Saponins from Swartzia langsdorffii: biological activities

The presence of saponins and the molluscicidal activity of the roots, leaves, seeds and fruits of Swartzia langsdorffii Raddi (Leguminosae) against Biomphalaria glabrata adults and eggs were investigated. The roots, seeds and fruits were macerated in 95% ethanol. These extracts exerted a significant molluscicidal activity against B. glabrata, up to a dilution of 100 mg/l. Four mixtures (A2, B2, C and D) of triterpenoid oleanane type saponins were chromatographically isolated from the seed and fruit extracts. Two known saponins (1 and 2) were identified as beta-D-glucopyranosyl-[alpha-L-rhamnopyranosyl-(1->3)- beta-D-glucuronopyranosyl-(1->3)]-3beta-hydroxyolean-12-ene-28 -oate, and beta-D-glucopyranosyl-(1->3)-beta-D-glucuronopyranosyl-(1 ->3)]-3beta-hydroxyolean-12-ene-28-oate, respectively. These two saponins were present in all the mixtures, together with other triterpenoid oleane type saponins, which were shown to be less polar, by reversed-phase HPLC. The saponin identifications were based on spectral evidence, including ¹H-¹H two-dimensional correlation spectroscopy, nuclear Overhauser and exchange spectroscopy, heteronuclear multiple quantum coherence, and heteronuclear multiple-bond connectivity experiments. The toxicity of S. langsdorffii saponins to non-target organisms was prescreened by the brine shrimp lethality test.

Allergens Induce the Release of Lactoferrin by Neutrophils from Asthmatic Patients.

BACKGROUND Despite the evidence that Lactoferrin (Lf) is involved in allergic asthma processes, it is unknown whether neutrophils can be one of the main cellular sources of this key inflammatory mediator directly in response of an IgE mediated stimulus. The present study was undertaken to analyze this question. METHODS Neutrophils from healthy subjects (n = 34) and neutrophils from allergic asthmatic patients (n = 102) were challenged in vitro with specific allergens to which the patients were sensitized, PAF, or agonist mAbs against IgE-receptors, and the levels of Lf were measured in the culture supernatant. The levels of serum IgE together with the severity of symptoms were also analyzed. RESULTS Lf was released into the culture supernatant of neutrophils from allergic asthmatic patients in response to allergens and PAF. This response was highly allergen-specific, and did not happen in neutrophils from healthy donors. Allergen effect was mimicked by Abs against FcεRI and galectin-3 but not by FcεRII. The levels of released Lf correlated well with the levels of serum specific IgE and severity of asthma symptoms. These observations represent a novel view of neutrophils as an important source of Lf in allergic asthma. Importantly, the levels of released Lf by neutrophils could therefore be used to evaluate disease severity in allergic asthmatic patients.

Reasons for discontinuation of recommended therapies according to the patients after acute coronary syndromes.

BACKGROUND: The prescription of recommended medical therapies is a key factor to improve prognosis after acute coronary syndromes (ACS). However, reasons for cardiovascular therapies discontinuation after hospital discharge are poorly reported in previous studies. METHODS: We enrolled 3055 consecutive patients hospitalized with a main diagnosis of ACS in four Swiss university hospitals with a prospective one-year follow-up. We assessed the self-reported use of recommended therapies and the reasons for medication discontinuation according to the patient interview performed at one-year follow-up. RESULTS: 3014 (99.3%) patients were discharged with aspirin, 2983 (98.4%) with statin, 2464 (81.2%) with beta-blocker, 2738 (90.3%) with ACE inhibitors/ARB and 2597 (100%) with P2Y12 inhibitors if treated with coronary stent. At the one-year follow-up, the discontinuation percentages were 2.9% for aspirin, 6.6% for statin, 11.6% for beta-blocker, 15.1% for ACE inhibitor/ARB and 17.8% for P2Y12 inhibitors. Most patients reported having discontinued their medication based on their physicians' decision: 64 (2.1%) for aspirin, 82 (2.7%) for statin, 212 (8.6%) for beta-blocker, 251 (9.1% for ACE inhibitor/ARB) and 293 (11.4%) for P2Y12 inhibitors, while side effect, perception that medication was unnecessary and medication costs were uncommon reported reasons (<2%) according to the patients. CONCLUSIONS: Discontinuation of recommended therapies after ACS differs according the class of medication with the lowest percentages for aspirin. According to patients, most stopped their cardiovascular medication based on their physician's decision, while spontaneous discontinuation was infrequent.

GW501516-activated PPARβ/δ promotes liver fibrosis via p38-JNK MAPK-induced hepatic stellate cell proliferation.

ABSTRACT: BACKGROUND: After liver injury, the repair process comprises activation and proliferation of hepatic stellate cells (HSCs), which produce extracellular matrix (ECM) proteins. Peroxisome proliferator-activated receptor beta/delta (PPARβ/δ) is highly expressed in these cells, but its function in liver repair remains incompletely understood. This study investigated whether activation of PPARβ/δ with the ligand GW501516 influenced the fibrotic response to injury from chronic carbon tetrachloride (CCl4) treatment in mice. Wild type and PPARβ/δ-null mice were treated with CCl4 alone or CCl4 co-administered with GW501516. To unveil mechanisms underlying the PPARβ/δ-dependent effects, we analyzed the proliferative response of human LX-2 HSCs to GW501516 in the presence or absence of PPARβ/δ. RESULTS: We found that GW501516 treatment enhanced the fibrotic response. Compared to the other experimental groups, CCl4/GW501516-treated wild type mice exhibited increased expression of various profibrotic and pro-inflammatory genes, such as those involved in extracellular matrix deposition and macrophage recruitment. Importantly, compared to healthy liver, hepatic fibrotic tissues from alcoholic patients showed increased expression of several PPAR target genes, including phosphoinositide-dependent kinase-1, transforming growth factor beta-1, and monocyte chemoattractant protein-1. GW501516 stimulated HSC proliferation that caused enhanced fibrotic and inflammatory responses, by increasing the phosphorylation of p38 and c-Jun N-terminal kinases through the phosphoinositide-3 kinase/protein kinase-C alpha/beta mixed lineage kinase-3 pathway. CONCLUSIONS: This study clarified the mechanism underlying GW501516-dependent promotion of hepatic repair by stimulating proliferation of HSCs via the p38 and JNK MAPK pathways.

Polimorfismo enzimático nos tecidos de pereira

O trabalho foi realizado com o objetivo de avaliar o polimorfismo enzimático em diferentes tecidos de oito cultivares de pereira Pyrus communis L. Os genótipos utilizados fazem parte da coleção de plantas disponíveis na Universidade de Estudos de Bolonha. Para as análises isoenzimáticas foram utilizadas gemas floríferas dormentes no inverno, casca de ramos de um ano obtida de plantas em pleno desenvolvimento, folhas obtidas no início da primavera e folhas de plantas mantidas in vitro. A corrida eletroforética foi realizada em gel de poliacrilamida a gradiente com (5% a 12,5%). Os resultados obtidos com os genótipos utilizados indicaram que o sistema enzimático beta-glucosidase (E.C.3.2.1.21) apresentou atividade apenas nas folhas das plantas in vitro, com uma banda na mesma posição para todas as cultivares, ao passo que os sistemas para as enzimas esterase (E.C.3.1.1.2) e peroxidase (E.C.1.11.1.7) apresentaram elevado polimorfismo. Nas gemas dormentes analisadas, o sistema peroxidase permitiu diferenciar todos os genótipos. As formas isoenzimáticas da esterase permitiram separar todos os genótipos independentemente dos tecidos utilizados.

Aplicação de metodologias FTIR de transmissão e fotoacústica à caracterização de materiais altamente energéticos: parte II

Materials obtained during the synthesis of octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) were characterized by Fourier transform infrared (FTIR) transmission spectroscopy and/or Fourier transform infrared photoacoustic spectroscopy (FTIR-PAS). By these techniques the spectrometric alterations that occurred during the process were observed. The characterized species during the synthesis of HMX were alpha-HMX, beta-HMX, hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and HMX/RDX mixtures. The FTIR-PAS was verified to be a promising technique of great usefulness of the characterization of highly energetic materials because it is fast, simple and requires no sample preparation unlike Fourier transform infrared transmission technique (KBr pellet). The FTIR-PAS analysis showed that with small sample quantity is possible to distinguish between thealpha-HMX and beta-HMX and to detect even in a qualitative way different HMX / RDX ratios.

Constituintes químicos e atividade antioxidante de Sida galheirensiS Ulbr. (Malvaceae)

The phytochemical investigation of Sida galheirensis led to the isolation of 5,4'-dihydroxy-3,7,3´-trimethoxyflavone, 17³-ethoxyphaeoforbide, a rare natural product, 6,7-dimethoxycoumarin, ortho-hydroxybenzoic acid, sitosterol-3-O-beta-D-glucopyranoside, stigmasterol-3-O-beta-D-glucopyranoside, 5,7,4'-trihydroxyflavone, 5,7,3',4'-tetrahydroxyflavone, kaempferol-3-O-beta-D-(6"-E-p-coumaroyl) glucopyranoside and luteolin 7-O-beta-D-glucopyranoside. Their structures were assigned based on spectroscopic analyses, including two-dimensional NMR techniques. Antioxidant activities of hexane, CHCl3, EtOAc, BuOH and EtOH extracts of Sida galheirensis were measured using the 1,2-diphenyl-2-picryl-hydrazyl (DPPH) free radical scavenging assay. This is also the first work reporting the chemical investigation of Sida galheirensis.

Constituintes químicos das cascas do caule de Cenostigma macrophyllum: ocorrência de colesterol

Phytochemical investigation of the bark of Cenostigma macrophyllum (Leguminosae-Caesapinioideae) resulted in the isolation and identification of valoneic acid dilactone, ellagic acid, lupeol, alkyl ferulate, four free sterols (cholesterol, campesterol, stigmasterol and sitosterol), a mixture of sitosteryl ester derivatives of fatty acids, sitosterol-3-O-beta-D-glucopyranoside, stigmasterol-3-O-beta-D-glucopyranoside and saturated and unsaturated fatty acids. The structures of the isolated compounds were identified by ¹H and 13C NMR spectral analysis and comparison with literature data. The mixtures of 3-beta-hydroxysterols and fatty acids were analysed by GC/MS.

Efeito da temperatura no crescimento micelial e patogenicidade de Pythium spp. que ocorrem em alface hidropônica

Doze isolados de Pythium foram obtidos de raízes de alface cultivada em sistemas hidropônicos comerciais, apresentando ou não sintomas de apodrecimento. Três desses isolados foram identificados como Pythium helicoides Drechsler (H1, H2 e H3), cinco como pertencentes ao grupo F (F1 a F5) e quatro ao grupo T (T1 a T4) de Pythium. A identificação das espécies foi realizada baseando-se nas características morfológicas. O efeito da temperatura (10, 15, 18, 21, 24, 27, 30, 37 e 40ºC) sobre o crescimento micelial foi determinado para todos os isolados. As temperaturas mínima e máxima, estimadas pela função beta generalizada, variaram de 3,5 a 10ºC e de 40 a 40,7ºC, respectivamente. A temperatura ótima foi de 24 a 37ºC para P. helicoides, de 25 a 35ºC para o isolado F4 e de 21 a 30ºC para os demais isolados. A patogenicidade e a agressividade dos isolados foram avaliadas, inoculando-se sementes de alface cv. Verônica, semeadas em ágar-água, a 21 e 30ºC. A 30ºC, os isolados de P. helicoides foram notadamente os mais agressivos, ocasionando 100 % de mortalidade das sementes logo após sua germinação. A 21ºC, todos os isolados induziram subdesenvolvimento de plântulas, acompanhado ou não de necrose dos tecidos radiculares. Trata-se do primeiro relato de P. helicoides para o Brasil e a primeira referência mundial da espécie em hidroponia.

Rôle de l’enzyme PAS kinase dans la régulation du facteur de transcription PDX-1 dans la cellule bêta pancréatique.

Le diabète de type 2 (DT2) se caractérise par une production insuffisante d'insuline par le pancréas ainsi qu'une résistance des tissus périphériques à l'action de l'insuline. Dans les cellules bêta pancréatiques, le glucose stimule la production de l'insuline en induisant la transcription de son gène et la traduction ainsi que la sécrétion de sa protéine. Paradoxalement, une exposition prolongée et simultanée de ces cellules à de hautes concentrations de glucose en présence d'acides gras conduit à la détérioration de la fonction bêta pancréatique et au développement du DT2. Toutefois, les mécanismes moléculaires responsables de ces effets du glucose ne sont que partiellement connus. L'objectif du travail décrit dans cette thèse est d'identifier les mécanismes responsables de la régulation de la transcription du gène de l'insuline. PDX-1 (de l’anglais pour pancreatic and duodenal homeobox 1) est un facteur de transcription majeur et essentiel tant pour le développement du pancréas que pour le maintien de sa fonction à l'état adulte. En réponse au glucose, PDX-1 se lie au promoteur du gène de l'insuline et induit sa transcription. Ceci est inhibé par l'acide gras palmitate. Dans la première partie des travaux effectués dans le cadre de cette thèse, nous avons identifié deux mécanismes de régulation de la transcription du gène de l'insuline: le premier via ERK1/2 (de l'anglais pour extracellular-signal-regulated protein kinases 1 and 2) et le second par l’enzyme PASK (pour per-arnt-sim kinase). Nous avons également mis en évidence l'existence d'un troisième mécanisme impliquant l'inhibition de l'expression du facteur de transcription MafA par le palmitate. Nos travaux indiquent que la contribution de la signalisation via PASK est majeure. L'expression de PASK est augmentée par le glucose et inhibée par le palmitate. Sa surexpression dans les cellules MIN6 et les îlots isolés de rats, mime les effets du glucose sur l'expression du gène de l'insuline ainsi que sur l'expression de PDX-1 et prévient les effets délétères du palmitate. Dans la deuxième partie de la thèse, nous avons identifié un nouveau mécanisme par lequel PASK augmente la stabilité protéique de PDX-1, soit via la phosphorylation et l'inactivation de la protéine kinase GSK3 bêta (de l'anglais pour glycogen synthase kinase 3 beta). Le glucose induit la translocation de PDX-1 du cytoplasme vers le noyau, ce qui est essentiel à sa liaison au promoteur de ses gènes cibles. L'exclusion nucléaire de PDX-1 a été observée dans plusieurs modèles ex vivo et in vivo de dysfonction de la cellule bêta pancréatique. Dans le dernier volet de cette thèse, nous avons démontré l'importance de l'utilisation de cellules primaires (îlots isolés et dispersés) pour étudier la translocation nucléaire de PDX-1 endogène étant donné que ce mode de régulation est absent dans les lignées insulino-sécrétrices MIN6 et HIT-T15. Ces études nous ont permis d'identifier et de mieux comprendre les mécanismes régulant la transcription du gène de l'insuline via le facteur de transcription PDX-1. Les cibles moléculaires ainsi identifiées pourraient contribuer au développement de nouvelles approches thérapeutiques pour le traitement du diabète de type 2. Mots-clés : Diabète, îlots de Langerhans, cellule bêta pancréatique, gène de l'insuline, PDX-1, PASK, GSK3 bêta, ERK1/2, PKB, glucose, palmitate.

A new molecular scaffold for the formation of supramolecular peptide double helices: the crystallographic insight

A series of water-soluble synthetic dipeptides (1-3) with an N-terminally located beta-alanine residue, beta-alanyl-L-valine (1), beta-alanyl-L-isoleucine (2), and beta-alanyl-L-phenylalanine (3, form hydrogen-bonded supramolecular double helices with a pitch length of 1 nm, whereas the C-terminally positioned beta-alanine containing dipeptide (4), L-phenylalanyl-beta-alanine, does not form a supramolecular double helical structure. beta-Ala-Xaa (Xaa = Val/Ile/Phe) can be regarded as a new motif for the formation of supramolecular double helical structures in the solid state.

Development of novel brush-type chiral stationary phases based on terpenoid selectors: HPLC evaluation and theoretical investigation of enantioselective binding interactions

The terpenoid chiral selectors dehydroabietic acid, 12,14-dinitrodehydroabietic acid and friedelin have been covalently linked to silica gel yielding three chiral stationary phases CSP 1, CSP 2 and CSP 3, respectively. The enantiodiscriminating capability of each one of these phases was evaluated by HPLC with four families of chiral aromatic compounds composed of alcohols, amines, phenylalanine and tryptophan amino acid derivatives and beta-lactams. The CSP 3 phase, containing a selector with a large friedelane backbone is particularly suitable for resolving free alcohols and their derivatives bearing fluorine substituents, while CSP 2 with a dehydroabietic architecture is the only phase that efficiently discriminates 1, 1'-binaphthol atropisomers. CSP 3 also gives efficient resolution of the free amines. All three phases resolve well the racemates of N-trifluoracetyl and N-3,5-dinitrobenzoyl phenylalanine amino acid ester derivatives. Good enantioseparation of beta-lactams and N-benzoyl tryptophan amino acid derivatives was achieved on CSP 1. In order to understand the structural factors that govern the chiral molecular recognition ability of these phases, molecular dynamics simulations were carried out in the gas phase with binary diastereomeric complexes formed by the selectors of CSP 1 and CSP 2 and several amino acid derivatives. Decomposition of molecular mechanics energies shows that van der Waals interactions dominate the formation of the diastereomeric transient complexes while the electrostatic binding interactions are primarily responsible for the enantioselective binding of the (R)- and (S)-analytes. Analysis of the hydrogen bonds shows that electrostatic interactions are mainly associated with the formation of N-(HO)-O-...=C enantio selective hydrogen bonds between the amide binding sites from the selectors and the carbonyl groups of the analytes. The role of mobile phase polarity, a mixture of n-hexane and propan-2-ol in different ratios, was also evaluated through molecular dynamics simulations in explicit solvent. (c) 2006 Elsevier Ltd. All rights reserved.

Dietary vitamin C down-regulates inflammatory gene expression in apoE4 smokers

The deleterious impact of cigarette smoking on cardiovascular health may be in part attributable to a free radical mediated proinflammatory response in circulating monocytes. In the current investigation, the impact of vitamin C supplementation on monocyte gene expression was determined in apoE4 smokers versus non-smokers. A total of 10 smokers and 11 non-smokers consumed 60 mg/day of vitamin C for four weeks and a fasting blood sample was taken at baseline and post-intervention for the determination of plasma vitamin C and monocyte gene expression profiles using cDNA array and real time PCR. In apoE4 smokers, supplementation resulted in a 43% increase in plasma vitamin C concentrations. Furthermore, a number of genes were differentially expressed more than 2-fold in response to treatment, including a downregulation of the proinflammatory mediators tumor necrosis factor (TNF) beta, TNF receptor, neurotrophin-3 growth factor receptor, and monocyte chemoattractant protein I receptor. The study has identified a number of molecular mechanisms underlying the benefit of vitamin C supplementation in smokers. (c) 2005 Elsevier Inc. All rights reserved.

Incorporation of a S-glycosidic linkage into a glyconucleoside changes the conformational preference of both furanose sugars

A glyconucleoside containing a thioglycoside linkage, namely 1-(3-S-beta-D-ribofuranosyl-2,3-dideoxy-3-thio-beta-D-ribofuranosyl)-thy mine, has been prepared through condensation of a suitably protected derivative of 3'-thiothymidine with an activated ribose sugar. NMR has been used to study the conformation of the S-disaccharide and the unmodified O-disaccharide. A full pseudorotational analysis showed that for the S-disaccharide, the ribose and deoxy ribose sugars have a preference for the south and north pucker, respectively; which is the reverse of what is seen for the O-disaccharide. (c) 2006 Elsevier Ltd. All rights reserved.

Evaluation of Quantitative RT-PCR Using Nonamplified and Amplified RNA

Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) is a standard assay in molecular medicine for gene expression analysis. Samples from incisional/needle biopsies, laser-microdissected tumor cells and other biologic sources, normally available in clinical cancer studies, generate very small amounts of RNA that are restrictive for expression analysis. As a consequence, an RNA amplification procedure is required to assess the gene expression levels of such sample types. The reproducibility and accuracy of relative gene expression data produced by sensitive methodology as qRT-PCR when cDNA converted from amplified (A) RNA is used as template has not yet been properly addressed. In this study, to properly evaluate this issue, we performed 1 round of linear RNA amplification in 2 breast cell lines (C5.2 and HB4a) and assessed the relative expression of 34 genes using cDNA converted from both nonamplified (NA) and A RNA. Relative gene expression was obtained from beta actin or glyceraldehyde 3-phosphate dehydrogenase normalized data using different dilutions of cDNA, wherein the variability and fold-change differences in the expression of the 2 methods were compared. Our data showed that 1 round of linear RNA amplification, even with suboptimal-quality RNA, is appropriate to generate reproducible and high-fidelity qRT-PCR relative expression data that have similar confidence levels as those from NA samples. The use of cDNA that is converted from both A and NA RNA in a single qRT-PCR experiment clearly creates bias in relative gene expression data.