Therapeutic effect of adipose-derived mesenchymal stem cells in a porcine model of abdominal sepsis.

BACKGROUND: The term sepsis refers to a complex and heterogeneous syndrome. Although great progress has been made in improving the diagnosis and treatment of this condition, it continues to have a huge impact on morbidity and mortality worldwide. Mesenchymal stem cells are a population of multipotent cells that have immunomodulatory properties, anti-apoptotic effects, and antimicrobial activity. We studied these capacities in a porcine model of peritoneal sepsis. METHODS: We infused human adipose-derived mesenchymal stem cells (ADSCs) into a porcine model of peritoneal sepsis. Twenty piglets were treated with antibiotics alone (control group) or antibiotics plus peritoneal infusion of ADSCs at a concentration of 2 × 106 cells/kg or 4 × 106 cells/kg (low- and high-dose experimental groups, respectively). The animals were evaluated at different time points to determine their clinical status, biochemical and hematologic parameters, presence of inflammatory cytokines and chemokines in blood and peritoneal fluid, and finally by histologic analysis of the organs of the peritoneal cavity. RESULTS: One day after sepsis induction, all animals presented peritonitis with bacterial infection as well as elevated C-reactive protein, haptoglobin, IL-1Ra, IL-6, and IL-1b. Xenogeneic ADSC infusion did not elicit an immune response, and peritoneal administration of the treatment was safe and feasible. One day after infusion, the two experimental groups showed a superior physical condition (e.g., mobility, feeding) and a significant increase of IL-10 and TGF-ß in blood and a decrease of IL-1Ra, IL-1b, and IL-6. After 7 days, all animals treated with ADSCs had better results concerning blood biomarkers, and histopathological analysis revealed a lower degree of inflammatory cell infiltration of the organs of the peritoneal cavity. CONCLUSIONS: Intraperitoneal administration of ADSCs as an adjuvant therapy for sepsis improves the outcome and diminishes the effects of peritonitis and associated organ damage by regulating the immune system and reducing intra-abdominal adhesions in a clinically relevant porcine model of abdominal sepsis.

Distribution of the stx1 and stx2 genes in Escherichia coli isolated from milk cattle according to season, age, and production scale in southwestern region of Goiás, Brazil / Distribuição dos genes stx1 e stx2 em Escherichia coli isoladas de bovinos de leite de acordo com a estação, idade e escala de produção na região sudoeste de Goiás, Brasil

This study determined the distribution of stx1 and stx2 genes in Escherichia coli isolated from dairy herds with regard to animal age, season, and farm production-scale, and analyzed the phylogenetic distribution of the groups A, B1, B2, and D of 276 isolates of bovine feces Shiga toxin-producing E. coli (STEC). The stx1 profile was the most common, detected in 20.4% (202/990) of the isolates, followed by stx2 (4.54%, 45/990) and stx1+stx2 (2.92%, 29/990). The stx1 gene was detected more frequently in calves than in adult animals. In the dry season (winter), the presence of stx1+stx2 profile in cattle feces was higher than in the rainy season (summer), while no significant changes were observed between seasons for the stx1 and stx2 profiles. The most predominant phylogenetic groups in adult animals were B1, A, and D, while groups A and B1 prevailed in calves. Our data highlight the importance of identifying STEC reservoirs, since 7.5% of the tested isolates were positive for stx2, the main profile responsible for the hemolytic-uremic syndrome (HUS). Moreover, these microorganisms are adapted to survive even in hostile environments and can contaminate the food production chain, posing a significant risk to consumers of animal products.(AU)

Esse estudo determinou a distribuição dos genes stx1 e stx2 em Escherichia coli isolados de rebanhos leiteiros em relação a idade, estação e produção, e analisaram a distribuição filogenética dos grupos A, B1, B2 e D de 276 E. coli produtoras de toxina Shiga (STEC). O perfil stx1 foi mais comum, detectado em 20,4% (202/990) dos isolados, seguido de stx2 (4,54%, 45/990) e stx1+stx2 (2,92%, 29/990). O gene stx1 foi detectado mais frequentemente em bezerros que animais adultos. No período de seca (inverno), a presença do perfil stx1+stx2 nas fezes dos bovinos foi mais prevalente que no período chuvoso (verão), apesar de não haver diferença significativa entre estações para os perfis stx1 e stx2. Os grupos filogenéticos mais predominantes em animais adultos foram B1, A e D, enquanto grupos A e B2 prevaleceram em bezerros. Nossos dados enfatizam a importância de se detectar reservatórios de STEC já que 7,5% dos isolados testados foram positivos para stx2, o perfil mais prevalente em casos de síndrome hemolítica-urêmica. Ademais, esses microorganismos são adaptados à sobreviver em ambientes hostis e contaminam a cadeia alimentar, levando a risco significativo para consumidores de alimentos de origem animal.(AU)

Validation of EST-derived microsatellite markers for two Cerrado-endemic Campomanesia (Myrtaceae) species.

We assessed the transferability of 120 EST-derived Eucalyptus microsatellite primers to Campomanesia adamantium and C. pubescens. Both species are berry trees native to the Brazilian Cerrado, and population genetic information is poor. Twelve markers were used to analyze the genetic variability of four sampled populations. Regarding DNA extraction, we sampled leaf tissues from two populations of each species (80 individuals). Of the 120 primers evaluated, 87 did not amplify any PCR products, and 21 rendered nonspecific amplification. Twelve primers were successfully transferred, providing a low combined probability of genetic identity for both species (5.718 x 10(-10) for C. adamantium; 1.182 x 10(-11) for C. pubescens) and a high probability of paternity exclusion (0.99939 for C. adamantium; 0.99982 for C. pubescens). The average number of alleles in the polymorphic loci was 6.8 for C. adamantium and 7.8 for C. pubescens, ranging from 2 to 16 alleles per locus. The observed heterozygosity values for C. adamantium and C. pubescens were 0.504 and 0.503, respectively, and the expected heterozygosity values for C. adamantium and C. pubescens were 0.517 and 0.579, respectively. The populations exhibited structured genetic variability with qP values of 0.105 for C. adamantium and 0.249 for C. pubescens. Thus, we concluded that these 12 microsatellite markers, transferred from Eucalyptus, were efficient for population genetic studies of C. adamantium and C. pubescens.

Characterization of laminar extrudates manufactured at room temperature in the absence of solvents for the delivery of drugs.

Extrudates with laminar shape for delivery of drugs were manufactured at room temperature and without solvents. Different lipidic materials, formulations and extrusion conditions were studied, as well as the influence of the size of particles of coumarin on the release from the extrudates. Materials were mixed prior to feeding an in-house built ram extruder with a rectangular shaped die. The process of extrusion was characterized for the force at steady state whereas extrudates were characterized immediately after production and over storage for density, porosity, bending strength, elasticity, stiffness, deformation, thermal behavior and release performance of coumarin, the model drug. The variety of lipid excipients and their proportions in the formulations directly influenced the properties of the extrudates, namely the bending strength, stiffness, deformation and elasticity and, the density and porosity, which changed over storage time: in general, the bending strength, stiffness and porosity increased over time, whereas deformation, elasticy and density decreased. The thermal analysis supported these observations as there was an increase in the enthalpies of fusion of the extrudates over time. The release of coumarin from the extrudates occured both by diffusion within the extrudates and by structural alterations of the extrudates and an increase of the coumarin particle size corresponded to a decrease of the release rate. The study has proved the ability to manufacture extrudates in a continuous fashion, with laminar shape using a green technology.

Genetic diversity of gabiroba based on random amplified polymorphic DNA markers and morphological characteristics.

The fragmentation of the original vegetation of the Cerrado biome, caused by the expansion of agricultural areas, mainly in central-west Brazil, calls for an assessment of the native population of this vegetation, especially of the species of interest for domestication and sustainable use. The purpose of this study was to characterize the genetic diversity of 140 gabiroba mother plants (Campomanesia spp) and their progenies from 17 locations in Goiás. The morphological characteristics of the mother plants were evaluated, and the leaflets were collected for molecular analysis using 12 random amplified polymorphic DNA primers. The seed progenies of these matrices were transplanted to the field and morphologically evaluated. Distance matrices of the morphological data of the mother plants and progenies as well as the molecular data of the mother plants were constructed, and groups were formed using the Tocher method and the unweighted pair-group method based on arithmetic averages. The polymorphism level in the matrix was 90.44%. The greatest molecular distance (0.66) was observed between mother plants from Santa Rita do Araguaia and Alexânia. By the Tocher method, 10, 13, and 17 groups were formed. The morphological evaluation of the mother plants and progenies as well as the molecular analysis of the mother plants showed genetic diversity. Significant genetic variability was detected in the progenies of the gabiroba base collection planted in Campus Jataí, Goiás.

Vaccine delivery carriers: insights and future perspectives.

Vaccination is undoubtedly the most effective health intervention for disease prevention and eradication. Nevertheless, currently there is still a need for improving immunization coverage worldwide. A promising strategy to achieve this goal nowadays relies on the use of delivery carriers capable of inducing an effective immune response and providing improved stability, safety and cost effectiveness. This article focuses on analyzing the critical aspects in the design of these carriers, and reviewing the state of the art of currently marketed formulations and those in advanced clinical development. These vaccine delivery carriers include emulsions, liposomes and polymeric particulate carriers. Finally, particular attention is given to the evolution in the design of polymeric nanocarriers, which have been receiving increasing attention and hold promise to generate novel platforms for needle-free administration and single-dose vaccination.

Alginate films containing Lactobacillus plantarum as wound dressing for prevention of burn infection.

The topical application of lactobacilli is receiving attention for the prevention of superficial skin and burn wound infections. We studied the properties of Lactobacillus plantarum immobilized in calcium alginate films and investigated the antibacterial activity of these films in a model burn wound in rats. A multiresistant clinical isolate, VIM-2-metallo-ß-lactamase-producing Pseudomonas aeruginosa, was used as the indicator strain. Films incorporating L. plantarum at cell concentrations of 10(8)cfu/mL caused a reduction of 5-6 log(10) in P. aeruginosa in the model burn wounds. L. plantarum immobilized in freeze-dried calcium alginate films remained viable during six months of storage at 4 °C.

Comparison of ibuprofen release from minitablets and capsules containing ibuprofen: ß-cyclodextrin complex.

Mixtures containing ibuprofen (IB) complexed with ß-cyclodextrin (ßCD) obtained by two complexation methods [suspension/solution (with water removed by air stream, spray- and freeze-drying) and kneading technique] were processed into pharmaceutical dosage forms (minitablets and capsules). Powders (IB, ßCD and IBßCD) were characterized for moisture content, densities (true and bulk), angle of repose and Carr's index, X-ray and NMR. From physical mixtures and IBßCD complexes without other excipients were prepared 2.5-mm-diameter minitablets and capsules. Minitablets were characterized for the energy of compaction, tensile strength, friability, density and IB release (at pH 1.0 and 7.2), whereby capsules were characterized for IB release. The results from the release of IB were analyzed using different parameters, namely, the similarity factor (f(2)), the dissolution efficiency (DE) and the amounts released at a certain time (30, 60 and 180 min) and compared statistically (α=0.05). The release of IB from the minitablets showed no dependency on the amount of water used in the formation of the complexes. Differences were due to the compaction force used or the presence of a shell for the capsules. The differences observed were mostly due to the characteristics of the particles (dependent on the method considered on the formation of the complexes) and neither to the dosage form nor to the complex of the IB.

Oral bioavailability of silymarin phytocomplex formulated as self-emulsifying pellets.

The objective of this study was to develop new solid self-emulsifying pellets to deliver milk thistle extract (silymarin). These pellets were prepared via extrusion/spheronisation procedure, using a self-emulsifying system or SES (Akoline MCM®, Miglyol®, Tween 80®, soy lecithin and propylene glycol), microcrystalline cellulose and lactose monohydrate. To select the most suitable formulations for extrusion and spheronisation, an experimental design of experiences was adopted. The screening amongst formulations (13 different blends) was performed preparing pellets and evaluating extrusion profiles and quality of the spheronised extrudates. The pellets were characterised for size and shape, density, force required to crush them. Although more than one type of pellets demonstrated adequate morphological and technological characteristics, pellets prepared from formulation 7 revealed the best properties and were selected for further biopharmaceutical investigations, including in vitro dissolution and in vivo trials on rats to study serum and lymph levels after oral administration of the pellets. These preliminary technological and pharmacokinetic data demonstrated that extrusion/spheronisation is a viable technology to produce self-emulsifying pellets of good quality and able to improve in vivo oral bioavailability of main components of a phytotherapeutic extract of more than 100 times by enhancing the lymphatic route of absorption.

Numerical simulation of in-vitro dispersion and deposition of nanoparticles in dry-powder-inhaler aerosols.

Aerosol dispersion and deposition inside an idealized mouth-throat has been numerically simulated using a stochastic Lagrangian model accounting for Brownian motion and particle-wall interaction. Delivery of nanoparticles to the lungs is extremely difficult, mainly due to their low inertia, and for this reason they are often loaded into larger carrier particles. Bearing in mind the potentialities of nanoparticles in advanced drug delivery, a set of monodisperse particles with diameters in the nanqsize range, as well as in the respirable and carrier ranges, were considered in the present simulations. Deposition patterns were obtained by tracking a total of 16,000 particles for each diameter. The results have shown that similar patterns were obtained in the mouth-throat for 400 nm particles and larger. A clear correspondence between secondary flow structures in the fluid and these deposition patterns was observed, demonstrating the role of the convective transport processes for this size range. In contrast, a much more uniform distribution of the particles adhering to the walls was noted for a size of 200 nm. It was also found that a very large amount of these particles (nearly 80%) is lost by deposition on the mouth-throat, thus recommending the use of larger carrier particles.

The influence of the preparation methods on the inclusion of model drugs in a beta-cyclodextrin cavity.

The work aims to prove the complexation of two model drugs (ibuprofen, IB and indomethacin, IN) by beta-cyclodextrin (betaCD), and the effect of water in such a process, and makes a comparison of their complexation yields. Two methods were considered: kneading of a binary mixture of the drug, betaCD, and inclusion of either IB or IN in aqueous solutions of betaCD. In the latter method water was removed by air stream, spray-drying and freeze-drying. To prove the formation of complexes in final products, optical microscopy, UV spectroscopy, IR spectroscopy, DSC, X-ray and NMR were considered. Each powder was added to an acidic solution (pH=2) to quantify the concentration of the drug inside betaCD cavity. Other media (pH=5 and 7) were used to prove the existence of drug not complexed in each powder, as the drugs solubility increases with the pH. It was observed that complexation occurred in all powders, and that the fraction of drug inside the betaCD did not depend neither on the method of complexation nor on the processes of drying considered.

Investigation on the co-extrudability and spheronization properties of wet masses.

In the present work, preliminary results of co-extrusion and spheronization of wet masses are reported. A ram co-extruder, manufactured in-house, was designed with two concentric single dies mounted on two concentric and independent chambers. This equipment has allowed the production of three types of extrudates (rod or solid extrudates, tubular or hollow extrudates and co-extrudates). Different wet mixtures of microcrystalline cellulose, lactose, a non-soluble in water dye and water were produced and used to feed the chambers of the ram co-extruder. Extrusions of the wet masses were carried out at different speeds of the ram (25-400 mm/min). The extrudates were evaluated according to surface characteristics (by visual inspection), force of extrusion and duration of steady-state (after recording the force applied to the ram and its displacement). Simultaneously, for each process of extrusion it was possible to assess the angles of convergence to the bottom of the chambers for both the external and internal chambers. These angles reflected the high complexity of the extrusion occurring on the external chamber in consequence of its annular geometry, in which the bisecting-line was not parallel to the axis of the extruder, by opposition to the converging angle in the internal chamber, where the bisecting-line was perfectly aligned to the axis of the extruder. Variations in the amount of water in the formulations and the speed of extrusion affected both the production and the quality of the extrudates and their ability to provide pellets. The rod extrudates were the easiest to produce and the relationships between the formulations, the processing conditions and the properties of the extrudates were immediately apparent. On the other hand, co-extrudates were more complex to characterise, although identical relationships between formulations, processing conditions and the properties of the co-extrudates were observed as for the rod extrudates. Different batches of extrudates (rod, tubular and co-extrudates) were spheronized to a maximum spheronization time of 10 min at 1000 rpm. The pellets were characterized with respect to size, size distribution, sphericity and density. Results have shown that for a larger diameter of the co-extrudates, the pellets produced were bigger ( approximately 3.38 mm) than the pellets produced from rod extrudates (1.22 mm). For longer times of spheronization, the aspect ratio and the density increased for both pellets produced from rod (0.95 and 1.46 g/cm(3)) and co-extrudates (0.90 and 1.47 g/cm(3)). The study has shown the potential of this new technology in providing a product with advantages over the traditional spheres produced by extrusion and spheronization.

Myelopathy in a previously asymptomatic HIV-1-infected patient.

A wide variety of disorders of diverse pathogenic mechanisms can trigger spinal cord dysfunction in HIV-1-infected patients. The most common such condition is HIV-1-associated myelopathy (HM) which characteristically complicates advanced HIV-1 disease in patients with low CD4 cell counts and previous AIDS-defining diagnoses. We describe an unusual presentation of HM in a previously asymptomatic patient with a relatively preserved CD4 cell count (458 cells/mm3) who was even unaware of his serological status. The patient presented with a clinically severe, slowly progressive myelopathy and could not walk unassisted. Significant neurological improvement could be obtained as rapidly as within 4 weeks after the institution of an antiretroviral combination of only two nucleoside analog HIV-1 reverse transcriptase inhibitors (zidovudine and didanosine). An HIV-1 protease inhibitor was also prescribed at that point but could only be added to intensify the regimen 3 months later, when significant neurological improvement had already been recorded. We also review the disorders reported to derange spinal cord function in previously asymptomatic HIV-1-infected patients.

Macrocytosis in patients on stavudine.

We retrospectively reviewed the effects on the erythrocyte mean corpuscular volume (MCV) of the use of stavudine-including antiretroviral regimens in both zidovudine-naive and zidovudine-experienced HIV-infected patients. Macrocytosis was commonly observed among patients on stavudine-based regimens although the MCV usually stabilized at a lower level than that observed with zidovudine.

Assessment of the extrudability of three different mixtures of saturated polyglycolysed glycerides by determination of the "specific work of extrusion" and by capillary rheometry.

PURPOSE: Assessment of the extrudability of mixtures of saturated polyglycolysed glycerides used as carriers for poorly soluble drugs. METHODS: Three different mixtures of saturated polyglycolysed glycerides were extruded in a ram extruder at different rates and using different dies. The force applied to the ram and its displacement were recorded and used to determine the "specific work of extrusion" and pursuit rheometric determinations, according to the Bagley's approach. RESULTS: As the melting range of the mixtures, or the length of the dies increased, the higher the "specific work of extrusion" observed. From the rheometric analysis, viscoelastic properties were identified and decomposed into plastic and elastic components. The elastic component contributed more to the total pressure loss, and for the majority of the cases, was higher than the plastic component. From the Bagley plot it was possible to calculate the wall shear stress (sigma w), that increased with the melting range of the materials and the shear rate applied (gamma). The viscosity (eta) decreased as the shear rate increased, reflecting a non-Newtonian behavior of the materials. Finally, materials showing lower viscosities required less work to produce the extrudates. CONCLUSIONS: This study has (a) shown the possibility of preparing extrudates from different mixtures of polyglycolysed glycerides alone in a non-aqueous environment, (b) demonstrated the usefulness of the "specific work of extrusion," and (c) revealed through the rheometric studies the viscoelastic nature of the materials quantifying the contribution of the elastic and the plastic components for the total pressure loss.

Extended Wavelength Coverage of a Ce(3+):LiCAF Laser Between 223 and 243 nm by Sum Frequency Mixing in nu-Barium Borate.

We obtained coherent radiation in the spectral region between 223 and 243 nm by frequency mixing the tunable output of a solid-state Ce(3+):LiCAF laser with 1.064-mum radiation from a Nd(3+):YAG pump laser in a beta-barium borate nonlinear crystal.

Unstable Cr:LiSAF laser resonator with a variable reflectivity output coupler.

The performance of a flash-lamp-pumped Cr:LiSAF unstable laser resonator utilizing a fourth-order super-Gaussian variable reflectivity mirror as an output coupler is described. The super-Gaussian mirror results in a smooth, flattop transverse beam profile in the near field that is advantageous for nonlinear frequency-conversion applications. Long-pulse and Q-switched operation of the Cr:LiSAF unstable laser resonator are described and compared with stable resonator operation. We obtained tunable ultraviolet radiation extending from 267 to 290 nm by frequency mixing theQ-switched Cr:LiSAF laser output with lithium triborate and beta-barium borate nonlinear crystals.

Absolute Emission Spectra from Bacillus subtilis and Escherichia coli Vegetative Cells in Solution.

Spectrally resolved emission (270-560 nm) from dilute suspensions of washed Bacillus subtilis and Escherichia coli were measured by use of tunable laser excitation between 270 and 300 nm. Integrated absolute emission cross sections increase with decreasing excitation wavelength and range from 1.8 x 10(-12) to 6.0 x 10(-11) cm(2)/(particle sr). An emission band near 340 nm dominates all observed spectra. At each excitation wavelength spectrally resolved emissions from the E. coli and B. subtilis suspensions are indistinguishable.

Tm(3+):YLF laser continuously tunable between 2.20 and 2.46 microm.

Room-temperature, continuous-wave laser action at 2.3 microm corresponding to the (3)H(4)-(3)H(5) transition in Tm(3+)- doped YLF is achieved. Output powers of 200 mW and a slope efficiency of 15% have been obtained for a pump power of 2 W at 0.78 microm. In addition, continuous tunability of this laser from 2.20 to 2.46 microm is obtained.