Hemojuvelin and Hepcidin Genes Sequencing in Brazilian Patients with Primary Iron Overload

Background: Most hereditary hemochromatosis (HH) patients are homozygous for the p. C282Y mutation in the HFE gene. Some studies reported that HH phenotypic expression could be modulated by genetic factors such as HJV and HAMP gene mutations. Aims: The aims of this study were to identify HJV and HAMP mutations and to analyze their impact on HH phenotype in non-p. C282Y homozygous individuals. Methods: Twenty-four Brazilian patients with primary iron overload and non-p. C282Y homozygous genotype (transferrin saturation >50% in women and >60% in men and absence of secondary causes) were selected. Subsequent bidirectional sequencing of the HJV and HAMP exons was performed. Results: Sequencing revealed a substitution in heterozygosis, c. 929C>G, which corresponds to p.A310G polymorphism in HJV exon 4 (rs7540883). In the same gene, in another individual, an IVS1-36C>G intronic variant was detected in heterozygosis. In the HAMP gene, an IVS3 + 42G>A intronic variant was identified. There were six (25.0%) patients carrying a heterozygous genotype for the HFE p. C282Y and nine (37.5%) patients carrying a heterozygous genotype for the HFE p. H63D. Conclusion: HJV p.A310G polymorphism and two intronic variants were found, but none of these alterations were associated with digenic inheritance with the HFE gene. Our data indicate that HJV and HAMP functional mutations are not frequent in these patients.

The oleoresin secretory system in seedlings and adult plants of copaiba (Copaifera langsdorffii Desf., Leguminosae-Caesalpinioideae)

The ecological and economic importance of oleoresin produced by Copaifera langsdorffii is well established. This study aims to investigate the ontogeny, anatomy and ultrastructure of the internal glands of C. langsdorffii during plant development. Samples were processed for light and electron microscopy and a specific technique was applied to impregnate endomembranes. Internal secretory glands were observed in the hypocotyl, epicotyl and eophylls of seedlings, and in the primary stem, pulvinus, petiole, rachis and leaf blade of adult plants. Canals and cavities show differential distribution. They arise from ground meristem cells, and the lumen is first formed by schizogenesis followed by later schizolysigenous development. The dense cytoplasm of epithelial cells shows mitochondria, plastids without thylakoids, polyribosomes and endoplasmic reticulum. A periplastidial reticulum was also observed. Secretion is released by eccrine, granulocrine and holocrine processes. Lipophilic and hydrophilic compounds were histochemically detected in both canals and cavities, whereas resin was detected only in canals. The presence of these substances has been associated with plants` defences against dehydration, as well as against attacks from herbivores and pathogens, from seedling stage onwards. (C) 2011 Elsevier GmbH. All rights reserved.

Lipase Production by Endophytic Fungus Cercospora kikuchii: Stability of Enzymatic Activity after Spray Drying in the Presence of Carbohydrates

The present work deals with improving the production and stabilization of lipases from Cercospora kikuchii. Maximum enzyme production (9.384 U/ml) was obtained after 6 days in a medium supplemented with 2% soybean oil. The lipases were spray dried with different adjuvants, and their stability was studied. The residual enzyme activity after drying with 10% (w/v) of lactose, b- cyclodextrin, maltodextrin, mannitol, gum arabic, and trehalose ranged from 63 to 100%. The enzyme activity was lost in the absence of adjuvants. Most of the adjuvants used kept up at least 50% of the enzymatic activity at 5 degrees C and 40% at 25 degrees C after 8 months. The lipase dried with 10% of beta-cyclodextrin retained 72% of activity at 5 degrees C. Lipases were separated by butyl-sepharose column into 4 pools, and pool 4 was partially purified (33.1%; 269.5 U/mg protein). This pool was also spray dried in maltodextrin DE10, and it maintained 100% of activity.

The Protective Role of Lychnophora ericoides Mart. (Brazilian Arnica) in 1,2-Dimethylhydrazine-Induced Experimental Colon Carcinogenesis

Aberrant crypt foci (ACF) and colon rectal mucosal epithelial cell proliferation have been shown to be increased in patients with colon cancer and have been largely used for early detection of factors that influence colorectal carcinogenesis in rats. Fifty male Wistar rats were randomly divided into 5 groups. The groups G1 to G4 were given 4 injections of the carcinogen 1,2-dimethylhydrazine (DMH). The G2 group received Lychnophora ericoides (LE) extracts for 6 wk. The groups G3 and G4 received LE for 4 wk and 2 wk, respectively, at the postinitiation and initiation phases of colonic carcinogenesis. The group G5 was the control. Forty-two days after the first injections of DMH for the neoplasic induction, we observed a statistically significant decrease in the number of aberrant crypt foci (ACF) and an attenuation of the increase in cell proliferation induced by DMH in all the LE-treated groups. Thus, we concluded that Lychnophora ericoides extracts were effective against the development of cancer. These data suggest that LE has a protective influence on the process of colon carcinogenesis, suppressing both the initiation and the promotion of colonic carcinogenesis.

Biotransformation using Mucor rouxii for the production of oleanolic acid derivatives and their antimicrobial activity against oral pathogens

The goal of this study is to produce oleanolic acid derivatives by biotransformation process using Mucor rouxii and evaluate their antimicrobial activity against oral pathogens. The microbial transformation was carried out in shake flasks at 30A degrees C for 216 h with shaking at 120 rpm. Three new derivatives, 7 beta-hydroxy-3-oxo-olean-12-en-28-oic acid, 7 beta,21 beta-dihydroxy-3-oxo-olean-12-en-28-oic acid, and 3 beta,7 beta,21 beta-trihydroxyolean-12-en-28-oic acid, and one know compound, 21 beta-hydroxy-3-oxo-olean-12-en-28-oic acid, were isolated, and the structures were elucidated on the basis of spectroscopic analyses. The antimicrobial activity of the substrate and its transformed products was evaluated against five oral pathogens. Among these compounds, the derivative 21 beta-hydroxy-3-oxo-olean-12-en-28-oic acid displayed the strongest activity against Porphyromonas gingivalis, which is a primary etiological agent of periodontal disease. In an attempt to improve the antimicrobial activity of the derivative 21 beta-hydroxy-3-oxo-olean-12-en-28-oic acid, its sodium salt was prepared, and the minimum inhibitory concentration against P. gingivalis was reduced by one-half. The biotransformation process using M. rouxii has potential to be applied to the production of oleanolic acid derivatives. Research and antimicrobial activity evaluation of new oleanolic acid derivatives may provide an important contribution to the discovery of new adjunct agents for treatment of dental diseases such as dental caries, gingivitis, and periodontitis.

Self-Similarity of Air-Water Flows in Skimming Flows on Stepped Spillways

Skimming flows on stepped spillways are characterised by a significant rate of turbulent dissipation on the chute. Herein an advanced signal processing of traditional conductivity probe signals is developed to provide further details on the turbulent time and length scales. The technique is applied to a 22° stepped chute operating with flow Reynolds numbers between 3.8 and 7.1 E+5. The new correlation analyses yielded a characterisation of large eddies advecting the bubbles. The turbulent length scales were related to the characteristic depth Y90. Some self-similar relationships were observed systematically at both macroscopic and microscopic levels. These included the distributions of void fraction, bubble count rate, interfacial velocity and turbulence level, and turbulence time and length scales. The self-similarity results were significant because they provided a picture general enough to be used to characterise the air-water flow field in prototype spillways.

Male Reproductive Apparatus and Spermatophore Morphology of the Hermit Crabs Pagurus brevidactylus and P. criniticornis (Anomura, Paguridae)

Spermatozoa of most crustacean species are nonmotile and are packed into spermatophores. In Decapoda, spermatophores are highly variable in morphology and can be useful in the solving of taxonomic and systematic questions, especially among the Anomura. In this study, the morphology and morphometry of the spermatophores of the western Atlantic hermit crabs Pagurus brevidactylus and P criniticornis are described. The abdomen of fresh male specimens was dissected to expose the reproductive system and to extract the spermatophores, which were analyzed by stereoscopic, light, and scanning electron microscopy. The vas deferens can be divided macroscopically in three regions, all of them containing spermatophores. Tripartite spermatophores are composed of an elongated cylindrical main ampulla, a triangular accessory ampulla, a narrow cylindrical peduncle, and a round pedestal. Dimensions of the spermatophore components are positively correlated to the size of the crab. Morphological patterns observed in this study resemble those of other pagurid hermit crabs investigated to date. The morphological character distribution confirms classifications based on adult morphology and molecular analysis. J. Morphol. 272:1271-1280, 2011. (C) 2011 Wiley-Liss, Inc.

Comparative reproductive biology of two southwestern Atlantic populations of the hermit crab Pagurus exilis (Crustacea: Anomura: Paguridae)

Pagurus exilis (Benedict, 1892) is an endemic South Atlantic hermit crab with a distribution ranging from Rio de Janeiro State, Brazil, to Buenos Aires Province, Argentina. The present study analyzed the reproduction of two populations at the extremes of this geographical distribution, and compared their reproductive period, fecundity, and changes in egg size and egg volume during the incubation period in order to assess the variability over this latitudinal range. Hermit crabs were collected monthly over a 2-year period. In total, 108 females were analyzed for Brazil (44 non-ovigerous and 64 ovigerous), and 141 for Argentina (87 non-ovigerous and 54 ovigerous). Reproduction in Brazil occurs year-round, with peaks in the fall and winter seasons; in Argentina reproduction occurs only in spring and summer. The Brazilian ovigerous females were significantly larger than the Argentina ones (Brazil: SL = 5.33 +/- 1.45 mm; Argentina: SL = 4.15 +/- 0.52 mm; P < 0.001). The fecundity was 1447 +/- 831 eggs (317 to 2885) in Brazil and 987 +/- 711 eggs (114 to 2665) in Argentina, with a trend towards higher fecundity in Brazil. Eggs in the Argentina population were larger than those in Brazil for all the three stages investigated, and no changes in egg volume were found during egg development for both populations. The reproductive traits of the two populations showed some important differences, which may reflect adaptations to local environmental conditions, demonstrating a high plasticity of reproductive features of the species in Brazilian and Argentine waters. The strategy adopted by the Argentina population involves a lower production of larger eggs compared to the population in Brazil; this lower production is associated with reproduction in cold-water regions.

Does Petrolisthes armatus (Anomura, Porcellanidae) form a Species Complex or Are We Dealing with Just One Widely Distributed Species?

Fernando L. Mantelatto, Leonardo G. Pileggi, Ivana Miranda, and Ingo S. Wehrtmann (2011) Does Petrolisthes armatus (Anomura, Porcellanidae) form a species complex or are we dealing with just one widely distributed species? Zoological Studies 50(3): 372-384. Petrolisthes armatus has the widest distribution known among members of the family Porcellanidae and is one of the most ubiquitous and locally abundant intertidal decapods along the Atlantic coast of the Americas. Considering its geographical distribution and morphological plasticity, several authors postulated the existence of a P. armatus species complex. In the present study we used genetic data from the mitochondrial 16S ribosomal gene to determine the genetic variability of P. armatus from selected locations within its eastern tropical Pacific and western Atlantic distributions. Our phylogenic analysis included 49 specimens represented by 26 species of the genus Petrolisthes and 16 specimens from 10 species and 4 related genera. Genetic distances estimated among the analyzed Petrolisthes species ranged from 2.6%-22.0%; varied between 0%-5.7% for 16S. Additionally, the revision of P. armatus specimens from Pacific Costa Rica and Brazilian Waters showed no geographically significant morphological variations among the analyzed specimens. Therefore, our morphological and genetic data do not support the hypothesis of a P. armatus complex within the specimens studied herein from the Americas, but convincingly confirm the monophyly and non-separateness of the members assigned as P. armatus. http://zoolstud.sinica.edu.tw/Journals/50.3/372.pdf

Reverse Microemulsion Synthesis, Structure, and Luminescence of Nanosized REPO(4):Ln(3+) (RE = La, Y, Gd, or Yb, and Ln = Eu, Tm, or Er)

A surfactant-mediated solution route for the obtainment of nanosized rare-earth orthophosphates of different compositions (LaPO(4):Eu(3+), (Y,Gd)PO(4):Eu(3+),LaPO(4):Tm(3+), YPO(4):Tm(3+), and YbPO(4):Er(3+)) is presented, and the implications of the morphology control on the solids properties are discussed. The solids are prepared in water-in-heptane microemulsions, using cetyltrimethylammonium bromide and 1-butanol as the surfactant and cosurfactant; the alteration of the starting microemulsion composition allows the obtainment of similar to 30 nm thick nanorods with variable length. The morphology and the structure of the solids were evaluated through scanning electron microscopy and through powder X-ray diffractometry; dynamic light scattering and thermal analyses were also performed. The obtained materials were also characterized through vibrational (FTIR) and luminescence spectroscopy (emission/excitation, luminescence lifetimes, chromaticity, and quantum efficiency), where the red, blue, and upconversion emissions of the prepared phosphors were evaluated.

Glutamatergic neurotransmission mediated by NMDA receptors in the inferior colliculus can modulate haloperidol-induced catalepsy

The inferior colliculus (IC) is primarily involved in the processing of auditory information, but it is distinguished from other auditory nuclei in the brainstem by its connections with structures of the motor system. Functional evidence relating the IC to motor behavior derives from experiments showing that activation of the IC by electrical stimulation or excitatory amino acid microinjection causes freezing, escape-like behavior, and immobility. However, the nature of this immobility is still unclear. The present study examined the influence of excitatory amino acid-mediated mechanisms in the IC on the catalepsy induced by the dopamine receptor blocker haloperidol administered systemically (1 or 0.5 mg/kg) in rats. Haloperidol-induced catalepsy was challenged with prior intracollicular microinjections of glutamate NMDA receptor antagonists, MK-801 (15 or 30 mmol/0.5 mu l) and AP7 (10 or 20 nmol/0.5 mu l), or of the NMDA receptor agonist N-methyl-D-aspartate (NMDA, 20 or 30 nmol/0.5 mu l). The results showed that intracollicular microinjection of MK-801 and AP7 previous to systemic injections of haloperidol significantly attenuated the catalepsy, as indicated by a reduced latency to step down from a horizontal bar. Accordingly, intracollicular microinjection of NMDA increased the latency to step down the bar. These findings suggest that glutamate-mediated mechanisms in the neural circuits at the IC level influence haloperidol-induced catalepsy and participate in the regulation of motor activity. (C) 2010 Published by Elsevier B.V.

L-Allylglycine dissociates the neural substrates of fear in the periaqueductal gray of rats

The dorsal (dPAG) and ventral (vPAG) regions of the periaqueductal gray are well known to contain the neural substrates of fear and anxiety. Chemical or electrical stimulation of the dPAG induces freezing, followed by a robust behavioral reaction that has been considered an animal model of panic attack. In contrast, the vPAG is part of a neural system, in which immobility is the usual response to its stimulation. The defense reaction induced by the stimulation of either region is accompanied by anti nociception. Although GABAergic mechanisms are known to exert tonic inhibitory control on the neural substrates of fear in the dPAG, the role of these mechanisms in the vPAG is still unclear. The present study examined defensive behaviors and antinociception induced by microinjections of an inhibitor of gamma-aminobutyric acid synthesis, L-allylglycine (L-AG; 1, 3, and 5 mu g/0.2 mu l), into either the dPAG or vPAG of rats subjected to the open field and tail-flick tests. Passive or tense immobility was the predominant behavior after L-AG (1 or 3 mu g) microinjection into the vPAG and dPAG, respectively, which was replaced with intense hyperactivity, including jumps or rearings, after injections of a higher dose (5 mu g/0.2 mu l) into the dPAG or vPAG. Moreover, whereas intra-dPAG injection of 3 mu g L-AG produced intense antinociception, only weak antinociception was induced by intra-vPAG injections of 5 mu g L-AG. These findings suggest that GABA mechanisms are involved in the mediation of antinociception and behavioral inhibition to aversive stimulation of the vPAG and exert powerful control over the neural substrates of fear in the dPAG to prevent a full-blown defense reaction possibly associated with panic disorder. (C) 2009 Elsevier Inc. All rights reserved.

Outcomes and safety of drug provocation tests

Drug provocation tests (DPTs) are considered the gold standard for identifying adverse drug reactions (ADRs). The aim of this study was to analyze DPT results and discuss severe systemic reactions associated with them. This was a retrospective analysis of 500 patients with ADRs who sought treatment and were submitted to DPTs when indicated between 2006 and 2010. We performed DPTs according to the European Network for Drug Allergy recommendations. Single-blind, placebo-controlled DPTs were performed with antibiotics, local anesthetics, and nonsteroidal anti-inflammatory drugs, as well as with other drugs. Patient characteristics, DPT results, and reactions were analyzed. The sample comprised 198 patients (80.8% of whom were female patients) submitted to 243 DPTs. Ages ranged from 9 to 84 years (mean, 39.9 years). The 243 DPTs were performed with local anesthetics (n = 93), antibiotics (n = 19), acetaminophen (n = 44), benzydamine (n = 33), COX-2 inhibitors (n = 26), dipyrone (n = 7), aspirin (n = 4), or other drugs (n = 17). The results of 4 tests (1.6%) were inconclusive, whereas those of 10 (4.1%) revealed positive reactions to antibiotics (2/19), COX-2 inhibitors (2/26), acetaminophen (3/44), and local anesthetics (3/93). Two severe reactions were observed: cephalexin-induced anaphylactic shock and bupivacaine-induced anaphylaxis without shock. Four patients (2.0%) reacted to the placebo before administration of the drug. Drug provocation tests are safe for use in clinical practice but they should be placebo-controlled and should be performed under the supervision of an allergist. To confirm a presumptive diagnosis and to manage allergies appropriately, it is crucial to perform DPTs. (Allergy Asthma Proc 32:301-306, 2011; doi: 10.2500/aap.2011.32.3450)

Protonectin (1-6): A novel chemotactic peptide from the venom of the social wasp Agelaia pallipes pallipes

Peptides constitute the largest group of Hymenoptera venom toxins; some of them interact with GPCR, being involved with the activation of different types of leukocytes, smooth muscle contraction and neurotoxicity. Most of these toxins vary from dodecapeptides to tetradecapeptides, amidated at their C-teminal amino acid residue. The venoms of social wasps can also contains some tetra-, penta-, hexa- and hepta-peptides, but just a few of them have been structurally and functionally characterized up to now. Protonectin (ILG-TILGLLKGL-NH(2)) is a polyfunctional peptide, presenting mast cell degranulation, release of lactate dehydrogenase (LDH) from mast cells, antibiosis against Gram-positive and Gram-negative bacteria and chemotaxis for polymorphonucleated leukocytes (PMNL), while Protonectin (1-6) (ILGTIL-NH(2)) only presents chemotaxis for PMNL However, the mixture of Protonectin (1-6) with Protonectin in the molar ratio of 1:1 seems to potentiate the biological activities dependent of the membrane perturbation caused by Protonectin, as observed in the increasing of the activities of mast cell degranulation, LDH releasing from mast cells, and antibiosis. Despite both peptides are able to induce PMNL chemotaxis, the mixture of them presents a reduced activity in comparison to the individual peptides. Apparently, when mixed both peptides seems to form a supra-molecular structure, which interact with the receptors responsible for PMNL chemotaxis, disturbing their individual docking with these receptors. In addition to this, a comparison of the sequences of both peptides suggests that the sequence ILGTIL is conserved, suggesting that it must constitute a linear motif for the structural recognition by the specific receptor which induces leukocytes migration. (C) 2010 Elsevier Ltd. All rights reserved.

Real-time quantitative PCR in cerebral toxoplasmosis diagnosis of Brazilian human immunodeficiency virus-infected patients

Cerebral toxoplasmosis is the most common cerebral mass lesion in AIDS patients in Brazil, and results in high mortality and morbidity, despite free access to HAART (highly active antiretroviral treatment). Molecular diagnosis based on conventional PCR (cnPCR) or real-time quantitative PCR (qrtPCR) has been indispensable for definitive diagnosis. We report here the evaluation of qrtPCR with blood and cerebrospinal fluid (CSF) samples from AIDS patients in Brazil. This prospective study was conducted for 2 years, analysing DNA samples extracted from 149 AIDS patients (98 blood and 51 CSF samples) with confirmed clinical and radiological diagnosis The laboratory diagnosis included cnPCR (with the B22/B23 primer set) and indirect immunofluorescence (IF). For qrtPCR, two primer sets were simultaneously designed based on described genes and using a 6-carboxyfluorescein dye-labelled TaqMan MGB (minor groove binder) probe One was Bug, which amplified a sequence from the B1 gene The other was the RETg, which amplified a PCR product of the 529 bp sequence. The overall cnPCR and qrtPCR results were positive results were observed in 33.6% (50) patients The sensitivities were 98% for cnPCR (B22/B23), and 86 and 98% for qrtPCR (B1Tg and RETg, respectively). Negative reactions were observed in 66 4% patients. The specificities were 97% for cnPCR and qrtPCR (B1Tg). and 88.8% for RETg These data show that RETg PCR is highly sensitive as it amplifies a repeat region with many copies; however, its specificity is lower than the other markers However, B1Tg PCR had good specificity, but lower sensitivity Among the patients, 20 had blood and CSF collected simultaneously Thus, their results permitted us to analyse and compare molecular, serological and clinical diagnosis for a better understanding of the different scenarios of laboratorial and clinical diagnosis. For nine patients with confirmed cerebral toxoplasmosis diagnosis, four scenarios were observed: (i) and (ii) negative molecular diagnosis for CSF and positive for blood with variable IF titres for the sera and CSF (negative or positive), (iii) positive molecular diagnosis with CSF and negative with blood, and (iv) positive molecular diagnosis in both samples. In the latter two situations, normally the IF titres in sera and CSF are variable. Other opportunistic infections were shown in 11 patients Despite the IF titres in sera and CSF being variable, all of them had negative molecular diagnosis for both samples qrtPCR allows for a rapid identification of Toxoplasma gondii DNA in patient samples; in a minority of cases discrepancies occur with the cnPCR.