Structural Characterization of Photopolymerizable Binary Liposomes Containing Diacetylenic and Saturated Phospholipids

The use of liposomes to encapsulate materials has received widespread attention for drug delivery, transfection, diagnostic reagent, and as immunoadjuvants. Phospholipid polymers form a new class of biomaterials with many potential applications in medicine and research. Of interest are polymeric phospholipids containing a diacetylene moiety along their acyl chain since these kinds of lipids can be polymerized by Ultra-Violet (UV) irradiation to form chains of covalently linked lipids in the bilayer. In particular the diacetylenic phosphatidylcholine 1,2-bis(10,12-tricosadiynoyl)- sn-glycero-3-phosphocholine (DC8,9PC) can form intermolecular cross-linking through the diacetylenic group to produce a conjugated polymer within the hydrocarbon region of the bilayer. As knowledge of liposome structures is certainly fundamental for system design improvement for new and better applications, this work focuses on the structural properties of polymerized DC8,9PC:1,2-dimyristoyl-sn-glycero-3-phusphocholine (DMPC) liposomes. Liposomes containing mixtures of DC8,9PC and DMPC, at different molar ratios, and exposed to different polymerization cycles, were studied through the analysis of the electron spin resonance (ESR) spectra of a spin label incorporated into the bilayer, and the calorimetric data obtained from differential scanning calorimetry (DSC) studies. Upon irradiation, if all lipids had been polymerized, no gel-fluid transition would be expected. However, even samples that went through 20 cycles of UV irradiation presented a DSC band, showing that around 80% of the DC8,9PC molecules were not polymerized. Both DSC and ESR indicated that the two different lipids scarcely mix at low temperatures, however few molecules of DMPC are present in DC8,9PC rich domains and vice versa. UV irradiation was found to affect the gel fluid transition of both DMPC and DC8,9PC rich regions, indicating the presence of polymeric units of DC8,9PC in both areas, A model explaining lipids rearrangement is proposed for this partially polymerized system.

Regression analysis of mean quality-adjusted survival time based on pseudo-observations

Regression models for the mean quality-adjusted survival time are specified from hazard functions of transitions between two states and the mean quality-adjusted survival time may be a complex function of covariates. We discuss a regression model for the mean quality-adjusted survival (QAS) time based on pseudo-observations, which has the advantage of directly modeling the effect of covariates in the QAS time. Both Monte Carlo Simulations and a real data set are studied. Copyright (C) 2009 John Wiley & Sons, Ltd.

Beta-binomial/Poisson regression models for repeated bivariate counts

We analyze data obtained from a study designed to evaluate training effects on the performance of certain motor activities of Parkinson`s disease patients. Maximum likelihood methods were used to fit beta-binomial/Poisson regression models tailored to evaluate the effects of training on the numbers of attempted and successful specified manual movements in 1 min periods, controlling for disease stage and use of the preferred hand. We extend models previously considered by other authors in univariate settings to account for the repeated measures nature of the data. The results suggest that the expected number of attempts and successes increase with training, except for patients with advanced stages of the disease using the non-preferred hand. Copyright (c) 2008 John Wiley & Sons, Ltd.

1,4-Dioxane enhances properties and biocompatibility of polyanionic collagen for tissue engineering applications

Polyanionic collagen obtained from bovine pericardial tissue submitted to alkaline hydrolysis is an acellular matrix with strong potential in tissue engineering. However, increasing the carboxyl content reduces fibril formation and thermal stability compared to the native tissues. In the present work, we propose a chemical protocol based on the association of alkaline hydrolysis with 1,4-dioxane treatment to either attenuate or revert the drastic structural modifications promoted by alkaline treatments. For the characterization of the polyanionic membranes treated with 1,4-dioxane, we found that (1) scanning electron microscopy (SEM) shows a stronger reorientation and aggregation of collagen microfibrils; (2) histological evaluation reveals recovering of the alignment of collagen fibers and reassociation with elastic fibers; (3) differential scanning calorimetry (DSC) shows an increase in thermal stability; and (4) in biocompatibility assays there is a normal attachment, morphology and proliferation associated with high survival of the mouse fibroblast cell line NIH3T3 in reconstituted membranes, which behave as native membranes. Our conclusions reinforce the ability of 1,4-dioxane to enhance the properties of negatively charged polyanionic collagen associated with its potential use as biomaterials for grafting, cationic drug- or cell-delivery systems and for the coating of cardiovascular devices.

Thermal and spectrophotometric studies of new crosslinking method for collagen matrix with glutaraldehyde acetals

Despite the many existing crosslinking procedures, glutaraldehyde (GA) is still the method of choice used in the manufacture of bioprosthesis. The major problems with GA are: (a) uncontrolled reactivity due to the chemical complexity or GA solutions; (b) toxicity due to the release of GA from polymeric crosslinks; and (c) tissue impermeabilization due to polymeric and heterogeneous crosslinks formation, partially responsible for the undesirable calcification of the bioprosthesis. A new method of crosslinking glutaraldehyde acetals has been developed with GA in acid ethanolic solution, and after the distribution inside de matrix, GA is released to crosslinking. Concentrations of hydrochloride acid in ethanolic solutions between 0.1 and 0.001 mol/L with GA concentration between 0.1 and 1.0% were measured in an ultraviolet spectrophotometer to verify the presence of free aldehyde groups and polymeric compounds of GA. After these measurements, the solutions were used to crosslink bovine pericardium. The spectrophotometric results showed that GA was better protected in acetal forms for acid ethanolic solution with HCl at 0.003 mol/L and GA 1.0%(v/v). The shrinkage temperature results of bovine pericardium crosslinked with acetal solutions showed values near 85 C after the exposure to triethylamine vapors.

Qualidade de vida em idosos

Introdução: Esta tese de doutorado é mais uma contribuição do Grupo de Qualidade de Vida (Centro Brasileiro) do Programa de Pós-Graduação em Ciências Médicas: Psiquiatria, da UFRGS. Esse grupo têm trabalhado em projetos transculturais de elaboração de instrumentos de qualidade de vida (QV) sob a coordenação da Organização Mundial de Saúde. Entre os instrumentos genéricos de QV já desenvolvidos estão o WHOQOL-100 e o WHOQOL-Bref e entre os instrumentos específicos, para populações especiais, estão o WHOQOL-HIV, o WHOQOL-SRPB e, mais recentemente, o WHOQOL-OLD. O desenvolvimento de uma escala de QV para idosos é especialmente importante tendo em vista especificidades deste grupo etário, bem como o aumento da proporção de idosos na população mundial. Objetivos: O objetivo maior deste estudo é desenvolver uma escala de QV para idosos (WHOQOL-OLD). Entretanto, como trata-se de um processo com longa duração de tempo, foram gerados objetivos específicos a partir da revisão da literatura e coleta de dados, que deram origem a 5 artigos, cada um com sua proposta, a saber: artigo 1. Apresentar a metodologia utilizada e os resultados dos grupos focais para avaliação de QV do idoso, artigo 2. Identificar variáveis relevantes na QV de pessoas idosas, artigo 3. Investigar a relação da percepção de QV do idoso com a percepção de QV do idoso na opinião de seu cuidador, artigo 4. Investigar variáveis associadas com percepção subjetiva de saúde em idosos internados e artigo 5. Pesquisar um possível viés nas respostas de idosos no Inventário de Depressão de Beck (BDI). Métodos: O primeiro estudo teve um desenho qualitativo enquanto os demais foram quantitativos. As amostras variaram para cada estudo. Em todos os estudos idosos(as) acima de 60 anos foram entrevistados. Para os estudos 1 e 2, a amostra contou com profissionais da área da saúde (1) e cuidadores (1 e 2). E, para o estudo 5, adultos acima de 18 anos também foram pesquisados. A coleta dos dados foi realizada em hospitais, lares e grupos comunitários, residências e recrutamento utilizando a técnica de "snow-ball" (bola de neve) em que cada sujeito indicava um outro sujeito. Todos os entrevistados preencheram o Termo de Consentimento Informado e, a partir daí, foram convidados a responder acerca de informações sociodemográficas, QV percebida (WHOQOL-100) e sintomatologia depressiva (BDI), com pequena variação para os cuidadores. O último estudo contou apenas com os dados sociodemográficos e com as respostas ao BDI. Resultados: De forma abreviada, os resultados dos 5 artigos confirmam as especificidades do idoso e portanto a necessidade de desenvolvimento de instrumentos específicos para esta população. O artigo 1 teve como resultado a sugestão de novos itens para idosos, a partir das respostas espontâneas e análise dos domínios e facetas do WHOQOL-100. O artigo 2, por sua vez, mostrou associações da percepção de QV geral com níveis de depressão, percepção subjetiva de saúde e sexo. No artigo 3, foi possível verificar uma tendência, em todos os domínios e na medida QV geral, de o cuidador responder pior percepção de QV do idoso do que o próprio idoso cuidado, apesar de algumas concordâncias (domínios físico, nível de independência, meio ambiente e espiritualidade/religião). Também observou-se que a intensidade de depressão do idoso exerceu forte influência tanto na sua própria percepção de QV quanto na percepção do cuidador sobre o idoso. O artigo 4 mostrou uma prevalência alta e não esperada de idosos internados que se percebiam como saudáveis. Foi possível observar, ainda, uma associação significativa entre percepção saudável e menor intensidade de sintomas depressivos, bem como melhor percepção de QV no domínio nível de independência. E por último, o artigo 5 discute o viés da subescala somático e de desempenho nas respostas do idoso ao BDI. Conclusões: Idosos constituem um grupo particular e, como tal, apresentam especificidades relevantes. A avaliação dos idosos em relação às suas percepções de QV está associada a sexo, idade, estado civil, classe social, percepção de saúde e mais fortemente associada a níveis de sintomas depressivos. Explorando o cuidador como avaliador da QV do idoso observou-se uma tendência de o cuidador perceber a QV do idoso pior do que a própria percepção do idoso, apesar de fortes correlações para todos os domínios e na medida QV geral na percepção do par idoso-cuidador. Já na avaliação de percepção de saúde em idosos foi verificada a influência da intensidade dos sintomas depressivos bem como da dimensão independência: quanto menor a intensidade de depressão e quanto maior o nível de independência, maior associação com percepção de saúde entre idosos. Além desses, características próprias da população idosa podem interferir nos resultados do BDI fazendo com que seus achados sejam maximizados por questões somáticas e de desempenho sugerindo pontos de corte especiais para os idosos. Novos estudos são sugeridos a fim de atender a demanda específica do idoso.

Caracterização parcial e atividades farmacológicas do extrato rico em polissacarídeos sulfatatos da angiosperma marinha Halodule wrightii

Sulfated polysaccharides (PS) are biomolecules with a great biotechnological potential. There are few data about PS from high plants. In addition, pharmacological activities of PS from plants have not been carrying out. The aim of this work was extract PS from the angiosperm Halodule wrightii and study their anticoagulant and antioxidant activities. Histological analysis showed the presence of the PS manly in the roots. A polysaccharide-rich extract was obtained from H. wrightii by proteolysis followed by methanol and TCA precipitation. Chemical, infra-red analysis and agarose gel electrophoresis in 1.3 diaminopropane acetate buffer confirmed the presence of sulfated polysaccharides made by glucose, galactose, xylose and sulfate residues in the proportion 1: 0,9: 1: 1. In addition polyacrilamide electrophoresis have shown that extract is mainly compose by 11kDa sulfated polysaccharides. Pharmacological analysis have shown total antioxidant capacity (CAT) that resulted in 15,21 μg for equivalent of ascorbic acid, scavenging activity of the DPPH radical with 41,36 % of scavenging, activity of reducing power with the maximum of 0,290 nm (50 % of vitamin C activity) and scavenging activity superoxide radical (O2-) with a maximum of 32,23 %. Chelating activity of metal less than 4% and scavenging activity of the radical hydroxyl (OH-) less than 2%. Time of activated partial tromboplastin (aPTT) doubling the time of coagulation from 20μg of and protrombin time (PT) was not present. The data indicate that PS from Halodule wrightii could be considered for future applications in medicine, food production or cosmetic industry

Caracterização parcial e atividades farmacológicas do extrato rico em polissacarídeos sulfatados da angiosperma marinha Halodule wrightii

Sulfated polysaccharides (PS) are biomolecules with a great biotechnological potential. There are few data about PS from high plants. In addition, pharmacological activities of PS from plants have not been carrying out. The aim of this work was extract PS from the angiosperm Halodule wrightii and study their anticoagulant and antioxidant activities. Histological analysis showed the presence of the PS manly in the roots. A polysaccharide-rich extract was obtained from H. wrightii by proteolysis followed by methanol and TCA precipitation. Chemical, infra-red analysis and agarose gel electrophoresis in 1.3 diaminopropane acetate buffer confirmed the presence of sulfated polysaccharides made by glucose, galactose, xylose and sulfate residues in the proportion 1: 0,9: 1: 1. In addition polyacrilamide electrophoresis have shown that extract is mainly compose by 11kDa sulfated polysaccharides. Pharmacological analysis have shown total antioxidant capacity (CAT) that resulted in 15,21 μg for equivalent of ascorbic acid, scavenging activity of the DPPH radical with 41,36 % of scavenging, activity of reducing power with the maximum of 0,290 nm (50 % of vitamin C activity) and scavenging activity superoxide radical (O2-) with a maximum of 32,23 %. Chelating activity of metal less than 4% and scavenging activity of the radical hydroxyl (OH-) less than 2%. Time of activated partial tromboplastin (aPTT) doubling the time of coagulation from 20μg of and protrombin time (PT) was not present. The data indicate that PS from Halodule wrightii could be considered for future applications in medicine, food production or cosmetic industry

Avaliação do efeito do composto tipo heparina isolado do caranguejo Chaceon fenneri na hemostasia e na morte celular

Heparin is a pharmaceutical animal widely used in medicine due to its potent anticoagulant effect. Furthermore, it has the ability to inhibit the proliferation, invasion and adhesion of cancer cells to vascular endothelium. However, its clinical applicability can be compromised by side effects such as bleeding. Thus, the search for natural compounds with low bleeding risk and possible therapeutic applicability has been targeted by several research groups. From this perspective, this study aims to evaluate the hemorrhagic and anticoagulant activities and citotoxic effect for different tumor cell lines (HeLa, B16-F10, HepG2, HS-5,) and fibroblast cells (3T3) of the Heparin-like from the crab Chaceon fenneri (HEP-like). The HEP-like was purified after proteolysis, ion-exchange chromatography, fractionation with acetone and characterized by electrophoresis (agarose gel) and enzymatic degradation. Hep-like showed eletroforetic behavior similar to mammalian heparin, and high trisulfated /Nacetylated disaccharides ratio. In addition, HEP-like presented low in vitro anticoagulant activity using aPTT and a minor hemorrhagic effect when compared to mammalian heparin. Furthermore, the HEP-like showed significant cytotoxic effect (p<0.001) on HeLa, HepG2 and B16-F10 tumor cells with IC50 values of 1000 ug/mL, after incubation for 72 hours. To assess the influence of heparin-like on the cell cycle in HeLa cells, analysis was performed by flow cytometry. The results of this analysis showed that HEP-like influence on the cell cycle increasing S phase and decreasing phase G2. Thus, these properties of HEP-like make these compounds potential therapeutic agents

Síntese e caracterização do poli(ácido láctico) para potencial uso em sistemas de liberação controlada de fármacos

With the advances in medicine, life expectancy of the world population has grown considerably in recent decades. Studies have been performed in order to maintain the quality of life through the development of new drugs and new surgical procedures. Biomaterials is an example of the researches to improve quality of life, and its use goes from the reconstruction of tissues and organs affected by diseases or other types of failure, to use in drug delivery system able to prolong the drug in the body and increase its bioavailability. Biopolymers are a class of biomaterials widely targeted by researchers since they have ideal properties for biomedical applications, such as high biocompatibility and biodegradability. Poly (lactic acid) (PLA) is a biopolymer used as a biomaterial and its monomer, lactic acid, is eliminated by the Krebs Cycle (citric acid cycle). It is possible to synthesize PLA through various synthesis routes, however, the direct polycondensation is cheaper due the use of few steps of polymerization. In this work we used experimental design (DOE) to produce PLAs with different molecular weight from the direct polycondensation of lactic acid, with characteristics suitable for use in drug delivery system (DDS). Through the experimental design it was noted that the time of esterification, in the direct polycondensation, is the most important stage to obtain a higher molecular weight. The Fourier Transform Infrared (FTIR) spectrograms obtained were equivalent to the PLAs available in the literature. Results of Differential Scanning Calorimetry (DSC) showed that all PLAs produced are semicrystalline with glass transition temperatures (Tgs) ranging between 36 - 48 °C, and melting temperatures (Tm) ranging from 117 to 130 °C. The PLAs molecular weight characterized from Size Exclusion Chromatography (SEC), varied from 1000 to 11,000 g/mol. PLAs obtained showed a fibrous morphology characterized by Scanning Electron Microscopy (SEM)

Mutagênese induzida por flavonóides presentes do decocto das cascas da aroeira (Schinus terebinthifolius, Raddi)

The decoction of Brazilian pepper tree barks (Schinus terebinthifolius, Raddi), is used in medicine as wound healing and antiinflamatory. Once extracts from this plant are used for acceleration of scar s process, it is important to study their mutagenic and genotoxic potential. In previous works in our laboratory, it was observed mutagenicity caused by the decoction when in high concentrations. Among the chemical compounds of this plant that could be able to induce mutation, the flavonoids were the only group that was referred to have either an oxidant or antioxidant potential. The flavonoids were isolated, purified and quantified by adsorptive column chromatography under silica gel, bacterial and in vitro genotoxic tests were realized to determine if the flavonoids were the responsible agents for this mutagenicity found. The tests realized with plasmidial DNA were indicative that the flavonoids are probably genotoxic, due to the presence of correlation between increase of the flavonoid concentration and in plasmidial DNA double strand breakage visualized in agarose gel, as well as they were capable to generated abasic sites shown by the in vitro treatment with exonuclease III. The same tests with plasmidial DNA in the presence of copper [10 µM] and of a Tris-HCl pH 7.5 [10 µM] buffer were realized with the isolated flavonoids to determine if there would be or not participation of reactive oxygen species (ROS). The transformation of plasmidial DNA in different bacterial strains proficient and deficient in DNA repair enzymes in the presence or not of a Tris-HCl buffer, suggests that the enzymes that repair oxidative lesions are necessary to repair the lesions generated by the flavonoids and that ROS are generated and are necessary to promote the lesions. Bacterial tests with Escherichia coli strains of the CC collection (deficient or not for DNA repair enzymes), showed that the flavonoids are able to increase the frequency of mutations, mainly in strains mutated in repair enzymes (MutM, MutY-glicosylases and double mutant), suggesting that these agents are responsible for the enhancement in the mutation rate. In order to determine the mutation spectrum caused by the flavonoids of the Brazilian pepper tree stem bark, plasmidial DNA previously treated with the flavonoids were transformed in bacterial strains deficient and proficient in the DNA repair enzymes, followed by a blue-white selection with X-gal, DNA amplification by PCR and sequencing the positive mutant clones. Analysis of the mutants obtained from strains CC104, CC104mutM, CC104mutY, CC104mutMmutY, BW9101, BW9109 indicated a predominance of some mutations like G:C to C:G that can be correlated with the origin of 8-oxoG, due to oxidative lesions caused by the flavonoids. So it can concluded that the flavonoid isolated or in fractions enriched on them are genotoxic and mutagenic, and their mutations are predominantly oxidative, mediated by ROS, and the lesions are recognized by the BER system. In this way it is proposed that the flavonoids can act in two different ways to generate the DNA lesion: 1. in a Fenton-like reaction, when the flavonoid are in the presence of metal ions and that together with the water generate ROS that promotes the DNA lesions; 2. in another way the lesions can be generated by the formation of ROS due to the internal chemical structure of the flavonoid molecule due to the quantity and location of hydroxyl groups, and so producing the DNA lesions, those lesions can be directly (suggested by the in vitro experiments) or indirectly done (supported by the experiments using the CC bacterial strains)