Both tumour-promoting and non-promoting phorbol esters inhibit 125I-EGF binding and stimulate the phosphorylation of a 80kd protein kinase C substrate protein in intact Swiss 3T3 cells

Sapintoxin A (SAP A) and 12-deoxyphorbol 13-phenylacetate (DOPP), are two biologically active but non-turnour-promoting phorbol esters that potently bind to and activate the phorbol ester receptor, protein kinase C (PKC). SAP A and DOPP cause a dose-dependent increase in the phosphorylation of an 80 kd (80K) substrate protein for PKC in Swiss 3T3 cells. A similar dose—response effect was seen with sapintoxin D (SAP D), the stage 2 promoting analogue of 12-O-tetradecanoylphorbol-13-acetate and the complete promoter phorbol 12,13-dibutyrate (PDB). The doses resulting in a half maximal phosphorylation of this protein (Ka were 20 nM (SAP A), 45 nM (DOPP), 23 nM (SAP D) and 37 nM (PDB). Both non-promoting and phorbol esters induced a dose-dependent inhibition of [125I]epidermal growth factor (EGF) binding to its receptor in Swiss 3T3 cells. The doses required for 50% inhibition of binding (Ki) were: 8 nM (SAP A), 16 nM (DOPP), 14 nM (SAP D) and 17 nM (PDB). The results clearly demonstrate that induction of phosphorylation of the Pu 80K phosphoprotein and inhibition of [125I]EGF binding in Swiss 3T3 cells following exposure to phorbol esters is independent of the tumour-promoting activity of these compounds. The fact that SAP A, DOPP, SAP D and PDB are mitogenic for a variety of cell types and that exposure to these compounds leads to 80K phosphorylation and inhibition of [125I]EGF binding, suggests that these early biological events may play a role in the mitogenic response induced by these compounds.

Surface structure of thin asymmetric PS-b-PMMA diblock copolymers investigated by atomic force microscopy

Asymmetric poly(styrene-b-methyl methacrylate) (PS-b-PMMA) diblock copolymers of molecular weight M-n = 29,700g mol(-1) (M-PS = 9300 g mol(-1) M-PMMA = 20,100 g mol(-1), PD = 1.15, chi(PS) = 0.323, chi(PMMA) = 0.677) and M-n = 63,900 g mol(-1) (M-PS = 50,500 g mol(-1), M-PMMA = 13,400 g mol(-1), PD = 1.18, chi(PS) = 0.790, chi(PMMA) = 0.210) were prepared via reversible addition-fragmentation chain transfer (RAFT) polymerization. Atomic force microscopy (AFM) was used to investigate the surface structure of thin films, prepared by spin-coating the diblock copolymers on a silicon substrate. We show that the nanostructure of the diblock copolymer depends on the molecular weight and volume fraction of the diblock copolymers. We observed a perpendicular lamellar structure for the high molar mass sample and a hexagonal-packed cylindrical patterning for the lower molar mass one. Small-angle X-ray scattering investigation of these samples without annealing did not reveal any ordered structure. Annealing of PS-b-PMMA samples at 160 degrees C for 24 h led to a change in surface structure.

Tin–osmium cluster formation with a monomeric tin(II) alkyl: the formation, crystal structure and reactivity of [Os3(µ-H)SnR(CO)10], a formal stannyne complex of the osmium triangle {R = C(SiMe3)2C5H4N-2}

The monomeric tin(II) species SnR2{R = C(SiMe3)2C5H4N-2} reacts with [Os3(H)2(CO)10] in hexane to give [Os3(µ-H)SnR(CO)10]1 quantitatively; 1 is the first formal stannyne complex of the triosmium nucleus, in which the picoline nitrogen is coordinated to the tin atom, and which is itself also reactive, being a potential precursor to high nuclearity SnOs clusters.

Frontiers and borderlands in Imperial perspectives: exploring Rome’s Egyptian frontier

Archaeological research has addressed imperial frontiers for more than a century. Romanists, in particular, have engaged in exploring frontiers from economic, militaristic, political, and (more recently) social vantages. This article suggests that we also consider the dialogue between space and social perception to understand imperial borderland developments. In addition to formulating new theoretical approaches to frontiers, this contribution represents the first comprehensive overview of both the documentary sources and the archaeological material found in Egypt's Great Oasis during the Roman period (ca. 30 B.C.E. to the sixth century C.E.). A holistic analysis of these sources reveals that Egypt's Great Oasis, which consisted of two separate but linked oases, served as a conceptual, physical, and human buffer zone for the Roman empire. This buffer zone protected the "ordered" Nile Valley inhabitants from the "chaotic" desert nomads, who lived just beyond the oases. This conclusion suggests that nomads required specific imperial frontier policies and that these policies may have been ideological as well as economic and militaristic.

One-pot three component reaction involving cyclohexyl isocyanide for the synthesis of furo[3,4-b]chromenes

On stirring an equimolar mixture of 4-oxo-4H-chromene-3-carbaldehyde, ninhydrin and cyclohexyl isocyanide in CH(2)Cl(2)-MeOH (7: 1) at room temperature produces 3-cyclohexylimino-1-(2-hydroxy-1,3-dioxo-2,3-dihydro-1H-inden-2-yl)-1,3-dihydro-9H-furo[3,4-b]chromen-9-one which on hydrolysis produces 1-(2-hydroxy-1,3-dioxo-2,3-dihydro-1H-inden-2-yl)-1H-furo[3,4-b]chromene-3,9-dione. The structure of the latter compound was confirmed by single crystal X-ray diffraction

Micelle size characterization of lipopeptides produced by B. subtilis and their recovery by the two-step ultrafiltration process

The aim of this work was to investigate the lipopeptides aggregation behavior in single and mixed solutions in a wide range of concentrations, in order to optimize their separation and purification following the two-step ultrafiltration process and using large pore size membranes (up to MWCO = 300 kDa). Micelle size was determined by dynamic light scattering. In single solutions of lipopeptide both surfactin and mycosubtilin formed micelles of different size depending on their concentration, micelles of average diameter = 5–105 nm for surfactin and 8–18 nm for mycosubtilin. However when the lipopeptides were in the same solution they formed mixed micelles of different size (d = 8 nm) and probably conformation to that formed by the individual lipopeptide, this prevents their separation according to size. These lipopeptides were purified from fermentation culture by the two-step ultrafiltration process using different MWCO membranes ranging from 10 to 300 kDa. This led to their effective rejection in the first ultrafiltration step by membranes with MCWO = 10–100 kDa but poor rejection by the 300 KDa membrane. The lipopeptides were recovered at 90% purity (in relation to protein) and with 2.34 enrichment in the permeate of the second ultrafiltration step with the 100 KDa membrane upon addition of 75% ethanol.

Genetic association analysis of vitamin D pathway with obesity traits

Objective:Observational studies have examined the link between vitamin D deficiency and obesity traits. Some studies have reported associations between vitamin D pathway genes such as VDR, GC and CYP27B1 with body mass index (BMI) and waist circumference (WC); however, the findings have been inconsistent. Therefore, we investigated the involvement of vitamin D metabolic pathway genes in obesity-related traits in a large population-based study.Methods:We undertook a comprehensive analysis between 100 tagging single nucleotide polymorphisms (tagSNPs) in genes encoding for DHCR7, CYP2R1, VDBP, CYP27B1, CYP27A1, CYP24A1, VDR and RXRG, and obesity traits in 5224 participants (aged 45 years) in the 1958 British birth cohort (1958BC). We further extended our analyses to investigate the associations between SNPs and obesity traits using the summary statistics from the GIANT (Genetic Investigation of Anthropometric Traits) consortium (n=123 865).Results:In the 1958BC (n=5224), after Bonferroni correction, none of the tagSNPs were associated with obesity traits except for one tagSNP from CYP24A1 that was associated with waist-hip ratio (WHR) (rs2296239, P=0.001). However, the CYP24A1 SNP was not associated with BMI-adjusted WHR (WHRadj) in the 1958BC (rs2296239, P=1.00) and GIANT results (n=123 865, P=0.18). There was also no evidence for an interaction between the tagSNPs and obesity on BMI, WC, WHR and WHRadj in the 1958BC. In the GIANT consortium, none of the tagSNPs were associated with obesity traits.Conclusions:Despite a very large study, our findings suggest that the vitamin D pathway genes are unlikely to have a major role in obesity-related traits in the general population.

The permeability parameter of the outer membrane of pseudomonas aeruginosa Varies with the concentration of a test substrate, cephalosporin C

The permeability parameter (C) for the movement of cephalosporin C across the outer membrane of Pseudomonas aeruginosa was measured using the widely accepted method of Zimmermann & Rosselet. In one experiment, the value of C varied continuously from 4·2 to 10·8 cm3 min-1 (mg dry wt cells)-1 over a range of concentrations of the test substrate, cephalosporin C, from 50 to 5 μm. Dependence of C on the concentration of test substrate was still observed when the effect of a possible electric potential difference across the outer membrane was corrected for. In quantitative studies of β-lactam permeation the dependence of C on the concentration of β-lactam should be taken into account.

State-resolved gas-surface reactivity of methane in the symmetric C-H stretch vibration on Ni(100)

The state-resolved reactivity of CH4 in its totally symmetric C-H stretch vibration (�1) has been measured on a Ni(100) surface. Methane molecules were accelerated to kinetic energies of 49 and 63:5 kJ=mol in a molecular beam and vibrationally excited to �1 by stimulated Raman pumping before surface impact at normal incidence. The reactivity of the symmetric-stretch excited CH4 is about an order of magnitude higher than that of methane excited to the antisymmetric stretch (�3) reported by Juurlink et al. [Phys. Rev. Lett. 83, 868 (1999)] and is similar to that we have previously observed for the excitation of the first overtone (2�3). The difference between the state-resolved reactivity for �1 and �3 is consistent with predictions of a vibrationally adiabatic model of the methane reaction dynamics and indicates that statistical models cannot correctly describe the chemisorption of CH4 on nickel.

Activation of glycoprotein VI (GPVI) and C-type lectin-like receptor-2 (CLEC-2) underlies platelet activation by diesel exhaust particles and other charged/hydrophobic ligands

Platelets are activated by a range of stimuli that share little or no resemblance in structure to each other or to recognized ligands, including diesel exhaust particles (DEP), small peptides [4N1-1, Champs (computed helical anti-membrane proteins), LSARLAF (Leu-Ser-Ala-Arg-Leu-Ala-Phe)], proteins (histones) and large polysaccharides (fucoidan, dextran sulfate). This miscellaneous group stimulate aggregation of human and mouse platelets through the glycoprotein VI (GPVI)-FcR γ-chain complex and/or C-type lectin-like receptor-2 (CLEC-2) as shown using platelets from mice deficient in either or both of these receptors. In addition, all of these ligands stimulate tyrosine phosphorylation in GPVI/CLEC-2-double-deficient platelets, indicating that they bind to additional surface receptors, although only in the case of dextran sulfate does this lead to activation. DEP, fucoidan and dextran sulfate, but not the other agonists, activate GPVI and CLEC-2 in transfected cell lines as shown using a sensitive reporter assay confirming a direct interaction with the two receptors. We conclude that this miscellaneous group of ligands bind to multiple proteins on the cell surface including GPVI and/or CLEC-2, inducing activation. These results have pathophysiological significance in a variety of conditions that involve exposure to activating charged/hydrophobic agents.

Stimulation of phosphatidylinositol hydrolysis, protein kinase C translocation, and mitogen-activated protein kinase activity by bradykinin in rat ventricular myocytes: dissociation from the hypertrophic response.

In ventricular myocytes cultured from neonatal rat hearts, bradykinin (BK), kallidin or BK(1-8) [(Des-Arg9)BK] stimulated PtdinsP2 hydrolysis by 3-4-fold. EC50 values were 6 nM (BK), 2 nM (kallidin), and 14 microM [BK(1-8)]. BK or kallidin stimulated the rapid (less than 30 s) translocation of more than 80% of the novel protein kinase C (PKC) isoforms nPKC-delta and nPKC-epsilon from the soluble to the particulate fraction. EC50 values for nPKC-delta translocation by BK or kallidin were 10 and 2 nM respectively. EC50 values for nPKC-epsilon translocation by BK or kallidin were 2 and 0.6 nM respectively. EC50 values for the translocation of nPKC-delta and nPKC-epsilon by BK(1-8) were more than 5 microM. The classical PKC, cPKC-alpha, and the atypical PKC, nPKC-zeta, did not translocate. BK caused activation and phosphorylation of p42-mitogen-activated protein kinase (MAPK) (maximal at 3-5 min, 30-35% of p42-MAPK phosphorylated). p44-MAPK was similarly activated. EC50 values for p42/p44-MAPK activation by BK were less than 1 nM whereas values for BK(1-8) were more than 10 microM. The order of potency [BK approximately equal to kallidin > BK (1-8)] for the stimulation of PtdInsP2 hydrolysis, nPKC-delta and nPKC-epsilon translocation, and p42/p44-MAPK activities suggests involvement of the B2 BK receptor subtype. In addition, stimulation of all three processes by BK was inhibited by the B2BK receptor-selective antagonist HOE140 but not by the B1-selective antagonist Leu8BK(1-8). Exposure of cells to phorbol 12-myristate 13-acetate for 24 h inhibited subsequent activation of p42/p44-MAPK by BK suggesting participation of nPKC (and possibly cPKC) isoforms in the activation process. Thus, like hypertrophic agents such as endothelin-1 (ET-1) and phenylephrine (PE), BK activates PtdInsP2 hydrolysis, translocates nPKC-delta, and nPKC-epsilon, and activates p42/p44-MAPK. However, in comparison with ET-1 and PE, BK was only weakly hypertrophic as assessed by cell morphology and patterns of gene expression. This difference could not be attributed to dissimilarities between the duration of activation of p42/p44-MAPK by BK or ET-1. Thus activation of these signalling pathways alone may be insufficient to induce a powerful hypertrophic response.

Prevalence of Vitamin D Insufficiency in Brazilian Adolescents

Background/Aims: Cutaneous sun exposure and dietary vitamin D intake are important determinants of vitamin D status. The objective of the present study was to evaluate the vitamin D status of a group of healthy adolescent students living in Brazil. Methods: One hundred and thirty-six adolescents, 64 boys and 72 girls, aged 16-20 years old, living in a rural town in the state of Sao Paulo, Brazil, participated in this study. Results: The mean dietary vitamin D intake was 140 (120-156) IU/day [3.5 (3.0-3.9) mu g/day]. Only 14.9% of the students met the daily adequate intake recommendation of vitamin D. Only 27.9% practice physical activity outdoors and 17.6% of the adolescents apply sunscreen daily. The mean 25(OH)D concentration was 73.0 (22.0) nmol/l [29.2 (8.8) ng/ml]. Vitamin D insufficiency was observed in 60% of adolescents. Conclusions: The present study suggests that even in a sunny climate like Brazil the prevalence of vitamin D insufficiency in adolescents is high. Most likely this is due to low intakes of vitamin D in this group. Due to the limited extent of natural dietary sources of vitamin D, a policy of vitamin D food fortification should be considered in the future, and in the meantime greater use of vitamin D supplements in this population group should be encouraged to provide the increased amounts of this essential nutrient for optimal health. Copyright (C) 2009 S. Karger AG, Basel