Pd-catalysed amidation of 2,6-dihalopurine nucleosides. Replacement of iodine at 0 ºC

Pd-catalysed reactions of 2-Cl, 2-Br and 2-I derivatives of a 6-chloropurine nucleoside with benzamide have been compared, using Pd2dba3, Xantphos and Cs2CO3 in toluene, between 20 and 80 °C. The reactivity order was 2-I > 2-Br > 6-Cl ≫ 2-Cl. The 2-I substituent could be replaced even at 0 °C, under conditions disclosed here for the first time. On the other hand, the replacement of the chlorine atom at position 2 (2-Cl) required 110 °C.

EEG reactivity to pain in comatose patients: Importance of the stimulus type.

INTRODUCTION: Electroencephalogram (EEG) background reactivity is a potentially interesting outcome predictor in comatose patients, especially after cardiac arrest, but recent studies report only fair interrater reliability. Furthermore, there are no definite guidelines for its testing. We therefore investigated the EEG effect of standardized noxious stimuli in comatose patients not reactive to auditory stimuli. METHODS: In this prospective study we applied a protocol using three different painful stimuli (bilateral nipple pinching, pinprick at the nose base, finger-nail compression on each side), grouped in three distinct clusters with an alternated sequence, during EEG recordings in comatose patients. We only analyzed recordings showing any reactivity to pain. Fisher and χ2 tests were used as needed to assess contingency tables. RESULTS: Of 42 studies, 12 did not show any background reactivity, 2 presented SIRPIDs, and 2 had massive artefacts; we thus analyzed 26 EEGs recorded in 17 patients (4 women, 24%). Nipple pinching more frequently induced a change in EEG background activity (p<0.001), with a sensitivity of 97.4% for reactivity. Neither the order of the stimuli in the cluster (p=0.723), nor the cluster order (p=0.901) influenced the results. CONCLUSION: In this pilot study, bilateral, synchronous nipple pinching seems to be the most efficient method to test nociceptive EEG reactivity in comatose patients. This approach may enhance interrater reliability, but deserves confirmation in larger cohorts.

Evaluation of a new serological test for the detection of anti-Coxiella and anti-Rickettsia antibodies.

Coxiella burnetii and members of the genus Rickettsia are obligate intracellular bacteria. Since cultivation of these organisms requires dedicated techniques, their diagnosis usually relies on serological or molecular biology methods. Immunofluorescence is considered the gold standard to detect antibody-reactivity towards these organisms. Here, we assessed the performance of a new automated epifluorescence immunoassay (InoDiag) to detect IgM and IgG against C. burnetii, Rickettsia typhi and Rickettsia conorii. Samples were tested with the InoDiag assay. A total of 213 sera were tested, of which 63 samples from Q fever, 20 from spotted fever rickettsiosis, 6 from murine typhus and 124 controls. InoDiag results were compared to micro-immunofluorescence. For acute Q fever, the sensitivity of phase 2 IgG was only of 30% with a cutoff of 1 arbitrary unit (AU). In patients with acute Q fever with positive IF IgM, sensitivity reached 83% with the same cutoff. Sensitivity for chronic Q fever was 100% whereas sensitivity for past Q fever was 65%. Sensitivity for spotted Mediterranean fever and murine typhus were 91% and 100%, respectively. Both assays exhibited a good specificity in control groups, ranging from 79% in sera from patients with unrelated diseases or EBV positivity to 100% in sera from healthy patients. In conclusion, the InoDiag assay exhibits an excellent performance for the diagnosis of chronic Q fever but a very low IgG sensitivity for acute Q fever likely due to low reactivity of phase 2 antigens present on the glass slide. This defect is partially compensated by the detection of IgM. Because it exhibits a good negative predictive value, the InoDiag assay is valuable to rule out a chronic Q fever. For the diagnosis of rickettsial diseases, the sensitivity of the InoDiag method is similar to conventional immunofluorescence.

AltitudeOmics : Resetting of Cerebrovascular CO2 Reactivity Following Acclimatization to High Altitude.

Previous studies reported enhanced cerebrovascular CO2 reactivity upon ascent to high altitude using linear models. However, there is evidence that this response may be sigmoidal in nature. Moreover, it was speculated that these changes at high altitude are mediated by alterations in acid-base buffering. Accordingly, we reanalyzed previously published data to assess middle cerebral blood flow velocity (MCAv) responses to modified rebreathing at sea level (SL), upon ascent (ALT1) and following 16 days of acclimatization (ALT16) to 5260 m in 21 lowlanders. Using sigmoid curve fitting of the MCAv responses to CO2, we found the amplitude (95 vs. 129%, SL vs. ALT1, 95% confidence intervals (CI) [77, 112], [111, 145], respectively, P = 0.024) and the slope of the sigmoid response (4.5 vs. 7.5%/mmHg, SL vs. ALT1, 95% CIs [3.1, 5.9], [6.0, 9.0], respectively, P = 0.026) to be enhanced at ALT1, which persisted with acclimatization at ALT16 (amplitude: 177, 95% CI [139, 215], P < 0.001; slope: 10.3%/mmHg, 95% CI [8.2, 12.5], P = 0.003) compared to SL. Meanwhile, the sigmoidal response midpoint was unchanged at ALT1 (SL: 36.5 mmHg; ALT1: 35.4 mmHg, 95% CIs [34.0, 39.0], [33.1, 37.7], respectively, P = 0.982), while it was reduced by ~7 mmHg at ALT16 (28.6 mmHg, 95% CI [26.4, 30.8], P = 0.001 vs. SL), indicating leftward shift of the cerebrovascular CO2 response to a lower arterial partial pressure of CO2 (PaCO2) following acclimatization to altitude. Sigmoid fitting revealed a leftward shift in the midpoint of the cerebrovascular response curve which could not be observed with linear fitting. These findings demonstrate that there is resetting of the cerebrovascular CO2 reactivity operating point to a lower PaCO2 following acclimatization to high altitude. This cerebrovascular resetting is likely the result of an altered acid-base buffer status resulting from prolonged exposure to the severe hypocapnia associated with ventilatory acclimatization to high altitude.

Direct Interrogation of Viral Peptides Presented by the Class I HLA of HIV-Infected T Cells

Identification of CD8+ cytotoxic T lymphocyte (CTL) epitopes has traditionally relied upon testing of overlapping peptide libraries for their reactivity with T cells in vitro. Here, we pursued deep ligand sequencing (DLS) as an alternative method of directly identifying those ligands that are epitopes presented to CTLs by the class I human leukocyte antigens (HLA) of infected cells. Soluble class I HLA-A*11:01 (sHLA) was gathered from HIV-1 NL4-3-infected human CD4+ SUP-T1 cells. HLA-A*11:01 harvested from infected cells was immunoaffinity purified and acid boiled to release heavy and light chains from peptide ligands that were then recovered by size-exclusion filtration. The ligands were first fractionated by high-pH high-pressure liquid chromatography and then subjected to separation by nano-liquid chromatography (nano-LC)–mass spectrometry (MS) at low pH. Approximately 10 million ions were selected for sequencing by tandem mass spectrometry (MS/MS). HLA-A*11:01 ligand sequences were determined with PEAKS software and confirmed by comparison to spectra generated from synthetic peptides. DLS identified 42 viral ligands presented by HLA-A*11:01, and 37 of these were previously undetected. These data demonstrate that (i) HIV-1 Gag and Nef are extensively sampled, (ii) ligand length variants are prevalent, particularly within Gag and Nef hot spots where ligand sequences overlap, (iii) noncanonical ligands are T cell reactive, and (iv) HIV-1 ligands are derived from de novo synthesis rather than endocytic sampling. Next-generation immunotherapies must factor these nascent HIV-1 ligand length variants and the finding that CTL-reactive epitopes may be absent during infection of CD4+ T cells into strategies designed to enhance T cell immunity.

Predictors of smoking craving during virtual reality exposure

Cue exposure treatment (CET) consists of controlled and repeated exposure to drugrelated stimuli in order to reduce cue-reactivity. Virtual reality (VR) has proved to be a promising tool for exposition. However, identifying the variables that can modulate the efficacy of this technique is essential for selecting the most appropriate exposure modality. The aim of this study was to determine the relation between several individual variables and self-reported craving in smokers exposed to VR environments. Fortysix smokers were exposed to seven complex virtual environments that reproduce typical situations in which people smoke. Self-reported craving was selected as the criterion variable and three types of variables were selected as the predictor variables: related to nicotine dependence, related to anxiety and impulsivity, and related to the sense of presence in the virtual environments. Sense of presence was the only predictor of self-reported craving in all the experimental virtual environments. Nicotine dependence variables added predictive power to the model only in the virtual breakfast at home. No relation was found between anxiety or impulsivity and self-reported craving. Virtual reality technology can be very helpful for improving CET for substance use disorders. However, the use of virtual environments would make sense only insofar as the sense of presence was high. Otherwise, the effectiveness of exposure might be affected. © 2012 by the Massachusetts Institute of Technology.

Singlet oxygen reactivity in water-rich solvent mixtures

The 3-methylindole (3MI) oxygenation sensitized by psoralen (PSO) has been investigated in 100%, 20% and 5% O2-saturated water/dioxane (H2O/Dx) mixtures. The lowering of the ¹O2* chemical rate when water (k chem∆3MI = 1.4 × 109 M-1 s-1) is replaced by deuterated water (k chem∆3MI = 1.9 × 108 M-1 s-1) suggests that hydrogen abstraction is involved in the rate determining step. A high dependence of the chemical rate constant on water concentration in H2O/Dx mixtures was found showing that water molecules are absolutely essential for the success of the 3MI substrate oxidation by ¹O2* in water-rich solvent mixtures.

Structural and reactivity analyses of 2-benzylamino-1,4-naphthoquinone by X-ray characterization, electrochemical measurements, and dft single-molecule calculations

This study represents an integrated approach towards understanding the electronic and structural aspects of 2-benzylamino-1,4-naphthalenedione, a representative 2-amino-napfthoquinone. To this end, theoretical calculations performed at the B3PW91/6-31+G(d) level of density functional theory, electrochemical and X-ray structural investigation were employed. Two intramolecular H-bonds and other two intermolecular H-bonds were observed, including non-classical interactions. Cyclic voltammogram (CV) and differential pulse voltammetry (DPV) show two pairs of peaks, being each one a monoelectronic process.

GLOBAL AND LOCAL REACTIVITY DESCRIPTORS FOR PICLORAM HERBICIDE: A THEORETICAL QUANTUM STUDY

In this work, we studied the reactivity of picloram in the aqueous phase at the B3LYP/6-311++G(2d,2p) and MP2/6-311++G(2d,2p) levels of theory through global and local reactivity descriptors. The results obtained at the MP2 level indicate that the cationic form of picloram exhibits the highest hardness while the anionic form is the most nucleophilic. From the Fukui function values, the most reactive site for electrophilic and free radical attacks are on the nitrogen in the pyridine ring. The more reactive sites for nucleophilic attacks are located on the nitrogen atom of the amide group and on the carbon atoms located at positions 2 and 3 in the pyridine ring.

Estudo da movimentação de plantas aquáticas imersas presentes no reservatório da UHE Eng. Souza Dias - Jupiá

O objetivo deste estudo foi desenvolver um método para determinação de pontos de reprodução e de suas respectivas importâncias na disseminação das espécies E. densa, E. najas e C. demersum no reservatório de Jupiá. Foram monitorados dez locais nos quais se encontrou registro de altas infestações dessas espécies. Dois desses locais situam-se no rio Paraná, nas lagoas denominadas de "Ferradura" e "Pernilongo"; os outros oito sítios estão localizados no rio Tietê, nas lagoas marginais "Testemunha", "Barrenta", "Vírgula" e "Flórida", e no leito do rio nos pontos "Acima da Ponte dos Barrageiros", "Abaixo da Ponte", "Baía ao Lado da Ponte" e em frente da "Praia de Itapura". Em cada um desses sítios de reprodução foram liberados e monitorados dez blocos de plantas aquáticas imersas por mês. Os blocos possuíam volume de 0,14 cm³ e receberam uma etiqueta plástica de identificação externa e uma bóia de cor laranja, com o objetivo de serem facilmente localizadas ou visualizadas à distância. Após a soltura, cada bloco foi georreferenciado por meio de um aparelho GPS, sendo o seu deslocamento avaliado a cada 15 dias. Os sítios "Lagoa Vírgula", "Lagoa Testemunha" e "Lagoa Barrenta" destacaram-se quanto ao número de blocos (30, 20 e 19 blocos, respectivamente), que percorreram distâncias superiores a 500 m e, conseqüentemente, saíram de seus respectivos locais de reprodução. A grande maioria dos blocos permaneceu dentro da "Lagoa Flórida"; somente 12 blocos saíram e alcançaram o leito do rio Tietê. O sítio denominado "Leito Acima da Ponte" foi o que mais se destacou dentre os três localizados no leito do rio Tietê, pois forneceu 18 blocos de plantas, enquanto os sítios "Abaixo da Ponte" e "Baía ao Lado da Ponte" forneceram 15 e 14 blocos, respectivamente. O sítio "Praia de Itapura" foi o que apresentou menor importância em relação ao fornecimento de plantas imersas no rio Tietê, sendo localizadas apenas 11 bóias com deslocamento superior a 500 m. Os sítios de reprodução localizados no rio Paraná também contribuíram para o fornecimento de plantas imersas ao reservatório de Jupiá; nove blocos saíram da "Lagoa Pernilongo" e apresentaram deslocamentos superiores a 500 m. Entretanto, os sítios "Lagoa Barrenta" e "Lagoa Ferradura" foram aqueles que mais contribuíram com a chegada de plantas na tomada de água da UHE Eng. Souza Dias, uma vez que foram recolhidos, respectivamente, seis e quatro blocos de plantas provenientes desses locais de dispersão. A metodologia utilizada mostrou-se eficaz quanto à avaliação da movimentação de plantas aquáticas imersas dentro do reservatório de Jupiá.

Effects of ouabain on vascular reactivity

Ouabain is an endogenous substance occurring in the plasma in the nanomolar range, that has been proposed to increase vascular resistance and induce hypertension. This substance acts on the a-subunit of Na+,K+-ATPase inhibiting the Na+-pump activity. In the vascular smooth muscle this effect leads to intracellular Na+ accumulation that reduces the activity of the Na+/Ca2+ exchanger and to an increased vascular tone. It was also suggested that circulating ouabain, even in the nanomolar range, sensitizes the vascular smooth muscle to vasopressor substances. We tested the latter hypothesis by studying the effects of ouabain in the micromolar and nanomolar range on phenylephrine (PE)-evoked pressor responses. The experiments were performed in normotensive and hypertensive rats in vivo, under anesthesia, and in perfused rat tail vascular beds. The results showed that ouabain pretreatment increased the vasopressor responses to PE in vitro and in vivo. This sensitization after ouabain treatment was also observed in hypertensive animals which presented an enhanced vasopressor response to PE in comparison to normotensive animals. It is suggested that ouabain at nanomolar concentrations can sensitize vascular smooth muscle to vasopressor stimuli possibly contributing to increased tone in hypertension

Reactivity of the isolated perfused rat tail vascular bed

Isolated segments of the perfused rat tail artery display a high basal tone when compared to other isolated arteries such as the mesenteric and are suitable for the assay of vasopressor agents. However, the perfusion of this artery in the entire tail has not yet been used for functional studies. The main purpose of the present study was to identify some aspects of the vascular reactivity of the rat tail vascular bed and validate this method to measure vascular reactivity. The tail severed from the body was perfused with Krebs solution containing different Ca2+ concentrations at different flow rates. Rats were anesthetized with sodium pentobarbital (65 mg/kg) and heparinized (500 U). The tail artery was dissected near the tail insertion, cannulated and perfused with Krebs solution plus 30 µM EDTA at 36oC and 2.5 ml/min and the procedures were started after equilibration of the perfusion pressure. In the first group a dose-response curve to phenylephrine (PE) (0.5, 1, 2 and 5 µg, bolus injection) was obtained at different flow rates (1.5, 2.5 and 3.5 ml/min). The mean perfusion pressure increased with flow as well as PE vasopressor responses. In a second group the flow was changed (1.5, 2, 2.5, 3 and 3.5 ml/min) at different Ca2+ concentrations (0.62, 1.25, 2.5 and 3.75 mM) in the Krebs solution. Increasing Ca2+ concentrations did not alter the flow-pressure relationship. In the third group a similar protocol was performed but the rat tail vascular bed was perfused with Krebs solution containing PE (0.1 µg/ml). There was an enhancement of the effect of PE with increasing external Ca2+ and flow. PE vasopressor responses increased after endothelial damage with air and CHAPS, suggesting an endothelial modulation of the tone of the rat tail vascular bed. These experiments validate the perfusion of the rat tail vascular bed as a method to investigate vascular reactivity

Reactivity of anti-thyroid antibodies to thyroglobulin tryptic fragments: comparison of autoimmune and non-autoimmune thyroid diseases

Studies concerning the antigenicity of thyroglobulin fragments allow the characterization of the epitopes but do not consider the role of heavier antigenic fragments that could result in vivo from the action of endoproteases. Here we assess the relative importance of the fragments obtained from thyroglobulin by limited proteolysis with trypsin and compare by immunoblotting their reactivity to serum from patients with autoimmune (Graves' disease and Hashimoto's thyroiditis) and non-autoimmune (subacute thyroiditis) disease. The results showed no difference in frequency of recognition of any peptide by sera from patients with autoimmune thyroiditis. In contrast, sera from patients with subacute thyroiditis reacted more frequently with a peptide of 80 kDa. These results suggest the presence of antibody subpopulations directed at fragments produced in vivo by enzymatic cleavage of thyroglobulin. This fragment and antibodies to it may represent markers for subacute thyroiditis.