Immune recovery following bronchiolitis is linked to a drop in cytokine and LTC4 levels.

BACKGROUND: Bronchiolitis is the main cause of hospitalization of children younger than 1 year; however, the immune mechanism of bronchiolitis is not completely understood. The aim of this study was to analyze the recovery of immune response after a bronchiolitis episode. METHODS: Forty-nine infants hospitalized with bronchiolitis diagnosis were enrolled. Nasopharyngeal aspirates (NPAs) were processed. Twenty-seven pro-inflammatory biomarkers linked to innate immunity, inflammation, and epithelial damage, as well as nitrites and lipid mediators, were evaluated in the NPA supernatant by ELISA (enzyme-linked immunosorbent assay) and Luminex. Also, 11 genes were analyzed in NPA cells by quantitative PCR. RESULTS: A widespread statistically significant decline of multiple pro-inflammatory parameters and cytokines were detected in the recovery period after respiratory infection: interferon-α2 (IFNα2), IFNγ, interleukin-10 (IL-10), IL-1ß, IL-8, IFN-γ-inducible protein-10, vascular endothelial growth factor, monocyte chemoattractant protein-1, macrophage inflammatory protein-1α (MIP-1α), and MIP-1ß. Supporting these results, a decreased nuclear factor-κB gene expression was observed (P = 0.0116). A significant diminution of cysteinyl leukotriene C4 (LTC4) soluble levels (P = 0.0319) and cyclooxygenase-2 (COX-2) gene expression were observed in the recovery sample. In children classified by post-bronchiolitis wheezing, LTC4 remains elevated in the NPA supernatant. CONCLUSIONS: After bronchiolitis, cytokines and biomarkers linked to innate immune response in NPA decrease significantly in the recovery period accompanied by a drop in LTC4 levels; however, this reduction was lower in infants with post-bronchiolitis wheezing.

Asthma diagnosis using integrated analysis of eosinophil microRNAs.

BACKGROUND: Asthma is a syndrome characterized by airway inflammation and obstruction. Due to its heterogeneity, the difficulties in asthma diagnosis and treatment make the discovery of new biomarkers a focus of research. So, we determined the differential miRNA expression of eosinophils between healthy and asthmatic patients and to establish a differentially expressed miRNA profile detectable in sera for use as biomarker. METHODS: MicroRNAs from peripheral eosinophils from healthy and asthmatic subjects were isolated and analyzed by next-generation sequencing and confirmed by quantitative PCR in 29 asthmatics and 10 healthy individuals. The levels of serum miRNAs were performed by quantitative PCR in 138 asthmatics and 39 healthy subjects. Regression analysis and Random Forest models were performed. RESULTS: We found a set of miRNAs whose expression differs between eosinophils from asthmatics and healthy subjects. These miRNAs can classify asthmatics into two clusters that differed in the number of eosinophils and periostin concentration in serum. Some of these miRNAs were also confirmed in sera, as miR-185-5p which discriminates asthmatics from healthy subjects. Together with other two miRNAs, miR-185-5p allowed us to create a logistic regression model to discriminate better both conditions and a Random Forest model that can even sort the asthmatics into intermittent, mild persistent, moderate persistent, and severe persistent asthma. CONCLUSION: Our data show that miRNAs profile in eosinophils can be used as asthma diagnosis biomarker in serum and that this profile is able to rank asthma severity.

Impaired erythropoietin synthesis in chronic kidney disease is caused by alterations in extracellular matrix composition.

Renal fibrosis and anaemia are two of the most relevant events in chronic kidney disease. Fibrosis is characterized by the accumulation of extracellular matrix proteins in the glomeruli and tubular interstitium. Anaemia is the consequence of a decrease in erythropoietin production in fibrotic kidneys. This work analyses the possibility that the accumulation of abnormal collagens in kidney interstitium could be one of the mechanisms responsible for erythropoietin decreased synthesis. In renal interstitial fibroblast grown on collagen I, erythropoietin mRNA expression and HIF-2α protein decreased, whereas focal adhesion kinase protein (FAK) phosphorylation and proteasome activity increased, compared to cells grown on collagen IV. Proteasome inhibition or FAK inactivation in cells plated on collagen I restored erythropoietin and HIF-2α expression. FAK inhibition also decreased the collagen I-dependent proteasome activation. In a model of tubulointerstitial fibrosis induced by unilateral ureteral obstruction in mice, increased collagen I protein content and an almost complete disappearance of erythropoietin mRNA expression were observed in the ureteral ligated kidney with respect to the contralateral control. Interestingly, erythropoietin synthesis was recovered in obstructed mice treated with proteasome inhibitor. These data suggest that reduced kidney erythropoietin synthesis could be caused by the accumulation of abnormal extracellular matrix proteins.

Corrigendum to "Glucose Oxidase Induces Cellular Senescence in Immortal Renal Cells through ILK by Downregulating Klotho Gene Expression".

[This corrects the article DOI: 10.1155/2015/416738.].

Glucose Oxidase Induces Cellular Senescence in Immortal Renal Cells through ILK by Downregulating Klotho Gene Expression.

Cellular senescence can be prematurely induced by oxidative stress involved in aging. In this work, we were searching for novel intermediaries in oxidative stress-induced senescence, focusing our interest on integrin-linked kinase (ILK), a scaffold protein at cell-extracellular matrix (ECM) adhesion sites, and on the Klotho gene. Cultured renal cells were treated with glucose oxidase (GOx) for long time periods. GOx induced senescence, increasing senescence associated ß-galactosidase activity and the expression of p16. In parallel, GOx increased ILK protein expression and activity. Ectopic overexpression of ILK in cells increased p16 expression, even in the absence of GOx, whereas downregulation of ILK inhibited the increase in p16 due to oxidative stress. Additionally, GOx reduced Klotho gene expression and cells overexpressing Klotho protein did not undergo senescence after GOx addition. We demonstrated a direct link between ILK and Klotho since silencing ILK expression in cells and mice increases Klotho expression and reduces p53 and p16 expression in renal cortex. In conclusion, oxidative stress induces cellular senescence in kidney cells by increasing ILK protein expression and activity, which in turn reduces Klotho expression. We hereby present ILK as a novel downregulator of Klotho gene expression.

Hyperphosphatemia induces cellular senescence in human aorta smooth muscle cells through integrin linked kinase (ILK) up-regulation.

Aging is conditioned by genetic and environmental factors. Hyperphosphatemia is related to some pathologies, affecting to vascular cells behavior. This work analyze whether high concentration of extracellular phosphate induces vascular smooth muscle cells senescence, exploring the intracellular mechanisms and highlighting the in vivo relevance of this phenomenon. Human aortic smooth muscle cells treated with ß-Glycerophosphate (BGP, 10mM) suffered cellular senescence by increasing p53, p21 and p16 expression and the senescence associated ß-galactosidase activity. In parallel, BGP induced ILK overexpression, dependent on the IGF-1 receptor activation, and oxidative stress. Down-regulating ILK expression prevented BGP-induced senescence and oxidative stress. Aortic rings from young rats treated with 10mM BGP for 48h, showed increased p53, p16 and ILK expression and SA-ß-gal activity. Seven/eight nephrectomized rats feeding a hyperphosphatemic diet and fifteenth- month old mice showed hyperphosphatemia and aortic ILK, p53 and p16 expression. In conclusion, we demonstrated that high extracellular concentration of phosphate induced senescence in cultured smooth muscle through the activation of IGF-1 receptor and ILK overexpression and provided solid evidences for the in vivo relevance of these results since aged animals showed high levels of serum phosphate linked to increased expression of ILK and senescence genes.

Relevant role of PKG in the progression of fibrosis induced by TNF-like weak inducer of apoptosis.

TNF-like weak inducer of apoptosis (TWEAK) is an inflammatory cytokine that activates the FGF-inducible 14 receptor. Both TWEAK and the FGF-inducible 14 receptor are constitutively expressed in the kidney. TWEAK has been shown to modulate several biological responses, such as inflammation, proliferation, differentiation, and apoptosis, that contribute to kidney injury. However, the role of TWEAK in fibrosis and TWEAK-activated intracellular signaling pathways remain poorly understood. We tested the hypothesis that TWEAK can be a potent inducer of renal fibrosis by increasing transforming growth factor (TGF)-ß1 expression (a well-known switch in the fibrosis process) through PKG-I downregulation. We showed that in human mesangial cells, TWEAK increased TGF-ß1 expression and activity, leading to higher levels of the extracellular matrix protein fibronectin and decreased PKG-I expression and activity via the Ras pathway. PKG-I activation with 8-bromo-cGMP, Ras inactivation with dominant negative Ras, or Ras pathway inhibition with the ERK1/2 inhibitor PD-98059 resulted in the prevention of TWEAK-induced TGF-ß1 upregulation. In vivo, exogenous administration of TWEAK to wild-type mice downregulated kidney PKG-I and increased kidney TGF-ß1 expression. These effects were blunted in H-Ras knockout mice. Together, these data demonstrate, for the first time, the key role of PKG-I in TGF-ß1 induction by TWEAK in kidney cells.

Intracellular redox equilibrium is essential for the constitutive expression of AP-1 dependent genes in resting cells: studies on TGF-ß1 regulation.

The mechanisms involved in the continuous expression of constitutive genes are unclear. We hypothesize that steady state intracellular reactive oxygen species (ROS), which their levels are tightly maintained, could be regulating the expression of these constitutive genes in resting cells. We analyzed the regulation of an important constitutive gene, TGF-ß1, after decreasing intracellular ROS concentration in human mesangial cells. Decreased intracellular hydrogen peroxide by catalase addition reduced TGF-ß1 protein, mRNA expression and promoter activity. Furthermore, catalase decreased the basal activity of Activated Protein-1 (AP-1) that regulates TGF-ß1 promoter activity. This effect disappeared when AP-1 binding site was removed. Similar results were observed with another protein containing AP-1 binding sites in its promoter, such as eNOS, but it was not the case in other constitutive genes without any AP-1 binding site, as COX1 or PKG1. The pharmacological inhibition of the different ROS synthesis sources by blocking NADPH oxidase, the mitochondrial respiratory chain or xanthine oxidase, or the use of human fibroblasts with genetically deficient mitochondrial activity, induced a similar, significant reduction of steady state ROS concentration as the one observed with catalase. Moreover, there was decreased TGF-ß1 expression in all the cases excepting the xanthine oxidase blockade. These findings suggest a novel role for the steady state intracellular ROS concentration, where the compartmentalized, different systems involved in the intracellular ROS production, could be essential for the expression of constitutive AP1-dependent genes, as TGF-ß1.

Start-up of a clinical sample processing, storage and management platform: organisation and development of the REDinREN Biobank.

BACKGROUND: The creation of the Biobank, a resource pertaining to the Spanish Renal Research Network (REDinREN) promotes advances in clinical research on kidney disease in Spain. The Biobank's aims are to generate an archive of clinical samples and associated data, furnish those samples to research teams, and coordinate with European biobanks. METHOD: Applicable legislation had to be complied with in order to launch the Biobank project (Biomedical Research Law, Data Protection Law and Biological Sample Transport Regulations). A strict work protocol and a new database for the Network's clinical data were also implemented. RESULTS: Over time, the Biobank has acquired additional infrastructure and qualified personnel. In 2010, 2953 new patient samples were collected, giving a total of 37,043 stored vials containing different types of samples. Furthermore, the Biobank is currently participating in eleven research projects. DISCUSSION: Although the Biobank was originally designed for REDinREN use, we must take joint action to make this biological sample storage system and the many possibilities it offers available to the entire nephrological community with a view to promoting kidney disease research.

Puesta en marcha de una plataforma de proceso, almacenamiento y gestión de muestras clínicas: organización y desarrollo del Biobanco de REDinREN / Start-up of a clinical sample processing, storage and management platform: organisation and development of the Biobank of REDinREN

Antecedentes: La creación del Biobanco, como una plataforma dentro de la Red de Investigación Renal (REDinREN), impulsa el avance de la investigación clínica de la enfermedad renal en España. Los objetivos del Biobanco son la generación del archivo de muestras clínicas y de datos asociados, para su cesión a los grupos de investigación, y la coordinación con biobancos europeos. Métodos: Para su puesta en marcha, fue necesaria la implementación de la normativa vigente (Ley de Investigación Biomédica y de Protección de Datos y la Normativa del Transporte de Sustancias Biológicas), un estricto protocolo de trabajo y la creación de una base de datos clínicos de la Red. Resultados: En su evolución, el Biobanco ha adquirido infraestructura y personal cualificado, lo que permitió que en el año 2010 se obtuviera un total de 2.953 pacientes, lo que hace un total de 37.043 viales almacenados con muestras de diferentes naturalezas. Además, hasta la fecha, el Biobanco está incluido en 11 proyectos de investigación. Discusión: Aunque el Biobanco fue diseñado como una plataforma de soporte de la REDinREN, es necesario con una acción conjunta poner a disposición de toda la comunidad científica nefrológica las posibilidades que otorga este sistema de almacenamiento de muestras biológicas para potenciar la investigación de la enfermedad renal (AU)

Background: The Biobank creation in Network for the Kidney Research (REDinREN) promotes the advance of kidney disease clinic research in Spain. The Biobank's aims are to generate an archive of clinical samples and associated data, furnish those samples to research teams, and coordinate with European biobanks. Methods: In the beginning were indispensable the implementation of in force normative (Bio-medic Investigation Law, Dates Protect Law and Biologic Samples Transport Normative), a work protocol and the creation of medical database of the Red. Results: The Biobank growth included infastructures and qualified personnel. In 2010 year, the patien samples increased in 2953 achieved 37,042 vials of differect nature stored. Mereover in this moment the Biobank participates in eleventh research projects. Discussion: Even if the Biobank was designed for the support to REDinREN is neccesary to opening his biologic samples to all scientific nephrology community for promote the kidney disease research (AU)

Brucella abortus INTA2, a novel strain 19 (Delta)bp26::luc (Delta)bmp18 double mutant lacking drug resistance markers.

Brucella abortus INTA2, a novel mutant strain, was constructed by inactivation of two B. abortus S19 genes: bp26 and bmp18, with the objective of obtaining a mutant strain that could be compatible with a diagnostic test and have less residual virulence than strain 19. The double mutant was constructed by replacing a large section of the bp26 coding region with the luciferase (luc) coding gene, resulting in mutant strain B. abortus M1luc, followed by partial deletion of bmp18 coding sequence. Both genes were inactivated by allelic replacement assisted by sacB counter-selection. Luciferase expression was evaluated and confirmed that it is a valid marker in the construction of mutant strains. When B. abortus INTA2 was inoculated in BALB/c mice, significantly fewer colony forming units (CFUs) were recovered from mice spleens during initial phase of infection. No splenomegaly was observed in strain INTA2-immunized mice at any time suggesting that strain INTA2 has lost some residual virulence of the parental strain. Nevertheless, similar protection levels against virulent challenge were observed in mice immunized with strains INTA2 or S19. Although strain INTA2 would still induce O-side antibodies, it does not express BP26. This would allow differentiation of INTA2-vaccinated animals from animals infected with field strains by measuring anti-BP26 antibodies, either by an agglutination test or ELISA using BP26 as antigen. Altogether these results indicate that B. abortus INTA2 might be a promising vaccine strain against brucellosis.

Monitoreo ambiental en instalaciones para el control de los residuos sólidos

Desde hace cuatro años se establecieron Programas de monitoreo ambiental en instalaciones del Sistema de aseo urbano de la ciudad de México para realizar una operación compatible con el ambiente

Monitoreo ambiental en instalaciones para el control de los residuos sólidos

Desde hace cuatro años se establecieron Programas de monitoreo ambiental en instalaciones del Sistema de aseo urbano de la ciudad de México para realizar una operación compatible con el ambiente