Design of a Modern Liposome and Bee Venom Formulation for the Traditional VIT-Venom Immunotherapy

Traditional venom immunotherapy uses injections of whole bee venom in buffer or adsorbed in Al (OH)(3) in an expensive, time-consuming way. New strategies to improve the safety and efficacy of this treatment with a reduction of injections would, therefore, be of general interest. It would improve patient compliance and provide socio-economic benefits. Liposomes have a long tradition in drug delivery because they increase the therapeutic index and avoid drug degradation and secondary effects. However, bee venom melittin (Mel) and phospholipase (PLA(2)) destroy the phospholipid membranes. Our central idea was to inhibit the PLA(2) and Mel activities through histidine alkylation and or tryptophan oxidation (with pbb, para-bromo-phenacyl bromide, and/or NBSN-bromosuccinimide, respectively) to make their encapsulations possible within stabilized liposomes. We strongly believe that this formulation will be nontoxic but immunogenic. In this paper, we present the whole bee venom conformation characterization during and after chemical modification and after interaction with liposome by ultraviolet, circular dichroism, and fluorescence spectroscopies. The PLA(2) and Mel activities were, measured indirectly by changes in turbidity at 400(nm), rhodamine leak-out, and hemolysis. The native whole bee venom (BV) presented 78.06% of alpha-helical content. The alkylation (A-BV) and succynilation (S-BV) of BV increased 0.44 and 0.20% of its alpha-helical content. The double-modified venom (S-A-BV) had a 0.74% increase of alpha-helical content. The BV chemical modification induced another change on protein conformations observed by Trp that became buried with respect to the native whole BV. It was demonstrated that the liposomal membranes must contain pbb (SPC:Cho:pbb, 26:7:1) as a component to protect them from aggregation and/or fusion. The membranes containing pbb maintained the same turbidity (100%) after incubation with modified venom, in contrast with pbb-free membranes that showed a 15% size decrease. This size decrease was interpreted as membrane degradation and was corroborated by a 50% rhodamine leak-out. Another fact that confirmed our interpretation was the observed 100% inhibition of the hemolytic activity after venom modification with pbb and NBS (S-A-BV). When S-A-BV interacted with liposomes, other protein conformational changes were observed and characterized by the increase of 1.93% on S-A-BV alpha-helical content and the presence of tryptophan residues in a more hydrophobic environment. In other words, the S-A-BV interacted with liposomal membranes, but this interaction was not effective to cause aggregation, leak-out, or fusion. A stable formulation composed by S-A-BV encapsulated within liposomes composed by SPC:Cho:pbb, at a ratio of 26:7:1, was devised. Large unilamellar vesicles of 202.5 nm with a negative surface charge (-24.29 mV) encapsulated 95% of S-A-BV. This formulation can, now, be assayed on VIT.

Abundance and seasonality of Diptera (Insecta) in a poultry house in the northeast region of the state of Sao Paulo, Brazil

Modem production systems accommodate broody hens in high densities, leading to the accumulation of excrement under the cages. This substrate is excellent for the development of sinantropic flies. Thus, the accomplishment of surveys in these places becomes essential, in order to plan better strategies of control. The present work aimed at studying the entornofauna and the seasonality of the species of dipterous present in the Crisdan poultry house located in the Municipality of Sao Joao da Boa Vista, the State of Sao Paulo, Brazil. In the period of January of 2001 to December of 2002, 1,012,595 flies were captured using the ""jug-trap"". The species were identified: Drosophi-la repleta (Wollaston, 1858), Musca domestica (Linnaeus, 1758), Ophyra spp., Hennetria illucens (Linnaeus, 1758), Fannia canicularis (Linnaeus, 1761), Chrysomya megacephala (Fabricius, 1794), and Sepsidae. More frequently D. repleta and M. domestica had added 99.47% of the dipterous. Increased rainfall and the collection months influenced the sampling of dipterous (P < 0.05). Drosophila repleta was the most abundant species, representing 91% of all captured flies. However, this diptera did not develop at the surveyed site since immatures were not captured therein.

Access to the frontal sinus and zygomatico frontal suture through the supratarsal fold

Precise repositioning of a fractured zygoma is difficult. The traditional approach is through an eyebrow incision, but it can produce a scar that causes aesthetic and psychological problems for the patient. We describe the supratarsal fold approach to expose the frontozygomatic suture and to reduce small displacements of frontal sinus anterior wall; it gives good access and excellent aesthetic results. (C) 2007 The British Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

The comparative field body temperature among Liolaemus lizards: Testing the static and the labile hypotheses

Two competing hypotheses have been suggested to explain thermal sensitivity of lizards to environmental conditions. These are the static and the labile hypotheses. The static hypothesis posits that thermal physiology is evolutionary conservative and consequently relatively insensitive to directional selection. Contrarily, the labile hypothesis states that thermal physiology does respond readily to directional selection in some lizard taxa. In this paper, we tested both hypotheses among species of Liolaemus lizards. The genus Liolaemus is diverse with about 200 species, being broadly distributed from central Peru to Tierra del Fuego at the southern end of South America. Data of field body temperature (T(b)) from Liolaemus species were collected from the literature. Based on the distributional range of the species we also collected data of mean annual ambient temperatures. We observed that both the traditional analysis and the phylogenetic approach indicate that in the genus Liolaemus T(b) of species varies in a manner that is consistent with ecological gradient of ambient temperature. The data suggest that the thermal physiology of Liolaemus lizards is evolutionarily flexible, and that this plasticity has been partially responsible for the colonization of a wide array of thermal environments. (C) 2009 Elsevier Ltd. All rights reserved.

The timing of Neotropical speciation dynamics: A reconstruction of Myiopagis flycatcher diversification using phylogenetic and paleogeographic data

Neotropical forests have brought forth a large proportion of the world`s terrestrial biodiversity, but the underlying evolutionary mechanisms and their timing require further elucidation. Despite insights gained from phylogenetic studies, uncertainties about molecular clock rates have hindered efforts to determine the timing of diversification processes. Moreover, most molecular research has been detached from the extensive body of data on Neotropical geology and paleogeography. We here examine phylogenetic relationships and the timing of speciation events in a Neotropical flycatcher genus (Myiopagis) by using calibrations from modern geologic data in conjunction with a number of recently developed DNA sequence dating algorithms and by comparing these estimates with those based on a range of previously proposed molecular clock rates. We present a well-supported hypothesis of systematic relationships within the genus. Our age estimates of Myiopagis speciation events based on paleogeographic data are in close agreement with nodal ages derived from a ""traditional"" avian mitochondrial 2%/My clock, while contradicting other clock rates. Our comparative approach corroborates the consistency of the traditional avian mitochondrial clock rate of 2%/My for tyrant-flycatchers. Nevertheless, our results argue against the indiscriminate use of molecular clock rates in evolutionary research and advocate the verification of the appropriateness of the traditional clock rate by means of independent calibrations in individual studies. (C) 2009 Elsevier Inc. All rights reserved.

A new proposal for the picture changing operators in the minimal pure spinor formalism

Using a new proposal for the ""picture lowering"" operators, we compute the tree level scattering amplitude in the minimal pure spinor formalism by performing the integration over the pure spinor space as a multidimensional Cauchy-type integral. The amplitude will be written in terms of the projective pure spinor variables, which turns out to be useful to relate rigorously the minimal and non-minimal versions of the pure spinor formalism. The natural language for relating these formalisms is the. Cech-Dolbeault isomorphism. Moreover, the Dolbeault cocycle corresponding to the tree-level scattering amplitude must be evaluated in SO(10)/SU(5) instead of the whole pure spinor space, which means that the origin is removed from this space. Also, the. Cech-Dolbeault language plays a key role for proving the invariance of the scattering amplitude under BRST, Lorentz and supersymmetry transformations, as well as the decoupling of unphysical states. We also relate the Green`s function for the massless scalar field in ten dimensions to the tree-level scattering amplitude and comment about the scattering amplitude at higher orders. In contrast with the traditional picture lowering operators, with our new proposal the tree level scattering amplitude is independent of the constant spinors introduced to define them and the BRST exact terms decouple without integrating over these constant spinors.