increased rolling flux. A relationship between ER stress activation and leukocyte-endothelial interactions was apparent, since the adaptive markers GRP78 and sXBP1 were negatively correlated with rolling velocity and the proapoptotic marker CHOP was positively correlated with leukocyte adhesion. These findings indicate that, when T2D is well controlled, the velocity of leukocytes along the endothelium slows down and, when there is poor control of the disease, leukocyte adhesion is exacerbated.
Finally, we assessed the role of autophagy in T2D by determining LC3-II and Beclin1 levels in leukocytes. We detected enhanced levels of these autophagic markers together with a chromatin condensation increase that indicated ongoing cell death. The same leukocytes displayed signs of mitochondrial dysfunction, including reduced mitochondrial mass, enhanced ROS production, and increased ER stress with a rise in GRP78, p-eIF2α and ATF6 levels. Furthermore, Beclin1 levels were positively associated with ROS production, while Beclin1 and LC3-II were associated with markers of ER stress. These results suggest that T2D alters autophagic function in leukocytes, and that this function is regulated in parallel with oxidative and ER stresses during T2D.
Overall, our findings reveal that mitochondrial dysfunction, oxidative stress, ER stress and autophagy oc
Oxobenzo[f]benzopyrans as new fluorescent photolabile protecting groups for the carboxylic function
The properties of three oxobenzo[f]benzopyrans as new fluorogenic photolabile protecting groups for the carboxylic function of amino acids were studied. Fluorescent amino acid conjugates were efficiently prepared and characterised. Photodeprotection of these compounds was carried out by irradiation at 300, 350 and 419 nm, the most suitable wavelength being 350 nm, on account of short irradiation times and good deprotection yields. (c) 2006 Elsevier Ltd. All fights reserved.We thank the Fundação para a Ciência e Tecnologia (Portugal) for its financial support to the Centro de Química (Universidade do Minho)
Metformin and its redox-related mechanisms of action in type 2 diabetes
Type 2 diabetes (T2D) is a long-term metabolic disease characterized by progressive β-cell functional decline and insulin resistance, which increases the risk of cardiovascular complications as well as associated-morbidity and mortality. Evidence suggests a strong relationship between hyperglycaemia, oxidative stress and the development and progression of T2D. Indeed, a hyperglycaemic state can reduce the activity of antioxidant enzymes and increase lipid peroxidation and protein oxidation products, as well as DNA damage. At present, metformin is the recommended first-line glucose-lowering agent in patients with T2D. Despite the vast clinical experience gained over several decades of use, several mechanisms of action of metformin have yet to be fully elucidated. This review provides an overview of the existing literature concerning the complicated interplay between oxidative stress and T2D and the molecular mechanisms underlying the redox-related mechanisms of action of metformin, which include (but are not limited to) interaction with AMP-activated protein kinase (AMPK)-dependent and AMPK-independent mechanisms, inhibition of gluconeogenesis and action on leukocyte–endothelium interactions
Relevance of secretor status genotype and microbiota composition in susceptibility to rotavirus and norovirus infections in humans
Host genetic factors, such as histo-blood group antigens (HBGAs), are associated with susceptibility to norovirus (NoV) and rotavirus (RV) infections. Recent advances point to the gut microbiome as a key player necessary for a viral pathogen to cause infection. In vitro NoV attachment to host cells and resulting infections have been linked to interactions with certain bacterial types in the gut microbiota. We investigated the relationship between host genotype, gut microbiota, and viral infections. Saliva and fecal samples from 35 adult volunteers were analysed for secretor status genotype, the gut microbiota composition by 16S rRNA gene sequencing, and salivary IgA titers to NoV and RV. Higher levels of IgA against NoV and RV were related to secretor-positive status. No significant differences were found between the FUT2 genotype groups, although the multivariate analysis showed a significant impact of host genotype on specific viral susceptibilities in the microbiome composition. A specific link was found between the abundance of certain bacterial groups, such as Faecalibacterium and Ruminococcus spp., and lower IgA titers against NoV and RV. As a conclusion, we can state that there is a link between host genetics, gut microbiota, and susceptibility to viral infections in humans
Procesos de regulación metacognitiva en las transformaciones de tratamiento y conversión para solucionar problemas en estructuras aditivas de los números enteros
En nuestra experiencia docente encontramos que los estudiantes tienen dificultades en la resolución de problemas matemáticos. La presente investigación, siguiendo un enfoque cualitativo, interpretativo y descriptivo, se centra en analizar de qué manera la regulación metacognitiva favorece las transformaciones de tratamiento y conversión en la solución de problemas en estructuras aditivas de los números enteros. Los resultados muestran que en la etapa de indagación los participantes muestras pocas habilidades acerca de los componentes de la regulación metacognitiva, pero luego se encontró que los participantes utilizaron progresivamente los procesos de planeación, monitoreo y control, así como las transformaciones de tratamiento y conversión ante los problemas planteados. En conclusión, se mostró la forma en que los procesos de regulación metacognitiva inciden positivamente en los participantes, al desarrollar la capacidad para autorregular su propio aprendizaje. Igualmente, se observó el fortalecimiento de las transformaciones de tratamiento y conversión en el proceso de resolución de problemas.In our teaching experience we found that students have difficulties in solving mathematical problems. This paper aims to follow a qualitative, interpretative and descriptive approach. It focuses on analyzing how metacognitive regulation favors the transformation of treatment and conversion in the solution of problems in additive structures of integer numbers. The results show that at a starting stage the participants show few skills about the components of metacognitive regulation, but then it was found that the participants progressively used the planning, monitoring and control processes, as well as the transformations of treatment and conversion before the proposed problems. As a conclusion, it was shown that processes of metacognitive regulation affected positively the participants, by developing the ability to self-regulate their own learning. In the same way, the strengthening of treatment and conversion transformations in problem-solving was sawn
Occurrence of arsenic species in algae and freshwater plants of an extreme arid region in northern Chile, the Loa River Basin
This study reports data on arsenic speciation in two green algae species (Cladophora sp. and Chara sp.) and in five aquatic plants (Azolla sp., Myriophyllum aquaticum, Phylloscirpus cf. desserticola, Potamogeton pectinatus, Ruppia filifolia and Zannichellia palustris) from the Loa River Basin in the Atacama Desert (northern Chile). Arsenic content was measured by Mass Spectrometry coupled with Inductively Coupled Plasma (ICP-MS), after acidic digestion. Liquid Chromatography coupled to ICP-MS was used for arsenic speciation, using both anionic and cationic chromatographic exchange systems. Inorganic arsenic compounds were the main arsenic species measured in all samples. The main arsenic species in the extracts of freshwater algae and plants were arsenite and arsenate, whereas glycerol-arsenosugar (gly-sug), dimethylarsinic acid (DMA) and methylarsonic acid (MA) were present only as minor constituents. Of the samples studied, algae species accumulated more arsenic than aquatic plants. Total arsenic content ranged from 182 to 11,100 and from 20 to 248 mg As kg-1 (d.w.) in algae and freshwater plants, respectively. In comparison with As concentration in water samples, there was hyper-accumulation (>0.1% d.w.) in Cladophora sp
Clinical characteristics and comorbidities of elderly asthmatics who attend allergy clinics
Background: To date, few studies have focused on the clinical and allergic characteristics of asthma in the elderly, defined as asthma in people aged 60 or over. Thus, we propose to identify and study the clinical and allergic characteristics and comorbidities of patients with asthma among the elderly.Methods: A retrospective, observational, descriptive study was developed in five clinics and hospitals in Argentina. Allergy Physicians analyzed their patients? medical records in 2014 and included those adults over the age of 60, who had been diagnosed with asthma according to the GINA guidelines. Clinical and allergic characteristics were analyzed.Results: A total of 152 patients diagnosed with asthma, of whom 73% were women and 11% ex-smokers, were included in this study, with a mean age of 66 years. Only 10.5% of the participants had onset asthma past the age of 60. Regarding asthma severity, 74.3% were diagnosed with moderate persistent asthma, and 7.2% with severe persistent asthma. Eighty-four percent of the patients were treated with an inhaled corticosteroid (ICS) along with a long-acting β 2-adrenergic agent (LABA). More than half of the patients had two or more comorbidities simultaneously. Allergic comorbidities were the most frequent comorbidities, followed by arterial hypertension. Among allergic comorbidities, most patients presented allergies at the nasal level. There were no significant differences between the subpopulations of patients with late-onset asthma (LOA) and asthma with onset before the age of 60, i.e. early onset asthma (EOA) in most of their clinical characteristics. However, it was observed that EOA accounted for a higher percentage of patients with nasal allergies as compared to LOA (71% vs 46%, p < 0.05).It is worth mentioning that almost half of the patients with LOA had allergies at the nasal level.Conclusion: These results may provide a better understanding of the clinical characteristics of asthma in the elderly in Argentina, thus, enabling the development of future therapeutic strategies and a better quality of life for our elderly asthma patients.Fil: Yáñez, Anahí. Investigaciones En Alergia y Enfermedades Respiratoria; ArgentinaFil: Soria, Marcela. Gobierno de la Ciudad Autónoma de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Departamento de Medicina; ArgentinaFil: de Barayazarra, Norma Susana. Gobierno de la Provincia de Cordoba. Nuevo Hospital San Roque.; ArgentinaFil: Recuero, Nancy. Gobierno de la Provincia de Cordoba. Nuevo Hospital San Roque.; ArgentinaFil: Rovira, Francisco. No especifíca;Fil: Jares, Edgardo. No especifíca;Fil: Stok, Ana María. Instituto de Investigaciones En Patologías Respiratorias; ArgentinaFil: Nemirovsky, Sergio Ivan. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Bueno, Carlos Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; Argentin
Interaction of intestinal bacteria with human rotavirus during infection in children
The gut microbiota has emerged as a key factor in the pathogenesis of intestinal viruses, including enteroviruses, noroviruses and rotaviruses (RVs), where stimulatory and inhibitory effects on infectivity have been reported. With the aim of determining whether members of the microbiota interact with RVs during infection, a combination of anti-RV antibody labeling, fluorescence-activated cell sorting and 16S rRNA amplicon sequencing was used to characterize the interaction between specific bacteria and RV in stool samples of children suffering from diarrhea produced by G1P[8] RV. The genera Ruminococcus and Oxalobacter were identified as RV binders in stools, displaying enrichments between 4.8- and 5.4-fold compared to samples nonlabeled with anti-RV antibodies. In vitro binding of the G1P[8] Wa human RV strain to two Ruminococcus gauvreauii human isolates was confirmed by fluorescence microscopy. Analysis in R. gauvreauii with antibodies directed to several histo-blood group antigens (HBGAs) indicated that these bacteria express HBGA-like substances on their surfaces, which can be the target for RV binding. Furthermore, in vitro infection of the Wa strain in differentiated Caco-2 cells was significantly reduced by incubation with R. gauvreauii. These data, together with previous findings showing a negative correlation between Ruminococcus levels and antibody titers to RV in healthy individuals, suggest a pivotal interaction between this bacterial group and human RV. These results reveal likely mechanisms of how specific bacterial taxa of the intestinal microbiota could negatively affect RV infection and open new possibilities for antiviral strategies
Recombinant Noroviruses Circulating in Spain from 2016 to 2020 and Proposal of Two Novel Genotypes within Genogroup I
Noroviruses are the leading cause of sporadic cases and outbreaks of viral gastroenteritis. For more than 20 years, most norovirus infections have been caused by the pandemic genotype GII.4, yet recent studies have reported the emergence of recombinant strains in many countries. In the present study, 4,950 stool samples collected between January 2016 and April 2020 in Valencia, Spain, from patients with acute gastroenteritis were analyzed to investigate the etiological agent. Norovirus was the most frequently detected enteric virus, with a positivity rate of 9.5% (471/4,950). Among 224 norovirus strains characterized, 175 belonged to genogroup II (GII) and 49 belonged to GI. Using dual genotyping based on sequencing of the open reading frame 1 (ORF1)/ORF2 junction region, we detected 25 different capsid-polymerase-type associations. The most common GII capsid genotype was GII.4 Sydney 2012, followed by GII.2, GII.3, GII.6, and GII.17. A high prevalence of recombinant strains (90.4%) was observed among GII infections between 2018 and 2020. GII.4 Sydney[P16] was the predominant genotype from 2019 to 2020. In addition, GII.P16 polymerase was found harbored within six different capsid genes. GI.4 and GI.3 were the predominant genotypes in genogroup I, in which recombinant strains were also found, such as GI.3[P10], GI.3[P13], and GI.5[P4]. Interestingly, applying the criterion of 2 times the standard deviation, we found that 12 sequences initially classified as GI.3 may represent two new tentative genotypes in genogroup I, designated GI.10 and GI.11. This study shows the extensive diversity of recombinant noroviruses circulating in Spain and highlights the role of recombination events in the spread of noroviruses
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