No description available.

Les dendrimères de type Janus (à deux faces) prennent une importance croissante du fait de leurs applications thérapeutiques ou diagnostiques potentielles, mais demandent des synthèses multi-étapes laborieuses. Les dendrigrafts de poly-L-Lysine (DGL) récemment découverts proposent une alternative "à faible coût" aux dendrimères. Ces DGL sont préparés par cycles successifs de polycondensation d'un N-carboxyanhydride (Lys(Tfa)NCA) dans l'eau suivie de la déprotection des chaînes latérales. Le spectre d'applications des DGL déjà identifiées s'élargira nettement si l'on parvient à construire des architectures DGL de type "Janus", ce qui requiert de fonctionnaliser le DGL (avec des groupes "clickables") de façon contrôlée, tant au cœur qu'en périphérie.Le chapitre I est une synthèse bibliographique qui montre les principales différences entre dendrimères et dendrigrafts, et qui résume l'état de l'art du domaine en ce qui concerne les matériaux à base de lysine.Le chapitre II étudie la fonctionnalisation périphérique du DGL par la chimie Click-Huisgen (CuAAC), et met en avant l'électrophorèse capillaire (EC) et l'analyse de la dispersion de Taylor (TDA), qui sont des outils efficaces pour caractériser le degré de fonctionnalisation du DGL, en montrant que celle-ci est homogène et régulière.Le Chapitre III est consacré à la synthèse et caractérisation de DGL fonctionnalisés à cœur (par un bras PEG portant un azoture terminal "clickable"), obtenus en modifiant la synthèse des DGL natifs, en particulier via l'amorçage de la condensation de NCA par une amine fonctionnelle. L'accessibilité de la fonction à cœur a été étudiée par réaction Click avec un chromophore, et par des tests de reconnaissance immunochimique en compétition.Enfin le chapitre IV résume notre stratégie de synthèse vers des DGL de type Janus (à deux face) et présente des résultats préliminaires qui valident le concept, avec comme perspective plus lointaine un accès éventuel à des DGL "Janus" à trois faces. / "Janus"-like (double-faced) dendrimers gain increasing attention for their high potential of therapeutic or diagnostic applications, however involving tedious, multistep synthesis. Recently discovered poly-L-lysine dendrigrafts (DGL), prepared through the alternation of N-carboxyanhydride (Lys(Tfa)-NCA) polycondensation in aqueous medium with deprotection of side chain amines, constitute promising "low-cost" equivalents of dendrimers. The already identified spectrum of DGL applications will benefit from an easy access to "Janus"-like DGL architectures, what requires controlled functionalisation (suitable for further click chemistry) of both core and periphery of DGLs.The chapter I is a bibliographic survey of the topic, highlighting the main differences between dendrigrafts and dendrimers, and summarising the state-of the-art about lysine-based materials in both domains.The chapter II investigates the surface functionalisation of DGL by Huisgen Click chemistry (CuAAC), and promotes capillary electrophoresis (CE) and Taylor Dispersion Analysis (TDA) as efficient analytic tools for characterising the functionalisation extent of DGL, thus proving the regularity and homogeneity of surface functionalisation.The chapter III is devoted to the synthesis and characterisation of core-functionalised DGL (bearing a PEG2–4 spacer with a clickable azido endgroup), through a modification of the "native" DGL synthetic route, involving initiation of NCA polycondensation in water by a functional amine. The DGL core group accessibility was assessed by click coupling with a chromophore group, and by immunochemical competition assays, concluding that a sufficiently long PEG linker ensures good core group accessibility.The chapter IV outlines the synthetic route toward double faced Janus DGL and presents preliminary results as a proof of the concept. Further, this synthetic strategy might potentially be extended to three-faced Janus DGL.

Les matières premières biosourcées s'avèrent être une possibilité de substitution du pétrole de plus en plus prisée dans le domaine des matériaux. De plus, l'utilisation de procédé propre limitant l'impact environnemental du développement de matériau est aujourd'hui incontournable. Dans ce contexte, l'acide aminé L-lysine, un nouveau synthon issu des biotechnologies blanches peu étudié dans le domaine de la chimie des matériaux, a été retenu. Ce synthon sera modifié pour la conception de macromonomères polymérisables sous irradiation UV. En effet, la technique de photopolymérisation est un procédé propre en plein essor, et qui permet le développement de revêtement.La L-lysine étant peu soluble dans les milieux organiques classiques, une étape de transformation est nécessaire pour améliorer sa processabilité. Ainsi, la polycondensation en masse de la L-lysine a conduit à des oligomères de poly-L-lysine de faibles masses molaires. La détermination de la structure obtenue a été réalisée par différentes techniques d'analyses. Ces oligomères sont alors greffés par des fonctions photopolymérisables dans des conditions douces. Le choix s'est porté sur des fonctions accepteur et donneur d'électron. Enfin, l'étude de la photopolymérisation des oligomères de L-lysine greffés accepteurs donneurs a été effectuée par la technique UV aqueuse, une technique innovante et respectueuse de l'environnement, dans différentes conditions pour en optimiser le système. / Biobased raw materials are an interesting and promising option for the substitution of fossil resources in material design. Moreover, using green processes which limit environmental impact of the material conception can't be avoided nowadays. In this context, the L-lysine amino acid, a building block made by white biotechnologies and poorly described in material field has been evaluated. As photopolymerization is a green process in great expansion and allowing coating design, this building block has been modified into a photocurable macromonomer.L-lysine is hardly soluble in usual organic solvents, a transformation step is necessary in order to improve its processability. Thus, L-lysine polycondensation has been tackled and led to oligomers of poly-L-lysine with low molar mass and improved solubility. In addition, the structure determination has been undertaken by different analytic technics. These oligomers can thus be grafted with photocurable functional groups in mild conditions. The chosen photocurable functional groups are donor acceptor of electron. At last, the photopolymerization of L-lysine based oligomers grafted with donor acceptor functional groups has been done by UV waterborne technic which is innovative and environmentally friendly. The photopolymerization has been carried out in different conditions in order to optimize the process.

A mist reactor was used to study plant growth and development under various environmental conditions towards the production of healthy plantlets ready for soil transplant in one step from inoculation. In addition, a 3D type of cultivation via surface attachment of explants to vertically hanging strips inside the mist reactor was also investigated to maximize productivity with minimal footprint. Using carrot as the model species, pre-embryogenic cell suspensions were successfully spray-inoculated onto hanging poly-L-lysine (PLL)-coated nylon mesh to which they then attached and remained for several weeks while they developed into rooted plantlets. To study single step micropropagation from shoot explants to fully acclimatized plantlets, Artemisia annua was used as the model species. Nodal cuttings of A. annua were inoculated onto PLL-coated mesh strips by briefly immersing the strips in the suspension of nodal cuttings. Investigation of medium, phytohormones, CO2, ventilation level and humidity ensued resulting in selection of a preferred final process that reduced physiological aberrations like hyperhydricity and was time efficient. The nodal cuttings that attached to the strips were first misted with half strength shooting medium for 7 days to develop new shoots. Then the new shoots were misted with the rooting medium supplemented with NAA for 12 days to develop roots. Rooted plantlets were acclimatized in the same rooting medium for 9 days to acquire fully functional stomata prior to planting into soil. Taken together this study suggested that fully developed plantlets ready for planting into soil could be obtained in a single step in a bioreactor from embryogenic cells or from nodal explants.

La thérapie génique de myopathies telles que la dystrophie musculaire de Duchenne nécessite une approche systémique afin de traiter l'ensemble de la musculature. Le vecteur AAV est actuellement le plus efficace pour transduire le muscle. Nous montrons que la biodistribution du vecteur AAV administré par voie veineuse peut être modifiée en utilisant diverses stratégies adjuvantes chez la souris saine. La pré-injection de polymères permet ainsi d'améliorer la transduction des muscles par le vecteur AAV, ou encore de baisser la réponse immune neutralisante induite par l'injection intraveineuse du vecteur. Nous abordons également l'impact de facteurs modulateurs exogènes ou endogènes – tels que la procédure d'administration ou certains facteurs sanguins – sur la transduction systémique de l'AAV. Dans une seconde approche, nous avons évalué le transfert de gènes dans le muscle dystrophique afin de sécréter dans la circulation sanguine une protéine transgénique fusionnant le récepteur soluble I du TNF-α avec le fragment constant d'une immunoglobuline (TNFR-Is/mIgG1). La comparaison des cinétiques de sécrétion obtenu après le transfert de gène dans le muscle de souris saines ou de souris dystrophiques mdx indique que le contexte inflammatoire du muscle dystrophique favorise une réponse immune contre le transgène. Nous montrons que l'expression et la sécrétion d'un variant murin peu immunogène du TNFR-Is/mIgG1 améliore la fonction musculaire de la souris mdx sans toutefois conférer un avantage sélectif aux fibres musculaires dystrophiques qui continuent leur cycle de nécrose et de régénération.

In this study we fabricated free standing multilayer films of polyelectrolyte complexes for potential use in tissue engineering of corneal stroma by using the layer-by-layer (LbL) approach. In the formation of these LbL films negatively charged, photocrosslinkable (methacrylated) hyaluronic acid (MA-HA) was used along with polycations polyethyleneimine (PEI) and poly(L-lysine) (PLL). Type I collagen (Col) was blended in with PLL for improving the water absorption and cell attachment properties of the films. It was shown that the LbL films could be easily peeled off from glass substrates due to the photocrosslinking of one of the LbL components, the hyaluronic acid. Film growth and composition were monitored with FTIR-ATR. Heights of peaks at 3383 cm-1, and 2958 cm-1increased along with the bilayer number confirming the polymer build-up. Film integrity and thickness were investigated by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Films thicker than 5 bilayers (BLs) were found to be uniform in appearance and 10 BL (PEI/MeHA) films were calculated to be ca. 6 &mu / m thick. Atomic force microscopy (AFM) revealed that as the number of BLs increased, surface roughness decreased. Activity of methacrylated hyaluronic acid was shown by the increased resistance of photocrosslinked multilayer films against hydrolysis by hyaluronidase. Patterns could be created on the films by photocrosslinking further proving that the crosslinking step is successful. Since the ultimate goal was to construct a corneal stroma PEI/MA-HA films were tested with corneal stroma cells, keratocytes. Cell proliferation on PEI/MA-HA films was quite poor in comparison to TCPS. In order to improve the cell adhesion the tests were repeated with PLL/MA-HA. Collagen was added to decrease the hydrophilicity and introduce cell adhesion sequences (Arg-Gly-Asp, RGD) to improve cell proliferation on the films and thus PLL+Col/MA-HA films were also tested. Introduction of collagen to the PLL/MA-HA films was found to decrease water retention of the multilayer films and improve cell viability and proliferation. Col+PLL/MA-HA LbL thus appear to be a promising platform for tissue engineering, especially of corneal stroma.

Thesis (Ph. D.)--Auburn University, 2008. / Abstract. Vita. The following patent resulted from the dissertation research: Davis, V., Simonian, A.L., Nepal, D., Balasubramanian, S, "Preparation of Precisely Controlled Thin Film Nanocomposites of Carbon Nanotubes and Biomaterials", U.S. Provisional Patent Application No. 61/000,938, filed on 30 October 2007. The following peer-reviewed publications resulted from the dissertation research: Dhriti Nepal, Shankar Balasubramanian, Aleksandr Simonian, and Virginia Davis, "Mechanically Strong Antibacterial Thin Film Based on Single-Walled Carbon Nanotubes Armored with Biopolymers", Nano Letters ASAP article, May 2008 (# equal contribution) -- Shankar Balasubramanian, Iryna B. Sorokulova, Vitaly J. Vodyanoy, and Aleksandr L. Simonian, "Lytic Phage as a Specific and Selective Probe For Detection of Staphylococcus Aureus: A Surface Plasmon Resonance Spectroscopic Study", Biosensors and Bioelectronics, 2007, 22, 948-955 -- Shankar Balasubramanian, Alexander Revzin, Aleksandr Simonian, "Electrochemical Desorption of Proteins from Gold Electrode Surface", Electroanalysis, 2006, 18, 1885-1892 (Invited article) -- Vishwaprakash Nanduri, Shankar Balasubramanian, Srinivas Sista, Vitaly J. Vodyanoy, and Aleksandr L. Simonian, "Highly Sensitive Phage-based Biosensor for the Detection of ß-galactosidase", Analytica Chimica Acta, 2007, 589, 166- 172 -- H. Luckarift, Shankar Balasubramanian, S. Paliwal, G. Johnson and A. Simonian, "Enzyme-Encapsulated Silica Monolayers For Rapid Functionalization of a Gold Surface", Colloids and Surfaces B: Biointerfaces, 2007, 58, 28-33 (Invited article) -- Dong Wei, Omar Oyarzabal, Tung-Shi Huang, Shankar Balasubramanian, Srinivas Sista, Aleksandr Simonian, "Development of Surface Plasmon Resonance Biosensor For The Identification of Campylobacter jejuni", Journal of Microbiological Methods, 2007, 69, 78-85. The following conferences presentations resulted from the dissertation research: Covalent Immobilization of Organophosphorus Hydrolase on Carbon Nanotubes for Biosensor Applications, accepted for oral presentation at 12th International Meeting on Chemical Sensors, Jul. 13-16, 2008, Columbus, OH -- Electrochemical characteristics of SWNT-biopolymer nanocomposites, accepted for 213th meeting of The Electrochemical Society, May 18-23, 2008, Phoenix, AR -- Mechanically Robust Antibacterial Thin Films Composed of Single-Walled Carbon Nanotubes and Biopolymers, 2008 AIChE Spring National Meeting, Apr. 6-10, New Orleans, LA -- Production and characterization of protein and DNA based single wall carbon nanocomposites by layer-by-layer assembly, MRS Fall Meeting, Nov. 26-30, 2007, Boston, MA -- Gold surface modified with enzyme-encapsulated silica monolayers for biosensor application, The 58th Southeast Regional Meeting of the American Chemical Society, Nov. 1-4, 2006, Augusta, GA -- Electrochemical modulation of biological interfaces, 209th meeting of The Electrochemical Society, May 7-12, 2006, Denver, CO -- SPR based biosensor using lytic phage as a specific and selective probe for staphylococcus aureus detection, 57th Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Mar. 12-17, 2006, Orlando, FL -- Specific & selective detection of staphylococcus aureus by lytic phage using SPR biosensor, 57th Southeast / 61st Southwest Joint Regional Meeting of the American Chemical Society, Nov. 1-4, 2005, Memphis, TN -- Prevention of non-specific binding as a way to increase sensitivity of SPR-based sensors, 206th meeting of The Electrochemical Society, October 3-8, 2004, Honolulu, HI. Includes bibliographical references.

Hollow fiber membranes (HFMs) formed through phase inversion methods exhibit specific physicochemical characteristics and generally favorable surface and mechanical properties, supporting their use in diverse applications including ultrafiltration, dialysis, cell culture, bioreactors, and tissue engineering. Characterization of, and modifications to, such membranes are important steps in achieving desired characteristics for specific applications. HFMs subject to gas, irradiation, and chemical sterilization techniques were characterized based on several analytical techniques. It was revealed that these common sterilization techniques can cause inadvertent changes to HFM properties. While these changes may cause detrimental effects to HFMs used in filtration, the methods of sterilization are also presented as a facile means of tuning properties toward specific applications. Modifications to HFM surface chemistries were also sought as a method of adsorbing bacterial lipopolysaccharide (LPS) from solutions used in hemodialysis treatments and bioprocessing applications. It was found that additives such as polyvinylpyrrolidone (PVP), polyethyleneglycol (PEG), and poly-L-lysine (PLL) can facilitate adsorption capacities of HFMs toward LPS. Additionally, chemical changes are presented as a means of preferentially adsorbing LPS to specific locations on the HFM surface.

The ultimate design of a microencapsulated AI dose is to continuously release sperm over a period of time in the female reproductive tract, thus alleviating the need for estrus detection. The objective of Trial 1 was to determine in vitro sperm release times for three microcapsule membranes. Semen was collected from four bulls, pooled, extended in 20% egg yolk TEST to a concentration of 80 = 10⁶ cells/ml, and encapsulated. Microcapsule membranes were constructed from isomers of polylysine: .1% poly-L-lysine (PLL), .1% poly-D-lysine (PDL), and a 50:50 mixture of the isomers (PLPD). Microcapsules were incubated at 37°C in a buffer containing .5% heparin or .5% trypsin and evaluated at 0.5, 1, 2, 4, 8, and 16 h post-encapsulation. For sperm encapsulated there were no significant differences in sperm motility. However, peak time of maximum sperm release differed between PLL and PDL membranes at 2 and 4 h of incubation. In Trial 2, sperm viability and microcapsule membrane stability were assessed post-thaw using PLL or PDL, two encapsulating temperatures (5°C or 23°C) and two times of glycerol addition (prior or post encapsulation at 5°C). Semen was extended to 80 = 10⁶ cells/ml and encapsulated. Capsules from all treatment combinations were incubated in .5% trypsin and evaluated as in Trial 1. In addition, motility was estimated at 1, 3, 6, and 9 h post-thaw. Motility from the unencapsulated control and capsules with glycerol addition prior to encapsulation, was superior (P < .05). Additionally, sperm release from capsules prepared at 5°C with glycerol addition post encapsulation was greater than all other treatments (P < .05). Time of peak sperm release for capsules was similar to the previous trial. There was a positive correlation between average capsule diameter and sperm release for both trials (P < .05). These data suggest that a combination of PLL and PDL capsules may complement each other in timing of sperm release and may be utilized in an inseminate mixture for extending the effective release in the female / Master of Science

L'immobilisation de biomacromolécules à la surface de polymères peu réactifs est une voie de synthèse de nanomatériaux qui fait actuellement l'objet de nombreuses recherches pour le développement d'applications biologiques et médicales. Nous avons synthétisé de nouveaux nanomatériaux à base de polypropylène (PP) greffé par des dendrimères de lysine (DGL). Les DGL sont parfaitement solubles dans l'eau, biocompatibles, polycationiques à pH neutre et leur structure dendritique particulière font d'eux des macromolécules de plus en plus étudiées en interactions avec les milieux biologiques. Différents traitements par plasma ont permis de fonctionnaliser la surface du PP et plusieurs stratégies ont été adoptées pour greffer les DGL sous forme de monocouche, multicouche ou à partir de brosses de polymères : le greffage direct, les polymérisations non contrôlée et contrôlée de type RAFT associées au plasma d'iode et à la chimie click de surface. L'aptitude des matériaux PP fonctionnalisés par le DGL à interagir avec les milieux biologiques a été étudiée, en particulier l'immobilisation de l'ATP et le comportement vis-à-vis des bactéries et des virus. Les propriétés de ces nanomatériaux sont liées à la réactivité des groupements amine des DGL ainsi qu'à la structure régulière et sphérique des dendrimères. Les résultats obtenus ouvrent de nombreuses applications potentielles pour le traitement des eaux, le diagnostic et la prévention du développement des micro-organismes. / Great attention has been focused these last years on tailoring polymer surfaces by immobilizationof suitable molecules for biological and medical applications such as tissue engineering, drug delivery systems, antibacterial supports, and biosensors. In that context, we report the preparation of an original hybrid material based on polypropylene and poly-L-Lysine dendrigrafts (DGL) which are perfectly water soluble, and biocompatible. First, activation of the polypropylene surface (PP) was achieved using plasma treatment. Then, several strategies have been developed to graft DGL onto the PP surface such as (i) direct grafting of DGL after surface activation, (ii) the use of conventional radical polymerization or (iii) RAFT polymerization of monomers from the PP surface. The last methodology favored the increase of the DGL grafts density onto the surface. The ability of PP surface functionalized with DGL to interact with biological media was studied and the modified surfaces open the way to many potential applications in water treatment, diagnosis and prevention of the development of microorganisms.