Membrane properties of structurally modified ceramides : effects on lipid lateral distribution and sphingomyelin-interactions in artificial bilayer membranes

Ceramides comprise a class of sphingolipids that exist only in small amounts in cellular membranes, but which have been associated with important roles in cellular signaling processes. The influences that ceramides have on the physical properties of bilayer membranes reach from altered thermodynamical behavior to significant impacts on the molecular order and lateral distribution of membrane lipids. Along with the idea that the membrane physical state could influence the physiological state of a cell, the membrane properties of ceramides have gained increasing interest. Therefore, membrane phenomena related to ceramides have become a subject of intense study both in cellular as well as in artificial membranes. Artificial bilayers, the so called model membranes, are substantially simpler in terms of contents and spatio-temporal variation than actual cellular membranes, and can be used to give detailed information about the properties of individual lipid species in different environments. This thesis focuses on investigating how the different parts of the ceramide molecule, i.e., the N-linked acyl chain, the long-chain sphingoid base and the membrane-water interface region, govern the interactions and lateral distribution of these lipids in bilayer membranes. With the emphasis on ceramide/sphingomyelin(SM)-interactions, the relevance of the size of the SMhead group for the interaction was also studied. Ceramides with methylbranched N-linked acyl chains, varying length sphingoid bases, or methylated 2N (amide-nitrogen) and 3O (C3-hydroxyl) at the interface region, as well as SMs with decreased head group size, were synthesized and their bilayer properties studied by calorimetric and fluorescence spectroscopic techniques. In brief, the results showed that the packing of the ceramide acyl chains was more sensitive to methyl-branching in the mid part than in the distal end of the N-linked chain, and that disrupting the interfacial structure at the amide-nitrogen, as opposed to the C3-hydroxyl, had greater effect on the interlipid interactions of ceramides. Interestingly, it appeared that the bilayer properties of ceramides could be more sensitive to small alterations in the length of the long-chain base than what was previously reported for the N-linked acyl chain. Furthermore, the data indicated that the SM-head group does not strongly influence the interactions between SMs and ceramides. The results in this thesis illustrate the pivotal role of some essential parts of the ceramide molecules in determining their bilayer properties. The thesis provides increased understanding of the molecular aspects of ceramides that possibly affect their functions in biological membranes, and could relate to distinct effects on cell physiology.

Studies on membrane properties of cholesterol and 3-beta modified sterol analogs

Cholesterol (Chol) is an important lipid in cellular membranes functioning both as a membrane fluidity regulator, permeability regulator and co-factor for some membrane proteins, e.g. G-protein coupled receptors. It also participates in the formation of signaling platforms and gives the membrane more mechanical strenght to prevent osmotic lysis of the cell. The sterol structure is very conserved and already minor structural modifications can completely abolish its membrane functions. The right interaction with adjacent lipids and the preference of certain lipid structures over others are also key factors in determining the membrane properties of cholesterol. Because of the many important properties of cholesterol it is of value to understand the forces and structural properties that govern the membrane behavior of this sterol. In this thesis we have used established fluorescence spectroscopy methods to study the membrane behavior of both cholesterol and some of its 3β-modified analogs. Using several fluorescent probes we have established how the acyl chain order of the two main lipid species, sphingomyelin (SM) and phosphatidylcholine (PC) affect sterol partitioning as well as characterized the membrane properties of 3β-aminocholesterol and cholesteryl phosphocholine. We concluded that cholesterol prefers SM over PC at equal acyl chain order, indicating that other structural properties besides the acyl chain order are important for sphingomyelin-sterol interactions. A positive charge at the 3β position only caused minor changes in the sterol membrane behavior compared to cholesterol. A large phosphocholine head group caused a disruption in membrane packing together with other membrane lipids with large head groups, but was also able to form stable fluid bilayers together with ceramide and cholesterol. The Ability of the large head group sterol to form bilayers together with ceramide was further explored in the last paper where cholesteryl phosphocholine/ceramide (Chol-PC/Cer) complexes were successfully used to transfer ceramide into cultured cells.

Responses of soil respiration and barley growth to modified supply of oxygen in the soil

Selostus: Maan hengityksen ja ohran kasvun reagointi hapensaannin muutoksiin maassa

Pressure to change and resistance against it: consumer perceptions and acceptance of nutritionally modified foods

Selostus: Muutospaineet ja muutosvastarinta: kuluttajien suhtautuminen ravitsemuksellisesti muunnettuihin elintarvikkeisiin

Oxygen adsorption on Cu(211) and structurally and chemically modified Cu(100)

Kuparipinnan hapettuminen on viimevuosina ollut suosittu tutkimuskohde materiaalitieteissä kuparin laajan teollisuuskäytön vuoksi. Teollisuussovellusten, kuten suojaavien pintaoksidien kehittäminen vaatii kuitenkin syvällistä tuntemusta hapettumisprosessista ja toisaalta myös normaaliolosuhteissa materiaalissa esiintyvien hilavirheiden vaikutuksesta siihen. Tässä työssä keskitytäänkin tutkimaan juuri niitä mekanismeja, joilla erilaiset pintavirheet ja porrastettu pintarakenne vaikuttavathapen adsorptioprosessiin kuparipinnalla. Tutkimus on tehty käyttämällä laskennallisia menetelmiä sekä VASP- ja SIESTA-ohjelmistoja. Työssätutkittiin kemiallisia ja rakenteellisia virheitä Cu(100)-pinnalla, joka on reaktiivisin matalanMillerin indeksin pinta ja porrastetun pinnan tutkimuksessa käytettiin Cu(211)-pintaa, joka puolestaan on yksinkertainen, stabiili ja aiemmissa tutkimuksissa usein käytetty pintarakenne. Työssä tutkitut hilavirheet, adatomit, vähentävät molekyylin dissosiaatiota kuparipinnalla, kun taas vakanssit toimivat dissosiaation keskuksina. Kemiallisena epäpuhtautena käytetty hopeakerros ei estä kuparin hapettumista, sillä happi aiheuttaa mielenkiintoisen segregaatioilmiön, jossa hopeatyöntyy syvemmälle pinnassa jättäen kuparipinnan suojaamattomaksi. Porrastetulla pinnalla (100)-hollow on todennäköisin paikka molekyylin dissosiaatiolle, kun taas portaan bridge-paikka on suotuisin molekulaariselle adsorptiolle. Lisäksi kuparin steppipinnan todettiin olevan reaktiivisempi kuin tasaiset kuparipinnat.

A modified structural stress method for fatigue assessment of welded structures

Fatigue life assessment of weldedstructures is commonly based on the nominal stress method, but more flexible and accurate methods have been introduced. In general, the assessment accuracy is improved as more localized information about the weld is incorporated. The structural hot spot stress method includes the influence of macro geometric effects and structural discontinuities on the design stress but excludes the local features of the weld. In this thesis, the limitations of the structural hot spot stress method are discussed and a modified structural stress method with improved accuracy is developed and verified for selected welded details. The fatigue life of structures in the as-welded state consists mainly of crack growth from pre-existing cracks or defects. Crack growth rate depends on crack geometry and the stress state on the crack face plane. This means that the stress level and shape of the stress distribution in the assumed crack path governs thetotal fatigue life. In many structural details the stress distribution is similar and adequate fatigue life estimates can be obtained just by adjusting the stress level based on a single stress value, i.e., the structural hot spot stress. There are, however, cases for which the structural stress approach is less appropriate because the stress distribution differs significantly from the more common cases. Plate edge attachments and plates on elastic foundations are some examples of structures with this type of stress distribution. The importance of fillet weld size and weld load variation on the stress distribution is another central topic in this thesis. Structural hot spot stress determination is generally based on a procedure that involves extrapolation of plate surface stresses. Other possibilities for determining the structural hot spot stress is to extrapolate stresses through the thickness at the weld toe or to use Dong's method which includes through-thickness extrapolation at some distance from the weld toe. Both of these latter methods are less sensitive to the FE mesh used. Structural stress based on surface extrapolation is sensitive to the extrapolation points selected and to the FE mesh used near these points. Rules for proper meshing, however, are well defined and not difficult to apply. To improve the accuracy of the traditional structural hot spot stress, a multi-linear stress distribution is introduced. The magnitude of the weld toe stress after linearization is dependent on the weld size, weld load and plate thickness. Simple equations have been derived by comparing assessment results based on the local linear stress distribution and LEFM based calculations. The proposed method is called the modified structural stress method (MSHS) since the structural hot spot stress (SHS) value is corrected using information on weld size andweld load. The correction procedure is verified using fatigue test results found in the literature. Also, a test case was conducted comparing the proposed method with other local fatigue assessment methods.

A Study on Fractionation and Ultrafiltration of Proteins with Characterized Modified and Unmodified Membranes

Membrane filtration has become increasingly attractive in the processing of both foodand biotechnological products. However, the poor selectivity of the membranes and fouling are the critical factors limiting the development of UF systems for the specific fractionation of protein mixtures. This thesis gives an overview on fractionation of proteins from model protein solutions or from biological solutions. An attempt was made to improve the selectivity of the available membranes by modifying the membranes and by exploiting the different electrostatic interactions between the proteins and the membrane pore surfaces. Fractionation and UF behavior of proteins in the model solutions and in the corresponding biological solutions were compared. Characterization of the membranes and protein adsorptionto the membrane were investigated with combined flux and streaming potential studies. It has been shown that fouling of the membranes can be reduced using "self-rejecting" membranes at pH values where electrostatic repulsion is achieved between the membrane and the proteins in solution. This effect is best shown in UF of dilute single protein solutions at low ionic strengths and low pressures. Fractionation of model proteins in single, binary, and ternary solutionshas been carried out. The results have been compared to the results obtained from fractination of biological solutions. It was generally observed that fractination of proteins from biological solutions are more difficult to carry out owingto the presence of non studied protein components with different properties. Itcan be generally concluded that it is easier to enrich the smaller protein in the permeate but it is also possible to enrich the larger protein in the permeateat pH values close to the isoelectric point of the protein. It should be possible to find an optimal flux and modification to effectively improve the fractination of proteins even with very similar molar masses.

Role of lipid reactions in quality of oat products

Selostus: Rasvojen reaktiot prosessoiduissa kauratuotteissa

Welding of Sheet metal using Modified short arc MIG/MAG welding process

In this research work, the results of an investigation dealing with welding of sheet metals with diverse air gap using FastROOT modified short arc welding method and short circuit MAG welding processes have been presented. Welding runs were made under different conditions and, during each run, the different process parameters were continuously monitored. It was found that maximum welding speed and less HAZ are reached under specific welding conditions with FastROOT method with the emphasis on arc stability. Welding results show that modified short arc exhibits a higher electrode melting coefficient and with virtually spatter free droplet transition. By adjusting the short circuit duration the penetration can be controlled with only a small change in electrode deposition. Furthermore, by mixing pulsed MIG welding with modified arc welding the working envelope of the process is greatly extended allowing thicker material sections to be welded with improved weld bead aesthetics. FastROOT is a modified short arc welding process using mechanized or automated welding process based on dip transfer welding, characterized by controlled material deposition during the short circuit of the wire electrode to the workpiece.

Characterization of novel genes predominantly expressed in the epididymis – from genome analyses to genetically modified mice

In mammals, post-testicular sperm maturation taking place in the epididymis is required for the spermatozoa to acquire the abilities required to fertilize the egg in vivo. The epididymal epithelial cells secrete proteins and other small molecules into the lumen, where they interact with the spermatozoa and enable necessary maturational changes. In this study different in silico, in vitro and in vivo approaches were utilized in order to find novel genes responsible for the function of the epididymis and post-testicular sperm maturation in the mouse. Available online genomic databases were analyzed to identify genes potentially expressed in the epididymis, gene expression profiling was performed by studying their expression in different mouse tissues, and significance of certain genes to fertility was assessed by generating genetically modified mouse models. A recently discovered Pate (prostate and testis expression) gene family was found to be predominantly expressed in the epididymis. It represents one of the largest known gene families expressed in the epididymis, and the members code for proteins potentially involved in defense against microorganisms. Through genetically modified mouse models CRISP4 (cysteine-rich secretory protein 4) was identified to regulate sperm acrosome reaction, and BMYC to inhibit the expression of the Myc proto-oncogene in the developing testis. A mouse line expressing iCre recombinase specifically in the epididymis was also generated. This model can be used to generate conditional, epididymis-specific knock-out models, and will be a valuable tool in fertility studies.

“IT’S ALL RIGHT”. Multimodal rightward spatial bias modified by age and praxis

The general goal of the present work was to study whether spatial perceptual asymmetry initially observed in linguistic dichotic listening studies is related to the linguistic nature of the stimuli and/or is modality-specific, as well as to investigate whether the spatial perceptual/attentional asymmetry changes as a function of age and sensory deficit via praxis. Several dichotic listening studies with linguistic stimuli have shown that the inherent perceptual right ear advantage (REA), which presumably results from the left lateralized linguistic functions (bottom-up processes), can be modified with executive functions (top-down control). Executive functions mature slowly during childhood, are well developed in adulthood, and decline as a function of ageing. In Study I, the purpose was to investigate with a cross-sectional experiment from a lifespan perspective the age-related changes in top-down control of REA for linguistic stimuli in dichotic listening with a forced-attention paradigm (DL). In Study II, the aim was to determine whether the REA is linguistic-stimulus-specific or not, and whether the lifespan changes in perceptual asymmetry observed in dichotic listening would exist also in auditory spatial attention tasks that put load on attentional control. In Study III, using visual spatial attention tasks, mimicking the auditory tasks applied in Study II, it was investigated whether or not the stimulus-non-specific rightward spatial bias found in auditory modality is a multimodal phenomenon. Finally, as it has been suggested that the absence of visual input in blind participants leads to improved auditory spatial perceptual and cognitive skills, the aim in Study IV was to determine, whether blindness modifies the ear advantage in DL. Altogether 180-190 right-handed participants between 5 and 79 years of age were studied in Studies I to III, and in Study IV the performance of 14 blind individuals was compared with that of 129 normally sighted individuals. The results showed that only rightward spatial bias was observed in tasks with intensive attentional load, independent of the type of stimuli (linguistic vs. non-linguistic) or the modality (auditory vs. visual). This multimodal rightward spatial bias probably results from a complex interaction of asymmetrical perceptual, attentional, and/or motor mechanisms. Most importantly, the strength of the rightward spatial bias changed as a function of age and augmented praxis due to sensory deficit. The efficiency of the performance in spatial attention tasks and the ability to overcome the rightward spatial bias increased during childhood, was at its best in young adulthood, and decreased as a function of ageing. Between the ages of 5 and 11 years probably at first develops movement and impulse control, followed by the gradual development of abilities to inhibit distractions and disengage attention. The errors especially in bilateral stimulus conditions suggest that a mild phenomenon resembling extinction can be observed throughout the lifespan, but especially the ability to distribute attention to multiple targets simultaneously decreases in the course of ageing. Blindness enhances the processing of auditory bilateral linguistic stimuli, the ability to overcome a stimulus-driven laterality effect related to speech sound perception, and the ability to direct attention to an appropriate spatial location. It was concluded that the ability to voluntarily suppress and inhibit the multimodal rightward spatial bias changes as a function of age and praxis due to sensory deficit and probably reflects the developmental level of executive functions.

Fractionation of Hen Egg and Oat Lipids with supercritical Fluids Chemical and functional Properties of Fractions

Hen eggs and oats (Avena Sativa) are important materials for the food industry. Today, instead of merely satisfying the feeling of hunger, consumers are asking for healthier, biologically active and environmentally friendly products. The growing awareness of consumers’ increasing demands presents a great challenge to the food industry to develop more sustainable products and utilise modern and effective techniques. The modification of yolk fatty acid composition by means of feed supplements is well understood. Egg yolk phospholipids are polar lipids and are used in several applications including food, cosmetics, pharmaceuticals, and special nutrients. Egg yolk phospholipids are excellent emulsifiers, typically sold as mixtures of phospholipids, triacylglycerols, and cholesterol. However, highly purified and characterised phospholipids are needed in several sophisticated applications. Industrial fractionation of phospholipids is usually based on organic solvents. With these fractionation techniques, some harmful residues of organic solvents may cause problems in further processing. The objective of the present study was to investigate the methods to improve the functional properties of eggs, to develop techniques to isolate the fractions responsible for the specific functional properties of egg yolk lipids, and to apply the developed techniques to plant-based materials, too. Fractionation techniques based on supercritical fluids were utilised for the separation of the lipid fractions of eggs and oats. The chemical and functional characterisation of the fractions were performed, and the produced oat polar lipid fractions were tested as protective barrier in encapsulation processes. Modifying the fatty acid compositions of egg yolks with different types of oil supplements in feed had no affect on their functional or sensory properties. Based on the results of functional and sensory analysis, it is evident that eggs with modified fatty acid compositions are usable in several industrial applications. These applications include liquid egg yolk products used in mayonnaise and salad dressings. Egg yolk powders were utilised in different kinds of fractionation processes. The precipitation method developed in this study resembles the supercritical anti-solvent method, which is typically used in the pharmaceutical industry. With pilot scale supercritical fluid processes, non-polar lipids and polar lipids were successfully separated from commercially produced egg yolk powder and oat flakes. The egg and oat-based polar lipid fractions showed high purities, and the corresponding delipidated fractions produced using supercritical techniques offer interesting starting materials for the further production of bioactive compounds. The oat polar lipid fraction contained especially digalactosyadiacylglycerol, which was shown to have valuable functional properties in the encapsulation of probiotics.