Surfactant vesicle-mediated delivery of DNA vaccines via the subcutaneous route

Compared to naked DNA immunisation, entrapment of plasmid-based DNA vaccines into liposomes by the dehydration-rehydration method has shown to enhance both humoural and cell-mediated immune responses to encoded antigens administered by a variety of routes. In this paper we have compared the potency of lipid-based and non-ionic surfactant based vesicle carrier systems for DNA vaccines after subcutaneous immunisation. Plasmid pI.18Sfi/NP containing the nucleoprotein (NP) gene of A/Sichuan/2/87 (H3N2) influenza virus in the pI.18 expression vector was incorporated by the dehydration-rehydration method into various vesicle formulations. The DRV method, entailing mixing of small unilamellar vesicles (SUV) with DNA, followed by dehydration and rehydration, yielded high DNA vaccine incorporation values (85-97% of the DNA used) in all formulations. Studies on vesicle size revealed lipid-based systems formed cationic submicron size vesicles whilst constructs containing a non-ionic surfactant had significantly large z-average diameters (>1500 nm). Subcutaneous vesicle-mediated DNA immunisation employing two DRV(DNA) formulations as well as naked DNA revealed that humoural responses (immunoglobulin total IgG, and subclasses IgG 1 and 1gG 2a) engendered by the plasmid encoded nucleoprotein were substantially higher after dosing twice, 28 days apart with 10 μg DRV-entrapped DNA compared to naked DNA. Comparison between the lipid and non-ionic based vesicle formulations revealed no significant difference in stimulated antibody production. These results suggest that, not only can DNA be effectively entrapped within a range of lipid and non-ionic based vesicle formulations using the DRV method but that such DRV vesicles containing DNA may be a useful system for subcutaneous delivery of DNA vaccines. © 2004 Elsevier B.V. All rights reserved.

Particulate delivery systems for vaccines

This review focuses on the use of particulate delivery systems for the purposes of immunization. This includes poly(lactide-co-glycolide) (PLGA), ISCOMs, liposomes, niosomes, virosomes, chitosan, and other biodegradable polymers. These systems are evaluated in terms of their use as carriers for protein subunit and DNA vaccines. There is an extensive focus on recent literature, the understanding of biological interactions, and relation of this to our present understanding of immunological mechanisms of action. In addition, there is consideration of formulation techniques including emulsification, solvent diffusion, DNA complexation, and entrapment. The diversity of formulation strategies presented is a testament to the exponential growth and interest in the area of vaccine delivery systems. A case study for the application of particulate vaccine carriers is assessed in terms of vaccine development and recent insights into the possible design and application of vaccines against two of the most important pathogens that threaten mankind and for which there is a significant need: Mycobacterium tuberculosis and human immunodeficiency virus. This review addresses the rationale for the use of particulate delivery systems in vaccine design in the context of the diversity of carriers for DNA- and protein-based vaccines and their potential for application in terms of the critical need for effective vaccines. © 2005 by Begell House, Inc.

3D culture of bone-derived cells immobilised in alginate following light-triggered gelation

Photoreactive liposomes have been exploited as a means of developing 3D tissue constructs. Liposomes formulated using the photosensitive lipid 1,2-bis(4-(n-butyl)phenylazo-4′-phenylbutyroyl)phosphatidylcholine (Bis Azo PC), which undergoes conformational change on stimulation with long wavelength ultraviolet light, were prepared with entrapped CaCl2 before being incorporated into a 4% alginate solution. It was shown that stimulation of the photosensitive lipid using a light emitting diode (LED) (peak emission at 385 nm, dose equivalent to 9 mJ/cm2) caused the release of liposome-entrapped CaCl2, resulting in cross-linking of the alginate solution and immobilisation of bone-derived cells over a range of seeding densities, approximately 97% of which remained viable for periods of up to 14 days in culture. Entrapment volumes of a variety of liposome types were evaluated and interdigitating fusion vesicles were identified as having the highest payload (24%), however the inclusion of cholesterol as a means of shifting Bis Azo PC sensitivity into the visible light wavelengths resulted in an approximately 10-fold reduction in calcium entrapment. This application of light-sensitised liposomes offers the potential to create complex tissue engineering substrates containing cells immobilised in precise locations, in contrast with substrates onto which cells are seeded post-production. © 2007 Elsevier B.V. All rights reserved.

FITC-lectin avidity of Cryptococcus neoformans cell wall and capsular components

Flow cytometry and confocal microscopy were used to quantify and visualize FITC-lectin binding to cell-surface carbohydrate ligands of log and stationary phase acapsular and capsular Cryptococcus neoformans strains. Cell populations demonstrated marked avidity for terminal a-linked mannose and glucose specific FITC-Con A, mannose specific FITC-GNL, as well as N-acetylglucosamine specific FITC-WGA. Exposure to other FITC-lectins specific for mannose, fucose and N-acetylgalactosamine resulted in little cell-surface fluorescence. The nature of cell-surface carbohydrates was investigated further by measurement of the fluorescence from surfaces of log and stationary phase cell populations after exposing them to increasing concentrations of FITC-Con A and FITC-WGA. Cell fluorescence increased significantly with small increases in FITC-Con A and FITC-WGA concentrations attaining reproducible maxima. Measurements of this nature supported calculation of the lectin binding determinants EC 50, Hn, Fmax and relative Bmax values. EC50 values indicated that the yeast-cell surfaces had greatest affinity for FITC-WGA, however, relative Bmax values indicated that greater numbers of Con A binding sites were present on these same cell surfaces. Hn values suggested a co-operative lectin-carbohydrate ligand interaction. Imaging of FITC-Con A and FITC-WGA cell-surface fluorescence by confocal microscopy demonstrated marked localization of both lectins to cell surfaces associated with cell division and maturation, indicative of dynamic carbohydrate ligand exposure and masking. Some fluorescence was associated with entrapment of FITC-Con A by capsular components, but FITC-Con A and FITC-WGA readily penetrated the capsule matrix to bind to the same cell surfaces labelled in acapsular cells.

Development of photosensitive liposomes for the controlled release of drugs

The facility to controlled triggered release from a “cage” system remains a key requirement for novel drug delivery. Earlier studies have shown that Bis-Azo PC based photosensitive liposomes are beneficial for drug delivery. Thus, the aim of this project was to develop photosensitive liposomes that can be used for the controlled release of drugs through UV irradiation, particularly therapeutic agents for the treatment of psoriasis. Bis-Azo PC was successfully synthesized and incorporated into a range of liposomal formulations, and these liposomes were applied for the controlled release of BSA-FITC. Bis-Azo PC sensitized liposomes were prepared via interdigitation fusion method. IFV containing optimum cholesterol amount in terms of protein loading, stability and photo-trigger release of protein was investigated. Further studies investigated the stability and triggered release of the HMT from IFV. Finally, permeation behavior of HMT and HMT-entrapped IFV through rat skin was examined using Franz cell. Results from protein study indicated that the stable entrapment of the model protein was feasible as shown through fluorescence spectroscopy and maximum of 84% protein release from IFV after 12 min of UV irradiation. Moreover, stability studies indicated that IFV were more stable at 4 0C as compared to 25 0C. Hence, DPPC:Chol:Bis-Azo PC (16:2:1) based IFV was chosen for the controlled release of HMT and these studies exhibited that photo-trigger release and stability data of HMT-entrapped IFV are in line with the protein results. Franz cell work inferred that HMT-entrapped IFV attributed to slower skin permeation as compared to HMT. CLSM also demonstrated that HMT can be used as a fluorescent label for the in vitro skin study. Overall, the work highlighted in this thesis has given useful insight into the potentials of Bis-Azo PC based IFV as a promising carrier for the treatment of psoriasis.

Spic'ing into Existence: Epitaph, Epithet, and the Ethnopoetic Imagination

Quién Es, Quién Somos? Spic’ing into Existence claims a four-fold close-reading: first, analysis of texts: from theoretical meditations to (prison) memoir and film. Second, a half dozen central figures appear, largely Latinx and black American. They cut across a score of registers, socio-economics, ideological reservations, but all are, as Carl Carlton sang, poetry in motion. Writers, poets, theologians, pathologists, artists, comedians, actors, students whose vocation is invocation, the inner surge of their calling. Third, the manuscript draws from a series of historical moments—from radical liberation of the late 60s, to contemporary student activism. Finally, this body of work is movement, in all its social, gestural, and kinesthetic viscera. From this last heading, we peel away layers of what I call the ethnopoet, the fascia undoing that reveals its bio-political anatomy, dressing its bare life with kinship speech. First, the social revolutions of the Civil Rights, Black Power, abolitionism, the Black Panthers and Young Lords, boycotts and jarring artistic performances. These events are superficial not in vain sense, but key epicenters of underground murmurings, the workings of a cunning assailant. She robs not lavish estates, but another day to breathe. Gesturally, as perhaps the interlocutor, lies this author, interspersing his own diatribes to conjure her presence. The final branch is admittedly the most intangible. Kinesthetically, we map the nimbleness, footwork lígera of what I call the ethnopoet. Ethnopoet is no mere aggregate of ethnicity and poetry, but like chemical reaction, the descriptor for its behavior under certain pressures, temperatures, and elements. Elusive and resisting confinement, and therefore definition, the ethnopoet is a shapeshifting figure of how racialized bodies [people of color] respond to hegemonic powers. She is, at bottom, however, a native translator, the plural-lensed subject whose loyalty is only to the imagination of a different world, one whose survival is not contingent upon her exploitation. The native translator’s constant re-calibrations of oppressive power apparatuses seem taxing at best, and near-impossible, at worst. To effectively navigate through these polarized loci, she must identify ideologies that in turn seek “affective liberatory sances” in relation to the dominant social order (43). In a kind of performative contradiction, she must marshall the knowledge necessary to “break with ideology” while speaking within it. Chicana Studies scholar, Chela Sandoval, describes this dual movement as “meta-ideologizing”: the appropriation of hegemonic ideological forms in order to transform them (82). Nuestros padres se subieron encima de La Bestia, y por eso somos pasageros a ese tren. Y ya, dentro su pansa, tenemos que ser vigilantes cuando plantamos las bombas. In Methodology of the Oppressed, Sandoval schematizes this oppositional consciousness around five principle categories: “equal rights,” “revolutionary,” “supremacist,” “separatist,” and “differential.” Taken by themselves, the first four modes appear mutually exclusive, incapable of occupying the same plane, until a fifth pillar emerges. Cinematographic in nature, differential consciousness, as Sandoval defines it, is “a kinetic motion that maneuvers, poetically transfigures, and orchestrates while demanding alienation, perversion, and reformation in both spectators and practitioners” (44). For Sandoval, then, differential consciousness is a methodology that privileges an incredible sense mobility, one reaching artistic sensibilities. Our fourth and final analytic of movement serves an apt example of this dual meaning. Lexically speaking, ‘movement’ may be regarded as a political mobilization of aggrieved populations (through sustained efforts), or the process of moving objects (people or otherwise) from one location to another. Praxis-wise, it is both action and ideal, content and form. Thus, an ethnic poetics must be regarded less as a series of stanzas, shortened lyric, or even arrangement of language, but as a lens through which peripheralized peoples kaleidecope ideological positions in an “original, eccentric, and queer sight” (43). Taking note of the advantages of postponing identifications, the thesis stands its ground on the term ethnopoet. Its abstraction is not dewey-eyed philosophy, but an anticipation of poetic justice, of what’s to come from callused hands. This thesis is divided into 7.5 chapters. The first maps out the ethnopoet’s cartographies of struggle. By revisiting that alleged Tío Tomas, Richard Rodriguez, we unearth the tensions that negatively, deny citizenship to one silo, but on the flipside, engender manifold ways of seeing, hearing, and moving . The second, through George Jackson’s prison memoirs, pans out from this ethnography of power, groping for an apparatus that feigns an impervious prestige: ‘the aesthetic regime of coercion.’ In half-way cut, the thesis sidesteps to spic into existence, formally announcing, through Aime Cesaire, myself, and Pedro Pietri, the poeticization of trauma. Such uplift denies New Age transcendence of self, but a rehearsal of our entrapment in these mortal envelopes. Thirdly, conscious of the bleeding ethnic body, we cut open the incipient corpse to observe her pathologist. Her native autopsies offer the ethnic body’s posthumous recognition, the ethnopoetics ability to speak for and through the dead. Chapter five examines prolific black artists—Beyonce and Kendrick Lamar—to elide the circumvention of their consumption via invoking radical black hi/her-stories, ones fragmenting the black body. Sixth, the paper compares the Black Power Salute of the 1968 Mexico City Olympics to Duke’s Mi Gente Boycott of their Latino Student Recruitment Weekend. Both wielded “silent gestures,” that shrewdly interfered with white noise of numbed negligence. Finally, ‘taking the mask off’ that are her functionalities, the CODA expounds on ethnopoet’s interiority, particularly after the rapid re-calibration of her politics. Through a rerun of El Chavo del Ocho, one of Mexican television’s most cherished shows, we tune into the heart-breaking indigence of barrio residents, only to marvel at the power of humor to, as Friday’s John Witherspoon put it, “fight another day.” This thesis is the tip of my tongue. Y por una vez, déjala que cante.

(Table 3) Reduced inorganic sulphur, d34S and nickel content of ODP Leg 165 sites

Leg 165 of the Ocean Drilling Program afforded a unique opportunity to investigate organic and inorganic geochemistry across a wide gradient of sediment compositions and corresponding chemical pathways. The solid fractions at Sites 998, 999, 1000, and 1001 reveal varying proportions of reactive carbonate species, a labile volcanic ash fraction occurring in discrete layers and as a dispersed component, and detrital fluxes that derive from continental weathering. The relative proportions and reactivities of these end-members strongly dictate the character of the diagenetic profiles observed during the pore-water work of Leg 165. In addition, alteration of the well-characterized basaltic basement at Site 1001 has provided a strong signal that is reflected in many of the dissolved components. The relative effects of basement alteration and diagenesis within the sediment column are discussed in terms of downcore relationships for dissolved calcium and magnesium. With the exception of Site 1002 in the Cariaco Basin, the sediments encountered during Leg 165 were uniformly deficient in organic carbon (typically <0.1 wt%). Consequently, rates of organic oxidation were generally low and dominated by suboxic pathways with subordinate levels of bacterial sulfate reduction and methanogenesis. The low rates of organic remineralization are supported by modeled rates of sulfate reduction. Site 1000 provided an exception to the generally low levels of microbially mediated redox cycling. At this site the sediment is slightly more enriched in organic phases, and externally derived thermogenic hydrocarbons appear to aid in driving enhanced levels of redox diagenesis at great depths below the seafloor. The entrapment of these volatiles corresponds with a permeability seal defined by a pronounced Miocene minimum in calcium carbonate concentration recognized throughout the basin and with a dramatic downcore increase in the magnitude of limestone lithification. The latter has been tentatively linked to increases in alkalinity associated with microbial oxidation of organic matter and gaseous hydrocarbons. Recognition and quantification of previously unconstrained large volumes and frequencies of Eocene and Miocene silicic volcanic ash within the Caribbean Basin is one of the major findings of Leg 165. High frequencies of volcanic ash layers manifest as varied but often dominant controls on pore-water chemistry. Sulfur isotope results are presented that speak to secondary metal and sulfur enrichments observed in ash layers sampled during Leg 165. Ultimately, a better mechanistic understanding of these processes and the extent to which they have varied spatially and temporally may bear on the global mass balances for a range of major and minor dissolved components of seawater.

Development of a Composite ion-exchange membrane for PDMS-based microfluidic devices

The purpose of this research is to investigate potential methods to produce an ion-exchange membrane that can be integrated directly into a polydimethylsiloxane Lab-on-a-Chip or Micro-Total-Analysis-System. The majority of microfluidic membranes are based on creating microporous structures, because it allows flexibility in the choice of material such that it can match the material of the microfluidic chip. This cohesion between the material of the microfluidic chip and membrane is an important feature to prevent bonding difficulties which can lead to leaking and other practical problems. However, of the materials commonly used to manufacture microfluidic chips, there are none that provide the ion-exchange capability. The DuPont product Nafion{TM} is chosen as the ion-exchange membrane, a copolymer with high conductivity and selectivity to cations and suitable for many applications such as electrolysis of water and the chlor-alkali process. The use of such an ion-exchange membrane in microfluidics could have multiple advantages, but there is no reversible/irreversible bonding that occurs between PDMS and Nafion{TM}. In this project multiple methods of physical entrapment of the ion-exchange material inside a film of PDMS are attempted. Through the use of the inherent properties of PDMS, very inexpensive sugar granulate can be used to make an inexpensive membrane mould which does not interfere with the PDMS crosslinking process. After dissolving away this sacrificial mould material, Nafion{TM} is solidified in the irregular granulate holes. Nafion{TM} in this membrane is confined in the irregular shape of the PDMS openings. The outer structure of the membrane is all PDMS and can be attached easily and securely to any PDMS-based microfluidic device through reversible or irreversible PDMS/PDMS bonding. Through impedance measurement, the effectiveness of these integrated membranes are compared against plain Nafion{TM} films in simple sodium chloride solutions.

Three-dimensional free surface modelling in an unstructured mesh environment for metal processing applications

In the casting of metals, tundish flow, welding, converters, and other metal processing applications, the behaviour of the fluid surface is important. In aluminium alloys, for example, oxides formed on the surface may be drawn into the body of the melt where they act as faults in the solidified product affecting cast quality. For this reason, accurate description of wave behaviour, air entrapment, and other effects need to be modelled, in the presence of heat transfer and possibly phase change. The authors have developed a single-phase algorithm for modelling this problem. The Scalar Equation Algorithm (SEA) (see Refs. 1 and 2), enables the transport of the property discontinuity representing the free surface through a fixed grid. An extension of this method to unstructured mesh codes is presented here, together with validation. The new method employs a TVD flux limiter in conjunction with a ray-tracing algorithm, to ensure a sharp bound interface. Applications of the method are in the filling and emptying of mould cavities, with heat transfer and phase change.

Superparamagnetic Liposomes for MRI Monitoring and External Magnetic Field-Induced Selective Targeting of Malignant Brain Tumors

International audience

Desenvolvimento e caracterização de nanocápsulas de poli (L-lactídeo) contendo benzocaína

In this paper we describe the preparation poly (L-lactide) (PLA) nanocapsules as a drug delivery system for the local anesthetic benzocaine. The characterization and in vitro release properties of the system were investigated. The characterization results showed a polydispersity index of 0.14, an average diameter of 190.1± 3 nm, zeta potential of -38.5 mV and an entrapment efficiency of 73%. The release profile of Benzocaine loaded in PLA nanocapsules showed a significant different behavior than that of the pure anesthetic in solution. This study is important to characterize a drug release system using benzocaine for application in pain treatment.

Physicochemical Stability of Poly(lactide-co-glycolide) Nanocapsules Containing the Local Anesthetic Bupivacaine

This paper describes the preparation of poly(DL-lactide-co-glicolide) (PLGA) nanocapsules as a drug carrier system for the local anesthetic bupivacaine. The system was characterized and its stability investigated. The results showed a size distribution with a polydispersity index of 0.12, an average diameter of 148 nm, a zeta potential of -43.5 mV and an entrapment efficiency of 75.8%. The physicochemical properties of polymeric nanocapsule suspensions (average diameter, polydispersity, zeta potential and drug association efficiency) were evaluated as a function of time to determine the formulation stability. The formulation did not display major changes in these properties over the time, and it was considered stable up to 120 days of storage at room temperature. The results reported here which refer to the initial characterization of these new formulations for the local anesthetic bupivacaine show a promising potential for future in vivo studies.

Development and evaluation of floating microspheres of curcumin in alloxan-induced diabetic rats

Purpose: To prepare and evaluate floating microspheres of curcumin for prolonged gastric residence and to study their effect on alloxan-induced diabetic rats. Methods: Floating microsphere were prepared by emulsion-solvent diffusion method, using hydroxylpropyl methylcellulose, chitosan and Eudragit S 100 polymer in varying proportions. Ethanol/dichloromethane blend was used as solvent in a ratio of 1:1. The floating microspheres were evaluated for flow properties, particle size, incorporation efficiency, as well as in-vitro floatability and drug release. The anti-diabetic activity of the floating microspheres of batch FM4 was performed on alloxaninduced diabetic rats. Result: The floating microspheres had particle size, buoyancy, drug entrapment efficiency and yield in the ranges of 255.32 - 365.65 μm, 75.58 - 89.59, 72.6 - 83.5, and 60.46 - 80.02 %, respectively. Maximum drug release after 24 h was 82.62 % for formulation FM4 and 73.879, 58.613 and 46.106 % for formulations FM1, FM2, and FM3 respectively. In-vivo data obtained over a 120-h period indicate that curcumin floating microspheres from batch FM4 showed the better glycemic control than control and a commercial brand of the drug. Conclusion: The developed floating curcumin delivery system seems economical and effective in diabetes management in rats, and enhances the bioavailability of the drug.

Microfluidics based manufacture of liposomes simultaneously entrapping hydrophilic and lipophilic drugs

Despite the substantial body of research investigating the use of liposomes, niosomes and other bilayer vesicles for drug delivery, the translation of these systems into licensed products remains limited. Indeed, recent shortages in the supply of liposomal products demonstrate the need for new scalable production methods for liposomes. Therefore, the aim of our research has been to consider the application of microfluidics in the manufacture of liposomes containing either or both a water soluble and a lipid soluble drug to promote co-delivery of drugs. For the first time, we demonstrate the entrapment of a hydrophilic and a lipophilic drug (metformin and glipizide respectively) both individually, and in combination, using a scalable microfluidics manufacturing system. In terms of the operating parameters, the choice of solvents, lipid concentration and aqueous:solvent ratio all impact on liposome size with vesicle diameter ranging from ∼90 to 300 nm. In terms of drug loading, microfluidics production promoted high loading within ∼100 nm vesicles for both the water soluble drug (20–25% of initial amount added) and the bilayer embedded drug (40–42% of initial amount added) with co-loading of the drugs making no impact on entrapment efficacy. However, co-loading of glipizide and metformin within the same liposome formulation did impact on the drug release profiles; in both instances the presence of both drugs in the one formulation promoted faster (up to 2 fold) release compared to liposomes containing a single drug alone. Overall, these results demonstrate the application of microfluidics to prepare liposomal systems incorporating either or both an aqueous soluble drug and a bilayer loaded drug.

Alternativas ao internamento juvenil através da colaboração de família madrinha

As intervenções alternativas são medidas que não devem ser rechaçadas por nenhum governante. O sistema de encarceramento penal existente no Brasil, assim como em muitas nações mundo afora, vem mostrando há décadas, ser um sistema falido, vez que é impossível (res)socializar alguém em cadeias que são verdadeiras masmorras. Não há condições de melhoramento, nem na esfera física ou psíquica deste sistema nefasto, que só funciona para pegar pobres, pretos, analfabetos e moradores das periferias. Dentro dos maiores problemas apontados por pesquisas atuais, a superlotação, a violência, o tráfico de drogas, o abuso sexual e a falta de uma ocupação que garanta a (res)socialização, forma verdadeiras faculdades do crime, onde o preso ou o adolescente chega como estagiário e sai como mestre na arte da criminalidade delinquente. Esta pesquisa em Criminologia e direito comparado, com foco no Brasil (Recife) e na Alemanha (Freiburg), faz uma contextualização histórica do tema, de forma geral e mais focada nos países comparados. Após esta contextualização, o trabalho retrata e analisa a situação dos jovens em conflito com a Lei, e explana as intervenções alternativas à internação, aconselhando a solução do acolhimento por famílias madrinhas.