Management Model For University Cooperation At Universidad Técnica De Manabí, Ecuador

In researching a management model for university cooperation at the Universidad Técnica de Manabí, with its components, phases, stages, input, and output information as well as the premises of its practical application, thereby achieving qualitative and quantitative changes proposed in the conception of university cooperation for sustainable development. The model for the management of cooperation, the assumptions of the model and the components are exposed, and a map with universities that are currently on the network and the issues raised in the strategy, setting out the results at national and International levels

Development of dioctadecyldimethylammonium bromide/monoolein liposomes for gene delivery

The artificial introduction of nucleic acids (NA) into mammalian cells (transfection) has become, in recent years, a well-established procedure in basic and applied research, which allowed the study of gene function and regulation. The advances in this area have made possible the use of these methods for gene-based medicines, which constitute alternative therapeutic approaches. One of the most prominent methods is lipofection that uses cationic liposome/NA complexes (a.k.a. lipoplexes) for the complexation, transport and release of therapeutic sequences into target cells. Although yielding lower transfection efficiencies compared with viral gene delivery, lipofection vectors are much safer for medical applications because no significant mutational or toxicological risk exist. Dioctadecyldimethylammonium Bromide (DODAB)/Monoolein (MO) liposomes have recently been described as a new promising alternative to common transfection reagents, due to the pioneering application of MO as helper lipid in lipoplex formulations. In this chapter, we will review the effect of MO on the physicochemical properties of DODAB/MO liposomes and pDNA/DODAB/MO lipoplexes. How lipoplex properties may affect the interaction with different extracellular components and their cell uptake and trafficking will be discussed. The importance of lipoplex biocompatibility towards efficient gene therapy will also be approached presenting pDNA/DODAB/MO system as a lipoplex model, supporting the use of MO as new helper lipid in lipofection.FCTCOMPETEThis work was supported by FCT research project PTDC/QUI/69795/2006, which is cofunded by the program COMPETE from QREN with co-participation from the European Community fund FEDER; CFUM [PEst-C/FIS/UI0607/2011]; CBMA [Pest C/BIA/UI4050/2011]; J.P.N. Silva holds a PhD Grant (SFRH/BD/46968/2008); A. C.N. Oliveira holds a PhD grant (SFRH/BD/68588/2010)

Application of computational quantum chemistry to chemical processes involved in mass spectrometry

The field of application of mass spectrometry (MS) has increased considerably due to the development of ionization techniques. Other factors that have stimulated the use of MS are the tandem mass spectrometry (MS/MS) and sequential mass spectrometry (MSn) techniques. However, the interpretation of the MS/MS and MSn data may lead to speculative conclusions. Thus, various quantum chemical methods have been applied for obtaining high quality thermochemical data in gas phase. In this review, we show some applications of computational quantum chemistry to understand the formation and fragmentation of gaseous ions of organic compounds in a MS analysis.Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES)FAPESPCNP

A novel approach for the production of DODAB:MO lipoplexes : the influence of temperature on the physicochemical characteristics and cell transfection efficiency

Cationic liposomes/ DNA complexes (lipoplexes) have been widely used as nano-carriers for animal cell transfection, with the neutral lipid (helper) playing a determinant role for the efficiency of this process due to the formation of non-lammellar intermediates that are akin to membrane fusion process [1]. We have developed a novel formulation containing the cationic agent dioctadecyldimethylammonium bromide (DODAB) and 1-monooleoyl-rac-glycerol (MO) as helper lipid [2]. In previous studies, we have demonstrated a strong dependence of the DNA complexation rate with several structural parameters such as the monoolein content or the DNA/cationic lipid charge ratio (+/-) [3]. The preparation method itself influences the structural properties of the lipoplexes formed as well as their lipofection capacity. This study addressed the effect of preparation temperature (25ºC or 50ºC) on DODAB:MO lipoplexes (molar ratios 4:1 and 2:1) physicochemical properties, as well as on their cell transfection efficiency (TE).Fundação para a Ciência e a Tecnologia (FCT) - PTDC/QUI/69795/2006, SFRH/BD/46968/200

LC-ESI-MS determination of quassinoids isobrucein B and neosergeolide in Picrolemma sprucei stem infusions

Infusions of the stems of Picrolemma sprucei (pseudonym: P. pseudocoffea) are used in the Amazon regions of Peru, Brazil and French Guiana as antimalarials among other uses. They contain the bitter quassinoids isobrucein B (1) and neosergeolide (2) that have important antimalarial and toxic properties among others. In this study, an LC-(+)-ESI-MS/MS method was developed and applied to the determination of 1 and 2 in a common remedy prepared by infusing 1 g of dry, powdered stems of P. sprucei in 1 L of boiling water. Isolated 1 and 2 were used in calibration ranges of 0.25 to 5 µg mL-1 and 0.5 to 10 µg mL-1, respectively, with the internal standard phloroglucinol at 4.0 µg mL-1. Good linearity, precision and accuracy were observed for both compounds. The concentrations of 1 and 2 in the stem infusions were found to be 60.1 and 774 µg L-1, respectively.Infusões dos caules de Picrolemma sprucei (pseudônimo: P. pseudocoffea) são principalmente utilizadas como antimaláricos em toda região amazônica. Desta espécie foram isolados os quassinoides isobruceína B e neosergeolida, os quais apresentam atividade antimalárica e citotóxica. Neste estudo, apresentamos o desenvolvimento de uma metodologia analítica por LC-(+)-ESI-MS/MS visando a determinação dos principais quassinoides desta espécie. O método desenvolvido foi empregado para análise de uma formulação artesanal (infusão de 1 g de matéria seca - caules pulverizados de P. sprucei em 1 L de água fervente). Padrões previamente isolados de isobruceína B e neosergeolida foram utilizados para a determinação da linearidade na faixa de calibração entre 0,25 to 5 µg mL-1 e 0,5 a 10 µg mL-1, respectivamente. Como padrão interno foi utilizada a substância fluoroglucinol na concentração de 4,0 µg mL-1. Ambos os compostos apresentaram boa linearidade, precisão e exatidão e as concentrações de isobruceína B e neosergeolida obtidas nas infusões foram de 60,1 e 774 µg L-1, respectivamente

Monoolein-based lipoplexes (DODAB/MO/DNA) as non-viral vector for transfection- from physicochemical characterization to biological application

Cationic liposomes/DNA (lipoplexes) have been widely used as non-viral vectors for transfection, the role of the neutral lipid in liposome formulation being determinant for the efficiency of this process [1,2]. In this work, we studied the potential of monoolein (MO) as helper lipid for cellular transfection. Lipoplexes composed of pDNA and dioctadecyldimethylammonium bromide (DODAB)/1-monooleoyl-rac-glycerol (MO) at different molar ratios (4:1, 2:1 and 1:1) were investigated, as well as different cationic lipid/DNA ratios. The physicochemical properties of the lipoplexes (size and charge), the formation of the lipoplexes, the effect of MO on pDNA condensation and the effect of heparin on percentage of pDNA release from the lipoplexes were also studied by Ethidium Bromide (EtBr) exclusion assays, Dynamic Light Scattering (DLS), Zeta Potential (æ) and electrophoresis. The cytotoxicity, transfection efficiency, as well as the intracellular localization of labeled DNA were evaluated on 293T cells. It was found that the presence of MO not only increases the efficiency of pDNA compactation, but also affects the physicochemical properties of lipoplexes, which could possibly interfere with lipoplex-cell interactions. The DODAB:MO (2:1) and (4:1) formulations were capable of efficiently mediate in vitro cell transfection. These results were consistent with fluorescence microscopy studies, which illustrated that lipoplexes were able to gain entry into the cytosol and deliver pDNA to the nucleus. Understanding the structure–activity relationship of MO based lipoplexes will give direction toward the development of safe and efficient gene delivery systems.Portuguese Foundation for Science and Technology (FCT) for financial support to Center of Physics and Center of Molecular & Environmental Biology and funding through projects PTDC/QUI/69795/2006 and SFRH/BD/46968/2009

Monoolein as helper lipid for non-viral transfection in mammals

Lipoplexes composed of pDNA and DODAB/MO at different molar ratios (4:1, 2:1 and 1:1) and different cationic lipid/DNA charge ratios were investigated. The physicochemical properties of the lipoplexes (size and charge), the pDNA complexation, and the effect of heparin on pDNA release, were studied by Dynamic Light Scattering, Zeta Potential, and Ethidium Bromide exclusion assays. The cytotoxicity, transfection efficiency and the intracellular localization of DNA were evaluated on 293T cells.The Portuguese Foundation for Science and Technology (FCT) for the financial support to the Center of Physics and Center of Molecular & Environmental Biology and funding through projects PTDC/QUI/69795/2006 and SFRH/BD/46968/2009 are acknowledged

Effects of preparation method on the physicochemical characteristics and cell transfection efficiency of non‐viral nanocarriers

The success of gene therapy is decisively dependent on the development of effective carrier systems, able to compact and thus protect the genetic material as to avoid both in vitro and in vivo barriers for transgene delivery [1]. Given the problems of safety encountered with viral vectors, non-viral carrier systems are a safer alternative for the therapeutic delivery of genes [2]. Cationic liposomes/DNA (lipoplexes) have been widely used as non-viral vectors for cell transfection, with the neutral lipid (helper) of the liposome formulation playing a determinant role for the efficiency of this process. The lipoplex preparation method itself influences the structural properties of the produced lipoplex, affecting its final lipofection capability [3, 4]. In this work, we have studied the potential for cell transfection of a new liposomal formulation containing monoolein (MO) as helper lipid, using various preparation methods. Lipoplexes composed of pSV-β-galactosidase and dioctadecyldimethylammonium bromide (DODAB)/1-monooleoyl-rac-glycerol (MO) at (4:1) molar ratio and at cationic lipid/DNA ratio of 4.0 were tested. Ethidium Bromide (EtBr) exclusion assays, Dynamic Light Scattering (DLS), and Zeta Potential (ζ) were used to study plasmidic DNA complexation and physicochemical properties of the formed lipoplexes (their size and electrical charge). Transfection efficiency was also evaluated on 293T cells by determining the activity of β-galactosidase, the reporter gene. The results indicate that the lipoplexes’ physicochemical properties are strongly dependent on the preparation method (one-step or multi-step complexation, at 25ºC or 50ºC), with resulting mean sizes varying from 350 to 1700 nm and superficial charge densities ranging from +9 mV to +30 mV. No clear correlation was found, however, between lipoplex physicochemical properties and observed cell transfection efficiencies, suggesting that other parameters, such as structure (degree of DNA condensation/compaction) of the complex and the mode of internalization by the cell may also weigh in on cell transfection. Nevertheless, it was possible to determine that lipoplex preparation methods can greatly affect DNA uptake by cells. Optimal nanotherapy requires the therapeutic molecule to be protected from degradation and reach its target cell and intracellular location. This work demonstrates that the MO-based system is a viable nanocarrier for plasmidic DNA and that the conditions to prepare the carriers for gene therapy application can be modulated to achieve maximal delivery efficiency