Isothermal Section of the V-Si-B System at 1600 A degrees C in the V-VSi(2)-VB Region

In recent years, the Me-Si-B (Me-metal) ternary systems have received considerable attention aiming at the development of high-temperature structural materials. Assuming that any real application of these materials will rely on multicomponent alloys, as is the case of Ni-base superalloys, phase equilibria data of these systems become very important. In this work, results are reported on phase equilibria in the V-Si-B system, and are summarized in the form of an isothermal section at 1600 A degrees C for the V-VSi(2)-VB region. Several alloys of different compositions were prepared via arc melting and then heat-treated at 1600 A degrees C under high vacuum. All the materials in both as-cast and heat-treated conditions were characterized through x-ray diffraction, scanning electron microscopy, and selected alloys via wavelength dispersive spectroscopy. A negligible solubility of B in the V(3)Si, V(5)Si(3) (T(1)), and V(6)Si(5) phases as well as of Si in V(3)B(2) and VB phases was noted. Two ternary phases presenting the structures known as T(2) (Cr(5)B(3)-prototype) and D8(8) (Mn(5)Si(3)-prototype) were observed in both as-cast and heat-treated samples. It is proposed that at 1600 A degrees C the homogeneity range of T(2) extends approximately from 5 at.% to 12 at.% Si at constant vanadium content and the composition of D8(8) phase is close to V(59.5)Si(33)B(7.5) (at.%).

Pharmacogenetics of OATP Transporters Reveals That SLCO1B1 c.388A > G Variant Is Determinant of Increased Atorvastatin Response

Aims: The relationship between variants in SLCO1B1 and SLCO2B1 genes and lipid-lowering response to atorvastatin was investigated. Material and Methods: One-hundred-thirty-six unrelated individuals with hypercholesterolemia were selected and treated with atorvastatin (10 mg/day/4 weeks). They were genotyped with a panel of ancestry informative markers for individual African component of ancestry (ACA) estimation by SNaPshot (R) and SLCO1B1 (c.388A>G, c.463C>A and c.521T>C) and SLCO2B1 (-71T>C) gene polymorphisms were identified by TaqMan (R) Real-time PCR. Results: Subjects carrying SLCO1B1 c.388GG genotype exhibited significantly high low-density lipoprotein (LDL) cholesterol reduction relative to c.388AA+c.388AG carriers (41 vs. 37%, p = 0.034). Haplotype analysis revealed that homozygous of SLCO1B1*15 (c.521C and c.388G) variant had similar response to statin relative to heterozygous and non-carriers. A multivariate logistic regression analysis confirmed that c.388GG genotype was associated with higher LDL cholesterol reduction in the study population (OR: 3.2, CI95%: 1.3-8.0, p < 0.05). Conclusion: SLCO1B1 c.388A>G polymorphism causes significant increase in atorvastatin response and may be an important marker for predicting efficacy of lipid-lowering therapy.

Adaptive Immunity against Leishmania Nucleoside Hydrolase Maps Its C-Terminal Domain as the Target of the CD4+ T Cell-Driven Protective Response

Nucleoside hydrolases (NHs) show homology among parasite protozoa, fungi and bacteria. They are vital protagonists in the establishment of early infection and, therefore, are excellent candidates for the pathogen recognition by adaptive immune responses. Immune protection against NHs would prevent disease at the early infection of several pathogens. We have identified the domain of the NH of L. donovani (NH36) responsible for its immunogenicity and protective efficacy against murine visceral leishmaniasis (VL). Using recombinant generated peptides covering the whole NH36 sequence and saponin we demonstrate that protection against L. chagasi is related to its C-terminal domain (amino-acids 199-314) and is mediated mainly by a CD4+ T cell driven response with a lower contribution of CD8+ T cells. Immunization with this peptide exceeds in 36.73 +/- 12.33% the protective response induced by the cognate NH36 protein. Increases in IgM, IgG2a, IgG1 and IgG2b antibodies, CD4+ T cell proportions, IFN-gamma secretion, ratios of IFN-gamma/IL-10 producing CD4+ and CD8+ T cells and percents of antibody binding inhibition by synthetic predicted epitopes were detected in F3 vaccinated mice. The increases in DTH and in ratios of TNF alpha/IL-10 CD4+ producing cells were however the strong correlates of protection which was confirmed by in vivo depletion with monoclonal antibodies, algorithm predicted CD4 and CD8 epitopes and a pronounced decrease in parasite load (90.5-88.23%; p = 0.011) that was long-lasting. No decrease in parasite load was detected after vaccination with the N-domain of NH36, in spite of the induction of IFN-gamma/IL-10 expression by CD4+ T cells after challenge. Both peptides reduced the size of footpad lesions, but only the C-domain reduced the parasite load of mice challenged with L. amazonensis. The identification of the target of the immune response to NH36 represents a basis for the rationale development of a bivalent vaccine against leishmaniasis and for multivalent vaccines against NHs-dependent pathogens.

Synthesis and total H-1- and C-13-NMR assignment of cephem derivatives for use in ADEPT approaches

We report the synthesis and total NMR characterization of 5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid-3-[[[(4''-nitrophenoxy)carbonyl]oxy]-methyl]-8-oxo-7[(2-thienyloxoacetyl)amino]-diphenylmethyl ester-5-dioxide (5), a new cephalosporin derivative. This compound can be used as the carrier of a wide range of drugs containing an amino group. The preparation of the intermediate product, 5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid-3-[methyl-4-(6-methoxyquinolin-8-ylamino) pentylcarbamate]-8-oxo-7-[(2-thienyloxoacetyl)amino]-diphenylmethyl ester-5-dioxide (6), as well as the synthesis of the antimalarial primaquine prodrug 5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid-3-[methyl-4-(6-methoxyquinolin-8-ylamino) pentylcarbamate]-8-oxo-7-[(2-thienyloxoacetyl)amino]-5-dioxide (7) are also described, together with their total H-1- and C-13-NMR assignments.

Chemometric Studies on Natural Products as Potential Inhibitors of the NADH Oxidase from Trypanosoma cruzi Using the VolSurf Approach

Natural products have widespread biological activities, including inhibition of mitochondrial enzyme systems. Some of these activities, for example cytotoxicity, may be the result of alteration of cellular bioenergetics. Based on previous computer-aided drug design (CADD) studies and considering reported data on structure-activity relationships (SAR), an assumption regarding the mechanism of action of natural products against parasitic infections involves the NADH-oxidase inhibition. In this study, chemometric tools, such as: Principal Component Analysis (PCA), Consensus PCA (CPCA), and partial least squares regression (PLS), were applied to a set of forty natural compounds, acting as NADH-oxidase inhibitors. The calculations were performed using the VolSurf+ program. The formalisms employed generated good exploratory and predictive results. The independent variables or descriptors having a hydrophobic profile were strongly correlated to the biological data.

Heterologous expression of glucose oxidase in the yeast Kluyveromyces marxianus

Background: In spite of its advantageous physiological properties for bioprocess applications, the use of the yeast Kluyveromyces marxianus as a host for heterologous protein production has been very limited, in constrast to its close relative Kluyveromyces lactis. In the present work, the model protein glucose oxidase (GOX) from Aspergillus niger was cloned into K. marxianus CBS 6556 and into K. lactis CBS 2359 using three different expression systems. We aimed at verifying how each expression system would affect protein expression, secretion/localization, post-translational modification, and biochemical properties. Results: The highest GOX expression levels (1552 units of secreted protein per gram dry cell weight) were achieved using an episomal system, in which the INU1 promoter and terminator were used to drive heterologous gene expression, together with the INU1 prepro sequence, which was employed to drive secretion of the enzyme. In all cases, GOX was mainly secreted, remaining either in the periplasmic space or in the culture supernatant. Whereas the use of genetic elements from Saccharomyces cerevisiae to drive heterologous protein expression led to higher expression levels in K. lactis than in K. marxianus, the use of INU1 genetic elements clearly led to the opposite result. The biochemical characterization of GOX confirmed the correct expression of the protein and showed that K. marxianus has a tendency to hyperglycosylate the protein, in a similar way as already observed for other yeasts, although this tendency seems to be smaller than the one of e. g. K. lactis and S. cerevisiae. Hyperglycosylation of GOX does not seem to affect its affinity for the substrate, nor its activity. Conclusions: Taken together, our results indicate that K. marxianus is indeed a good host for the expression of heterologous proteins, not only for its physiological properties, but also because it correctly secretes and folds these proteins.

Biochemical characterization of pulp of banana fruit: measurement of soluble sugars, organic acids, free ACC and in vitro ACC oxidase

Introduction. We present some protocols aiming at partially characterizing banana fruit quality through measurement of some key biochemical parameters. The principle, key advantages, starting plant material, time required and expected results are presented. Materials and methods. This part describes the required laboratory materials and the steps necessary for achieving four protocols making it possible to measure sugar, organic acids and free ACC contents, and in vitro ACC oxidase activity. Results. Standard results obtained by using the protocols described are presented in the figures.

Pluripotent Reprogramming of Fibroblasts by Lentiviralmediated Insertion of SOX2, C-MYC, and TCL-1A

Reprogramming of somatic cells to pluripotency promises to boost cellular therapy. Most instances of direct reprogramming have been achieved by forced expression of defined exogenous factors using multiple viral vectors. The most used 4 transcription factors, octamer-binding transcription factor 4 (OCT4), (sex determining region Y)-box 2 (SOX2), Kruppel-like factor 4 (KLF4), and v-myc myelocytomatosis viral oncogene homolog (C-MYC), can induce pluripotency in mouse and human fibroblasts. Here, we report that forced expression of a new combination of transcription factors (T-cell leukemia/lymphoma protein 1A [TCL-1A], C-MYC, and SOX2) is sufficient to promote the reprogramming of human fibroblasts into pluripotent cells. These 3-factor pluripotent cells are similar to human embryonic stem cells in morphology, in the ability to differentiate into cells of the 3 embryonic layers, and at the level of global gene expression. Induced pluripotent human cells generated by a combination of other factors will be of great help for the understanding of reprogramming pathways. This, in turn, will allow us to better control cell-fate and apply this knowledge to cell therapy.

Blood Meal-Derived Heme Decreases ROS Levels in the Midgut of Aedes aegypti and Allows Proliferation of Intestinal Microbiota

The presence of bacteria in the midgut of mosquitoes antagonizes infectious agents, such as Dengue and Plasmodium, acting as a negative factor in the vectorial competence of the mosquito. Therefore, knowledge of the molecular mechanisms involved in the control of midgut microbiota could help in the development of new tools to reduce transmission. We hypothesized that toxic reactive oxygen species (ROS) generated by epithelial cells control bacterial growth in the midgut of Aedes aegypti, the vector of Yellow fever and Dengue viruses. We show that ROS are continuously present in the midgut of sugar-fed (SF) mosquitoes and a blood-meal immediately decreased ROS through a mechanism involving heme-mediated activation of PKC. This event occurred in parallel with an expansion of gut bacteria. Treatment of sugar-fed mosquitoes with increased concentrations of heme led to a dose dependent decrease in ROS levels and a consequent increase in midgut endogenous bacteria. In addition, gene silencing of dual oxidase (Duox) reduced ROS levels and also increased gut flora. Using a model of bacterial oral infection in the gut, we show that the absence of ROS resulted in decreased mosquito resistance to infection, increased midgut epithelial damage, transcriptional modulation of immune-related genes and mortality. As heme is a pro-oxidant molecule released in large amounts upon hemoglobin degradation, oxidative killing of bacteria in the gut would represent a burden to the insect, thereby creating an extra oxidative challenge to the mosquito. We propose that a controlled decrease in ROS levels in the midgut of Aedes aegypti is an adaptation to compensate for the ingestion of heme.

Comparison of C-reactive protein and serum amyloid A protein in septic shock patients

Septic shock is a severe inflammatory state caused by an infectious agent. Our purpose was to investigate serum amyloid A (SAA) protein and C-reactive protein (CRP) as inflammatory markers of septic shock patients. Here we evaluate 29 patients in postoperative period, with septic shock, in a prospective study developed in a surgical intensive care unit. All eligible patients were monitored over a 7-day period by sequential organ failure assessment (SOFA) score, daily CRP, SAA, and lactate measurements. CRP and SAA strongly correlated up to the fifth day of observation but were not good predictors of mortality in septic shock. Copyright (C) 2008.

Prevalence, Incidence Density, and Genotype Distribution of GB Virus C Infection in a Cohort of Recently HIV-1-Infected Subjects in Sao Paulo, Brazil

Background: The results of previous studies elsewhere have indicated that GB virus C (GBV-C) infection is frequent in patients infected with the human immunodeficiency virus type 1 (HIV-1) due to similar transmission routes of both viruses. The aim of this study was to determine the prevalence, incidence density and genotypic characteristics of GBV-C in this population. Methodology/Principal Findings: The study population included 233 patients from a cohort primarily comprised of homosexual men recently infected with HIV-1 in Sao Paulo, Brazil. The presence of GBV-C RNA was determined in plasma samples by reverse transcriptase-nested polymerase chain reaction and quantified by real-time PCR. GBV-C genotypes were determined by direct sequencing. HIV viral load, CD4+ T lymphocyte and CD8+ T lymphocyte count were also tested in all patients. The overall prevalence of GBV-C infection was 0.23 (95% CI: 0.18 to 0.29) in the study group. There was no significant difference between patients with and without GBV-C infection and Glycoprotein E2 antibody presence regarding age, sex, HIV-1 viral load, CD4+ and CD8+ T cell counts and treatment with antiretroviral drugs. An inverse correlation was observed between GBV-C and HIV-1 loads at enrollment and after one year. Also, a positive but not significant correlation was observed between GBV-C load and CD4+ T lymphocyte. Phylogenetic analysis of the GBV-C isolates revealed the presence of genotype 1 and genotype 2, these sub classified into subtype 2a and 2b. Conclusion/Significance: GBV-C infection is common in recently HIV -1 infected patients in Sao Paulo, Brazil and the predominant genotype is 2b. This study provides the first report of the GBV-C prevalence at the time of diagnosis of HIV-1 and the incidence density of GBV-C infection in one year.

The C242T polymorphism of the p22-phox gene (CYBA) is associated with higher left ventricular mass in Brazilian hypertensive patients

Background: Reactive oxygen species have been implicated in the physiopathogenesis of hypertensive end-organ damage. This study investigated the impact of the C242T polymorphism of the p22-phox gene (CYBA) on left ventricular structure in Brazilian hypertensive subjects. Methods: We cross-sectionally evaluated 561 patients from 2 independent centers [Campinas (n = 441) and Vitoria (n = 120)] by clinical history, physical examination, anthropometry, analysis of metabolic and echocardiography parameters as well as p22-phox C242T polymorphism genotyping. In addition, NADPH-oxidase activity was quantified in peripheral mononuclear cells from a subgroup of Campinas sample. Results: Genotype frequencies in both samples were consistent with the Hardy-Weinberg equilibrium. Subjects with the T allele presented higher left ventricular mass/height(2.7) than those carrying the CC genotype in Campinas (76.8 +/- 1.6 vs 70.9 +/- 1.4 g/m(2.7); p = 0.009), and in Vitoria (45.6 +/- 1.9 vs 39.9 +/- 1.4 g/m(2.7); p = 0.023) samples. These results were confirmed by stepwise regression analyses adjusted for age, gender, blood pressure, metabolic variables and use of anti-hypertensive medications. In addition, increased NADPH-oxidase activity was detected in peripheral mononuclear cells from T allele carriers compared with CC genotype carriers (p = 0.03). Conclusions: The T allele of the p22-phox C242T polymorphism is associated with higher left ventricular mass/height(2.7) and increased NADPH-oxidase activity in Brazilian hypertensive patients. These data suggest that genetic variation within NADPH-oxidase components may modulate left ventricular remodeling in subjects with systemic hypertension.

Methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism and high plasma homocysteine in chronic hepatitis C (CHC) infected patients from the Northeast of Brazil

Background/Aim: Hyperhomocysteinemia due to Methylenetetrahydrofolate Reductase (MTHFR) gene, in particular the C677T (Ala222Val) polymorphism were recently associated to steatosis and fibrosis. We analyzed the frequency of MTHFR gene in a cross-sectional study of patients affected by Chronic Hepatitis C (CHC) from Northeast of Brazil. Method: One hundred seven-four untreated patients with CHC were genotyped for the C677T MTHFR. Genomic DNA was extracted from peripheral blood cells and the C677T MTHFR polymorphism was identified by PCR-RFLP. The homocysteine (Hcy) levels were determined by chemiluminescence method. All patients were negative for markers of Wilson's disease, hemochromatosis and autoimmune diseases and have current and past daily alcohol intake less than 100 g/week. Results: Among subjects infected with CHC genotype non-1 the frequency of MTHFR genotypes TT was 9.8% versus 4.4% genotype 1 (p = 0.01). Nevertheless, association was found between the MTHFR genotype TT x CT/CC polymorphism and the degree of steatosis and fibrosis in both hepatitis C genotype (p < 0.05). A significant difference was found on plasma Hcy levels in patients with steatosis regardless of HCV genotype (p = 0.03). Conclusion: Our results indicate that plasma Hcy levels is highly prevalent in subjects with chronic hepatits C with steatosis regardless of HCV genotype and vitamin deficiency. The presence of genotype TT of MTHFR C677T polymorphism was more common in CHC genotype non-1 infected patient regardless of histopathological classification and genotype TT+CT frequencies were significant in the presence of fibrosis grade 1+2 and of steatosis in CHC infected patients from the northeast of Brazil regardless of HCV genotype. The genetic susceptibility of MTHFR C677T polymorphism should be confirmed in a large population.

Prognostic factors for progression of liver structural lesions in chronic hepatitis C patients

AIM: To evaluate the epidemiological, clinical, laboratory and histological variables capable of predicting the progression of hepatic structural disturbances in chronic hepatitis C patients during the time interval between two liver biopsies. METHODS: Clinical charts of 112 chronic hepatitis C patients were retrospectively analyzed, whereas liver biopsies were revised. Immunohistochemical detection of interferon receptor was based on the Envision-Peroxidase System. RESULTS: In the multivariate analysis, the variables in the age at first biopsy, ALT levels, presence of lymphoid aggregates and siderosis were the determinants of the best model for predicting the severity of the disease. The direct progression rate of hepatic structural lesions was significantly higher in untreated patients, intermediate in treated non-responders and lower in treated responders to antiviral therapy (non-treated vs responders, 0.22 +/- 0.50 vs -0.15 +/- 0.46, P = 0.0053). Immuno-expression of interferon receptor is not a relevant factor. CONCLUSION: The best predictors of the progression of fibrosis are age at the first liver biopsy, extent of ALT elevation, inflammation at liver histology and hepatic siderosis. Antiviral treatment is effective in preventing the progression of liver structural lesions in chronic hepatitis C patients. (C) 2008 WJG. All rights reserved.

Distribution of hepatitis c virus (hcv) genotypes in patients with chronic infection from Rondonia, Brazil

Background: Hepatitis C virus (HCV) is an important human pathogen affecting around 3% of the human population. In Brazil, it is estimated that there are approximately 2 to 3 million HCV chronic carriers. There are few reports of HCV prevalence in Rondonia State (RO), but it was estimated in 9.7% from 1999 to 2005. The aim of this study was to characterize HCV genotypes in 58 chronic HCV infected patients from Porto Velho, Rondonia (RO), Brazil. Methods: A fragment of 380 bp of NS5B region was amplified by nested PCR for genotyping analysis. Viral sequences were characterized by phylogenetic analysis using reference sequences obtained from the GenBank (n = 173). Sequences were aligned using Muscle software and edited in the SE-AL software. Phylogenetic analyses were conducted using Bayesian Markov chain Monte Carlo simulation (MCMC) to obtain the MCC tree using BEAST v. 1.5.3. Results: From 58 anti-HCV positive samples, 22 were positive to the NS5B fragment and successfully sequenced. Genotype 1b was the most prevalent in this population (50%), followed by 1a (27.2%), 2b (13.6%) and 3a (9.0%). Conclusions: This study is the first report of HCV genotypes from Rondonia State and subtype 1b was found to be the most prevalent. This subtype is mostly found among people who have a previous history of blood transfusion but more detailed studies with a larger number of patients are necessary to understand the HCV dynamics in the population of Rondonia State, Brazil.