Plasma membranes have a structural property, commonly referred to as membrane fluidity, that is compositionally regulated. The two main features of plasma membrane lipid composition that determine membrane fluidity are the ratio of cholesterol to phospholipids and the ratio of saturated to unsaturated fatty acids that are incorporated into the phospholipids. These ratios are determined, at least in part, by regulation of membrane lipid biosynthesis-particularly that of cholesterol and oleate. It now appears that cholesterol and oleate biosynthesis are feedback regulated by a common transcriptional mechanism which is governed by the maturation of the SREBP transcription factors. In this article, we briefly review our current understanding of transcriptional regulation of plasma membrane lipid biosynthesis by sterols and oleate. We also discuss studies related to the mechanism by which the physical state of membrane lipids signals the transcriptional regulatory machinery to control the rates of synthesis of these structural components of the lipid bilayer.

"Tautologically, castability is a critical requirement in any casting process. The two most important factors impacting castability are the susceptibility of a metal to hot tearing and the degree of casting fluidity a material possesses. This work concerns itself with fluidity of molten metal. Since experimental investigations into casting fluidity began, researchers have sought to maximize fluidity through superheat, mold temperature, alloy chemistry, melt cleanliness, and mold design. Researchers who have examined the published results in the field have remarked on the difficulty of making quantitative comparisons and drawing conclusions from the data. Ragone developed a horizontal vacuum fluidity apparatus and an analytical expression for fluid length to help resolve these issues. This was expanded on by Flemings et al. Still, the comparison of results is complicated by experimental uncertainties and a plurality of experimental procedures. This work seeks to resolve these issues through an analysis of experimental uncertainties present in existing fluidity tests and the development of an improved test and procedure which is very precise, accurate, and reliable. Certain existing tests and software packages have been shown to be unsuitable for quantitative fluidity measurement. Expressions for experimental uncertainty in fluidity testing have been derived. The capability to predict variations in fluidity as a function of alloy chemistry and other variables whose range of values are intrinsic to the economics of the process will help to more accurately determine the superheat needed for successful castings and will in turn lead to a decrease in scrap rates. This will enable metal casters to more reliably cast thin sections, and to reduce cycle time or scrap rate to achieve productivity goals. Superheat was shown to remain the dominant factor in fluidity, but the test allowed investigation of alloy modifications within an alloy specification in this alloy system. Factors known to have negative effects on structural properties were found often to have neutral or positive impacts on fluidity. A deep understanding of variations in fluidity measurements is the next necessary step in a century-long quest to understand how best to make metal castings through the use of fluidity experiments."

In this thesis optical microscopy techniques are used to consider aspects of viral and bacterial infections. In part 1, the physical effects of cytomegalovirus on lipid droplet dynamics in live cells are studied; in part 2, the effects of an antimicrobial peptide on the fluidity of model bacterial membranes are studied. The optical microscopy techniques used to study the effects of murine-cytomegalovirus (mCMV) on lipid droplets in live NIH/3T3 fibroblast cells in real-time are coherent anti- Stokes Raman scattering (CARS), two-photon fluorescence (TPF) and differential interference contrast (DIC) microscopies. Using a multimodal CARS and TPF imaging system, the infection process was monitored by imaging the TPF signal caused by a green fluorescent protein (GFP)-expressing strain of mCMV, where the amount of TPF detected allowed distinct stages of infection to be identified. Meanwhile, changes to lipid droplet configuration were observed using CARS microscopy. Quantitative analysis of lipid droplet numbers and size distributions were obtained from live cells, which showed significant perturbations as the infection progressed. The CARS and TPF images were acquired simultaneously and the experimental design allowed incorporation of an environmental control chamber to maintain cell viability. Photodamage to the live cell population was also assessed, which indicated that alternative imaging methods must be adopted to study a single cell over longer periods of time. To this end, DIC microscopy was used to study the lipid droplet dynamics, allowing lipid droplet motion to be tracked during infection. In this way, the effects of viral infection on the mobility and arrangement of the lipid droplets were analysed and quantified. It was found that the diffusion coefficient of the lipid droplets undergoing diffusive motion increased, and the droplets undergoing directed motion tended to move at greater speeds as the infection progressed. In addition, the droplets were found to accumulate and cluster in infected cells. The second part of this thesis presents a study on the effects of an antimicrobial peptide on model bacterial membranes. Giant unilamellar vesicles (GUVs) were produced as a simple model of E. Coli membrane using a 3:1 mixture of DPPC and POPG lipids. Incorporating Laurdan fluorescent dye into the lipid membrane of the GUVs allowed the membrane fluidity to be probed and visualised using TPF microscopy, whereby the fluidity was quantified by determining the general polarization (GP) values. Studying GUVs comprising single lipid and mixed lipid compositions over a temperature range from 25 C to 55 C enabled the lipid phase bands to be identified on the basis of GP value as gel phase and liquid crystalline phase. As such, the changes in lipid phase as a result of interaction with AMP were quantified, and phase domains were identified. It was found that the amount of liquid crystalline phase domains increased significantly as a result of AMP interaction.

Fluorescence spectroscopy and imaging have a very broad spectrum of applicationswithin the life sciences, in particular for detection and characterization ofbiomolecular dynamics and interactions in different environments. This thesis comprisesprojects that strive to further expand the information content extracted fromthe detected fluorescence, leading to sensitive readout parameters for studies ofbiomolecular dynamics and interactions. Two major strategies are presented toachieve this aim. The first strategy is based on the expansion of the availablereadout parameters beyond the "traditional" fluorescence parameters: intensity,wavelength, polarization and fluorescence lifetime. The additional parameters arebased on blinking properties of fluorescent labels. In particular on transitions betweensinglet and triplet states, and transitions between the trans- and cis-isomersof fluorophores. Two publications in the thesis are based on this strategy (paperI and IV). The second strategy is based on the utilization of fluorescence intensityfluctuations in order to detect the oligomerization mechanisms of fluorescentlylabeled peptides and proteins. This strategy combines the intensity fluctuationanalysis and the readout of distance dependent energy transfer between fluorescentmolecules together with the correlation analysis of fluorescence from two labeledproteins emitting at different wavelengths. Another two publications presented inthe thesis are based on the second comprehensive strategy (papers II and III).The work presented in this thesis shows that the blinking kinetics of fluorescentlabels contain significant information that can be exploited by a combination of fluctuationsanalysis with distance dependent excitation energy transfer between thefluorescent molecules, or by analysis of fluorescence covariance between moleculesthat emit at different wavelengths. These fluorescence-based methods have a significantpotential for molecular interaction studies in the biomedical field. / <p>QC 20160527</p>

The lipid bilayer membrane of enveloped viruses, such as human immunodeficiency virus type 1 (HIV-1), plays an important role in key steps of the infection, including cell binding and uptake. Phosphatidylserine (PS) and monosialotetrahexosylganglioside (GM1) are examples of two host-derived lipids in the membrane of enveloped virus particles that are known to contribute to virus attachment, uptake, and ultimately dissemination. A quantitative characterization of their contribution to the functionality of the virus requires information about their relative concentrations in the viral membrane. In this dissertation, a gold nanoparticle (NP) binding assay for probing relative PS and GM1 lipid concentrations in the outer leaflet of different virus-like particles (VLPs) using small sample sizes is introduced. The assay evaluates both scattering intensity and resonance wavelength and determines relative NP densities through plasmon coupling as a measure for the target lipid concentrations in the NP-labeled VLP membrane. The performed studies reveal significant differences in the membrane of HIV-1 and Ebola VLPs that assemble at different intracellular sites and pave the way to an optical quantification of lipid concentration in virus particles at physiological titers. In addition, this technique was used in another application to improve the understanding of the relationship between the membrane PS lipid and the infectivity of HIV-2 and murine leukemia virus (MLV). The composition of the membrane, in particular the cholesterol (chol) content, determines its fluidity. As differences in the membrane composition of individual virus particles can lead to different intracellular fates, biophysical tools capable of probing the membrane fluidity on the single-virus level are required. In this dissertation, we demonstrate that fluctuations in the polarization of light scattered off gold or silver nanoparticle (NP)-labeled virus-like-particles (VLPs) encode information about the membrane fluidity of individual VLPs. We developed a plasmonic polarization fluctuation tracking microscopy (PFTM) which facilitated, for the first time, the investigation of the effect of chol content on the membrane fluidity and its dependence on temperature on the single-VLP level. Chol extraction studies with different methyl-β-cyclodextrin (MβCD) concentrations yielded a gradual decrease in polarization fluctuations as function of time. The PFTM revealed chol content and fluidity heterogeneities of an HIV-1 VLP population.

Lintégrité de la structure et de la dynamique de la membrane plasmatique est essentielle à la fonction de la cellule. Cette intégrité peut être évaluée par la mesure de la fluidité membranaire globale, reflet de lensemble des mouvements des éléments membranaires au sein de la bicouche phospholipidique. Or lintégrité de la membrane est menacée, entre autre, par les modifications de la structure lipidique résultant de lipoperoxidations. Ces peroxidations lipidiques résultent des attaques radicalaires par des espèces oxygénées activées (EOA) produites lors dagression oxydante sur les acides gras membranaires. Nous posons lhypothèse que les conditions doxygénations extrêmes, qui peuvent être rencontrées lors dune anesthésie ou lors dun stress oxydant induit par lexercice chez le cheval, peuvent affecter la fluidité membranaire des érythrocytes et que ces variations peuvent être modulées par la modification de la structure membranaire du globule rouge par un supplément antioxydant oral adéquat. Lobjectif de ce travail est donc dévaluer les effets de différentes conditions doxygénation et doxydation in vitro (par contact avec différents mélanges gazeux), puis in vivo sous anesthésie générale (en faisant varier la fraction inspirée en oxygène) et à lexercice, et enfin dévaluer les effets dune supplémentation enrichie en acides gras de type oméga-3 sur la fluidité membranaire du globule rouge. Les faibles pressions partielles en oxygène dans le sang artériel (PaO2), obtenues in vitro par contact du sang avec un gaz anoxique et in vivo sous anesthésie par inspiration dair ambiant (<45mmHg et <60mmHg respectivement), nont pas eu deffet sur la fluidité ni sur la structure de la membrane érythrocytaire. On peut supposer que le stimulus est insuffisant ou que la protection de la membrane résulte dune capacité antioxydante du plasma et de défenses cellulaires suffisantes. Les pressions partielles élevées en oxygène dans le sang, obtenues in vitro par contact du sang avec de loxygène pur (PaO2>500mmHg), ont induit un stress oxydant modéré qui na pas affecté la structure phospholipidique de la membrane malgré la peroxidation des acides gras de type oméga-6. La fluidité membranaire na pas été affectée par ces facteurs. In vivo, les pressions partielles élevées en oxygène observées dans le sang (>200mmHg) ont été insuffisantes pour induire des peroxidations significatives et des modifications de la fluidité membranaire. En revanche, les valeurs élevées de PaO2 ont augmenté la sensibilité du sang à lhémolyse dans un premier temps, puis sa résistance 24 heures après un retour à la normoxie. Dans ces conditions aucun effet na été noté sur la viscosité du sang ni la perfusion musculaire. Par ailleurs, lexercice intense semble diminuer la fluidité membranaire du globule rouge chez le cheval de sport. Cette diminution sobserve dès 15 minutes après larrêt de lexercice et persiste 24 heures après. Il existe également des corrélations entre certains de ces marqueurs indirects et la fluidité membranaire. La supplémentation na pas eu deffet significatif direct sur lévolution de la fluidité membranaire observée au repos. Mais elle a pourtant influencé la structure de la membrane. En effet, la complémentation a induit une augmentation du pourcentage dacides gras de type oméga-3 contenus dans la membrane érythrocytaire ainsi que du ratio oméga-3/oméga-6 pendant la période de repos. Cela résulte de lincorporation sélective dans la membrane de lacide eicosapentaénoïque (EPA) et de lacide docosahéxaénoïque (DHA) apportés par voie orale. Mais aucune corrélation na été observée dans notre étude entre la composition en acides gras de la membrane et le marqueur de la fluidité membranaire. La supplémentation na pas eu deffet significatif direct sur lévolution de la fluidité membranaire observée à lexercice, mais en a limité la diminution immédiate. Il résulte des études menées que : les conditions doxygénation les plus extrêmes qui peuvent être rencontrées en conditions atmosphériques ne semblent pas affecter la fluidité de la membrane. En revanche, un exercice intense, associé à une demande énergétique accrue, peut induire une diminution de la fluidité membranaire en corrélation avec les marqueurs du stress oxydant. Des modifications de la structure membranaire en acides gras polyinsaturés à longue chaîne de type oméga-3 naffectent pas la fluidité membranaire mais modulent les effets du stress oxydant lors de lexercice. La fluidité membranaire des érythrocytes pourrait être considérée comme un marqueur direct du stress oxydant dans certaines conditions. Mais ce marqueur semble moins sensible et global que dautres marqueurs du stress cellulaire tels que le test dhémolyse ou la mesure de la concentration plasmatique de peroxydes lipidiques spécifiques. The maintenance of plasmatic membrane integrity is mandatory for cell function. This integrity can be assessed by the measurement of global membrane fluidity which is proportional to the whole rotational and lateral diffusion rates of membrane components within the phospholipid bilayer. Membrane integrity could be threatened by changes in lipid structure as a result of lipid peroxidation by free radical species during oxidative stress. We hypothesize that extreme oxygenation status present during anesthesia or during exercise-induced oxidative stress in the horse can alter erythrocyte membrane fluidity (EMF), and that these changes in fluidity depend on variations in erythrocyte membrane structure under the action of an appropriate oral anti-oxidant supplementation. The aims of the study was: to assess the effect(s) of various oxygenation and oxidative conditions firstly created in vitro (by contact between erythrocyte and different gaz mixtures), and secondly in vivo during general anesthesia (with varying inspired oxygen fractions) as well as during exercise. To assess the effects of an omega-3 fatty acid-enriched supplementation on EMF. Low partial oxygen pressures, both obtained in vitro and in vivo under anesthesia (respectively <45 and <60 mmHg) did not have any effect on EMF or membrane structure. Erythrocyte membrane may have been protected by an increase in plasmatic anti-oxidative capacity and cellular defenses. High partial oxygen pressures (>500 mm Hg) obtained in vitro induced a moderate oxidative stress which did not alter the phospholipidic structure of the membrane despite peroxidation of omega 6 fatty acids. Partial oxygen pressures obtained in vivo (>200 mm Hg) were unable to induce significant peroxidation and alteration in membrane fluidity. However, high PaO2 values initially increased sensitivity of blood to hemolysis, followed by a tendency towards resistance to hemolysis after 24hours. Intense exercise decreases EMF in the sports horse. This was observed as soon as 15 minutes after exercise and persisted during the recovery period 24 hours later. Correlations were found between oxidative stress indirect markers and membrane fluidity. Supplementation did not affect membrane fluidity but influenced membrane structure by increasing the pourcentage of omega-3 fatty acids and the omega3/omega6 ratio at rest. These changes resulted from selective incorporation into the membrane of orally provided EPA and DHA . However, we could not evidence a correlation between membrane composition and the marker of membrane fluidity (correlation-relaxation time Tc). During exercise, supplementation had no direct effect on variations of membrane fluidity but tapered its immediate decrease. In conclusion, our studies show that the most extreme conditions encountered under atmospheric conditions do not appear to affect EMF. However intense exercise combined with increased energetic requirements induces a decrease in EMF which correlates with variations in markers of oxidative stress. Modifications of membrane composition in long-chain omega-3 polyinsaturated fatty acids do not affect EMF but modulate oxidative stress during exercise. EMF could be a direct marker of oxidative stress under certain conditions but appears less sensitive and comprehensive than other markers of celllular stress such as the hemolysis test or the concentration in specific lipidic peroxidation products.

Polymers in concrete have received considerable attention over the past 30 years. Superplasticizers are one of the admixtures which have polymeric structure. In this study, polycarboxylate type slump-releasing dispersant, which is a copolymer of 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPS) and methacrylic acid (MAA), was synthesized in different feed compositions. The synthesis procedure of this copolymer was retrieved from literature. The derivatives of this water-soluble copolymer (AMPS-MAA) were synthesized by a macromonomer which was synthesized by the esterification of poly(ethylene glycol) methyl ether (PEG) with MAA (PEGMA) and then copolymerizing this macromonomer with AMPS monomer, the resulting copolymer is AMPS-PEGMA. In order to study the methyl group effect on fluidity, the other type of macromonomer (PEGA), composed of PEG and acrylic acid (AA), was synthesized and copolymerized with AMPS monomer, giving AMPS-PEGA. The structures of synthesized polymers were verified by NMR and FT-IR analysis. The slump-releasing effect of the synthesized copolymers was studied in terms of reaction pH, composition, molecular weight, amount of PEG side chains, and molecular weight of PEG side chains. The AMPS-MAA copolymer with 40% AMPS content was the most effective in promoting the fluidity of cement pastes. In scope of reaction pH, the AMPS-MAA copolymer, synthesized at a pH of 11, gave the most effective result on fluidity of the cement pastes. In copolymers of PEG acrylate macromonomers and AMPS monomers, copolymers with 5% PEG acrylate content showed the highest fluidity both in copolymers of PEGA and PEGMA. In copolymers with PEG side chains, the 15% AMPS-PEGA copolymer synthesized at pH of 6 gave the most effective result on fluidity of cement pastes. In the study of mechanical properties of the mortar samples prepared by the copolymers selected, AMPS-PEGA copolymer with 25% PEG content showed the highest flexural strength, and AMPS-MAA copolymer with 60% AMPS content and a reaction pH of 11 gave the highest compressive strength. In this study, zeta potential measurements were also performed to analyze the fluidity behavior of the copolymers.

A pintura como geradora da reflexão sobre as imagens produzidas: da sutileza e da dúvida. Partindo do suporte, tomar o material como determinante de uma visualidade delicada, de vazios, escorridos, vapores e condensações. Imagens que resistem em se mostrar, propostas tímidas, como pontuação de delicadeza em um cotidiano saturado de imagens. O corpo como medida: da ação artística e da fluidez que dela resulta. / Painting as a reflection on generating images produced: the subtlety and doubt. Starting from the support, taking the material as determinant of a visual delicate, empty, drained, vapors and condensations. Images that resist showing off, timid proposals, as score of delicacy on a daily saturated images. The body as measure: from the artistic action and fluidity that it brings.

A pintura como geradora da reflexão sobre as imagens produzidas: da sutileza e da dúvida. Partindo do suporte, tomar o material como determinante de uma visualidade delicada, de vazios, escorridos, vapores e condensações. Imagens que resistem em se mostrar, propostas tímidas, como pontuação de delicadeza em um cotidiano saturado de imagens. O corpo como medida: da ação artística e da fluidez que dela resulta. / Painting as a reflection on generating images produced: the subtlety and doubt. Starting from the support, taking the material as determinant of a visual delicate, empty, drained, vapors and condensations. Images that resist showing off, timid proposals, as score of delicacy on a daily saturated images. The body as measure: from the artistic action and fluidity that it brings.

Thesis (MTech (BioMedical Technology))--Cape Peninsula University of Technology, 2006 / In multiple sclerosis (MS) the myelin sheaths surrounding the axons in the brain are mainly affected by the disease process. Myelin consists for the most part of lipids and proteins. An abnormality in essential fatty acid metabolism is known to be present in patients with MS (Horrobin, 1979), reflected in a high ratio of n-6 to n-3 fatty acids in cell membranes. It has also been established previously that the pathogenesis of inflammatory disorders is aggravated by excessive consumption of n-6 fatty acids relative to n-3 fatty acids (Guesnet et al., 2005),and it has been shown that ingesting a larger proportion of n-3 fatty acids could be crucial in the regulation of cellular physiology and in the prevention of pathologies such as autoimmune and inflammatory diseases. Modern Western medical treatment for autoimmune diseases, which includes MS, involves the administration of immunosuppressive drugs, such as beta interferon, cortisone (prednisone), methotrexate and cytoxan, which reduce the effectiveness of the entire Immune system, and can have serious, sometimes life threatening, side effec1s (Perlmutter, 2006, htlp:/Iwww.msfac1s.org). It would therefore be of interest to investigate other options for treatment Although there is an extensive literature on fatly acids in MS, the actual details of the mechanisms of fatly add imbalances in MS have not been established. It would therefore be advisable to Investigate the abnormality of the MS cell membrane fatly acid profile. Previous studies focused on individual fatty acids, but it would be more relevant to investigate the relationships within and between the n-6 and n-3 series, and their effect on outcome, and to establish any possible cumulative effects, because the metabolism of fatty adds within the two series does have an effect on one another.