Solvatochromism in urea/water and urea-derivative/water solutions.

This work reports the experimental measurements of solvent acidity (SA), basicity (SB), and solvent dipolarity and polarizability (SPP) for water solutions with urea (U) and its molecular derivatives, monomethyl-urea (MU), 1,3-dimethyl-urea (DMU) and tetramethyl-urea (TMU). These solvatochromic parameters are applied to understanding the variation of indexes of refraction and densities and other physico-chemical properties reported for these solutions. These properties are well correlated to the SA, SB, and SPP solvent parameters of these solutions. As a result, from the characterization of the physico-chemical properties, one can infer that urea and its molecular derivatives are mainly modifiers in the structure of liquid water. The solvatochromic parameters indicate the possible existence of different mechanisms in the denaturation process of proteins in these urea/water solutions.

Sevoflurane for Short Painful Procedures in the Neonatal Intensive Care Unit.

OBJECTIVE: The aim of this study is to evaluate the use and safety of a sedation protocol with sevoflurane for short painful procedures in newborns. STUDY DESIGN: This was a prospective and observational study conducted in a tertiary neonatal intensive care unit. Sevoflurane was recommended in patients undergoing an invasive procedure of short length, especially in those with spontaneous breathing or without venous access. Its safety and efficacy was assessed by continuous monitoring of respiratory and hemodynamic variables and clinical data recording. RESULTS: Sevoflurane was used for 39 procedures, the main indications were: intravitreal bevacizumab injection (12), central venous catheterization (11), and biopsy (6). The median administration length was 14 minutes (range: 5-65 minutes). The median minimum dose was 1.5% (range: 1-3%). The median maximum dose was 2.5% (range: 1-6%). An effective control of nociceptive manifestations was achieved in 35 cases (90%). No major adverse effects were noticed. Main adverse effects were hypotension (8), desaturation (4), and apnea (3). All of them were solved by decreasing (14) or discontinuing (1) the administration of sevoflurane. CONCLUSION: Sevoflurane is relatively easy to use and provides an optimal control of pain-related symptoms. Its prescription should be individualized and more long-term follow-up data are needed.

Underdosing of Surfactant for Preterm Babies with Respiratory Distress Syndrome in Clinical Practice: A Retrospective Cohort Study.

OBJECTIVE: To evaluate the initial doses of surfactant administered to preterm infants with respiratory distress syndrome. STUDY DESIGN: This is a retrospective cohort study of 206 preterm infants admitted in four level III neonatal intensive care units of acute tertiary care hospitals in Spain between 2013 and 2015. RESULTS: The mean initial dose of surfactant was 173.9 (37.3) mg/kg, and 47.5% of infants received a dose of 200 mg/kg ± 10% (180-220 mg/kg), 47% less than 180 mg/kg (-10%), and 5.4% more than 220 mg/kg (+10%). Very preterm infants (<28 weeks) received higher initial doses than more mature infants, but in all cases, the mean doses were below the recommended 200 mg/kg (by 9.2% in gestational age 23-28 weeks, by 15.9% in 29-32 weeks, and by 24.3% in >32 weeks). CONCLUSION: Administration of surfactant below the prescribed dose is a frequent error in clinical practice. Inadvertently rounding down doses seems a plausible explanation.

Pooling/bootstrap-based GWAS (pbGWAS) identifies new loci modifying the age of onset in PSEN1 p.Glu280Ala Alzheimer's disease.

The literature on GWAS (genome-wide association studies) data suggests that very large sample sizes (for example, 50,000 cases and 50,000 controls) may be required to detect significant associations of genomic regions for complex disorders such as Alzheimer's disease (AD). Because of the challenges of obtaining such large cohorts, we describe here a novel sequential strategy that combines pooling of DNA and bootstrapping (pbGWAS) in order to significantly increase the statistical power and exponentially reduce expenses. We applied this method to a very homogeneous sample of patients belonging to a unique and clinically well-characterized multigenerational pedigree with one of the most severe forms of early onset AD, carrying the PSEN1 p.Glu280Ala mutation (often referred to as E280A mutation), which originated as a consequence of a founder effect. In this cohort, we identified novel loci genome-wide significantly associated as modifiers of the age of onset of AD (CD44, rs187116, P=1.29 × 10⁻¹²; NPHP1, rs10173717, P=1.74 × 10⁻¹²; CADPS2, rs3757536, P=1.54 × 10⁻¹°; GREM2, rs12129547, P=1.69 × 10⁻¹³, among others) as well as other loci known to be associated with AD. Regions identified by pbGWAS were confirmed by subsequent individual genotyping. The pbGWAS methodology and the genes it targeted could provide important insights in determining the genetic causes of AD and other complex conditions.

Demonstration of event position reconstruction based on diffusion in the NEXT-white detector.

Noble element time projection chambers are a leading technology for rare event detection in physics, such as for dark matter and neutrinoless double beta decay searches. Time projection chambers typically assign event position in the drift direction using the relative timing of prompt scintillation and delayed charge collection signals, allowing for reconstruction of an absolute position in the drift direction. In this paper, alternate methods for assigning event drift distance via quantification of electron diffusion in a pure high pressure xenon gas time projection chamber are explored. Data from the NEXT-White detector demonstrate the ability to achieve good position assignment accuracy for both high- and low-energy events. Using point-like energy deposits from 83mKr calibration electron captures (E∼45 keV), the position of origin of low-energy events is determined to 2 cm precision with bias <1mm. A convolutional neural network approach is then used to quantify diffusion for longer tracks (E≥1.5 MeV), from radiogenic electrons, yielding a precision of 3 cm on the event barycenter. The precision achieved with these methods indicates the feasibility energy calibrations of better than 1% FWHM at Qßß in pure xenon, as well as the potential for event fiducialization in large future detectors using an alternate method that does not rely on primary scintillation.

Genetic predisposition to fetal and neonatal cancer.

BACKGROUND: Neonatal tumors represent an extremely rare and heterogeneous disease with an unknown etiology. Due to its early onset, it has been proposed that genetic factors could play a critical role; however, germline genetic analysis is not usually performed in neonatal cancer patients PATIENTS AND METHODS: To improve the identification of cancer genetic predisposition syndromes, we retrospectively review clinical characteristics in 45 patients with confirmed tumor diagnosis before 28 days of age, and we carried out germline genetic analysis in 20 patients using next-generation sequencing and directed sequencing. RESULTS: The genetic studies did not find any germline mutation except patients diagnosed with bilateral retinoblastoma who harbored RB1 germline mutations. CONCLUSIONS: Our results suggest that genetic factors have almost no higher impact in most neonatal tumors. However, since the heterogeneity of the tumors and the small sample size analyzed, we recommend complementary and centralized germline studies to discard the early onset as an additional criterion to take into account to improve the identification of cancer genetic predisposition syndromes in neonates.

Standardized Hypericum perforatum reduces oxidative stress and increases gene expression of antioxidant enzymes on rotenone-exposed rats.

Since oxidative stress is implicated in the pathophysiology of dementia and depression, this study was designed to investigate the pro-oxidant activity of rotenone, the protective role of standardized extract of Hypericum perforatum (SHP), as well as the mRNA levels of antioxidant enzymes, in brain homogenates of rats following exposure to rotenone and SHP extract. Quercetin in liposomes, one active constituent, was tested in the same experimental conditions to serve as a positive control. The animals received pretreatment with SHP (4 mg/kg) or quercetin liposomes (25 and 100 mg/kg) 60 min before of rotenone injection (2 mg/kg). All treatments were given intraperitoneally in a volume of 0.5 ml/kg body weight, for 45 days. Rotenone treatment increased activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and levels of malondialdehyde (MDA). The content of reduced glutathione (GSH) was decreased due to chronic rotenone treatment. Rotenone significantly induced the gene expression of CuZnSOD, MnSOD; CAT and GPx in brain. In contrast, SHP extract exerted an antioxidant action which was related with a decreased of MnSOD activity and mRNA levels of some antioxidant enzymes evaluated. Liposomal quercetin treatment resulted in a significant preservation of the activities of antioxidant enzymes and a decreased in the mRNA levels of these antioxidant enzymes. One possible mechanism of action of SHP extract may be related to quercetin in protecting neurons from oxidative damage. Therefore standardized extract of H. perforatum could be a better alternative for depressed elderly patients with degenerative disorder exhibiting elevated oxidative stress status.

Complicaciones obstétricas relacionadas con la terapia antirretroviral en gestantes infectadas por el VIH / Obstetric complications related to antiretroviral therapy in pregnant women with HIV infection

IntroducciónEl porcentaje de gestantes infectadas por VIH que demandan embarazo ha aumentado en los países desarrollados debido a la estabilidad de la infección y la disminución de la transmisión vertical por los tratamientos antirretrovirales (TAR) y las medidas preventivas. Sin embargo, existe poca información respecto al efecto del TAR sobre el embarazo.MétodosEstudio retrospectivo de las gestantes infectadas por VIH con TAR controladas en el Hospital La Paz entre los años 2000-2017. Se estudiaron las complicaciones maternofetales.ResultadosSe recogieron 141 gestaciones en 112 mujeres infectadas por VIH. El TAR más utilizado fue la combinación de 2 inhibidores de la transcriptasa inversa análogos nucleosídicos+1 inhibidor de la proteasa (58,1%), con diferencias significativas entre los distintos tratamientos en cuanto a la carga viral plasmática, siendo indetectable en mayor medida con 2 inhibidores de la transcriptasa inversa análogos nucleosídicos +1 inhibidor de la transcriptasa inversa no nucleosídico, segunda pauta más utilizada. Las tasas de neonatos a término de bajo peso (<2.500g) (11,3%), partos pretérmino (11,1%) y rotura prematura de membranas pretérmino (5,6%) fueron mayores que en la población general. Aunque no hubo asociación, estas complicaciones fueron más frecuentes en gestantes con un inhibidor de la proteasa/ritonavir. No se relacionaron con la carga viral plasmática. No se encontró aumento en la tasa de interrupciones gestacionales, malformaciones ni diabetes gestacional.ConclusiónEn las gestantes infectadas por VIH con TAR está aumentada la tasa de neonatos a término de bajo peso, prematuridad y rotura prematura de membranas pretérmino en comparación con la población general, relacionándose especialmente con el tratamiento con un inhibidor de la proteasa, aunque sin asociación significativa.

IntroductionThe percentage of HIV-infected pregnant women seeking pregnancy has increased in developed countries due to the stability of the infection and the decrease in vertical transmission due to antiretroviral treatment (ART) and preventive measures. However, there is little information regarding the effect of ART on pregnancy.MethodsRetrospective study of HIV-infected pregnant women on ART monitored at Hospital La Paz between 2000-2017. Maternal-foetal complications were studied.ResultsOne hundred and forty-one gestations were collected in 112 HIV-infected women. The most commonly used ART was the combination of 2 nucleoside reverse transcriptase inhibitor analogues+1 protease inhibitor (58.1%), with significant differences between the different treatments in terms of plasma viral load being undetectable to a greater extent with 2 nucleoside reverse transcriptase inhibitor analogues+1 non-nucleoside reverse transcriptase inhibitor, the second most used regimen. The rates of low birth weight (<2,500g) term neonates (11.3%), preterm delivery (11.1%) and preterm premature rupture of membranes (5.6%) were higher than in the general population. Although there was no association, these complications were more frequent in pregnant women with a protease inhibitor/ritonavir. They were not related to plasma viral load. No increase in the rate of gestational terminations, malformations or gestational diabetes was found.ConclusionHIV-infected pregnant women on ART have an increased rate of low birth weight, prematurity, and preterm premature rupture of membranes at term compared to the general population, especially related to treatment with protease inhibitor, although without significant association.

The mechanism of catabolite inhibition of invertase by glucose in Saccharomyces cerevisiae.

Saccharomyces cerevisiae -136ts synthesized invertase in media containing maltose and sucrose. In the presence of glucose synthesis of enzyme took place when the sugar concentration was lower than 1%. At higher concentrations enzyme formation was repressed. Analysis of the glucose effect before RNA inhibition showed that the hexose interfered with the transcription of DNA into invertase messenger RNA. Translation of invertase messenger already formed was also inhibited and the kinetics of this effect was similar to that produced by cycloheximide. Invertase activity was independent of glucose suggesting that the hexose produces no catabolite inhibition of invertase activity. Inhibition of invertase translation by glucose turned out to be reversible but the amount of enzyme produced was dependent on duration of treatment. It is suggested that the catabolite repression of invertase synthesis produced by glucose operates at the levels of transcription and translation and produces an increase in the rate of mRNA degradation. The catabolite repression has no effect on secretion and does not interfere with the catalytic activity of invertase.

Effect of papulacandin B and calcofluor white on the incorporation of mannoproteins in the wall of Candida albicans blastospores.

Incorporation of mannoproteins into the walls of Candida albicans blastospores (yeast phase) was followed by continuous labelling and pulse-chase experiments. The effect in the process of compounds that interfere with synthesis (papulacandin B) or assembly (calcofluor white) of structural polymers was also assessed. Mannoproteins which are kept in place by non-covalent bonds (mainly hydrogen bonds) were incorporated rapidly after their release into the periplasmic space, this process being blocked by calcofluor white. The stain had no effect on the incorporation of covalently linked mannoproteins. Papulacandin B inhibited formation of beta-glucans and incorporation of covalently linked mannoprotein molecules, whereas incorporation of hydrogen-bonded species took place normally. The results suggest that the formation of the non-covalent bonds between the mannoproteins occurs once they are secreted into the periplasmic space, whereas the formation of covalent connections between mannoproteins and wall glucan takes place at the level of the plasma membrane.

Tunicamycin and papulacandin B inhibit incorporation of specific mannoproteins into the wall of Candida albicans regenerating protoplasts.

Regeneration of Candida albicans protoplasts began with the formation of a chitin network which was complemented after a lag of about 60 min by the deposition of beta-glucan. Proteins were incorporated early to the growing structure, beginning with the mannoproteins which are kept in place by non-covalent bonds. Incorporation of covalently linked mannoproteins took place only after deposition of glucan. The incorporation of these mannoproteins did not occur when protoplasts were incubated with papulacandin B which inhibited glucan formation, or with tunicamycin which blocked N-glycosylation of mannoproteins. In the presence of papulacandin B, large amounts of native mannoproteins accumulated in the medium. However, in the presence of tunicamycin, the large mannoprotein material found was of smaller apparent molecular weight, suggesting that it was deficient in glycosylation. Partially regenerated walls were able to incorporate 'in vitro' non-covalently bound mannoproteins, indicating that some components of very large cellular structures such as walls are capable of being articulated by a self-assembly process.

Metabolic relationship between invertase and acid phosphatase isoenzymes in Saccharomyces cerevisiae.

Repressed cells of Saccharomyces cerevisiae, subjected to inhibition of both RNA and protein synthesis, showed a pattern of membrane-bound and cytosol acid phosphatase to the external enzyme which seemed to be linked through a precursor-product relationship. Gel exclusion chromatography did not indicate clear differences between the isoenzymes. Moreover, centrifugation experiments in CsCl and precipitation with concanavalin A suggested that there were no acid phosphatase molecules devoid of carbohydrate. Membrane-bound invertase displayed a molecular weight and a carbohydrate to protein ratio smaller than those of the exocellular enzyme. The values of molecular weight and buoyant density of the membrane-bound enzyme were closer to those found for the cytosol invertase. The stability of the level of the soluble invertase detected in the cytoplasm under derepression conditions, or after RNA or protein synthesis inhibition was found to be only apparent and represented the result of an equilibrium between synthesis and degradation.

Characterization of 5-fluorouracil loaded liposomes prepared by reverse-phase evaporation or freezing-thawing extrusion methods: study of drug release.

Entrapment of the anti-tumoral drug 5-fluorouracil (5-FU) in unilamellar liposomes prepared by freeze-thawing extrusion technique (FATVET) and the reverse-phase evaporation method (REV) from natural (bovine brain) sphingomyelin (SM) and synthetic distearoylphosphatidylcholine (DSPC) phospholipids was studied. Reverse-phase evaporation vesicles obtained from DSPC sized through polycarbonate membranes of 0.2 micron pore size were found to entrap roughly double amounts of drug than did extruded liposomes (0.1 micron pore size); however, s-REV in these preparations were more heterogenous in vesicle size than FATVET. The rate of in vitro drug release from the liposomes was found to be dependent of the bilayer composition and the method used to prepare the vesicles. The permeability coefficient P obtained was approx. 10(-11) m/s. The results suggest that 5-FU release is kinetically controlled by an interfacial process seemingly dependent on the surface activity of the drug. Also, the physical state of the bilayer determines the retention capacity of the vesicles. Thus, liposomes consisting of distearoylphosphatidylcholine whose acyl chains were in a gel state at the working temperature (37 degrees C) retained 70% of encapsulated 5-FU after 1 h, whereas liposomes composed of natural bovine brain sphingomyelin retained only 15% over the same period.

Invertase messenger ribonucleic acid in Saccharomyces cerevisiae. Kinetics of formation and decay.

Saccharomyces cerevisiae -136ts (Hutchison, H.T., Hartwell, L.H. and McLaughlin, C.S. (1969) J. Bacteriol. 99, 807-814) incubated in the presence of maltose at 23 degrees C (permissive temperature) synthesized the RNA messengers which codify derepressed invertase (an external mannoprotein) and induced alpha-glucosidase (a non-glycosylated internal enzyme). The enzymes were not synthesized if the mutant was transferred to the maltose-containing medium at the moment of incubation at 37 degrees C indicating that the cells had no pools of the specific RNA messengers and that transcription of the DNA was a prerequisite to enzyme synthesis. Cycloheximide inhibited syntheses of the enzymes both at 37 and at 23 degrees C suggesting that the enzymic activities were the result of "de novo" synthesis of the proteins and did not result from the activation of proenzymes. In derepressed cells the number of invertase mRNA molecules is probably larger than that actually being translated. The half-life of the derepressed invertase mRNA was calculated from the moment that the molecules of RNA messenger were limiting the enzyme synthesis and a value of 30-35 min was estimated. The value found for the basal (repression independent) invertase mRNA was of 45-50 min. The half-life of alpha-glucosidase mRNA was computed following the mathematical procedure described in the Appendix, and a value of 23 min was obtained. These results are consistent with the existence of relatively long-lived RNA messengers involved in the synthesis of extracellular macromolecules.

Cell wall synthesis regulation in Saccharomyces cerevisiae. Effect of RNA and protein inhibition.

In this investigation the regulation of wall formation in Saccharomyces cerevisiae ts-136 (Hutchison, H.T., Hartwell, L.H. and McLaughlin, C.S. (1969) J. Bacteriol. 99, 807-814) was analyzed by following the inhibition of RNA and protein synthesis. Lomofungin, thiolutin and 8-hydroxyquinoline at the concentrations needed to inhibit RNA synthesis also produced inhibition of glucan and mannan synthetases. The synthesis of RNA was also blocked in S. cerevisiae ts-136 by incubation at the non-permissive temperature (37 degrees C). Mannan formation decreased steadily but glucan synthesis remained after 4 to 5 h. After a few minutes of blocking protein synthesis with cycloheximide mannan synthesis was also blocked whereas glucan formation was unaffected by the presence of the drug. These results suggest a high degree of stability for glucan synthetases. S. cerevisiae ts-136 after 2 h of incubation at the non-permissive temperature (37 degrees C) showed a preferential formation of wall materials (mannan and glucan) indicating that the RNA messengers which codify wall mannan peptides have a slower decay rate than those of the cytoplasmic proteins. The data presented indicate that the existence of stable glucan synthetases and RNA messengers of the wall mannan peptides of slow decay rate results in the continuous synthesis of glucans and mannoproteins of the yeast wall throughout the cell cycle.

Regulation of alpha-galactosidase synthesis in Saccharomyces cerevisiae and effect of cerulenin on the secretion of this enzyme.

Saccharomyces cerevisiae -136ts MEL10 (a thermosensitive mutant whose RNA synthesis is inhibited at 37 degrees C but is normal at 23 degrees C), when grown at 23 degrees C in the presence of galactose, melibiose or L-arabinose, these cells synthesize alpha-galactosidase mRNA. In the simultaneous presence of both galactose and glucose the transcription of alpha-galactosidase mRNA is blocked. Glucose also interferes with mRNA translation, but the degree of inhibition depends on concentration and time of addition of the hexose to induced cells. It has been found that the final concentration of alpha-galactosidase produced by induced cells when transferred at the non-permissive temperature (37 degrees C) is inversely proportional to the incubation time in glucose. Cerulenin inhibits lipid formation on growing yeasts, but protein synthesis and selective permeability are not affected. The antibiotic partially inhibits secretion of alpha-galactosidase with a parallel accumulation of this enzyme in membranous structures, specially at the level of the plasma membrane. Induction of alpha-galactosidase or cerulenin addition to growing cells, results in changes in the polypeptide composition of the plasma membrane.

Regulation of acid phosphatase synthesis in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae-136ts (Hutchison, H.T., Hartwell, L.H. and McLaughlin, C.S. (1969) J. Bacteriol. 99, 807--814) derepressed acid phosphatase was almost exclusively located outside the permeability barrier. Only a minor part of the activity was associated with the protoplasts; about half of it (48%) in the soluble fraction, the rest bound to the internal (45%) and plasma (7%) membranes. The activity found in the membranes of derepressed cells decreased by 30--40% after addition of inorganic phosphate or cycloheximide suggesting that this activity is the precursor of the external enzyme. The alkaline phosphatase activity level could not be modified by changes in the concentration of inorganic phosphate. Acid phosphatase was not synthesized if the cells were transferred to a low phosphate medium at the moment of incubation at 37 degrees C or in the presence of cycloheximide at 23 degrees C. The data suggested that enzyme formation is the result of the transcription and translation of a specific gene(s) and not the activation of a proenzyme. Inorganic phosphate did not inhibit the translation of mRNA though it may act at the level of the transcription.

Critical steps in fungal cell wall synthesis: strategies for their inhibition.

Development of new effective antifungal drugs is limited by the absence of specific target sites in the fungal cells. Knowledge of the fungal cell wall structure and biosynthesis is of interest in searching for a potential target site for new chemotherapeutic agents. Our group has demonstrated that the fungal cell wall is a metabolically active structure where interaction between distinct components occurs to give rise to the mature cell wall structure. Mannoproteins play an essential role in the cell wall organization, and there is evidence for the formation of covalent bonds between these molecules and the structural polymers (glucans and chitin) outside the plasma membrane. Such interactions, which specifically occur at the fungal cell wall, are of great interest in defining target sites for potential new chemotherapeutic agents, which may inhibit the interactions and, thus, lead to a defective cell wall formation and cell death.

Cloning of a DNA fragment encoding part of a 70-kDa heat shock protein of Candida albicans.

Immunoscreening of a mycelial expression library with polyclonal antibodies raised against mycelial cell wall resulted in the detection of a cDNA encoding a heat shock protein of Candida albicans. Sequence analysis of a 0.8-kb cDNA subclone, 2M-1, revealed an open reading frame encoding 244 amino acids. Southern blot analysis with this fragment as a probe demonstrated hybridization to C. albicans DNA. Northern analysis showed a substantial increase in 2M RNA expression levels after cells were subjected to heat shock. Western blot analysis with 2M monospecific antibodies recognized a 70-kDa protein which was present in membrane particles and cytosolic fractions.