Using AnABlast for intergenic sORF prediction in the Caenorhabditis elegans genome.

MOTIVATION: Short bioactive peptides encoded by small open reading frames (sORFs) play important roles in eukaryotes. Bioinformatics prediction of ORFs is an early step in a genome sequence analysis, but sORFs encoding short peptides, often using non-AUG initiation codons, are not easily discriminated from false ORFs occurring by chance. RESULTS: AnABlast is a computational tool designed to highlight putative protein-coding regions in genomic DNA sequences. This protein-coding finder is independent of ORF length and reading frame shifts, thus making of AnABlast a potentially useful tool to predict sORFs. Using this algorithm, here, we report the identification of 82 putative new intergenic sORFs in the Caenorhabditis elegans genome. Sequence similarity, motif presence, expression data and RNA interference experiments support that the underlined sORFs likely encode functional peptides, encouraging the use of AnABlast as a new approach for the accurate prediction of intergenic sORFs in annotated eukaryotic genomes. AVAILABILITY AND IMPLEMENTATION: AnABlast is freely available at http://www.bioinfocabd.upo.es/ab/. The C.elegans genome browser with AnABlast results, annotated genes and all data used in this study is available at http://www.bioinfocabd.upo.es/celegans. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Perinatal cardiopulmonary adaptation to the thin air of the Alto Andino by a native Altiplano dweller, the llama.

Most mammals have a poor tolerance to hypoxia, and prolonged O2 restriction can lead to organ injury, particularly during fetal and early postnatal life. Nevertheless, the llama (Lama Glama) has evolved efficient mechanisms to adapt to acute and chronic perinatal hypoxia. One striking adaptation is the marked peripheral vasoconstriction measured in the llama fetus in response to acute hypoxia, which allows efficient redistribution of cardiac output toward the fetal heart and adrenal glands. This strong peripheral vasoconstrictor tone is triggered by a carotid body reflex and critically depends on α-adrenergic signaling. A second adaptation is the ability of the llama fetus to protect its brain against hypoxic damage. During hypoxia, in the llama fetus there is no significant increase in brain blood flow. Instead, there is a fall in brain O2 consumption and temperature, together with a decrease of Na+-K+-ATPase activity and Na+ channels expression, protecting against seizures and neuronal death. Finally, the newborn llama does not develop pulmonary hypertension in response to chronic hypoxia. In addition to maintaining basal pulmonary arterial pressure at normal levels the pulmonary arterial pressor response to acute hypoxia is lower in highland than in lowland llamas. The protection against hypoxic pulmonary arterial hypertension and pulmonary contractile hyperreactivity is partly due to increased hemoxygenase-carbon monoxide signaling and decreased Ca2+ sensitization in the newborn llama pulmonary vasculature. These three striking physiological adaptations of the llama allow this species to live and thrive under the chronic influence of the hypobaric hypoxia of life at high altitude.

Relationship between health-related quality of life, disease activity and disease damage in a prospective international multicenter cohort of childhood onset systemic lupus erythematosus patients.

Previously, we described associations between health-related quality of life (HRQOL) and disease-related factors among childhood onset systemic lupus erythematosus (cSLE) patients. Here we determined the relationship between HRQOL, disease activity and damage in a large prospective international cohort of cSLE. We compared HRQOL, disease activity and disease damage across different continents and examined the relationship between children's and parents' assessments of HRQOL. Patients with cSLE and their parents completed HRQOL measures at enrollment and ≥4 follow-up visits. Physicians assessed disease activity and damage. The multinational cohort ( n = 467) had relatively low disease activity and damage. Patient and parent HRQOL scores were significantly correlated. Asian and European patients had the highest HRQOL, while South and North American patients had lower HRQOL scores. Renal, CNS, skin and musculoskeletal systems exhibited the highest levels of damage. North and South American and Asian patients were more likely to have disease damage and activity scores above median values, compared with Europeans. Asians were more likely to use cyclophosphamide/rituximab. Female gender, high disease activity and damage, non-White ethnicity, and use of cyclophosphamide and/rituximab were related to lower HRQOL. HRQOL domain scores continue to emphasize that SLE has widespread impact on all aspects of children's and parents' lives.

Pharmacological approaches in either intermittent or permanent hypoxia: A tale of two exposures.

Hypoxia induces several responses at cardiovascular, pulmonary and reproductive levels, which may lead to chronic diseases. This is relevant in human populations exposed to high altitude (HA), in either chronic continuous (permanent inhabitants) or intermittent fashion (HA workers, tourists and mountaineers). In Chile, it is estimated that 1.000.000 people live at highlands and more than 55.000 work in HA shifts. Initial responses to hypoxia are compensatory and induce activation of cardioprotective mechanisms, such as those seen under intermittent hypobaric (IH) hypoxia, events that could mediate preconditioning. However, whenever hypoxia is prolonged, the chronic activation of cellular responses induces long-lasting modifications that may result in acclimatization or produce maladaptive changes with increase in cardiovascular risk. HA exposure during pregnancy induces hypoxia and oxidative stress, which in turn may promote cellular responses and epigenetic modifications resulting in severe impairment in growth and development. Sadly, this condition is accompanied with an increased fetal and neonatal morbi-mortality. Further, developmental hypoxia may program cardio-pulmonary circulations later in postnatal life, ending in vascular structural and functional alterations with augmented risk on pulmonary and cardiovascular failure. Additionally, permanent HA inhabitants have augmented risk and prevalence of chronic hypoxic pulmonary hypertension, right ventricular hypertrophy and cardiopulmonary remodeling. Similar responses are seen in adults that are intermittently exposed to chronic hypoxia (CH) such as shift workers in HA areas. The mechanisms involved determining the immediate, short and long-lasting effects are still unclear. For several years, the study of the responses to hypoxic insults and pharmacological targets has been the motivation of our group. This review describes some of the mechanisms underlying hypoxic responses and potential therapeutic approaches with antioxidants such as melatonin, ascorbate, omega 3 (Ω3) or compounds that increase the nitric oxide (NO) bioavailability.

Enhanced MCR-ALS modeling of HPLC with fast scan fluorimetric detection second-order data for quantitation of metabolic disorder marker pteridines in urine.

This work presents the development of a liquid chromatographic method based on modeling entire fast scan fluorimetric detection second-order data with the multivariate curve resolution alternating least squares algorithm, for the simultaneous determination of five marker pteridines in urine samples. The modeling strategy involves the building of a single MCR-ALS model composed of matrices augmented in the spectral mode, i.e. time profiles remain invariant while spectra may change from sample to sample. This approach allowed us to separate and determine the whole analytes at once. The developed approach enabled us to determine five of the most important metabolic disorder marker pteridines: biopterin, neopterin, isoxanthopterin, pterin and xanthopterin, three of them presenting emission spectra with the same emission wavelength maxima. In addition, some of these analytes present overlapped time profiles. As a consequence of using the entire data sets, a considerable reduction of the data processing experimental time can be achieved. Results are compared with a previous strategy in which data were split in five different regions, and information about the figures of merit of the new strategy compared with the previously reported strategy is reported.

Fetal and postnatal pulmonary circulation in the Alto Andino.

Lowland mammals at high altitude constrict the pulmonary vessels, augmenting vascular resistance and developing pulmonary arterial hypertension. In contrast, highland mammals, like the llama, do not present pulmonary arterial hypertension. Using wire myography, we studied the sensitivity to norepinephrine (NE) and NO of small pulmonary arteries of fetal llamas and sheep at high altitudes. The sensitivity of the contractile responses to NE was decreased whereas the relaxation sensitivity to NO was augmented in the llama fetus compared to the sheep fetus. Altogether these data show that the fetal llama has a lower sensitivity to a vasoconstrictor (NE) and a higher sensitivity to a vasodilator (NO), than the fetal sheep, consistent with a lower pulmonary arterial pressure found in the neonatal llama in the Andean altiplano. Additionally, we investigated carbon monoxide (CO) in the pulmonary circulation in lowland and highland newborn sheep and llamas. Pulmonary arterial pressure was augmented in neonatal sheep but not in llamas. These sheep had reduced soluble guanylate cyclase and heme oxygenase expression and CO production than at lowland. In contrast, neonatal llamas increased markedly pulmonary CO production and HO expression at high altitude. Thus, enhanced pulmonary CO protects against pulmonary hypertension in the highland neonate. Further, we compared pulmonary vascular responses to acute hypoxia in the adult llama versus the adult sheep. The rise in pulmonary arterial pressure was more marked in the sheep than in the llama. The llama pulmonary dilator strategy may provide insights into new treatments for pulmonary arterial hypertension of the neonate and adult.

Determination of marker pteridines in urine by HPLC with fluorimetric detection and second-order multivariate calibration using MCR-ALS.

A liquid chromatographic method has been developed, in combination with the multivariate curve resolution-alternating least squares algorithm (MCR-ALS), for the simultaneous determination of marker pteridines in urine samples. A central composite design has been applied to optimize the factors influencing the separation (buffer concentration, buffer pH, flow rate, oven temperature, mobile-phase composition). A set of 15 calibration samples were randomly prepared, in a concentration range of 0.5-10.5 ng mL(-1) for neopterin, biopterin, and pterin; 4.0-8.0 ng mL(-1) for xanthopterin; and 0.5-4.5 ng mL(-1) for isoxanthopterin. The validation was carried out with fortified urine samples from healthy adults. The optimized conditions were a mobile-phase composition of 10 mM citric buffer at pH 5.44 and acetonitrile (94.5/5.5, v/v), a flow rate of 1.0 mL min(-1), and an oven temperature of 25 °C. The detection system consisted of a fast-scanning spectrofluorimeter, which allows obtaining of second-order data matrices containing the fluorescence intensity as a function of retention time and emission wavelength. In this work, MCR-ALS was used to cope with coeluting interferences, on account of the second-order advantage inherent to this algorithm which, in addition, is able to handle data sets deviating from trilinearity, like the high-performance liquid chromatography data analyzed in the present report. The developed approach enabled us to determine five pteridines, some of them with overlapped profiles, reducing the experimental time and reagent consumption. Ratio values for pteridines/creatinine in urine, for infected children with different pathologies, are reported in this work.

Reliability of the E-test method for detection of colistin resistance in clinical isolates of Acinetobacter baumannii.

We compared the E-test to the broth microdilution method for testing the susceptibility of 115 clinical isolates of Acinetobacter baumannii to colistin. Twenty-two (19.1%) strains were resistant to colistin and 93 (80.8%) strains were susceptible according to the reference broth microdilution method. A categorical agreement of 98.2% was found, with only two (1.7%) very major errors. Agreement within 1 twofold dilution between the E-test and the broth microdilution was 16.5%. Complete agreement was found for the strains for which MICs fell within the range of 0.25 to 1 microg of colistin/ml. However, there was poor concordance, particularly in extreme dilutions with higher MICs by the E-test method.

American tegumentary leishmaniasis: antigen-gene polymorphism, taxonomy and clinical pleomorphism.

Multi-locus enzyme electrophoresis is the current gold standard for the genetic characterisation of Leishmania. However, this method is time-consuming and, more importantly, cannot be directly applied to parasites present in host tissue. PCR-based methods represent an ideal alternative but, to date, a multi-locus analysis has not been applied to the same sample. This has now been achieved with a sample of 55 neotropical isolates (Leishmania (Viannia) braziliensis, L. (V.) peruviana, L. (V.) guyanensis, L. (V.) lainsoni and L. (L.) amazonensis), using five different genes as targets, four of which encoded major Leishmania antigens (gp63, Hsp70, H2B and Cpb). Our multi-locus approach strongly supports the current taxonomy and demonstrates a highly robust method of distinguishing different strains. Within L. (V.) braziliensis, we did not encounter so far specific genetic differences between parasites isolated from cutaneous and mucosal lesions. Interestingly, results provided by each of the different antigen-genes in the species considered, were different, suggesting different selective pressures. Our work emphasises the need for a multi-disciplinary approach to study the clinical pleomorphism of leishmaniasis.

[Quantitative cultures from catheters, using the Brun Buisson technique]. / Cultivos cuantitativos de catéteres por la técnica de Brun Buisson.

The objective of this collaborative work carried out in the Fundación Favaloro and the Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia, was to determine optimal conditions for incubation (time and atmosphere) of quantitative cultures of catheters processed according to the technique of vortex agitation (Brun Buisson method). From 689 processed catheters, 551 yielded negative cultures. From the 138 positive cultures, 125 yielded monomicrobial cultures and 13 polimicrobial cultures (total number of microorganisms was 151). In the last situation each micoorganism was considered on an individual basis. A total of 58 episodes of catheter related bacteremias occurred, being 52 monomicrobial and 6 polimicrobial (total number of microorganisms was 64). When colony counts were compared in aerobic and in 5-10% CO2 atmospheres, a very good correlation was obtained (p = 0.27; r2 = 0.9268). No advantage was observed by incubating plates for more than 48 hours. Colony counts performed at the second versus the third day, and at the second day versus the seventh, gave very good correlation (p = 0.10 and r2 = 0.9996; p = 0.31 and r2 = 0.9995, respectively).

Cultivos cuantitativos de catéteres por la técnica de Brun Buisson / Quantitative cultures from catheters, using the Brun Buisson technique

The objective of this collaborative work carried out in the Fundación Favaloro and the Fundación para la Lucha contra las Enfermedades Neurológicas de la Infancia, was to determine optimal conditions for incubation (time and atmosphere) of quantitative cultures of catheters processed according to the technique of vortex agitation (Brun Buisson method). From 689 processed catheters, 551 yielded negative cultures. From the 138 positive cultures, 125 yielded monomicrobial cultures and 13 polimicrobial cultures (total number of microorganisms was 151). In the last situation each micoorganism was considered on an individual basis. A total of 58 episodes of catheter related bacteremias occurred, being 52 monomicrobial and 6 polimicrobial (total number of microorganisms was 64). When colony counts were compared in aerobic and in 5-10 CO2 atmospheres, a very good correlation was obtained (p = 0.27; r2 = 0.9268). No advantage was observed by incubating plates for more than 48 hours. Colony counts performed at the second versus the third day, and at the second day versus the seventh, gave very good correlation (p = 0.10 and r2 = 0.9996; p = 0.31 and r2 = 0.9995, respectively).

Low-dose inhaled carbon monoxide reduces pulmonary vascular resistance during acute hypoxemia in adult sheep.

Carbon monoxide (CO) is produced by the action of the heme oxygenase (HO) complex through the oxidation of heme. CO, like nitric oxide (NO), is a molecular gas that among other actions stimulates guanylyl cyclase and increases cGMP levels in smooth muscle cells, regulating the vascular tone. Acute hypoxia generates pulmonary hypertension and increases the expression of inducible HO isoform (HO-1) in the vascular endothelium. Inhaled NO causes a potent pulmonary vasodilation. We hypothesized that inhaled CO might produce similar actions as NO on pulmonary vascular resistance (PVR). To test our contention, we studied the effects of inhaled CO (40 ppm) in the augmented PVR observed during hypoxemia. Five chronically instrumented German Merino sheep were submitted to a protocol consisting of 20 min of normoxemia (N), 20 min of isocapnic hypoxemia (H20), 20 min of isocapnic hypoxemia plus CO 40 ppm (H40), and 20 min of recovery (R). In the control protocol, we did not administer inhaled CO. Arterial gases and pH, percentage of carboxyhemoglobin (COHb), systemic and pulmonary arterial pressure, systemic and pulmonary vascular resistance, and cardiac output were measured during each period. During H20 period, there was a significant increase in cardiac output and PVR in sheep submitted to both protocols. The sheep treated with inhaled CO (H40 + CO) showed a modest but significant decrease (16%) in the elevated PVR. Our data indicate that inhaled CO decreases pulmonary vascular resistance associated with acute hypoxemia in adult sheep.

Nitric oxide synthase activity in brain tissues from llama fetuses submitted to hypoxemia.

The fetal llama (Lama glama; a species adapted to live in chronic hypoxia in the highlands of the Andes) did not increase cerebral blood flow and reduce the brain oxygen uptake during hypoxemia. Although nitric oxide (NO) is a normal mediator in the regulation of vascular tone and synaptic transmission, NO overproduction by hypoxemia could produce neuronal damage. We hypothesized that nitric oxide synthase (NOS) activity is either maintained or reduced in the central nervous system of the llama fetuses submitted to chronic hypoxemia. Approximately 85% of the Ca(2+)-dependent NOS activity was soluble, at least 12% was associated with the mitochondrial fraction, and less than 5% remains associated with microsomes. To understand the role of NO in chronic hypoxemia, we determined the effect of 24-h hypoxemia on NOS activity in the central nervous system. No changes in activity or the subcellular distribution of NOS activity in brain tissues after hypoxemia were found. We proposed that the lack of changes in NOS activity in the llama under hypoxemia could be a cytoprotective mechanism inherent to the llama, against possible toxic effects of NO.

Insulin homeostasis in the extremely low birth weight infant.

The need to improve the nutritional status of extremely low birth weight infants has resulted in a higher incidence of problems related to glucose intolerance. The inability of the newborn to inhibit gluconeogenesis in response to a glucose infusion has been postulated as an important determinant of the hyperglycemia observed in extremely low birth weight infants. The 2 proposed mechanisms to explain this finding include inappropriate secretion of insulin by the pancreas and decrease sensitivity of the liver to the gluco-regulatory effect of insulin. The capacity of extremely low birth weight infants to oxidize glucose at higher rates, and the positive effect that insulin may have in glucose utilization and tolerance, support the use of insulin in the prevention and treatment of hyperglycemia. Continuous infusion of insulin appears to be safe for the treatment of hyperglycemia, based on the available studies. However, the effectiveness of insulin treatment needs to be critically tested further before it can be implemented in routine clinical practice.

[Intercultural adaptations at hospitals for Indians in New Spain]. / Adecuaciones interculturales en los hospitales para indios en la Nueva España.

Exclusive hospitals for attention to sick natives were founded in New Spain. The most important of these were established by Franciscan missionaries at the Valley of Mexico and by the Agustinians in Western Mexico. In addition, foundation of the hospitals towns at Michoacan's bishop, designed by Don Vasco de Quiroga, were based on the utopic and humanistic ideas of Thomas More. In this article we offer a revision of their origins, their multiple forms of economic support, the administrative operation, hospital organization with the extraordinary participation of the Indian natives the trust deposited in these institutions by the users, the peak and consolidation in the 16th and 17th centuries, as well as decadence and cancellation in the 19th century. Additionally, we carried out the review of the organization and operation with the goal of assessing the degree of realistic achievement in these institutions to render them more accessible to the appropriate social and cultural characteristics of the indigenous patients. The lessons of the past helped us not to similar mistakes again. In this vein, the knowledge and experience of those who preceded us can help us resolve actual problems.

Cardiovascular responses to arginine vasopressin blockade during acute hypoxemia in the llama fetus.

The fetal llama has a marked increase in the peripheral vascular resistance and no augmentation of brain blood flow during hypoxemia. In spite of the substantial plasma arginine-vasopressin (AVP) increase during hypoxemia, up to 8 times greater than in fetal sheep, there are no changes of carotid and femoral blood flows during hypoxemia with a V1 receptor blockade, as is seen in the fetal sheep. The aim of this study was to assess the role of AVP function in mediating the combined ventricular output and organ blood flow in the hypoxemic llama fetus. Six fetal llamas at 0.65 of gestation were instrumented under general anesthesia, and cardiorespiratory responses and blood flows determined under normoxemic and hypoxemic conditions. The AVP effect was determined using a V1 antagonist during normoxemic and hypoxemic conditions. Organ blood flows were measured with the radioactive microsphere technique. No significant differences in organ blood flow or in their vascular resistances were seen between the control and treated fetuses during hypoxemia. We conclude that V1 blockade did not have any important role in the cardiovascular response to acute hypoxemia in the llama fetus, in contrast with lowland fetuses. AVP may be playing a role in other regions, possibly in kidney or lung, during hypoxemia.

Adrenergic and vasopressinergic contributions to the cardiovascular response to acute hypoxaemia in the llama fetus.

1. The effects of fetal intravenous treatment with phentolamine or a vasopressinergic V1-receptor antagonist on the fetal cardiovascular responses to acute hypoxaemia in the llama were investigated. 2. Six llama fetuses were surgically prepared between 60 and 70 % of gestation under general halothane anaesthesia with vascular catheters and transit-time ultrasonic flow probes around a carotid artery and a femoral artery. At least 4 days after surgery all fetuses were subjected to a 3 h experiment: 1 h of normoxia, 1 h of hypoxaemia and 1 h of recovery while on slow i.v. infusion with saline. On separate days this experiment was repeated with fetal i.v. treatment with either phentolamine or a V1-receptor antagonist dissolved in saline. 3. During saline infusion all llama fetuses responded to acute hypoxaemia with intense femoral vasoconstriction. Phentolamine during normoxia produced hypotension, tachycardia and vasodilatation in both the carotid and the femoral circulations. During hypoxaemia, fetuses treated with phentolamine did not elicit the pronounced femoral vasoconstriction and all died within 20 min of the onset of hypoxaemia. A V1-receptor antagonist produced a femoral vasodilatation during normoxia but did not affect the fetal cardiovascular responses to acute hypoxaemia. 4. In conclusion, alpha-adrenergic and V1-vasopressinergic mechanisms contribute to a basal vasoconstrictor tone in the femoral circulation in the llama fetus. The enhanced femoral vasoconstriction during acute hypoxaemia in the llama fetus is not mediated by stimulation of V1-vasopressin receptors, but is dependent on alpha-adrenergic receptor stimulation. Such alpha-adrenergic efferent mechanisms are indispensable to fetal survival during hypoxaemia in the llama since their abolition leads to cardiovascular collapse and death.

Cardiorespiratory responses to acute hypoxemia in the chronically catheterized fetal llama at 0.7-0.9 of gestation.

The adult llama (Lama glama) has several compensatory mechanisms that allow it to successfully survive at high altitude. Llama fetuses at 0.6-0.7 of gestation, and near-term llama fetuses studied close to surgery, did not increase cerebral blood flow and decreased cerebral oxygen delivery during acute hypoxemia. It is not known whether these responses were the result of immaturity or surgical stress. The aim of this study was to determine whether the lack of increase in cerebral blood flow and the decrease in cerebral oxygen delivery during hypoxemia in the fetal llama is characteristic of this high-altitude species near term, and under nonstressed conditions. We chronically catheterized 7 llamas and their fetuses near to term, at 0.7-0.9 of gestation. Fetal cardiac output, cerebral and regional blood flows, systemic blood pressure, heart rate, pH, and blood gases, organ vascular resistances and organ oxygen deliveries were determined at least 4 days after surgery, both during the basal state and after 1 hr of acute fetal hypoxemia. During hypoxemia the llama fetus did not increase cerebral blood flow and markedly decreased its cerebral oxygen delivery. There was also a marked decrease in kidney blood flow and oxygen delivery. These results indicate that, in contrast to fetuses of lowland species, the fetal llama does not increase the cerebral blood flow during hypoxemia, suggesting specific cellular mechanisms to preserve brain integrity during oxygen limitation.