Descrição de uma nova espécie de Rhodnius (Hemiptera, Reduviidae, Triatominae) da Amazônia

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Avaliação toxicogenética e atividade antitumoral in vitro de complexos heterolépticos de Rutênio(II): Atividades citotóxicas, genotóxicas e interação com biomoléculas

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Estudo do nível de infecção por Babesia bovis e Babesia bigemina em bovinos da raça Canchim naturalmente infestados com o carrapato Rhipicephalus (Boophilus) microplus

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Cytochrome c-promoted cardiolipin oxidation generates singlet molecular oxygen

The interaction of cytochrome c (cyt c) with cardiolipin (CL) induces protein conformational changes that favor peroxidase activity. This process has been correlated with CL oxidation and the induction of cell death. Here we report evidence demonstrating the generation of singlet molecular oxygen [O-2((1)Delta(g))] by a cyt c-CL complex in a model membrane containing CL. The formation of singlet oxygen was directly evidenced by luminescence measurements at 1270 nm and by chemical trapping experiments. Singlet oxygen generation required cyt c-CL binding and occurred at pH values higher than 6, consistent with lipid-protein interactions involving fully deprotonated CL species and positively charged residues in the protein. Moreover, singlet oxygen formation was specifically observed for tetralinoleoyl CL species and was not observed with monounsaturated and saturated CL species. Our results show that there are at least two mechanisms leading to singlet oxygen formation: one with fast kinetics involving the generation of singlet oxygen directly from CL hydroperoxide decomposition and the other involving CL oxidation. The contribution of the first mechanism was clearly evidenced by the detection of labeled singlet oxygen [O-18(2)((1)Delta(g))] from liposomes supplemented with 18-oxygen-labeled CL hydroperoxides. However quantitative analysis showed that singlet oxygen yield from CL hydroperoxides was minor (<5%) and that most of the singlet oxygen is formed from the second mechanism. Based on these data and previous findings we propose a mechanism of singlet oxygen generation through reactions involving peroxyl radicals (Russell mechanism) and excited triplet carbonyl intermediates (energy transfer mechanism).

A relict new species of Oreobates (Anura, Strabomantidae) from the Seasonally Dry Tropical Forests of Minas Gerais, Brazil, and its implication to the biogeography of the genus and that of South American Dry Forests

A new species of Oreobates is described from Cavernas do Peruacu National Park, Januaria, Minas Gerais state, in the Atlantic Dry Forests of Brazil. The new species is distinguished from all other Oreobates by having the following combination of characters: large tympanum, discs broadly enlarged and truncate on Fingers III and IV, smooth dorsal skin, nuptial pads absent, snout subacuminate, and a very short pulsatile (2-3 pulses) single-noted advertisement call with dominant frequency of about 3150 Hz, and no harmonic structure. Molecular phylogenetic analyses using partial sequences of the mitochondrial genes cytochrome b (cyt b) and 16S using multiple outgroups recovered the new species within Oreobates and sister to O. heterodactylus. The latter species inhabits the Dry Forests of Mato Grosso (Cerrado) and Bolivia (Chiquitano forests), and is strictly associated to these habitats, which suggests a preterit connection between Chiquitano and Atlantic Dry Forests. The discovery of a new Oreobates in the Atlantic Dry Forest is of great importance for the conservation of these dry forests, as it is known only from this type of habitat.

Evolutionary Insights from Bat Trypanosomes: Morphological, Developmental and Phylogenetic Evidence of a New Species, Trypanosoma (Schizotrypanum) erneyi sp nov., in African Bats Closely Related to Trypanosoma (Schizotrypanum) cruzi and Allied Species

Parasites of the genus Trypanosoma are common in bats and those of the subgenus Schizotrypanum are restricted to bats throughout the world, with the exception of Trypanosoma (Schizotrypanum) cruzi that also infects other mammals and is restricted to the American Continent. We have characterized trypanosome isolates from Molossidae bats captured in Mozambique, Africa. Morphology and behaviour in culture, supported by phylogenetic inferences using SSU (small subunit) rRNA, gGAPDH (glycosomal glyceraldehyde 3-phosphate dehydrogenase) and Cyt b (cytochrome b) genes, allowed to classify the isolates as a new Schizotrypanum species named Trypanosoma (Schizotrypanum) erneyi sp. nov. This is the first report of a Schizotrypanum species from African bats cultured, characterized morphologically and biologically, and positioned in phylogenetic trees. The unprecedented finding of a new species of the subgenus Schizotrypanum from Africa that is closest related to the America-restricted Trypanosoma (Schizotrypanum) cruzi marinkellei and T. cruzi provides new insights into the origin and evolutionary history of T. cruzi and closely related bat trypanosomes. Altogether, data from our study support the hypothesis of an ancestor trypanosome parasite of bats evolving to infect other mammals, even humans, and adapted to transmission by triatomine bugs in the evolutionary history of T. cruzi in the New World. (c) 2012 Elsevier GmbH. All rights reserved.

Genotoxic and mutagenic effects of erythrosine B, a xanthene food dye, on HepG2 cells

Erythrosine (ErB) is a xanthene and an US Food and Drug Administration approved dye used in foods, drugs and cosmetics. Although its utilization is permitted, ErB is described as inhibitor of enzymes and protein-protein interactions and is toxic to pituitary and spermatogenesis processes. However, the genotoxicity and mutagenicity of ErB is inconclusive in the literature. This study aimed to analyze the genotoxicity of this dye using the alkaline comet assay and is the first investigation to evaluate ErB mutagenicity using the cytokinesis block micronucleus cytome (CBMN-Cyt) assay in HepG2 cells. These cells were chosen because they produce phase I and phase II enzymes that can mimic in vivo metabolism. The cells were treated with seven concentrations (0.1-70.0 mu g mL(-1)) of ErB, and the results showed genotoxicity at the two highest concentrations and mutagenicity at six concentrations. Furthermore, as micronuclei result from clastogenic and aneugenic processes, while comet assay is often considered more sensitive and detects DNA single strain breaks, we suggest that an aneugenic is responsible for the observed damage. Although ErB is approved for use in the food, cosmetic and pharmaceutical industries, it must be used carefully because it damages the DNA structure. (C) 2012 Elsevier Ltd. All rights reserved.

Description of Rhodnius montenegrensis n. sp (Hemiptera: Reduviidae: Triatominae) from the state of Rondonia, Brazil

We present here a multisource approach that takes advantage of several disciplines to address a taxonomic issue. A triatomine related to Rhodnius robustus Larrousse, 1927 was recently found in the state of Rondonia, Brazil. The name Rhodnius montenegrensis n. sp. is suggested because it was found in the municipality of Monte Negro. The main differences between these two species can be detected in the female and male genitalia, but there are also noticeable differences in their eggs. Molecular analysis using PCR-RFLP technique and Bayesian inferences based on a fragment of the Cytochrome b (Cyt b) gene corroborated the morphological findings. We used this integrative approach to address the taxonomic decision for a new Rhodnius species and its relationship with others of this genus. Results obtained herein stress that morphology must be used as the major approach for obtaining phenotypic information, and molecular data should be taken as a complementary tool.

In-vitro synthesis and reconstitution of cytochrome bo 3 ubiquinol oxidase in artificial membranes

Forschung über Membranenproteine stellt strenge Hindernisse, seit ruhigem gerade wenige Beispiele der Membranenproteinsorten sind gekennzeichnet worden in den verwendbaren experimentellen Plattformen gegenüber. Die Hauptherausforderung ist, ihre ausgezeichnete entworfene strukturelle Vollständigkeit zu konservieren, während die Ausdruck-, Lokalisierungs- und Wiederherstellungprozesse auftreten. In-vitro übersetzungssysteme können Vorteile über auf Zellenbasisgenausdruck zum Beispiel haben, wenn das über-ausgedrückte Produkt zur Wirtszelle giftig ist oder wenn fehlende Pfosten-Übersetzungsänderung in den bakteriellen Ausdrucksystemen die Funktionalität der Säugetier- Proteine oder Mangel an vorhandenem Membranenraum verdirbt, Funktionsausdruck verbieten.rn Der Nachahmer von biologische Membranen wie feste gestützte Lipidmembranen sind als Plattform am meisten benutzt, Proteinmembraneninteraktionen nachzuforschen. Wir sind in der Lage, Membranenproteinsorte, da wir eine Plattform für Membranenproteinsynthese vorstellen, nämlich die in-vitrosynthese der Membranenproteine in ein Peptid gestütztes Membranensystem zu adressieren. Die Wiederherstellung der Membranenproteine in den Lipid bilayers resultiert im Allgemeinen mit verschiedenen Proteinanpassungen. Als Alternative erforschen wir dieses System zum ersten Mal, um genaueres Modell zu den zellularen Membranen zu verursachen und ihre Funktion, wie Proteineinfügung, Proteinfunktion und Ligandinteraktionen nachzuahmen.rn In dieser Arbeit ist unser Ziel, komplizierte Transmembraneproteine, wie des Cytochrome bo3-ubiquinol Oxydase (Cyt-bo3) direkt innerhalb der biomimetic vorbildlichen Membrane zu synthetisieren. In unserem System wird festes gestütztes tBLM wie, P19/DMPE/PC als Plattform benutzt. Dieses künstliche Membranensystem mimiks die amphiphile Architektur eines Zelle-abgeleiteten Membranensystems.rn

Characterization of the cytochrome P-450 drug metabolism system of LS174T human colon tumor cell line

The human colon tumor cell line, LS174T, has been shown to have four major components of the drug metabolizing system; cytochrome b$\sb5$ reductase, cytochrome b$\sb5$, cytochrome P450 reductase and cytochrome P450, by activity measurements, spectral studies and antibody cross-reactivity. Cytochrome P450IA1 is induced by benzanthracene in these cells as shown by activity with the specific substrate, ethoxyresorufin, cross-reactivity with rabbit antibodies to rat IA1, and by a hybridizing band on a Northern blot to a rat IA1 probe.^ Further, this system has proven responsive to various inducers and conditions of growth. The enzyme activities were found stable over limited cell passages with control values of 0.03 and 0.13 $\mu$mol/min/mg protein for NADPH and NADH cytochrome c (cyt c) reducing activity, 0.05 nmol cyt b$\sb5$ per milligram and 0.013 nmol cytochrome P450 per milligram of microsomal protein. Phenobarbital/hydrocortisone treatment showed a consistent, but not always significant increase in the NADPH and NADH cyt c reducing activity and benzanthracene treatment an increase in the NADH cyt c reducing activity. Delta-aminolevulinic acid (0.5mM) caused a significant decrease in the specific activity of all enzyme contents and activities tested.^ Finally, the cytochrome b$\sb5$ to cytochrome P450, by the coordinate induction of the cytochrome b$\sb5$ pathway by P450 inducers, by the high ratio of NADH to NADPH ethoxycoumarin deethylase activity in uninduced cell microsomes, and by the increase in NADH and NADPH ethoxycoumarin deethylase activity when the microsomes were treated with potassium cyanide, a desaturase inhibitor. ^

CYTOCHROME P450 MEDIATED REDUCTIVE DEHALOGENATION OF HEXACHLOROBENZENE

The role of the cytochrome (CYT) P-450 mixed-function oxidase (MFO) in the biotransformation of hexachlorobenzene (HCB) was investigated, since in vivo interaction between this enzyme and chemical is very probable. HCB is a type I substrate with (Fe('3+)) CYT P-450 isozymes present in untreated, b-naphthoflavone (BNF) and phenobarbital (PB) induced rat liver microsomes. HCB dependent and saturable type I binding titrations yield spectral dissociation constants (K(,s)) of 180 and 83 uM for the isozymes present in untreated and PB induced microsomes, respectively. Purified CYT P-450b, the major isozyme induced by PB, produces HCB dependent and saturable type I spectra with a K(,s) of 0.38 uM.^ CYT P-450 mediated reductive dehalogenation occurs in microsomes and purified/reconstituted MFO systems and produces pentachlorobenzene (PCB) as the initial and major metabolite under both aerobic and anaerobic conditions. In microsomal reactions secondary metabolism of PCB occurs in the presence of oxygen. Pentachlorophenol (PCP) is produced only in aerobic reactions with PB induced microsomes with a concomitant decrease in PCB production. PCP is not detected in aerobic reactions with BNF induced microsomes, although PCB production is decreased compared to anaerobic conditions. A reaction scheme for the production of phenolic metabolities from PCB is deduced.^ CYT P-450 dependent and NADPH independent modes of PCB production occur with purified/reconstituted MFO systems and are consistent with dehalogenation pathways observed with microsomal experiments. The NADPH independent production of PCB requires native microsomal or purified MFO protein components and may be the result of nucleophilic displacement of a chlorine atom from HCB mediated or coupled with redox active functions (primary, secondary, tertiary and quarternary structures) of the proteins. CYT P-450 dependent production of PCB from HCB is isozyme dependent: CYT P-450c = CYT P-450d > CYT P-450a > CYT 450b. The low apparent specific activity may be due to non-optimal reconstitution conditions (e.g., isozyme choice and requirement of other microsomal elecron transport components) and secondary metabolism of PCB and the phenols derived from PCB. CYT P-450 mediated dehalogenation may be catalyzed through attack, by the iron oxene (postulated intermediate of CYT P-450 monooxygenations), at the chlorines of HCB instead of the aromatic nucleus. (Abstract shortened with permission of author.) ^

Mechanism of molecular regulation of nuclear -encoded mitochondrial gene expression by electrical stimulation in the cultured neonatal rat cardiac myocytes

To answer the question whether increased energy demand resulting from myocyte hypertrophy and enhanced $\beta$-myosin heavy chain mRNA, contractile protein synthesis and assembly leads to mitochondrial proliferation and differentiation, we set up an electrical stimulation model of cultured neonatal rat cardiac myocytes. We describe, as a result of increased contractile activity, increased mitochondrial profiles, cytochrome oxidase mRNA, and activity, as well as a switch in mitochondrial carnitine palmitoyltransferase-I (CPT-I) from the liver to muscle isoform. We investigate physiological pathways that lead to accumulation of gene transcripts for nuclear encoded mitochondrial proteins in the heart. Cardiomyocytes were stimulated for varying times up to 72 hr in serum-free culture. The mRNA contents for genes associated with transcriptional activation (c-fos, c-jun, junB, nuclear respiratory factor 1 (Nrf-1)), mitochondrial proliferation (cytochrome c (Cyt c), cytochrome oxidase), and mitochondrial differentiation (carnitine palmitonyltransferase I (CPT-I) isoforms) were measured. The results establish a temporal pattern of mRNA induction beginning with c-fos (0.25-3 hr) and followed by c-jun (0.5-3 hr), junB (0.5-6 hr), NRF-1 (1-12 hr), Cyt c (12-72 hr), cytochrome c oxidase (12-72 hr). Induction of the latter was accompanied by a marked decrease in the liver-specific CPT-I mRNA. Electrical stimulation increased c-fos, $\beta$-myosin heavy chain, and Cyt c promoter activities. These increases coincided with a rise in their respective endogenous gene transcripts. NRF-1, cAMP response element (CRE), and Sp-1 site mutations within the Cyt c promoter reduced luciferase expression in both stimulated and nonstimulated myocytes. Mutations in the Nrf-1 and CRE sites inhibited the induction by electrical stimulation or by transfection of c-jun into non-paced cardiac myocytes whereas mutation of the Sp-1 site maintained or increased the fold induction. This is consistent with the appearance of NRF-1 and fos/jun mRNAs prior to that of Cyt c. Overexpression of c-jun by transfection also activates the Nrf-1 and Cyt c mRNA sequentially. Electrical stimulation of cardiac myocytes activates the c-Jun-N-terminal kinase so that the fold-activation of the cyt c promoter is increased by pacing when either c-jun or c-fos/c-jun are cotransfected. We have identified physical association of Nrf-1 protein with the Nrf-1 enhancer element and of c-Jun with the CRE binding sites on the Cyt c promoter. This is the first demonstration that induction of Nrf-1 and c-Jun by pacing of cardiac myocytes directly mediates Cyt c gene expression and mitochondrial proliferation in response to hypertrophic stimuli in the heart.^ Subsequent to gene activation pathways that lead to mitochondrial proliferation, we observed an isoform switch in CPT-I from the liver to muscle mRNA. We have found that the half-life for the muscle CPT-I is not affected by electrical stimulation, but electrical decrease the T1/2 in the liver CPT-I by greater than 50%. This suggests that the liver CPT-I switch to muscle isoform is due to (1) a decrease in T1/2 of liver CPT-I and (2) activation of muscle CPT-Itranscripts by electrical stimulation. (Abstract shortened by UMI.) ^

Una mirada multinivel de la cooperación científica : La colaboración de Argentina con la Unión Europea en los niveles formal, instrumental y de las prácticas

Este trabajo tiene por objeto explorar las relaciones existentes entre diferentes formas de expresión de la cooperación científica entre Argentina y la Unión Europea (UE). Por un lado, los tratados bilaterales firmados con la región (nivel formal); por otro, los Programas Marco -PM- (nivel instrumental), y por otro, las publicaciones coautoradas (nivel de las prácticas). Los resultados revelan que la cooperación científica de Argentina con la UE se sustenta en relaciones establecidas en los tres niveles de análisis. La UE es la región con mayor cantidad de tratados firmados con Argentina, además de haberlo hecho más tempranamente que cualquier otra región o país del mundo. Los países de la UE que impulsaron los tratados (Alemania, Italia, Francia y España) son junto con Inglaterra los de mayor presencia en los PM y en las coautorías, además de que con ellos Argentina alcanza el mayor potencial investigador respecto del resto de los países de la Unión. Sin embargo no hay una relación directa entre los Tratados CyT y la participación argentina en los PM y en las coautorías. Estos niveles de análisis responden a lógicas diferentes. Mientras los primeros cumplen una función ornamental y expresan la intención de acercamiento entre los países, los PM y las coautorías responden a una lógica propia del campo científico determinada por la búsqueda de la excelencia

Una mirada multinivel de la cooperación científica : La colaboración de Argentina con la Unión Europea en los niveles formal, instrumental y de las prácticas

Este trabajo tiene por objeto explorar las relaciones existentes entre diferentes formas de expresión de la cooperación científica entre Argentina y la Unión Europea (UE). Por un lado, los tratados bilaterales firmados con la región (nivel formal); por otro, los Programas Marco -PM- (nivel instrumental), y por otro, las publicaciones coautoradas (nivel de las prácticas). Los resultados revelan que la cooperación científica de Argentina con la UE se sustenta en relaciones establecidas en los tres niveles de análisis. La UE es la región con mayor cantidad de tratados firmados con Argentina, además de haberlo hecho más tempranamente que cualquier otra región o país del mundo. Los países de la UE que impulsaron los tratados (Alemania, Italia, Francia y España) son junto con Inglaterra los de mayor presencia en los PM y en las coautorías, además de que con ellos Argentina alcanza el mayor potencial investigador respecto del resto de los países de la Unión. Sin embargo no hay una relación directa entre los Tratados CyT y la participación argentina en los PM y en las coautorías. Estos niveles de análisis responden a lógicas diferentes. Mientras los primeros cumplen una función ornamental y expresan la intención de acercamiento entre los países, los PM y las coautorías responden a una lógica propia del campo científico determinada por la búsqueda de la excelencia

Una mirada multinivel de la cooperación científica : La colaboración de Argentina con la Unión Europea en los niveles formal, instrumental y de las prácticas

Este trabajo tiene por objeto explorar las relaciones existentes entre diferentes formas de expresión de la cooperación científica entre Argentina y la Unión Europea (UE). Por un lado, los tratados bilaterales firmados con la región (nivel formal); por otro, los Programas Marco -PM- (nivel instrumental), y por otro, las publicaciones coautoradas (nivel de las prácticas). Los resultados revelan que la cooperación científica de Argentina con la UE se sustenta en relaciones establecidas en los tres niveles de análisis. La UE es la región con mayor cantidad de tratados firmados con Argentina, además de haberlo hecho más tempranamente que cualquier otra región o país del mundo. Los países de la UE que impulsaron los tratados (Alemania, Italia, Francia y España) son junto con Inglaterra los de mayor presencia en los PM y en las coautorías, además de que con ellos Argentina alcanza el mayor potencial investigador respecto del resto de los países de la Unión. Sin embargo no hay una relación directa entre los Tratados CyT y la participación argentina en los PM y en las coautorías. Estos niveles de análisis responden a lógicas diferentes. Mientras los primeros cumplen una función ornamental y expresan la intención de acercamiento entre los países, los PM y las coautorías responden a una lógica propia del campo científico determinada por la búsqueda de la excelencia