Toxicity of synthetic and natural compounds on Tetranychus urticae and the predator Phytoseiulus macropilis

The objective of this work was to evaluate the toxicity of synthetic and natural compounds on Tetranychus urticae and the predator Phytoseiulus macropilis. Mortality and growth rates of T. urticae and its predator were evaluated after applications of: abamectin, clofentezine, fenpropathrin, fenpyroximate, propargite, sulfur and spiromesifen, at their recommended concentrations; neem oils (Natuneem and Sempre Verde Killer Neem at 1%); and aqueous extracts at 10% of Dieffenbachia brasiliensis, Annona squamosa, Ruta graveolens, Agave angustifolia, Melia azedarach, Sonchus oleraceus, Mentha spicata x M. suaveolens, Allium cepa, Laurus nobilis, and Eucalyptus saligna. The acute toxicity and the influence of the compounds on the instantaneous growth rate of the mites were carried out in laboratory. Extracts of A. cepa, A. angustifolia, neem oil-based products, spiromesifen, propargite, fenpyroximate, abamectin and fenpropathrin caused mortality higher than 83% on T. urticae. Extract of A. angustifolia, Natuneem and clofentezine did not cause significant mortality rates on P. macropilis. Agave angustifolia and Natuneem did not affect significantly the growth rate of this predator. Propargite, fenpyroximate, abamectin, fenpropathrin, spiromesifen and extract of L. nobilis severely affected P. macropilis population.

A comparative thermoluminescence and electron spin resonance study of synthetic carbonated A-type hydroxyapatite

Intensity of the 150 degrees C thermoluminescence peak of beta-irradiated carbonated synthetic A-type hydroxyapatite is approximately 12 times higher than that of the noncarbonated material. Deconvolution of the glow curve showed that this peak is a result of a trap distribution. An attempt was made to relate this thermoluminescence peak enhanced by carbonation with the ESR signal of the CO2- radical in natural or synthetic hydroxyapatite. (C) 2011 Elsevier Ltd. All rights reserved.

Identification of intracellular peptides in rat adipose tissue: Insights into insulin resistance

Intracellular peptides generated by the proteasome and oligopeptidases have been suggested to function in signal transduction and to improve insulin resistance in mice fed a high-caloric diet. The aim of this study was to identify specific intracellular peptides in the adipose tissue of Wistar rats that could be associated with the physiological and therapeutic control of glucose uptake. Using semiquantitative mass spectrometry and LC/MS/MS analyses, we identified ten peptides in the epididymal adipose tissue of the Wistar rats; three of these peptides were present at increased levels in rats that were fed a high-caloric Western diet (WD) compared with rats fed a control diet (CD). The results of affinity chromatography suggested that in the cytoplasm of epididymal adipose tissue from either WD or CD rats, distinctive proteins bind to these peptides. However, despite the observed increase in the WD animals, the evaluated peptides increased insulin-stimulated glucose uptake in 3T3-L1 adipocytes treated with palmitate. Thus, intracellular peptides from the adipose tissue of Wistar rats can bind to specific proteins and facilitate insulin-induced glucose uptake in 3T3-L1 adipocytes.

Synthetic phosphoethanolamine a precursor of membrane phospholipids reduce tumor growth in mice bearing melanoma B16-F10 and in vitro induce apoptosis and arrest in G2/M phase

Phosphoethanolamine (Pho-s) is a compound involved in phospholipid turnover, acting as a substrate for many phospholipids of the cell membranes, especially phosphatidylcholine. We recently reported that synthetic Pho-s has potent effects on a wide variety of tumor cells. To determine if Pho-s has a potential antitumor activity, in this study we evaluated the activity of Pho-s against the B16-F10 melanoma both in vitro and in mice bearing a dorsal tumor. The treatment of B16F10 cells with Pho-s resulted in a dose-dependent inhibition of cell proliferation. At low concentrations, this activity appears to be involved in the arrest of the cell cycle at G2/M, while at high concentrations Pho-s induces apoptosis. In accordance with these results, the loss of mitochondrial potential and increased caspase-3 activity suggest that Phos has dual antitumor effects; i.e. it induces apoptosis at high concentrations and modulates the cell cycle at lower concentrations. In vivo, we evaluated the effect of Pho-s in mice bearing B16-F10 melanoma. The results show that Pho-s reduces the tumoral volume increasing survival rate. Furthermore, the tumor doubling time and tumor delays were substantially reduced when compared with untreated mice. Histological analyses reveal that Pho-s induces changes in cell morphology, typical characteristics of apoptosis, in addition the large areas of necrosis correlating with a reduction of tumor size. The results presented here support the hypothesis that Pho-s has antitumor effects by the induction of apoptosis as well as the inhibition of cell proliferation by arrest at G2/M. Thus, Pho-s can be regarded as a promising agent for the treatment of melanoma. Published by Elsevier Masson SAS.

Anticancer Effects of Synthetic Phosphoethanolamine on Ehrlich Ascites Tumor: An Experimental Study

Background: Antineoplastic phospholipids (ALPs) represent a promising class of drugs with a novel mode of action undergoes rapid turnover in the cell membrane of tumors, interfering with lipid signal transduction, inducing cell death. The aim of this study was to investigate the synthetic phosphoethanolamine (Pho-s) as a new anticancer agent. Materials and Methods: Cell viability and morphology were assessed by (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Hoechst and rhodamine staining. Apoptosis was assessed by Annexin V and propidium iodide (PI) staining, caspase-3 activity, mitochondrial membrane potential (Delta m psi) and cell cycle analysis, combined with evaluation of tumor growth in Ehrlich Ascites Tumor (EAT) bearing mice. Results: We found that Pho-s 2.30 mg/ml induced cytotoxicity in all tumor cell lines studied without affecting normal cells. In vitro studies with EAT cells indicated that Pho-s induced apoptosis, demonstrated by an increase in Annexin-V positive cells, loss of mitochondrial potential (Delta m psi) and increased caspase-3 activity. It was also shown to increase the sub-G(1) apoptotic fraction and inhibit progression to the S phase of the cell cycle. Additionally, antitumor effects on the EAT-bearing mice showed that Pho-s, at a concentration of 35 and 70 mg/kg, inhibited tumor growth and increased the lifespan of animals without causing liver toxicity. Conclusion: These findings suggest that Pho-s is a potential anticancer candidate drug.

Natriuretic Peptides and Long-Term Mortality in Patients with Severe Aortic Stenosis

Background and aim of the study: The natriuretic peptides, brain natriuretic peptide (BNP) and its N-terminal prohormone (NT-proBNP), can be used as diagnostic and prognostic markers for aortic stenosis (AS). However, the association between BNP, NT-proBNP, and long-term clinical outcomes in patients with severe AS remains uncertain. Methods: A total of 64 patients with severe AS was prospectively enrolled into the study, and underwent clinical and echocardiographic assessments at baseline. Blood samples were drawn for plasma BNP and NT-proBNP analyses. The primary outcome was death from any cause, through a six-year follow up period. Cox proportional hazards modeling was used to examine the association between natriuretic peptides and long-term mortality, adjusting for important clinical factors. Results: During a mean period of 1,520 681 days, 51 patients (80%) were submitted to aortic valve replacement, and 13 patients (20%) were medically managed without surgical interventions. Mortality rates were 13.7% in the surgical group and 62% in the medically managed group (p <0.001). Patients with higher plasma BNP (>135 pg/ml) and NT-proBNP (>1,150 pg/ml) levels at baseline had a greater risk of long-term mortality (hazard ratio [HR] 3.2, 95% confidence interval [CI] 1.1-9.1; HR 4.3, 95% CI 1.4-13.5, respectively). After adjusting for important covariates, both BNP and NT-proBNP remained independently associated with long-term mortality (HR 2.9, 95%CI 1.5-5.7; HR 1.8, 95%CI 1.1-3.1, respectively). Conclusion: In patients with severe AS, plasma BNP and NT-proBNP levels were associated with long-term mortality. The use of these biomarkers to guide treatment might represent an interesting approach that deserves further evaluation. The Journal of Heart Valve Disease 2012;21:331-336

Peptidomic Analysis of HEK293T Cells: Effect of the Proteasome Inhibitor Epoxomicin on Intracellular Peptides

Peptides derived from cytosolic, mitochondrial, and nuclear proteins have been detected in extracts of animal tissues and cell lines. To test whether the proteasome is involved in their formation, HEK293T cells were treated with epoxomicin (0.2 or 2 mu M) for 1 h and quantitative peptidomics analysis was performed. Altogether, 147 unique peptides were identified by mass spectrometry sequence analysis. Epoxomicin treatment decreased the levels of the majority of intracellular peptides, consistent with inhibition of the proteasome beta-2 and beta-5 subunits. Treatment with the higher concentration of epoxomicin elevated the levels of some peptides. Most of the elevated peptides resulted from cleavages at acidic residues, suggesting that epoxomicin increased the processing of proteins through the beta-1 subunit. Interestingly, some of the peptides that were elevated by the epoxomicin treatment had hydrophobic residues in P1 cleavage sites. Taken together, these findings suggest that, while the proteasome is the major source of intracellular peptides, other peptide-generating mechanisms exist. Because intracellular peptides are likely to perform intracellular functions, studies using proteasome inhibitors need to be interpreted with caution, as it is possible that the effects of these inhibitors are due to a change in the peptide levels rather than inhibition of protein degradation.

Detailed H-1 and C-13 NMR structural assignment and relative stereochemistry determination for three new and one known semi-synthetic sesquiterpene lactones

In this work is described a complete H-1 and C-13 NMR analysis for a group of four sesquiterpene lactones, three previously unknown. The unequivocal assignments were achieved by H-1 NMR, C-13{H-1} NMR, gCOSY. gHMQC, gHMBC and NOESY experiments and no ambiguities were left behind. All hydrogen coupling constants were measured, clarifying all hydrogen signals multiplicities. (C) 2011 Elsevier B.V. All rights reserved.

Metabolic engineering of beta-oxidation in Penicillium chrysogenum for improved semi-synthetic cephalosporin biosynthesis

Industrial production of semi-synthetic cephalosporins by Penicillium chrysogenum requires supplementation of the growth media with the side-chain precursor adipic acid. In glucose-limited chemostat cultures of P. chrysogenum, up to 88% of the consumed adipic acid was not recovered in cephalosporinrelated products, but used as an additional carbon and energy source for growth. This low efficiency of side-chain precursor incorporation provides an economic incentive for studying and engineering the metabolism of adipic acid in P. cluysogenum. Chemostat-based transcriptome analysis in the presence and absence of adipic acid confirmed that adipic acid metabolism in this fungus occurs via beta-oxidation. A set of 52 adipate-responsive genes included six putative genes for acyl-CoA oxidases and dehydrogenases, enzymes responsible for the first step of beta-oxidation. Subcellular localization of the differentially expressed acyl-CoA oxidases and dehydrogenases revealed that the oxidases were exclusively targeted to peroxisomes, while the dehydrogenases were found either in peroxisomes or in mitochondria. Deletion of the genes encoding the peroxisomal acyl-CoA oxidase Pc20g01800 and the mitochondrial acyl-CoA dehydrogenase Pc20g07920 resulted in a 1.6- and 3.7-fold increase in the production of the semi-synthetic cephalosporin intermediate adipoyl-6-APA, respectively. The deletion strains also showed reduced adipate consumption compared to the reference strain, indicating that engineering of the first step of beta-oxidation successfully redirected a larger fraction of adipic acid towards cephalosporin biosynthesis. (C) 2012 Elsevier Inc. All rights reserved.

P16-39. T cell recognition of autologous and non-autologous HIV-1 protease peptides by HIV-1 infected patients undergoing PI therapy

NIH, ICGEB, FAPESP, CNPq

Antimicrobial peptides and nitric oxide production by neutrophils from periodontitis subjects

Neutrophils play an important role in periodontitis by producing nitric oxide (NO) and antimicrobial peptides, molecules with microbicidal activity via oxygen-dependent and -independent mechanisms, respectively. It is unknown whether variation in the production of antimicrobial peptides such as LL-37, human neutrophil peptides (HNP) 1-3, and NO by neutrophils influences the pathogenesis of periodontal diseases. We compared the production of these peptides and NO by lipopolysaccharide (LPS)-stimulated neutrophils isolated from healthy subjects and from patients with periodontitis. Peripheral blood neutrophils were cultured with or without Aggregatibacter actinomycetemcomitans-LPS (Aa-LPS), Porphyromonas gingivalis-LPS (Pg-LPS) and Escherichia coli-LPS (Ec-LPS). qRT-PCR was used to determine quantities of HNP 1-3 and LL-37 mRNA in neutrophils. Amounts of HNP 1-3 and LL-37 proteins in the cell culture supernatants were also determined by ELISA. In addition, NO levels in neutrophil culture supernatants were quantitated by the Griess reaction. Neutrophils from periodontitis patients cultured with Aa-LPS, Pg-LPS and Ec-LPS expressed higher HNP 1-3 mRNA than neutrophils from healthy subjects. LL-37 mRNA expression was higher in neutrophils from patients stimulated with Aa-LPS. Neutrophils from periodontitis patients produced significantly higher LL-37 protein levels than neutrophils from healthy subjects when stimulated with Pg-LPS and Ec-LPS, but no difference was observed in HNP 1-3 production. Neutrophils from periodontitis patients cultured or not with Pg-LPS and Ec-LPS produced significantly lower NO levels than neutrophils from healthy subjects. The significant differences in the production of LL-37 and NO between neutrophils from healthy and periodontitis subjects indicate that production of these molecules might influence individual susceptibility to important periodontal pathogens.

'Synthetic lipase' production from a newly isolated Sporidiobolus pararoseus strain by submerged fermentation

The lipase produced by a newly isolate Sporidiobolus pararoseus strain has potential catalysis ability for esterification reactions. In order to improve its synthetic activity, this work aimed at optimizing 'synthetic lipase' production by submerged fermentation of a conventional media based on peptone, yeast extract, NaCl and olive oil using experimental design technique. According to the results obtained in the first experimental design (2(4-1)), yeast extract and NaCl concentrations were tested to further optimization by response surface methodology. The maximum 'synthetic lipase' activity obtained was 26.9 U/mL in the optimized media (5.0, 6.8, 7.0 and 1.0% (wt/v) of peptone, yeast extract, NaCl and olive oil, respectively), representing a 6.36-fold increase compared to the initial medium. The time course of 'synthetic lipase' production in the optimized condition was evaluated in terms of synthetic activity, protease activity, biomass and total carbon and the maximum synthetic activity was observed during the stationary phase of growth.

Synthetic biology: a novel approach for the construction of industrial microorganisms

The recent achievement of synthesising a functioning bacterial chromosome marks a coming of age for engineering living organisms. In the future this should allow the construction of novel organisms to help solve the problems facing the human race, including health care, food, energy and environmental protection. In this minireview, the current state of the field is described and the role of synthetic biology in biotechnology in the short and medium term is discussed. It is particularly aimed at the needs of food technologists, nutritionists and other biotechnologists, who might not be aware of the potential significance of synthetic biology to the research and development in their fields. The potential of synthetic biology to produce interesting new polyketide compounds is discussed in detail.