Effect of homeostatic T-cell proliferation in the vaccine responsiveness against influenza in elderly people.

BACKGROUND: Seasonal influenza virus infection is a significant cause of morbimortality in the elderly. However, there is poor vaccine efficacy in this population due to immunosenescence. We aimed to explore several homeostatic parameters in the elderly that could impact influenza vaccine responsiveness. METHODS: Subjects (> 60 years old) who were vaccinated against influenza virus were included, and the vaccine response was measured by a haemagglutination inhibition (HAI) test. At baseline, peripheral CD4 and CD8 T-cells were phenotypically characterized. Thymic function and the levels of different inflammation-related biomarkers, including Lipopolysaccharide Binding Protein (LBP) and anti-cytomegalovirus (CMV) IgG antibodies, were also measured. RESULTS: Influenza vaccine non-responders showed a tendency of higher frequency of regulatory T-cells (Tregs) before vaccination than responders (1.49 [1.08-1.85] vs. 1.12 [0.94-1.63], respectively, p = 0.061), as well as higher expression of the proliferation marker Ki67 in Tregs and different CD4 and CD8 T-cell maturational subsets. The levels of inflammation-related biomarkers correlated with the frequencies of different proliferating T-cell subsets and with thymic function (e.g., thymic function with D-dimers, r = - 0.442, p = 0.001). CONCLUSIONS: Age-related homeostatic dysregulation involving the proliferation of CD4 and CD8 T-cell subsets, including Tregs, was related to a limited responsiveness to influenza vaccination and a higher inflammatory status in a cohort of elderly people.

Transcriptomic changes in Cucurbita pepo fruit after cold storage: differential response between two cultivars contrasting in chilling sensitivity.

BACKGROUND: Zucchini fruit is susceptible to chilling injury (CI), but the response to low storage temperature is cultivar dependent. Previous reports about the response of zucchini fruit to chilling storage have been focused on the physiology and biochemistry of this process, with little information about the molecular mechanisms underlying it. In this work, we present a comprehensive analysis of transcriptomic changes that take place after cold storage in zucchini fruit of two commercial cultivars with contrasting response to chilling stress. RESULTS: RNA-Seq analysis was conducted in exocarp of fruit at harvest and after 14 days of storage at 4 and 20 °C. Differential expressed genes (DEGs) were obtained comparing fruit stored at 4 °C with their control at 20 °C, and then specific and common up and down-regulated DEGs of each cultivar were identified. Functional analysis of these DEGs identified similarities between the response of zucchini fruit to low temperature and other stresses, with an important number of GO terms related to biotic and abiotic stresses overrepresented in both cultivars. This study also revealed several molecular mechanisms that could be related to chilling tolerance, since they were up-regulated in cv. Natura (CI tolerant) or down-regulated in cv. Sinatra (CI sensitive). These mechanisms were mainly those related to carbohydrate and energy metabolism, transcription, signal transduction, and protein transport and degradation. Among DEGs belonging to these pathways, we selected candidate genes that could regulate or promote chilling tolerance in zucchini fruit including the transcription factors MYB76-like, ZAT10-like, DELLA protein GAIP, and AP2/ERF domain-containing protein. CONCLUSIONS: This study provides a broader understanding of the important mechanisms and processes related to coping with low temperature stress in zucchini fruit and allowed the identification of some candidate genes that may be involved in the acquisition of chilling tolerance in this crop. These genes will be the basis of future studies aimed to identify markers involved in cold tolerance and aid in zucchini breeding programs.

Phenotype and Polyfunctional Deregulation Involving Interleukin 6 (IL-6)- and IL-10-Producing Monocytes in HIV-Infected Patients Receiving Combination Antiretroviral Therapy Differ From Those in Healthy Older Individuals.

BACKGROUND: Despite the relevance of monocytes as promoters of the inflammatory response, whether human immunodeficiency virus (HIV) infection induces premature age-related changes to the phenotype and function of monocytes or whether these alterations are different and/or specifically driven by HIV remains to be mechanistically determined. METHODS: We assayed the activation phenotype and the responsiveness in vitro to Toll-like receptor (TLR) agonists in classical, intermediate, and nonclassical subsets of monocytes by assessing intracellular interleukin 1α (IL-1α), IL-1ß, interleukin 6 (IL-6), interleukin 8, tumor necrosis factor α, and interleukin 10 (IL-10) production in 20 HIV-infected patients receiving combination antiretroviral therapy (cART) and 2 groups of uninfected controls (20 age-matched young individuals and 20 older individuals aged >65 years). RESULTS: HIV-infected patients showed a more activated phenotype of monocytes than older controls. Regarding functionality, under unstimulated conditions HIV-infected patients showed a higher percentage of classical monocytes producing IL-6 and IL-10 than control subjects. The percentage of cells with production of multiple cytokines (polyfunctionality), including IL-10, in response to TLR agonists was greater among HIV-infected patients than among control subjects. CONCLUSIONS: Inflammatory alterations associated with monocytes during HIV infection are different from those in aging individuals. This monocyte dysfunction, mainly characterized by high levels of IL-6- and IL-10-producing monocytes, may have clinical implications in HIV-infected patients that are different from those in aging individuals.

Friedreich's ataxia: autosomal recessive disease caused by an intronic GAA triplet repeat expansion.

Friedreich's ataxia (FRDA) is an autosomal recessive, degenerative disease that involves the central and peripheral nervous systems and the heart. A gene, X25, was identified in the critical region for the FRDA locus on chromosome 9q13. This gene encodes a 210-amino acid protein, frataxin, that has homologs in distant species such as Caenorhabditis elegans and yeast. A few FRDA patients were found to have point mutations in X25, but the majority were homozygous for an unstable GAA trinucleotide expansion in the first X25 intron.

SOX7 transcription factor: sequence, chromosomal localisation, expression, transactivation and interference with Wnt signalling.

The Sox gene family consists of several genes related by encoding a 79 amino acid DNA-binding domain known as the HMG box. This box shares strong sequence similarity to that of the testis determining protein SRY. SOX proteins are transcription factors having critical roles in the regulation of diverse developmental processes in the animal kingdom. We have characterised the human SOX7 gene and compared it to its mouse orthologue. Chromosomal mapping analyses localised mouse Sox7 on band D of mouse chromosome 14, and assigned human SOX7 in a region of shared synteny on human chromosome 8 (8p22). A detailed expression analysis was performed in both species. Sox7 mRNA was detected during embryonic development in many tissues, most abundantly in brain, heart, lung, kidney, prostate, colon and spleen, suggesting a role in their respective differentiation and development. In addition, mouse Sox7 expression was shown to parallel mouse Sox18 mRNA localisation in diverse situations. Our studies also demonstrate the presence of a functional transactivation domain in SOX7 protein C-terminus, as well as the ability of SOX7 protein to significantly reduce Wnt/beta-catenin-stimulated transcription. In view of these and other findings, we suggest different modes of action for SOX7 inside the cell including repression of Wnt signalling.

[Screening for retinopathy of prematurity. Our experience about limits of birth weight, post-conceptional age and other risk factors]. / Despistaje de la retinopatía del prematuro. Nuestra experiencia sobre los límites de peso al nacer, edad gestacional y otros factores de riesgo.

PURPOSE: To assess the features of Retinopathy of Prematurity (ROP) in a Neonatal Intensive Care Unit during 8 years, analyzing the usefulness of the criteria in the screening protocol and the risk factors relating to the disease. METHODS: A retrospective study of the infants included in the screening program for ROP. The sensitivity of the criteria of birth weight (BW) < 1,500 g, post-conceptional age (PCA) < 32 weeks and subjective pediatric criteria (SPC) in the screening program were evaluated and compared with the incidence of ROP, and the need for treatment in these patient groups. Statistical analysis for ROP and no-ROP was then applied to the risk factors BW, PCA, oxygen therapy, and intercurrent diseases. RESULTS: Forty of 303 infants studied had ROP (13.2%). In ROP cases, 31 (77.5%) had spontaneous regression and 9 (22.5%) needed treatment. The screening program included 144 children with BW < 1,500 g and 159 children with BW > 1,500 g. The incidence of ROP was 26.4% in the first group and 1.3% in the second group (p < 0.001). Two cases were detected with a BW > 1,500 g but with a PCA < 32 weeks, and neither required treatment. There were 84 cases included because of SPC (27.7%); no cases of ROP were detected in these. The only independent risk factor found in a multivariant analysis was birth weight. CONCLUSIONS: None of the children included with a BW > 1500 g required treatment for ROP, but several cases of ROP could be missed by using this criteria only. SPC must be restricted in the screening program.

Mapping of Friedreich's ataxia locus by identification of recombination events in patients homozygous by descent.

The Friedreich's ataxia locus (FRDA) maps on chromosome 9q13. Genetic data, obtained from a small number of recombination events, indicated that the FRDA locus might be located centromeric to the D9S15/D9S5 linkage group, the most probable order being cen-FRDA-D9S5-D9S111-D9S15-D9S110-qter. Recently, new centromeric markers have been reported. Analysis of these markers allowed us to localize the recombination breakpoint in some of the recombinant families. However, only one proximal recombination has been found with these markers. To increase the genetic information from FRDA families, we have analyzed the centromeric markers FR1, FR2, FR7, FR8, and FR5 in patients homozygous by descent. These were ascertained because parents were consanguineous or because they were homozygous for the entire haplotype D9S15 or D9S111-D9S5-D9S411E-D9S202. Haplotype divergence for, at least, two contiguous markers was observed in two patients homozygous for the core D9S111-FR2 haplotype and in one third-degree consanguineous family homozygous for haplotype D9S411E-FR5. Interpretation of divergence as the result of ancient meiotic crossovers allowed the definition of three new recombination events which place the FRDA locus within the interval defined by markers D9S411E and FR8. A consanguineous family with first-cousin parents showed homozygosity only at D9S202 and FR2. Further investigations are needed to discern whether two different mutations are segregating in the family or whether two recombinations, one distal and one proximal, have taken place.

Evidence for a common origin of most Friedreich ataxia chromosomes in the Spanish population.

Haplotype analysis is a powerful approach to understand the spectrum of mutations accounting for a disease in a homogeneous population. We show that haplotype variation for 10 markers linked to the Friedreich ataxia locus (FRDA) argues in favor of an important mutation homogeneity in the Spanish population, and positions the FRDA locus in the region where it has been recently isolated. We also report the finding of a new single nucleotide polymorphism called FAD1. The new marker shows a very strong linkage disequilibrium with Friedreich ataxia (FA) in both the Spanish and French populations. suggesting the existence of an ancient and widespread FRDA mutations. Inclusion of FAD1 in the extended haplotype analysis has allowed to postulate that this main FRDA mutation could account for 50-90% of the disease chromosomes. The results indicate that FA, despite clinical heterogeneity, could have originated from a few initial mutations.

dfh is a Drosophila homolog of the Friedreich's ataxia disease gene.

A putative Drosophila homolog of the Friedreich's ataxia disease gene (FRDA) has been cloned and characterized; it has been named Drosophila frataxin homolog (dfh). It is located at 8C/D position on X chromosome and is spread over 1kb, a much smaller genomic region than the human gene. Its genomic organization is simple, with a single intron dividing the coding region into two exons. The predicted encoded product has 190 amino acids, being considered a frataxin-like protein on the basis of the sequence and secondary structure conservation when compared with human frataxin and related proteins from other eukaryotes. The closest match between the Drosophila and the human proteins involved a stretch of 38 amino acids at C-terminus, encoded by dfh exon 2, and exons 4 and 5a of the FRDA gene, respectively. This highly conserved region is very likely to form a functional domain with a beta sheet structure flanked by alpha-helices where the sequence is less conserved. A signal peptide for mitochondrial import has also been predicted in the Drosophila frataxin-like protein, suggesting its mitochondrial localization, as occurs for human frataxin and other frataxin-like proteins described in eukaryotes. The Drosophila gene is expressed throughout the development of this organism, with a peak of expression in 6-12h embryos, and showing a spatial ubiquitous pattern from 4h embryos to the last embryonic stage examined. The isolation of dfh will soon make available specific dfh mutants that help in understanding the pathogenesis of FRDA.

Suppression of pigment interference in the gas chromatographic analysis of proteinaceous binding media in paintings with EDTA.

A method to suppress the interference of pigments in the analysis of proteinaceous media used in paintings is presented in this paper. This method is based on the formation of metallic ion-ethylenediaminetetraacetic acid (EDTA) complexes previous to the derivatisation process, using ethyl chloroformate (ECF), to transform the amino acids in N(O,S)-ethoxycarbonyl (EOC) ethyl esters. Test specimens, containing different proteinaceous media such as albumin, porcine gelatine and casein mixed with lead white, chalk, verdigris and raw Sienna have been prepared for carrying out this study. Different pH conditions have been probed for the different pigments studied. Values of peak area ratio of amino acids relative to the alanine, obtained using the proposed method on a series of protein-pigment test specimens, have been compared to those from specimens of pure protein in which direct method of derivatisation was applied. Finally, the method has been successfully applied to the analysis of 18th century wall paintings in which animal glue was used as binding medium.

Dietary virgin olive oil triacylglycerols as an independent determinant of very low-density lipoprotein composition.

OBJECTIVE: We examined the effects of virgin olive oil (VOO) triacylglycerols (TGs) on the lipid composition of human very low-density lipoprotein (VLDL). METHODS: Twenty-one normocholesterolemic, normotensive, non-diabetic elderly subjects were recruited for the study. Two VOOs (VOO1 and VOO2) of the same variety, with an equivalent composition in minor components and differing only in the oleic and linoleic acid concentrations, were administered for 4 wk each to assess the effect of their TG molecular species compositions. Blood was collected after an overnight fast, VLDLs were isolated by ultracentrifugation, and lipid classes, TG molecular species, and TG fatty acid composition were determined. RESULTS: Dietary VOOs significantly differed in TG molecular species composition. VOO1 represented larger amounts of triolein (P < 0.01), whereas VOO2 was significantly enriched with dilinoleoyl-oleoyl-glycerol, linoleoyl-dioleoyl-glycerol, and linoleoyl-oleoyl-palmitoyl-glycerol (P < 0.01). For VLDL, intake of VOO1 caused an increase of total TG (P < 0.01) due mainly to increases in triolein and linoleoyl-dioleoyl-glycerol. Conversely, VOO2 increased VLDL cholesteryl esters (P < 0.01) and TG rich in arachidonic acid (P < 0.01). CONCLUSIONS: The different TG molecular species compositions of dietary oils may be an independent determinant of the lipid composition of VLDL in elderly people and therefore may play a role in regulating lipoprotein metabolism in these subjects.

Otoconial mineralization in post-natal development.

Otoconial mineralization has been studied in the utricle of post-natal rats, i.e. from birth to the 20th day. Calcium microanalytically detected is seen to decrease progressively until, after 20 days, when the level is rather lower than at birth. In the adult animal the calcium value is much higher, which leads us to believe that there is an immediate calcium loss that is later recovered. Calcium in the peripheral areas of the macula is proportionately less than in the central part.

Mineralization in the newborn rat statoconia. An EDAX study.

Several types of statoconia, differing in size and shape, have been observed in macula utriculi, the smaller ones in the macular periphery. The larger statoconia seem to be formed by the association of smaller units and calcium presence is comparatively higher: In some parts of the marginal zone there exists a granular material in which the presence of calcium is significant.

Meconium concentration and amniotic fluid index influence the outcome of amnioinfusion.

BACKGROUND: To determine the usefulness of amnioinfusion as a function of meconium concentration and amniotic fluid index. METHODS: This was a prospective study of 206 pregnant women in whom amniotic fluid was moderately or heavily stained with meconium, according to subjective evaluation. The women were assigned randomly to receive amnioinfusion (n=103) or no amnioinfusion (control group, n=103). The results were compared in women with =/<15 % or >15 % meconium in the amniotic fluid (measured by centrifugation), and in women in whom the amniotic fluid index calculated 60 min after insertion of the amnioinfusion catheter was <10 or =/>10. RESULTS: In women with >15% meconium, amnioinfusion decreased the rate of cesarian sections motivated by fetal distress (2.5% vs 22.2%), and in women with =/<15% meconium, amnioinfusion decreased the presence of meconium below the vocal cords (6.4% vs 25.9%). Greater benefits after amnioinfusion were seen in women with an amniotic fluid index =/>10: the rate of cesarian sections was lower (1.3% vs 13.3%), as was the frequency of meconium below the vocal cords (10.1% vs 33.3%). CONCLUSIONS: Beneficial effects of amnioinfusion were seen in women with high and low concentrations of meconium, and with high and low amniotic fluid indexes. These criteria should therefore not be used to decide whether amnioinfusion is indicated when the amniotic fluid is moderately or heavily stained with meconium.

Application of genomic tools in plant breeding.

Plant breeding has been very successful in developing improved varieties using conventional tools and methodologies. Nowadays, the availability of genomic tools and resources is leading to a new revolution of plant breeding, as they facilitate the study of the genotype and its relationship with the phenotype, in particular for complex traits. Next Generation Sequencing (NGS) technologies are allowing the mass sequencing of genomes and transcriptomes, which is producing a vast array of genomic information. The analysis of NGS data by means of bioinformatics developments allows discovering new genes and regulatory sequences and their positions, and makes available large collections of molecular markers. Genome-wide expression studies provide breeders with an understanding of the molecular basis of complex traits. Genomic approaches include TILLING and EcoTILLING, which make possible to screen mutant and germplasm collections for allelic variants in target genes. Re-sequencing of genomes is very useful for the genome-wide discovery of markers amenable for high-throughput genotyping platforms, like SSRs and SNPs, or the construction of high density genetic maps. All these tools and resources facilitate studying the genetic diversity, which is important for germplasm management, enhancement and use. Also, they allow the identification of markers linked to genes and QTLs, using a diversity of techniques like bulked segregant analysis (BSA), fine genetic mapping, or association mapping. These new markers are used for marker assisted selection, including marker assisted backcross selection, 'breeding by design', or new strategies, like genomic selection. In conclusion, advances in genomics are providing breeders with new tools and methodologies that allow a great leap forward in plant breeding, including the 'superdomestication' of crops and the genetic dissection and breeding for complex traits.

Sonography for assessment of haemophilic arthropathy in children: a systematic protocol.

Radiological imaging of joints in children with haemophilia is important to detect abnormalities, grade their severity and monitor the effects of treatment. Scoring systems for staging haemophilic arthropathy have been developed based on plain film or magnetic resonance imaging (MRI) findings. Radiographs alone may be inadequate for evaluating joint disease in children with haemophilia on prophylaxis while MRI may be difficult to access and require the child to be sedated. Sonography can be a useful complementary modality in the evaluation of haemophilic arthropathy that is readily available and does not require the child to be sedated. In this paper, we briefly review the current imaging scales available for the assessment of haemophilic arthropathy and present a systematic protocol for sonographic assessment of the knee and ankle in haemophilic children along with examples of findings in joint effusion/hemarthrosis, synovial hypertrophy and cartilage loss. Also, we correlate the ultrasound findings with the corresponding MRI images demonstrating the anatomic planes used for imaging acquisition. Sonography is a promising technique for the assessment of soft tissue changes which are the earliest findings in haemophilic arthropathy. Further investigation is required for evaluation of osteochondral changes given limitations of sonography in this regard and in minimizing operator dependency, especially if applied in multicentric clinical trials.

Biphasic behavior of changes in elemental composition during staurosporine-induced apoptosis.

Although the identification of events that occur during apoptosis is a fundamental goal of apoptotic cell death research, little is know about the precise sequence of changes in total elemental composition during apoptosis. We evaluated total elemental composition (Na, Mg, P, Cl, S, and K) in relation to molecular and morphological features in human U937 cells induced to undergo apoptosis with staurosporine, an intrinsic pathway activator. To evaluate total elemental content we used electron probe X-ray microanalysis to measure simultaneously all elements from single, individual cells. We observed two phases in the changes in elemental composition (mainly Na, Cl and K). The early phase was characterized by a decrease in intracellular K (P<0.001) and Cl (P<0.001) content concomitant with cell shrinkage, and preceded the increase in proteolytic activity associated with the activation of caspase-3. The later phase started with caspase-3 activation, and was characterized by a decrease in the K/Na ratio (P<0.001) as a consequence of a significant decrease in K and increase in Na content. The inversion of intracellular K and Na content was related with the inhibition of Na+/K+ ATPase. This later phase was also characterized by a significant increase (P<0.001) in intracellular Cl with respect to the early phase. In addition, we found a decrease in S content and an increase in the P/S ratio. These distinctive changes coincided with chromatin condensation and DNA fragmentation. Together, these findings support the concept that changes in total elemental composition take place in two phases related with molecular and morphological features during staurosporine-induced apoptosis.

Tirant is a new member of the gypsy family of retrotransposons in Drosophila melanogaster.

In this paper, we propose a consensus sequence for a putative complete Tirant retrotransposon. Several defective copies, as well as relevant sequences available in databases have been analyzed. The putative complete Tirant element is 8533 bp long, and presents all the structural features of a retrovirus-like transposable element of the gypsy family. It contains three ORFs (open reading frames) that encode putative products resembling the retroviral Gag, Pol, and Env proteins. Southern blot analyses show that complete and defective Tirant elements are widespread in Drosophila melanogaster. The different hybridization patterns observed in several natural populations of this species suggest that Tirant is an active element.

Isolation and characterization of an avirulent Candida albicans yeast monomorphic mutant.

Mutagenesis of Candida albicans strain ATCC 26555 with N-methyl-nitro-N-nitrosoguanidine followed by plating on solid yeast nitrogen base-N-acetylglucosamine medium at 37 degrees C yielded colony morphology variants that were characterized as forming smooth colonies, in contrast to the rough colonies formed by the parental strain. One yeast monomorphic mutant, CAL4, was studied in detail. Strain CAL4 is defective in filamentous growth, unable to form hyphae or pseudohyphae in vivo and in vitro. These filamentous structures are not elicited by commonly used external stimuli such as serum. The mutant had no obvious alterations in its mannan, glucan or chitin content. The total quantity of non-covalently linked wall proteins was reduced in the mutant strain, but the electrophoretic pattern shown by these proteins was identical to that of proteins from the parental strain. CAL4 showed major differences from the parental strain in its formation of covalently linked wall proteins. An important aspect of these differences lay in the practical absence of proteins recognized by two monoclonal antibodies, 1B12 and 3H8, which are considered valuable tools in the diagnosis of candidiasis in part because they normally react strongly with all strains. The C. albicans mutant, blocked in yeast-mycelium transition, was avirulent in a mouse model, although it was able to grow in animal tissues.