Forum Scientium Seminar Abstracts

May 15, 2013

 

Spinning an electroactive web towards cardiac tissue engineering
Amy Gelmi

This research is focused on developing implantable electroactive fiber scaffolds that will increase the differentiation ratio of mesenchymal stem cells into cardiomyocytes and thus increase the formation of novel cardiac tissue to repair or replace the damaged cardiac tissue after MI. Composite nanofibrous scaffold of poly(dl-lactide-co-glycolide) (PLGA) have been coated with biodoped polypyrrole to create an electroactive fiber scaffold, with controllable fiber dimensions and alignment.  The electrical properties of the polymers are an integral factor in creating these ‘intelligent’ 3-D materials; not only does the inherent conductivity provide a platform for electrical stimulation, but the ionic actuation of the polymer can also provide mechanical stimulation to the seeded cells. The biocompatibility of the polymer, PLGA scaffolds, and coated PLGA scaffolds has been investigated using primary cardiovascular progenitor cells.

Presentation of BILS (Bioinformatics Infrastructure for Life Sciences)
 

Malin Larsson, bioinformatician in BILS

BILS (Bioinformatics Infrastructure for Life Sciences) is a distributed national research infrastructure with support from the Swedish Research Council. The aim of BILS is to make bioinformatics support available for life science researchers in Sweden. BILS can provide bioinformatics expertise in several different areas, as well as give access to bioinformatics tools and data storage. The presentation will contain a brief presentation of BILS, the areas of expertise of BILS’s staff and the bioinformatics support forum (http://biosupport.se).

Photolithography for Bioelectronics
Erik Gabrielsson

Charged biomolecules can be transported in polymer matrixes by electrophoresis. In our group, we have exploited this feature to create miniaturized electrophoresis devices, which consists of micrometer-sized channels of ion exchange membranes. These devices are used to deliver specific biomolecules to cells and tissues both in vitro and in vivo. To build complex circuits for biomolecule delivery we use photolithography. This method allows for patterning of features down to the micrometer level on smooth surfaces. By adopting standard photolithography processing techniques, for example spin coating, UV-exposure, development and dry etching, to unconventional materials such as organic semiconductors and polyelectrolytes, we fabricate ionic equivalences to electronic components. The ionic diodes and transistors are useful as they combine non-linear current-voltage characteristics with the delivery of ions, thus enabling biomolecule delivery in an electronically addressed 2D array of pixels.

The Quest for Synthesizing Fucoidan-mimics and the Quandary of Multiple Analysis Methods
Mattias Tengdelius

For biomaterials used in regenerative medicine there is often need for biofunctionalisation to yield a material with biological properties tailored for its application. There are mainly two methods for this: Either to produce a material with inherent biological properties or to insert an additive exhibiting the desired properties. As a tool for the latter method we have produced a series of glycopolymers mimicking biological properties of a class of natural polysaccharides known as Fucoidan. Fucoidan possess desirable properties such as being anti-inflammatory, antiviral, anticancer and antithrombotic. During these syntheses I came upon analysis results that turned out to be conflicting in several aspects raising my thoughts on the possible benefits and disadvantages of using multiple analysis methods.
 

April 17, 2013

 

A chelator based sensor
Robert Selegård

Solid phase peptide synthesis (SPPS) is the standard chemical technique for synthesis of peptides and small proteins. Apart from SPPS offering a strategy to synthesize de novo designed sequences, it allows for an alternative way to produce naturally occurring peptides which are difficult to express in bacteria. The versatility of the technique allows for incorporation of L-, D- and unnatural amino acids. Alteration to peptide backbone is also made possible through this technique enabling the synthesis of branched and even circular peptides.

This presentation outlines the general principles of SPPS and how polypeptides that were derived from this technique were used in a chelator-based sensor for Phytic acid.

Mathematical Modeling Reveal Mechanisms of Insulin Resistance in Human Adipocytes
Elin Nyman

Insulin controls glucose homeostasis and failure in this control leads to insulin resistance and type 2 diabetes (T2D). Insulin act through its receptor at the cell surface of insulin responding cells. We study the network of proteins activated by insulin in adipocytes from patients diagnosed with T2D and compare with normal subjects. The diabetic state shows a reduced response to maximal insulin concentrations throughout the network of proteins. We see a reduced sensitivity to insulin for many of the proteins. We show with a mathematical modeling approach that this reduced sensitivity can be explained by a reduced feedback signal in the protein network. We combine this new insight with all our available data and are able to simulate the dynamic insulin response throughout the network and identify differences between normal and T2D insulin signaling in adipocytes. 

Development of mathematical models for operator training and simulation
Inga Gerlach

Training simulators are today successfully used for the training of technicians and engineers in a number of industrial and medical applications. We have recently shown their applicability in bioprocessing for training of plant operation, control and decision-making during production of important bioproducts such as recombinant proteins and ethanol.
The operator training simulator (BioProcessTrainer) uses structured mechanistic models that are integrated into the process control and simulation system WinErs. The simulator relied on the creation of a virtual plant operator environment based on the developed models where time can be accelerated or stopped. Simulator realism is highly dependent on predictability of the biological model part. In this presentation the model application is discussed and preliminary virtual simulation data for production of recombinant green fluorescent protein (GFP) are shown.

Mechanical and inflammatory effects on tendon healing
Malin Hammerman

Tendon rupture is a very big problem because tendons heal very slowly and a good treatment has not yet been established.  Mechanical loading seems to have positive effects on tendon healing because unloading of healing tendons results in weaker tendons. Loading creates deformation of the extracellular matrix and cells in tendons which give rise to intracellular signaling, increased gene expression and protein synthesis. Previous studies in our group have shown that short daily loading episodes during early tendon healing could potentially be beneficial for rehabilitation. Additionally, gene expression studies have shown that loading might have a role in regulating the inflammatory response during tendon healing. The aim of my project is to investigate the effect of mechanical loading, inflammation and the connection between mechanical loading and inflammation in tendon healing.
 

March 20, 2013

Collagen in-vitro studies for the myocardial extracellular matrix
Abeni Wickham

It has been a well developed thought that telopeptides are necessary for fibril formation in collagenous materials. These peptides are found at the N and the C terminus of the collagen chain and when are intact are hypothesized to initiate fibril formation. If these telopeptides are removed it should make fibril formation much more difficult. Our initial results show that this is not so.
Three different collagens, rat-tail tendon, Nippon Ham porcine, and fish collagen were used and all had the same amount of fibril formation within the same conditions. Nippon Ham is known to be without telopeptides, as it would elicit immune reasponses. This signals that there is something more influential than telopeptides availability for fibril formation. Homogenous fibrous mats consisting of 70nm fiber diameter were fabricated. These initial results illustrate that using both plastic compression with simplified buffer solutions and crosslinking system we can produce a resilient scaffold for regenerative medicine purposes. SEM images depict nanostructured fibrous materials formed from phosphate buffer saline solutions. They also show the lack of morphological changes when crosslinked which would help to keep the influence on changes in morphology on the cells at a minimum. This technique, since it can form fibrils in a mild environment can also allow for cell encapsulation during the compression. Here we intend to use HL-1 cells expressing lysly oxidase to crosslink the material as the cells are encapsulated, replicating the exact crosslinking technique that occurs naturally. Since there are little morphological changes when crosslinked this material should provide a porous and stable environment for diffusion of nutrients and general upkeep of the cells while encapsulated.

Switchable Bioelectronics
Onur Parlak

Natural biochemical processes are controlled and regulated by different mechanisms at various levels of complexity involving numerous biochemical and physiological reactions. Artificial biocatalytic and particularly bioelectrocatalytic processes used in various bioelectronic devices, such as biosensors, biofuel cells, and bioactuators, are suffering from the lack of the biochemical control and regulation. Recently developed switchable and tunable bio-electrocatalytic systems are controlled by various physical input signals: electrical, magnetic, light, etc. allowing their applications in novel biosensors and biofuel cells with an adjustable performance. However, this approach does not provide biochemical/ physiological control over their operation. There is an obvious need of another kind of their functional regulation based on an interface between bioelectronic systems and their biochemical environment. The present study addresses the challenging aim of biochemical control over bioelectronic systems by integrating a bioelectrocatalytic system with a stimuli-responsive material and enzyme-catalysed stimuli-generating reactions.

Needle in the haystack: Discovering Biomarkers for disease diagnostics
Jacob Kuruvilla

The three most important barriers that prevent from providing a potential clinical diagnosis of a diseased condition are:  the lack of a strong disease-associated protein or genetic markers; absence of an easy, riskless and affordable sampling method; lack of an accurate, portable and ease to use diagnostic platform. Human saliva is a non-invasive body fluid with enormous potential to reveal hints what is happening in the whole organism. The salivatory proteome is a complex mixture from proteins secreted from the salivary glands and additional component from the oral environment such as epithelial cell, and microorganisms. In the last years, the salivatory proteome has started to be explored as a possible source for novel diagnosis for systemic disease including cancer and cardiovascular diseases.
The utilization of saliva for the diagnosis of infectious diseases has been successful for Malaria diagnosis. However, other common infectious diseases worldwide, such as dengue fever, have not been explored deeply. The developing of early diagnostic tools for those infectious diseases in developing countries should fulfill additional requirement as such: minimal infectious risk in sample collection for the patients, cost-effective approach for large populations, and minimal requirement of instrumentation that ideally, not electrical source dependent. The aim of this project is to apply saliva proteomics as a source of knowledge to develop low-cost diagnostic tools for infectious diseases.

 

February 20, 2013

 

 

 

 

An alternative method to produce microfluidic devices using an open source 3D printer
Katarina Bengtsson

Microfluidic devices for doing biomedical analysis and diagnosis show big potential. They are traditionally made by combining several techniques such as photolithography and in the end assembled to a final device. We are instead of the more traditional technique trying to use an open-source 3D printer to produce microsystems. The type of printer used in this project has a syringe-based printing tool which allows for a wide range of materials. This method allows a bottom-up approach and more individual design of microsystems containing several different parts. For example allowing the printer to first place electrodes followed by a channel and casting of the structure. The first trials are promising and structures down to 400 µm in width and 20 µm in height have been produced. At present the focus is to develop materials suitable for the3D- printer to produce entire microfluidic device.

 

Building up on the mathematical model for insulin signaling in Type 2 Diabetes
Meenu Rajan

Insulin resistance outlines defect in insulin signaling where the insulin responsiveness of tissues namely, adipose tissue, muscle and liver is diminished. However, due to fragmented research, the mechanisms involved are still unclear. Our group has recently reported a systems-level mechanistic understanding of insulin resistance, based on which a dynamic mathematical model of insulin signaling has been developed. Our aim now is to expand the existing model to include the mitogenic effects of insulin and also add another level to the model by including the dynamics of the transcription factors such as Foxo1 and Elk-1. Inclusion of both metabolic and mitogenic effects of insulin in the model will not only help in identification of novel drug targets but also enable different pharmaceutical companies to assess carcinogenic potential of different insulin analogues.

Electrochemically Synthesis Imprinted Polymer for Quorum Sensing Molecule Detection
Mohsen Golabi

Quorum sensing (Q.S) molecules are small biomolecules produced by bacteria and used as a chemical language to evaluate their population density. Many of biofilm formation factors and virulence genes are express by the presence threshold amount of Q.S molecule in the local environment.
Electrochemical synthesis of molecular imprinting is a process in which a target molecule (the imprint molecule) plays a role as a substrate during electropolymerization of functional monomers in a solvent. Subsequently, the target is removed. So, cavities remain in the polymer network, capable to rebind specifically to the target molecule, due to matching in shape, size and functionality. 
The preliminary aim of the project is to fabricate molecular imprinting polymer with high affinity and specificity for Q.S molecule. In this step, both conductive and nonconductive polymers are being used. Target recognition efficiency of imprinted polymer could be evaluated electrochemically and also by using SEM, AFM and FTIR.

Nanotoxicology: Human dermal microvascular endothelial cells exposed to TiO2 nanoparticles and Single walled carbon nanotubes (SWCNTs)
Narges Bayat

In The field of nanotoxicology, the study of the effects of engineered nanoparticles on the vascular system as a key route of exposure is still in its infancy. Also, currently there are no standard methods for the study of the effects of nanoparticles on cells and scientist rely on assays previously used for assessing pharmaceutical and others chemicals’ toxicity. However due to the unique properties of NPs these methods usually are prone to errors. There is therefore urgent need for more accurate and detailed methods that can provide a global description of changes in the cells and define inter-relationship between the cellular elements. This research project is designed to address a number of important issues regarding nanoparticle toxicity, e.g. effects on cellular ultrastructure, adverse effects on fundamental cellular responses related to membrane integrity, proteins S-nitrosylation, DNA damage, mitochondrial function and oxidative damage as well as effects on the endothelial cells tube formation ability. In this presentation I will describe the methods we use to characterize the physicochemical properties of the nanoparticles as well as some of the methods we use to assess the effects TiO2 nanoparticles (1-3nm and 30 nm) and Single walled carbon nanotubes (SWCNTs) on human dermal microvascular endothelial cells.
 

January 23, 2013

Immunological characterization of a type 1 diabetes high risk group
Linda Åkerman

The etiology and pathogenesis of type 1 diabetes is still not fully established. Autoantibody screening among 17000 Swedish children has identified a group of 22 individuals that have a high risk to get the disease, and by studying this group we want to get a better picture of what is really going on in their immune system during the period preceding disease onset. We may also have a chance to find new markers of risk, and to find ways of distinguishing risk individuals that eventually get the disease from those who don’t. This information could be very useful in prevention and intervention trials. There are numerous ways by which this group can be studied, but the first step will be to analyze gene expression in circulating immune cells from the risk individuals and compare it to that of healthy children and type 1 diabetics. This is done by multivariate qPCR, measuring the expression of 88 different genes simultaneously. Based on the results, more analyses and methods will be added. 

Bioresponsive Polypeptide-based Hybrid Nanomaterials for Regenerative Medicine
Christopher Aronsson

The development of biologically active scaffolds that interact with the host’s tissues to induce regeneration of damaged organs has become the strategy of choice in regenerative medicine, as it is now recognized that stem cells alone are unable to reconstitute damaged tissue ablated by disease, trauma or surgical intervention. Different biological materials can provide an environment that allows stem cells to differentiate into desired cell types and thereby repair and regenerate lost tissue.

In my research I aim to develop a modular approach for fabrication of responsive and tunable biomaterials based on polypeptides that self-assemble into chemically and structurally well-defined fibers and hydrogels that mimic the extra cellular matrix. The purpose of this approach is to obtain a tool-box of materials that can be utilized for restoring damaged and dysfunctional tissues.

In today’s presentation I will present my research, show some preliminary results and give a briefly introduction to RP-HPLC – one of the most important technique used in my research.

From Flow Cytometry to Mass Cytometry – new multiparameter cytometry technique coming to LIU
Mikael Pihl
Traditional fluorescence based flow cytometry allows enumeration and analysis of single cells at high speed. Fluorescently tagged antibodies or intercalating molecules facilitates quantitation of surface protein expression, DNA content and protein phosphorylation. An inherent problem with fluorescence based measurements is the wide emission spectra of fluorescent molecules, which inevitably overlap when several are present simultaneously.
Atomic Mass Spectrometry allows detection of individual isotopes. The new Mass Cytometry technique utilizes this by replacing fluorescent tags with chelated lanthanoid atoms as tags for monoclonal antibodies. This removes the problem of spectral overlap completely, as atoms with different mass can be distinguished. The amount of stable rare-earth isotopes available also allows for 40 simultaneous tags at present, while the theoretical number of tags is close to 100.
LIU will become a national center for this new technique, which should be up and running this autumn or early in 2014. 

November 7, 2012

THE INFLUENCE OF POLYMER THICKNESS ON THE SETTLEMENT AND ADHESION BEHAVIOUR OF MARINE ALGA ULVA
Wetra Yandi

Marine biofouling is a worldwide phenomenon which has become more and more important due to its great economic and environmental impact.  The fouling behavior and adhesion mechanisms of marine organisms on different substrata is an important issue to address in order to open up new strategies for designing effective antifouling materials. Surface properties, for example topography, wettability, stiffness, thickness, charge and so on determine the surface selection by organisms prior to settlement on the substrate. In this research work we are focusing on the role of thickness and charge of polymer surfaces on the adhesion behavior of marine alga Ulva as there are few studies have been done on these two properties in correlation to marine biofouling field.

Different thicknesses (50, 100, 200, 400, 600 and 1000 Å) of poly(HEMA-co-PEGMA) and differently charged (positive, negative, neutral and zwitterionic) polymer brushes have been succesfully prepared via Surface Initiated Atomic Transfer Radical Polymerization (SI-ATRP).  Positively, negatively and zwitterionic charged polymer surfaces were prepared from 2-dimethylaminoethyl methacrylate (DMAEMA), 3-Sulfopropyl methacrylate (SPMA), and sulfobetaine methacrylate (SBMA), while for neutral charged polymer surfaces were prepared from HEMA and PEGMA. These samples were prepared at an identical thickness (approximately 100 Å).

The spore settlement and adhesion strength assays of marine alga Ulva have been used in this work. Massive, rapid and anomalous settlement of spores occurred on the positively charged polymer surfaces. The adhesion strength assay also revealed that the interactions between spores and this surface were relatively strong. The characteristic of spore settlement and adhesion strength on negatively, neutral and zwitterionic charged polymer surfaces were expressed by lower settlement and weak adhesion strength. We also found that Polymer thickness influences the adhesion behavior of marine alga Ulva. The optimum polymer thicknesses, where Ulva spore settlement is lower, and spore removal is higher, was achieved in the range 200-400 Å. The settlement experiments are complemented with physical characterization of the prepared surfaces, using Fourier-transform infrared reflection-absorption spectroscopic (FT-IRAS), null ellipsometry , atomic force microscopy, and contact angle measurements.

Histone H3 methylation at lysine 4 in human adipocytes in correlation to owerweight and insulin resistance
Åsa Jufvas

Epidemiological evidence indicates yet unknown epigenetic mechanisms underlying a propensity for overweight and type 2 diabetes. Specific histone modifications have been proposed to be involved in the pathogenesis of these disorders. In adipocytes the  methylation of lysine 4 in histone H3, a modification associated with transcriptional activation,  was recently shown to be differently regulated at metabolically important genes depending on nutritional status. We analyzed the extent of lysine 4 of histone H3 in primary human adipocytes from 27 subjects using modification-specific antibodies. The level of lysine 4 dimethylation correlated inversely with body mass index of non-obese subjects. Moreover, overweight and type 2 diabetic subjects exhibited close to half the level of dimethylation compared with control subjects. In contrast, trimethylation of lysine 4 was more abundant in adipocytes from diabetic subjects and correlated with insulin resistance of non-diabetic subjects. The findings define genome wide molecular modifications of histones in adipocytes that are directly associated with obesity and diabetes, and thus suggest a molecular basis for existing epidemiological evidence of epigenetic inheritance

Fucose on a chip - Challenges when three groups combine three techniques!
Per Erlandsson

Here we demonstrate a simple microfluidic device and imaging system to measure the concentration of fucose in urine samples. Fucose in urine is a well-established indicator of liver disease and several forms of cancer, and accordingly an attractive candidate for routine screening if suitable instrumentation was available.
The method uses active transport to complete the preparatory stages of a fucose fluorescence assay, and it is measured with a simple optical arrangement based on a blue laser and disposable PDMS optics for total internal reflection excitation. The disposable chip and optics are integrated in a regular glass slide and configured for imaging evaluation using cell phone cameras.

Microbial Agents Stimulate Aquaporin 9 Expression in Human Primary Inflammatory Cells
Angelika Holm

The aquaporins (AQPs) are a family of water channels involved in cell volume and shape regulation through water transport in and out of the cell, and they are thereby of importance in a number of cellular events. The aquaglyceroporin AQP9 has been suggested to play a pivotal role in inflammatory cell migration, differentiation and metabolism as well as in chronic inflammation like Rheumatoid Arthritis (RA) and Inflammatory Bowel Disease (IBD). Thus, we investigated the role and regulation of AQP9 in human primary immune cells in response to inflammatory stimuli, especially the effects of three distinct microbial agents, viz. Salmonella lipopolysaccharide (LPS), Rotavirus enterotoxin NSP4 and quorum-sensing molecule N-3-oxo-dodecanoyl-homoserine lactone (3O-C12-HSL) as analyzed with molecular techniques. qPCR analyses of human primary macrophages obtained from healthy blood donors showed a significant increase in the relative mRNA expression of AQP9 upon LPS stimulation. Western blots further confirmed this effect at the protein level. Human primary neutrophils responded in a similar way. Stimulation with 3O-C12-HSL also increased the AQP9 mRNA expression significantly and preliminary data show a slight effect on the AQP9 expression after NSP4 stimulation. These data implies a role for AQP9 in inflammatory responses.

October 10, 2012

Functionalized protein fibers
Fredrik Bäcklund

 

When heated in aqueous acid, the protein insulin self-assembles into nano-wire structures known as amyloid fibrils. Working with a previously developed method of preparing amyloid fibrils functionalized by hydrophobic molecules, we have extended that procedure to include several different functionalizing guest compounds, thus enabling the creation of numerous hybrid materials with distinctly varying properties. Recently, special attention has been put on preparing fibers functionalized with the fluorescent oligomer α-sexithiophene (6T). Specifically, we have investigated the distinct changes in emission properties that 6T displays during the self assembling formation of the amyloid fibrils and the effects 6T has on amyloid self assembly. In addition, variations on the preparation methodology have revealed interesting properties about the process of protein fibril formation and aggregation.
 

 

Transgenic mice - elucidating the molecular mechanisms in hematopoietic stem cells regulating oxidative stress-resistance in hypoxia: a key role for HIF-1α?
Camilla Halvarsson

Hematopoietic stem cells (HSCs) are thought to reside in specialized niches within the bone marrow, which protects the stem cells from depletion of exaggerated cell cycle progression, allowing them to stay in a relatively quiescent state. Indirect evidence suggests that these niches create a low-oxygen, hypoxic, environment that limits the production of reactive oxygen species (ROS), thus protecting HSCs from excessive ROS stress. In recent years, others and we have also demonstrated that hypoxia favors a slow turnover of HSCs and sustains survival. We have shown that hypoxia protects HSCs from oxidative stress-induced cell death by challenging Lineage^-Sca-1⁺c-kit⁺ (LSK) cells with ROS increase, but the exact mechanisms are still unknown.

Transgenic animals are widely used to study the effects of deleting or overexpressing a specific gene. Trying to elucidate the underlying molecular mechanisms of hypoxia-induced protection, we have used a mouse model lacking expression of HIF-1α, a major regulator of the cellular response to hypoxia. In this knockout mouse, HIF-1α is conditionally ablated after inducing the cre-loxP system specifically in hematopoietic cells. I will describe this technique more in detail and present our recent data on the role of HIF-1α in hematopoietic stem cells.

 

September 12, 2012

Fishing for viruses in biology’s soup - A metagenomic approach to virus discovery
Fredrik Lysholm

Virus infections impose a huge disease burden on humanity. Currently, diagnostic tools and methods employed to find viruses are biased and require knowledge about the potential viruses present in the sample. Therefore, we present a new unbiased method to virus discovery based on metagenomic sequencing and bioinformatic analysis. Through the use of a bioinformatic pipeline based on comparisons towards known sequence a wide range of known and unknown virial pathogens can be discovered in patient samples. Furthermore, we have developed several new methods which improve the characterization accuracy as well as the ability to distinguish even very distant homologs to known viruses. We have employed this method on various samples resulting in an increased knowledge about the viruses present as well as the discovery of several new viral pathogens.

 

GPCR models in yeast as an example of solving problems in biological research
Daniel Sanchez

There will be an introduction about my background and a short view about my career. It will follow a summary about what I can give in terms of personal experience: tools to manage disappointments, strategies to solve problems, I would briefly give some famous examples of (Einstein, Watson & Crick), philosophy of science, auto criticism,….

The rest of the presentation would be about expression methods of GPCR (G protein coupled receptors) in yeast as a solution to study a group of human proteins in isolated models of high interest in pharmaceutical research. This will connect with the first part of the speech as a way to solve problems and the search of backdoors to make valid approaches.

 

Future developments of ion delivery devices - where to go?
Amanda Jonsson

We have been working with a technology for ion delivery, for more than 7 years in our group. You have probably heard Klas Tybrandt talking about his ion transistors, or Erik talking about his ion diodes, or any of us talking about ion pumps. What it is all about is the development of a technology that can deliver charged molecules. We want to be able to control the delivery rate with an electrical signal, so that we can switch the delivery on and off, and so that we can achieve different delivery rates.
What is special about our technology is that there is no fluid flow along with the molecules. The molecules are released at a small outlet (typically 10 um), and diffuse from that point. We have been aiming at turning the ion delivery technology into a tool for cell biology, where you can culture cells on top of the device. In parallel, we develop devices that can be used in vivo, for example as a drug delivery system.
What I would like to do today though, is to ask you if you can forsee any potential applications for our ion delivery technology? Maybe this could inspire us where to take this technology in the future.

A new locomotor assay for Drosophila melanogaster
Linda Helmfors

Fly behavior, such as climbing, can be an important tool for understanding disease and I will present a locomotor assay that in a three dimensional way detects early differences in for example velocity, height reached and turn tightness between fly variants.

April 18, 2012

Circular Dichroism (CD) spectroscopy: a method for study secondary structure of proteins
Sara Helander

Investigation of the structure and the biophysical properties of human proteins are of major importance and may lead to improved understanding of various diseases. A range of techniques are used to address questions related to protein structure and function. One of the most used methods is Circular dichroism (CD) spectroscopy, which can be used to study secondary and tertiary structures of biomolecules. This presentation will cover the basic principle behind CD; together with some practical examples from a previous study where CD spectroscopy has been used to investigate the secondary structure for c-Myc95-158wt and two different mutants. Our data shows that there is no structural change between the wild type protein and the different mutations. This confirms that the regulatory effect that these mutations have on different cell lines not is a consequence of loss or change of structure. Further, this study is an example of how biophysical studies of proteins, in this case CD spectroscopy, can be used to solve different questions related to cell biology studies.
 

Modeling of insulin signaling in primary human fat cells
Cecilia Brännmark

The rates of type 2 diabetes are rapidly rising all over the world and especially the less developed world with poor healthcare is facing massive problems with the consequences. The onset of type 2 diabetes is always preceded by insulin resistance in the fat cells and this seems to give rise to insulin resistance in other tissues as well. We study the differences in insulin signaling between fat cells from people with and without type 2 diabetes using a combined experimental and mathematical modeling approach. We are currently working on the first mathematical insulin signaling model for humans with both normal and type 2 diabetes data. The main experimental method is quantitative western blot (WB) and this method will be the focus of my presentation.

Multifunctional nanoparticles for MRI and fluorescence bioimaging
Linnéa Selegård

Multifunctional nanoparticles comprising both luminescent and magnetic properties have been produced. The particles are composed of a luminescent ZnO core decorated with magnetic Gadolinium (III) ions. The nanoparticles are promising for use as contrast agents in MRI as well as fluorescent probes in fluorescence imaging. Nanoparticles with Gd/Zn ratio ranging from 0.067 to 0.67 has been studied. The fluorescence and realxivity properties have been studied as a function of Gd/Zn ratio. It turned out that the quantum yield increased with increasing amount of Gd. The multifunctional nanoparticles also showed greatly enhanced relaxivity compared to commercial contrast agents as well as pure Gd2O3 nanoparticles. Photoemission Electron Microscopy was used to investigate the distribution of Zn and Gd in nanoparticle aggregates. It turned out that both Gd and Zn is homogeniously distributed in the aggregates indicating that the ZnO nanoparticles are decorated with Gd (III) ions and that pure Gd2O3 nanoparticles are not formed.

Strain-specific characterization of prion aggregates using conformation-sensitive probes
Karin Magnusson

Prions are infectious proteins composed of the abnormal disease-causing isoform PrPSc, a variant of the cellular prion protein PrPC. The conformation of PrPSc varies and this phenomenon is most likely giving rise to distinct prion strains. Prion strains are derived from transmissible spongiform encephalopathy (TSE) isolates that even after inoculation into genetically identical hosts, cause disease with distinct protein aggregate deposition, incubation times, and pathology. We have previously used Luminescent Conjugated Polythiophenes (LCPs), which emit conformation-dependent fluorescence spectra, for characterizing prion strains. When the LCP is bound to entities with conformational variation, the fluorescence spectra reflects differences in the local environment via shifts in wavelength, intensity, and vibrational substructure. Together with biochemical and histopathologically features, LCPs can be used to investigate disease pathogenesis through strain competition and interaction. The studies show that prion strain interactions are selective and strain dependent. In addition, we have synthesized novel, chemically defined Luminescent Conjugated Oligothiophenes (LCOs) that can been used for in vivo labeling of prion deposits and for staining of prion deposits in other organs such as the spleen.

March 14, 2012

Proteomic studies on mixed microbial communities
Jutta Speda
The identification of new enzymes for use in industrial biotechnology is an important goal in enzyme discovery. The need for bio-based energy and materials especially give rise to new strategy opportunities of proteomic research. We are currently developing a meta-proteomic methodology for bio prospecting of secreted proteins from complex microbial communities. Those extracellular proteins found can be expected to be highly active and stable in their natural environment. As a model system, we are using a biogas producing methanogenic community. We have developed a multistep sample preparation method that has enabled us to visualize the secretome of the microbial community despite the many disturbing metabolic substances present.

MOLECULAR PROBES FOR DETECTION OF PROTEIN AGGREGATES USING PET AND SPR
Leif Johansson

The formation and accumulation of protein aggregates, amyloid, give rise to distinct pathological conditions known as amyloidosis. Alzheimer’s and Parkinson’s diseases are both examples of neurodegenerative conditions resolving on local amyloid accumulation in the brain. To be able to study these diseases, molecular probes that selectively identify amyloid are needed as a research tool. Luminescent conjugated oligothiophenes (LCOs) have earlier been proven to be utilized for in vivo imaging of protein aggregates using fluorescence microscopy.
In this part of my project I report the synthesis of a novel asymmetric functionalized thiophene backbone. This new molecule will provide the possibility to use other techniques than fluorescence, such as Positron Emission Tomography (PET), Surface Plasmon Resonance (SPR) and immuno assays for detecting the interaction between the LCO and amyloid fibrils.

How I grow my research tree and cut off the branches to complete a thesis
Feng-I Tai

As planning to have a dissertation in 8 month, this is probably the last presentation I’ll make for Forum monthly meeting. I would like to take this chance to discuss about different point of view of how we actually spend time in our research life as a phd student.
Back to the time when I started my projects on Day1 of my phd, honestly I had no confidence to believe these projects would work, neither a clear trace to achieve the goal. As long as keeping growing the tree of my research, eventually I have come to a point of making decisions regarding what should be included in my thesis. It is hard to leave some parts which were not completed, because of the time and effort that has been spent. And more, is the fun and joy brought from some good days that I thought I was improved.
However, everything comes to an end, and that is the phd thesis. We all have to make decision to achieve this final goal. I am on my way to make a plan for it, and would like to share experiences of cutting branches of my research tree.

An Activating Conformational Change Introduces Flexibility in the Kinase Domain of Ephrin type-B Receptor 2
Alexandra Ahlner

Kinases are a group of proteins that phosphorylate other proteins and are as such important parts of several complex cell-signaling paths.
Kinase structures have been mostly studied by X-ray crystallography, which generally generates a relatively static picture, but it has been suggested that the kinases need flexibility to be active. We have used nuclear magnetic resonance (NMR) spectroscopy and Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion experiments to investigate the dynamical properties of the kinase domain of Ephrin type-B Receptor 2 (EphB2). I will present atomic resolved dynamical data that together with secondary structural propensity (ssp) analysis suggests that EphB2 is allosterically regulated.

February 15, 2012

A great product is worthless if you cannot convince others about its value!
Sebastian Schultz

I defended my PhD thesis in September 2012 and thereafter became a “Transformer”. During my presentation I will share some of my experiences from my time as PhD. I will mainly focus on the importance of building up your network and some personal reflections on how to introduce yourself and/or your ideas to others (which you latest have to do during a job interview).

Investigating the potential link between IRS1 and S6K1
Siri Aili Fagerholm

In fat cells, insulin signaling controls glucose uptake from the blood as well as storage of triglycerides in its central lipid droplet. Patients with type 2 diabetes exhibit insulin resistance which involves a changed activity of the proteins in the signaling pathways. To gain further knowledge of the signaling mechanisms will improve our understanding of the disease. One of the key players in insulin signaling is the insulin receptor substrate 1 (IRS1). The activity of IRS1 is regulated by a number of other proteins in the insulin signaling pathways. A protein called S6K1 has from studies in murine cell lines been suggested to regulate IRS1 via phosphorylation of the site serin307. However, we have done time-studies and over-expression experiments in primary human fat cells and we believe that this is not the case for human primary cells, i.e. S6K1 does not phosphorylate IRS1 on ser307. Further, we will also use a lentivirus – based knock-down of S6K1 to establish the lack of connection between S6K1 and phosphorylation of ser307-IRS1.
 

Happy together? How to start, keep and deliver in projects together
with industry
Fredrik Agholme

Some personal reflections on how to collaborate with industry as an
academic researcher.

Nanocellulose
Anders Elfwing

Nanocellulose is an emerging material with some interesting properties. I will show some exciting novel procedures to functionalize the material. 

January 25, 2012

Clinical trials
Sara Bergstrand

There are many definitions of clinical trials, they are generally considered to be biomedical or health-related research studies in human beings that follow a pre-defined protocol. To perform a clinical trial there are many laws, regulations and standards that need to be followed. When planning a clinical trial there are issues of international, national, regional and local character and an overview will be presented. The base is Good Clinical Practice (GCP), an international quality standard that is provided by International Conference on Harmonisation (ICH). Good Clinical Practice guidelines include protection of human rights as a subject in clinical trial and provide assurance of the safety and efficacy of the newly developed compounds. Good Clinical Practice Guidelines include standards on how clinical trials should be conducted, and define the roles and responsibilities of clinical trial sponsors, clinical research investigators, and monitors. 

Ser 307 phosphorylation of insulin receptor substrate 1 in human fat cells
Cecilia Karlsson

Insulin receptor substrate 1 (IRS1) is an important protein in insulin signaling in human fat cells. By adding or removing phosphate groups on the protein you can alter its activity. When stimulating with insulin the phosphorylation of the IRS1 serine residue 307 gives rise to a positive feedback that retains the signal downstream in the cell. In cells from type 2 diabetes patients this phosphorylation is attenuated and it is therefore of great interest to study the regulation of this specific site. The kinase responsible is still unknown but we know that the phosphorylation is affected by the mTOR-complex 1 inhibitor rapamycin. We want to find out if mTOR, the catalytically active protein in the complex, is the kinase. This we do with interaction studies with co-immunoprecipitation and with an in vitro kinase assay.

November 30, 2011

Characterization of mutated FMS-like tyrosine kinase 3 (FLT3) receptor, a molecular target for anti-leukemic therapy
Amanda Nordigården

Each year, 300-400 persons are diagnosed with acute myeloid leukemia (AML) in Sweden, a severe illness that may emerge in hematopoietic stem cells. In one third of all these patients activating mutations of the FLT3 gene is detected (FLT3-ITD) which is associated with a poor prognosis for the patient. The tyrosine kinase receptor FLT3 (FMS like tyrosine kinase 3 receptor) is normally expressed on early myeloid and lymphoid progenitor cells and plays an important role in regulating survival, self-renewal and proliferation. FLT3-ITD mutations are duplications in the FLT3 receptor that can vary in size from 3 up to hundreds of nucleotides and result in a constituently active receptor. FLT3 is an attractive target gene in treatment of AML patients and a number of small tyrosine kinas inhibitors have been developed to prevent its activity. A main problem is that resistance against these inhibitors is developed in many patients. By mapping out the signaling pathways downstream of and towards FLT3 that contribute to the uncontrolled cell expansion it might be possible to overcome the problem of resistance using inhibitors that target these key molecules. We have earlier studied the role of FLT3-ITD in uncontrolled growth of leukemic cells and the effect of various recently developed pharmaceuticals that inhibit the activity of FLT3. Our findings can be used for developing more effective pharmaceuticals for AML patients. In our new project we have cloned FLT3-ITDs from various AML patients and plan to express these in hematopoietic cells to elucidate if there are any differences between these in the signaling pattern or surface/intracellular expressions.

Investigating the excitatory properties of cortical layer 6 cells
Sofie Sundberg

Different cells of the neocortical layers have somewhat different functions. Layers 4 and 6 in the primary visual cortex receive sensory information from dorsal lateral geniculate nucleus (dLGN) of the thalamus. The layer 6 cells can further act via a feedback mechanism on the dLGN neurons, regulating the flow of visual information through the dLGN. However, the pyramidal layer 6 cells could also be of pathophysiological interest, and have been suggested as a key player in cortical seizure generation. We are studying this alternative role of the layer 6 cells and want to investigate what properties these cells have that make them potentially important for seizure activity. Our experimental approach involves in vitro acute slices of visual cortex from rats and mice, where we measure the firing characteristics, the response to synaptic input and the oscillatory ability of the pyramidal layer 6 cells. These cells can fire action potentials in the range of 30-40 Hz and when activated they fire action potentials in a regular manner that does not adapt in the timescale of minutes. These properties indicate that once activated they will stay active long enough to feed excitatory input to other cells within the circuitry. This circuit activity could trigger recurrent excitation, one hallmark of epileptiform activity.

Optical Biosensing based on Localized Surface Plasmon Resonance
Erik Martinsson

Surface plasmon resonance (SPR) is a well-established technique for studying label-free biomolecular interactions and molecular adsorption in real time. The traditional sensing principal is based on detecting refractive index changes at a thin gold film using propagating surface plasmon polaritons. A similar sensing strategy can also be applied on metal nanoparticles, where another type of plasmons are created and utilized, so called localized surface plasmons. The frequency of these confined electron oscillations are highly dependent on the size and structure of the nanoparticles as well as the refractive index of the surrounding media. These unique optical properties make metal nanoparticles suitable elements for biosensing based on refractive index sensing and due to their size, much easier to minimize and integrate into other systems compared to the traditional SPR setup.

Development of virtual models for large-scale simualtion of industrial biotechnology processes
Inga Gerlach

Biotechnological processes have an increasing relevance on the market. Next to this important branch the demand for qualified personal is increasing. Therefore the education in practical operation procedures may be improved by using Operator Training Simulators (OTS). The structure of these simulators bases on mathematical models that can describe a biotechnological process. These models have been developing during the research project and have been implementing into a suitable process system in form of a Dynamic Link Library. The OTS can be described as computer programs that represent a virtual laboratory where the user can train there competences in general operation techniques. The aims of OTS are cost and time reduction for the practical training and the adequate extension of the academic education. A new version for the virtual bioethanol production was tested at Linköpings Universitet in Novemeber 2011. The first results showed the tendency that the students could improve their practical knowhow by using the OTS. 

November 2, 2011

Luminescent Conjugated Oligomers: Molecular probes
Rozalyn Simon

Conjugated polymers have been utilized in a variety of applications ranging from
organic electronic devices to chemical sensors. Delocalization of electrons over
the conjugated backbone allows for the emergence of a variety of useful features
such as conductivity and luminescence. Polythiophene is one such polymer that
has been exploited for just these features. Upon interaction with neighboring
molecules, changes in the planarity of the thiophene rings result in differences in
conjugation lengths over the polymer backbone and thus a spectral shift.
Functionalization of the backbone can also be used to alter conjugation length.
Utilizing functionalization of this type, one can tailor the spectral response upon
interaction with specific molecules. While the potential applications for this
technique are innumerable, our immediate aim is to synthesize a variety of
oligomeric thiophenes tailored for the detection of misfolded protein aggregates.
Via the synthesis of a small group of similar oligomers with slight variations, it is
possible to study the importance of the functionalization at specific positions on
the polymer and in this way, optimize for the intended applications in molecular
detection.

Rare earth nanoparticles as contrast agent in MRI: Nanomaterial design and functionalization
Maria Ahrén

Chelated gadolinium ions are used as contrast agents for magnetic resonance imaging. An alternative contrast agent composed of gadolinium oxide nanoparticles is expected to yield higher contrast and more easily made molecular capping. In this study we aim to equip 4-5 nm sized gadolinium oxide nanoparticles with molecules increasing the stability and water solubility and reducing the toxicity. This biofunctionalization strategy also introduces targeting capabilities, which means that a contrast agent for molecular imaging is to be produced. The chemical and physical properties of the nanomaterial are investigated with Fourier Transform Infrared Spectroscopy, X-ray Photoelectron Spectroscopy, Near-Edge X-ray Absorbtion Fine Structure Spectroscopy, X-ray Diffraction, Transmission Electron Microscopy and Dynamic Light Scattering. Relaxation measurements are performed on a NMR analyzer using a magnetic field of 1.4 T.

An idiots Guide to: Gene expression analysis of human dermal fibroblasts
Jonathan Rakar

Standard stainings and marker assays are often used to verify the identity of cells in culture, for example Oil Red O to show lipid droplets in adipocytes, von Kossa or Alizarin Red to show calcified matrix of bone. We have seen that such specific assays are positive in fibroblasts induced toward adipogenesis, osteogenesis and chondrogenesis. To investigate the induced cell types further, we employed full gene expression microarray (Affymetrix GeneChip HuGene 1.0ST) and compared the profiles to normal cultured fibroblasts as well as cultured reference cells (adipocytes, osteoblasts, chondrocytes). The datasets were compared using various statistical techniques, including Gene Set Enrichment Analysis (GSEA). Similarities and differences were interpreted from a putative functional perspective in an attempt to describe the degree of similarity between induced fibroblasts and their reference counterparts. The overarching aim is to investigate the feasibility of using primary human dermal fibroblasts for autologous tissue engineering. The gene expression analysis shows that the fibroblast phenotype is surprisingly receptive to induction stimuli, with key functions up-regulated, but that the phenotypes based on the monolayer culture and media induction system are distinct from the reference counterparts.

Interplay of AQP9 and the Motility Machinery
Thommie Karlsson

Cell migration is an evolutionary conserved process that occurs in numerous of events within the human body. The course of cell migration occurs through a specific pattern that is initiated by cell polarization and induction of membrane protrusions. Hence, in order to migrate, the cell requires a highly dynamic membrane and a highly dynamic cytoskeleton. Recently several membrane-anchored water channels known as aquaporins (AQPs) have been shown to augment cell migration. By using various imaging techniques, we assessed whether expression of AQP9 affected the cells ability to create membrane protrusions. We found that AQP9 induces various kinds of membrane protrusions that are highly associated with cell migration. Furthermore, we investigated the localization of AQP9 in these structures. Here, we propose a model for interplay of water fluxes through AQP9 with the motility machinery that enables rapid cellular shape changes.

October 5, 2011

Modified surfaces using tailor-made oligosaccharides having orthogonal attachments
Timmy Fyrner

The use of an appropriate spacer molecule is of great importance in many systems, ranging from biosensor applications to membrane-anchoring or as a crosslinker between biomolecules. Studies have shown that variations in the spacer length influence the mobility of the attached moieties (i.e. proteins, carbohydrates). To avoid a complicated synthesis of natural occurring carbohydrate containing linkers such as GPI anchors we turned our attention to the synthesis of a various oligosaccharide units of variable length.

Ion bipolar junction transistors for addressable delivery of biomolecules
Klas Tybrandt

Delivery of biomolecules at high spatiotemporal resolution is often desirable when interacting with biological systems. With electrophoretic delivery, temporal control can be achieved through the applied electrode potentials. For spatial control, many delivery points are required, e.g. in a matrix, where each point is individually addressable. As the basis for such addressable release of ions and charge biomolecules, we reported on the development of an ion bipolar junction transistor (IBJT) based on conducting polymers and selective membranes. The IBJT exhibit many similarities to the conventional BJT, with the major difference that electrons and holes are replaced by anions and cations. The IBJT is functional at physiological conditions and has successfully been applied to modulate the delivery of the neurotransmitter acetylcholine to neuronal cells. The first IBJT was of the pnp-type, i.e. the device modulated cationic currents. More recently the complementary version of the transistor, the npn-IBJT, has been developed. The characteristics of the npn-IBJT are very similar to those of the pnp-IBJT and both types of transistors can be fabricated onto the same substrate without additional fabrication steps. The npn-IBJT enables transport of anions and the realization of complementary ionic circuits. As a first step towards an addressable delivery matrix, an addressable delivery point has been realized by utilizing both pnp- and npn-IBJTs.

Ion Bipolar Membrane Diodes
Erik Gabrielsson

Cation and anion exchange membranes can be regarded as the ionic analog to p- and n-doped semiconductors, in the sense that they only conduct one type of charge. A bipolar membrane, consisting of a sandwich of a cation and a anion exchange membrane, is therefore expected to behave similar to a semiconductor pn-junction. Thus, bipolar membranes can be used for creating ionic transistors and diodes.
Our group’s method for fabrication ionic devices based on bipolar membranes utilizes standard microfabrication techniques such as spin coating, photolithography and removal of material using dry etching. As a typical device can contain up to 4 layers it is critical to evaluate the result of each added layer during the manufacturing process in order to find defects. A recent addition to our labs resources for measurement of topographies is an optical profilometer. This instrument is similar to an ordinary light microscope, but can also measure the depth of a sample using either confocal or interferometry techniques. Optical profilometry offers quick measurements and vertical resolution down to 1 nm, while the numerical aperture of the objective limits the lateral resolution. The possibility of using different measurement techniques is an advantage as some techniques might give erroneous result for certain materials and patterns.
I mainly use optical profilometry to study microfabricated structures down to the micrometer scale. We also have an objective for using in aqueous media, suitable for in situ measurement on bioelectronic devices.

Characterization of mutated FMS-like tyrosine kinase 3 (FLT3) receptor, a molecular target for anti-leukemic therapy
Amanda Nordigården

Each year, 300-400 persons are diagnosed with acute myeloid leukemia (AML) in Sweden, a severe illness that may emerge in hematopoietic stem cells. In one third of all these patients activating mutations of the FLT3 gene is detected (FLT3-ITD) which is associated with a poor prognosis for the patient. The tyrosine kinase receptor FLT3 (FMS like tyrosine kinase 3 receptor) is normally expressed on early myeloid and lymphoid progenitor cells and plays an important role in regulating survival, self-renewal and proliferation. FLT3-ITD mutations are duplications in the FLT3 receptor that can vary in size from 3 up to hundreds of nucleotides and result in a constituently active receptor. FLT3 is an attractive target gene in treatment of AML patients and a number of small tyrosine kinas inhibitors have been developed to prevent its activity. A main problem is that resistance against these inhibitors is developed in many patients. By mapping out the signaling pathways downstream of and towards FLT3 that contribute to the uncontrolled cell expansion it might be possible to overcome the problem of resistance using inhibitors that target these key molecules. We have earlier studied the role of FLT3-ITD in uncontrolled growth of leukemic cells and the effect of various recently developed pharmaceuticals that inhibit the activity of FLT3. Our findings can be used for developing more effective pharmaceuticals for AML patients. In our new project we have cloned FLT3-ITDs from various AML patients and plan to express these in hematopoietic cells to elucidate if there are any differences between these in the signaling pattern or surface/intracellular –expressions.

September 7, 2011

Design and Synthesis of Anti-Inflammatory Agents for Artificial Corneas
Mattias Tengdelius

Herpes simplex virus-1 (HSV-1) is the leading infectious cause of impaired vision in people in developed countries, affecting millions. Due to its complex epidemiology no treatment for HSV-1 exists, stressing the need for artificial corneal transplants to avoid the recurrence associated with transplants of human corneas. Although huge progresses have been made in this field over the last decades, numerous patients with artificial corneas still suffer from inflammation due to the transplantation itself and the recurrence of HSV-1. This presentation describes the design and synthesis of anti-inflammatory agents, based on the naturally occurring substance fucoidan, for use in artificial corneas.

Using nuclear magnetic resonance spectroscopy in protein science
Annica Theresia Johnsson

Proteins are essential for life. They manage and supervise life processes ranging from catalysis to cell signaling and cell contacts. In order to perform these functions, proteins need to be able to interact with each other and with their environment. Studying these events and the proteins themselves is therefore necessary to understand the mechanisms of both health and disease.
Nuclear magnetic resonance (NMR) spectroscopy provides the means to study biomolecular properties that are inaccessible to most other spectroscopic techniques. For instance, the structural and dynamical properties of proteins can be characterized at atomic resolution. Besides from determining the structural, dynamical and conformational properties of a protein, it is also possible to (among other things) study molecular events such as where and how a protein binds its substrate and the kinetics of binding.
We use NMR spectroscopy to study protein structure, function and characteristics. So far, we have confirmed a folding-on-binding event that occurs when adding the E6 protein of human papilloma virus (HPV) to the SAP97 PDZ2 I342W protein domain. We have also calculated the structure of the unbound PDZ domain and studied its dynamical properties. 

Molecular Assemblies for Formation of Tethered Lipid Membranes
Hung-Hsun Lee

Supported lipid membrane is not only a powerful model system but also a promising technological platform in biosensing and pharmaceutical screening. In particular, tethered lipid membranes (TLMs) on self-assembled monolayers (SAMs) allow controlling the composition and geometry of biomimetic assemblies down to molecular scale. However, there are only few examples of systematic investigations on the effect of the supporting assemblies to the TLM formation. The aim of this study is to develop a stable SAM as an underlying flexible surface for TLM attachment. Our model SAM surface comprised a mixture of a matrix molecule and an anchor molecule. We have prepared and characterized the structures of the model surface by means of ellipsometry, contact angle goniometry, and infrared reflection-absorption spectrometry (IRRAS). Moreover, quartz crystal microbalance with dissipation monitoring (QCM-D) analysis revealed a critical vesicle concentration on the surface is required for vesicle rupture and membrane formation.

Our investigation paves the way for micro- and nano-structured molecular architectures, as well as model surfaces for mimic of complex cell membrane assemblies.

Increasing our knowledge of cancellous bone healing
Olof Sandberg

Bone tissue can be structured in two ways. It can either be compact, this type is called cortical, or it can be of a more spongy constitution, in which case it is termed cancellous.
Most of the knowledge of bone healing has been gathered on the more accessible cortical bone. This is unfortunate for two reasons:
- the major health issue concerns fractures in cancellous bone
- cortical and cancellous bone have quite different healing mechanisms
My PhD thesis aims for increasing our knowledge of cancellous bone healing using methods of mechanical evaluation, gene expression analysis and µCT analysis, primarily in rat and mouse models.

April 20, 2011

Electrospinning biomaterials for Cardiac Tissue Engineering
Abeni Wickham

After myocardial infarction the heart muscle is usually damaged and does not self-repair leading to scarring, thinning of the muscle wall, and decreased function or possible heart failure. The use of a scaffold that can buttress or replace the scar tissue and induce the regeneration of the myocardium would help sustain the quality life of patients.
The scaffolds that will provide the architectural supports are fabricated by electrostatic or electrohydrodynamic atomization, aka electrospinning. This fabrication technique is a versatile and ‘cheaper’ method to produce homogenous scaffolds for tissue engineering. In conjunction to this technique, we are using different natural and synthetic polymers to help induce differentiation of, e.g. mesenchymal stem cells.

The role of hypoxia in preserving the stem cell potential of hematopoietic stem cells
Camilla Halvarsson

Hematopoietic stem cells (HSCs) give rise to all blood cells throughout the life and reside in specialized areas, or niches, of the bone marrow (BM), protected from oxidative stress and depletion caused by excessive proliferation. These niches are believed to be hypoxic, and it is suggested that this low oxygen level is supporting the HSCs to stay in a relatively low proliferative, quiescent state. By understanding the molecular properties of HSCs and how hypoxia influences self-renewal and long-term maintenance it would, among other things, be possible to improve ex vivo expansion of HSCs for BM transplantations. Our hypothesis is that hypoxic niches in the BM are essential to maintain stem cell potential. The aim of my study is to investigate this role of hypoxia by setting up limiting dilution series of murine Lineage-Sca-1+c-kit+ (LSK) cells, cultured in vitro in normoxia or hypoxia, transplanting the cells into mice and analysing the peripheral blood for reconstitution capacity.

Electrolyte Gated Organic Field Effect Transistors as Biosensors
Henrik Toss

Biosensing and bioanalytical instruments are often expensive, complicated to use and in need of laboratory analysis to yield interpretable results. Many also need quite large analyte quantities. It has been demonstrated that Electrolyte Gated Field-Effect Transistors (EGOFETs) can function using aqueous media as the gate electrolyte. This provides the opportunity to introduce water soluble analytes between the gate electrode and the semiconductor of the EGOFET. Any disturbance of the gate signal will yield an amplified response in the output current and some reaction of an analyte in the electrolyte could introduce such a disturbance. This means that with proper biorecognition elements an EGOFET could work as a biosensor with high sensitivity and specificity with a simple current response result that does not need any extra lab work to be analyzed. Because this is done with organic electronics a final device should be possible to produce by printing techniques and thuc be cheap and disposable.

Influence of a single loading episode on gene expression in healing rat Achilles tendons
Pernilla Eliasson

Mechanical loading stimulates tendon healing via mechanisms that are largely unknown. Genes will be differently regulated in loaded healing tendons, compared to unloaded, just because of the fact that healing processes have been changed. In order avoid such secondary effects and study the effect of loading per se, we therefore studied the gene expression response shortly after a single loading episode in otherwise unstimulated healing tendons.
The Achilles tendon was transected in 30 tail-suspended rats. The animals were let down from the suspension and the tendons loaded by treadmill running for 30 min once, 5 days after tendon transection. Gene expression was studied by Affymetrix microarray before, and 3, 12, 24 and 48 h after loading. The strongest response in gene expression levels (most regulated genes) was seen 3 hours after loading, when 150 genes were up- or down-regulated (fold change ≤2, p≥0.05). 12 hours after loading, only 3 genes were up-regulated, while 38 were down-regulated. Less than 7 genes were regulated after 24 and 48 hours. Genes involved in the inflammatory response were strongly regulated at 3 and 12 hours after loading; this included up-regulation of iNOS, ptges, Il-1b, and other interleukins. Also genes involved in wound healing/coagulation, angiogenesis and production of reactive oxygen species were strongly regulated by loading. Microarray results were confirmed for 14 selected genes in a repeat experiment (N=30 rats) using real-time PCR. It was also confirmed that a single loading episode on day 5 increased the strength of the healing tendon on day 12 by mechanical evaluation. The fact that there were hardly any regulated genes 24 h after loading suggests that optimal mechanical stimulation of healing requires a loading stimulus every day.

March 23, 2011

Conjugated Polymer Surfaces for Electronic Control of Cell Detachment
Kristin Persson

Adherent eukaryotic cells are commonly cultivated in cell culture dishes and it is of great interest to gain control over different parameters related to attachment, proliferation, spreading, detachment etc by means of external stimuli applied to the dish. We report a new method for control of cell detachment based on a conjugated polymer, PEDOT-S:H. This polymer is water soluble and intermolecularly self-doped to a high extent in its oxidised state. When PEDOT-S:H is further oxidised by applying an electric potential it disintegrates and detaches from the underlying substrate. Adherent epithelial cells were cultivated on PEDOT-S:H. When oxidising PEDOT-S:H with cells growing on top, the polymer detached and disintegrated, leaving cells free in the electrolyte. The polymer can also be patterned using standard photolithography into individually addressable areas, enabling spatial control of detachment.

An integrative experimental-modeling approach to study insulin signaling in adipocytes
Elin Nyman

Insulin controls glucose homeostasis and failure in this control leads to insulin resistance and type 2 diabetes. Insulin act through its receptor at the cell surface of insulin responding cells. After insulin binding the intracellular parts of the transmembrane receptor autophosphorylate and a downstream signaling cascade is initiated with metabolic and mitogenic responses inside the cell. We study the dynamic behavior of insulin signaling proteins in adipocytes experimentally and with mathematical modeling to unravel the essential mechanisms of the signaling network. We use an integrative approach and iterate between experimental work and modeling. In our approach we have a formalized way to test hypotheses and then either reject or further analyze these. From the so far acceptable hypotheses we make predictions that guide us to which new experiments to perform.

Multicomponent hydrogels to control adhesion and settlement of fouling organisms
Wetra Yandi

Preparation of surface material to prevent the adhesion of marine organism on the hull of the ship has become an important issue in marine coatings field. Antifouling surfaces, which resist the adhesion of marine organims, can be prepared via several ways. Material Surface modification has been used for several years to prepare marine antifouling surface materials.

In this work poly ethylene glycol methacrylate (PEGMA) and hydroxy ethyl methacrylate (HEMA) were grafted on the SAMs-immobilized gold surfaces via self and surface initiated Atom Transfer Radical Polymerization (SI-ATRP). FT-IRRAS, Ellipsometry, water contact angle, diidomethane contact angle and surface topography were studied in this work. Protein adsorption tests on the samples surface were performed to see the surface-biomolecule interaction.

FT-IRRAS spectra confirmed that poly(HEMA-co-PEGMA) were successfully polymerized on SAMs-immobilized gold surface. In this works, different thickness of polymer film were fabricated and observed by ellipsometry. Contact angles of Poly(HEMA-co-PEGMA) at different time were observe in this work and CH3 SAMs and Br SAMs were used as comparator. Compared to Br SAMs, poly(HEMA-co-PEGMA) shows a lower contact angle means that they are more hydrophilic than Br SAMs due to the hydrophilic properties of HEMA and PEGMA. The contact angle of poly(HEMA-co-PEGMA) tend to increase by the increase of ATRP time. Protein adsoption test showed that the polymer surface significantly reduced the amount of protein adsorbed on its surface.

UNDERSTANDING MECHANISMS BEHIND SETTLEMENT AND ADHESION ON SYNTHETIC PEPTIDE AND OLIGOSACCHARIDE SAMS
Pitsiri Sukkaew

Biofouling is a problem commonly observed on a surface submerged under water. To prevent such undesired settlement in marine environment without using biocides, an understanding regarding settlement and adhesion mechanisms including factors that induce/hinder the settlement become important.

Surface exploration stage and its effects on the subsequent settlement in the barnacle’s life cycle will be studied. Imaging surface plasmon resonance (iSPR) will be used for monitoring the barnacle cyprids’ exploration on self assembled monolayers (SAMs) with different surface chemistries. The number of deposited footprints on each surface will be counted and compared. Settlement assays on SAM surfaces of the same chemistries will also be performed. A correlation between the number of settlements and the number of deposited footprints observed during the exploration stage will be examined. Settlement inducing/hindering factors will be drawn based on surface characterization results obtained from IR spectroscopy, ellipsometry and water contact angle measurement. Those characterization methods will also be used for degrading evaluation of SAM surfaces after been submerged in artificial seawater.

February 23, 2011

Luminescent conjugated oligothiophenes can be used to spectrally discriminate between A plaques composed of different APP variants
Therése Klingstedt

Luminescent conjugated oligothiophenes (LCOs) is a novel class of fluorescent amyloid probes. They have a flexible backbone that dictates their optical properties and their binding to protein aggregates gives rise to conformation-dependent emission spectra.
This property has been utilized in several platforms such as cell systems, tissue samples and for real-time visualization of cerebral protein aggregates in a mouse model.

Alzheimer¿s disease (AD) is one of the most well-known protein misfolding diseases. It is histopathological characterized by Aβ plaques and tau tangles found in the brain of the patients, however, their involvement in the disease is still unknown.
The pentameric LCO p-FTAA has been used to investigate the composition of Aβ plaques found in mice harboring different mutations in the APP gene. APP is the precursor protein to Aβ and different variants of this protein increase the risk for developing AD at an early stage. The results revealed that p-FTAA can be used to spectrally discriminate between plaques containing the Swedish mutation and the Arctic mutation indicating that these mutations cause Aβ to aggregate in different ways.

Wall shear stress and vessel wall remodelling
Hanna Björck

Atherosclerosis is a chronic inflammatory disease affecting the walls of large arteries. The underlying disease mechanism is not fully understood, but characteristics are retention of lipids, accumulation of smooth muscle cells and macrophages, extracellular matrix remodeling, eventually leading to plaque formation. Atherosclerotic lesions predominately form at specific regions, such as arterial branch points or the inner curvature of the aortic arch. In these regions the blood flow is low, disturbed or oscillating, resulting in low or oscillatory wall shear stress (WSS). In comparison, linear vessel segments, which are exposed to steady, high laminar shear stress, tend to be lesion free.
In the present study we investigate the role of WSS on vessel wall remodelling, with special emphasis on the activation and expression of inflammatory mediators in the aortic arch of rat. Aortic geometry and blood flow velocities are determined by MRI, enabling calculation of WSS magnitude, as well as direction of flow using computational fluid dynamics. Local shear dependent gene expression is studied in low- and high shear regions.

A perspective on being a PhD student, the early years
Fredrik Agholme

For the new graduate student I will give a short overview over my experience as a PhD student and relay some of my thoughts and some practical advice on things that I wished someone told me when I started. Being a graduate student is an interesting and sometimes demanding time. You are on your way to become an independent researcher but the road towards a thesis is not necessary a straight and well paved one. As with any other job you will obtain new skills but also undergo personal development. Sometimes things will go your way and sometimes you will fail. The main theme of my talk is that it is important to have fun both in and outside the lab. But it is also important to realize that there are obstacles on the way. There are methods to overcome these available and the sooner you realize this, the easier your journey towards a great thesis becomes.

Electrical characterization of screen printed electrodes for ECG measurements
Linda Rattfält

In this study, electrical properties of screen printed electrodes were explored. The electrodes were made out of conductive ink and tested in a standardized measurement setup. The electrodes were printed on a plastic film substrate using several layers of conductive ink and insulating lacquer. The electrode area had a diameter of 1 cm and was printed using a bottom layer of Ag-based ink and a second layer of Ag/AgCl-based ink. The transmission lines were either a 2 cm broad strip of Carbon based ink or a 1 mm wide line of Ag. The transmission lines were coated with one or two layers of insulating lacquer. The electrodes were mounted on a rigid fixture and submerged in an electrochemical cell. Half cell measurements were performed with a potentiometer, a Ag/AgCl reference electrode and an active electrode of Palladium. Polarization potentials were registered during the first 30 minutes. Impedance measurements were done in the range of 0.05 to 2000 Hz (50 samples logarithmically distributed) with a sine curve, dc at open circuit potential and 10 mV ac. Results show that the polarization potential in average decreased 2 mV during 30 minutes where the largest decrease was during the first 5 minutes. No difference was shown between the different types of samples. The impedance of the electrodes at 10 Hz, show that electrodes with Ag conductors and 1 layer of lacquer had the lowest impedance, about 100 Ohms and 400 Ohms lower that Ag/AgCl and 2 layers of lacquer and Carbon conductor respectively. Phase angles are limited to 20 deg at 10 Hz. The results are coherent to previous measurements with textile electrodes. Furthermore, adhesion and the ionic transmission layer need to be addressed. Future tests also include changing the printing substrate to fabric.

January 26, 2010

Characterization of self assembled protein hybrid nanomaterials
Fredrik Bäcklund

When heated in aqueous acid, the protein insulin self-assembles into nano-wire structures known as amyloid fibrils. By simply co-grinding insulin with a given guest complex, a protein-guest molecule composite material can be formed that retains the protein-specific ability to self-assemble into fibrillar structures when exposed to heat. One possible application using a protein-iridium composite material was previously demonstrated by the generation of white light in OLEDs. Now, due to the simplicity and potentially extensive scope of the method of preparation, we are investigating several alternative functionalizing agents. Using various techniques such as AFM, fluorescence microscopy and polarized light microscopy, we are investigating the effect of using different guest molecules on the morphology of the amyloid nanostructures.

Detection of IgG subclasses in serum from type 1 diabetic patients participating in a phase II study with GAD65
Mikael Chéramy

In autoimmune diseases, like type 1 diabetes (T1D), the immune cells attack and destroy substances and tissues normally present in the body. This is partially mediated by B-cells which produce large amounts of antibodies directed towards certain autoantigens. In T1D one of the major autoantigens is Glutamic acid decarboxylase 65 (GAD65), which is present in the insulin producing β-cells. Approx 70-80% of newly diagnosed T1D patients have autoantibodies against GAD65 (GADA).
Immune modulation using GAD65 has been tested in a phase II trial conducted by our group. We show that the treatment maintain β-cell preservation and increase GADA titers. The GADA immunoglobulin is mainly constituted of the IgG isoform, which itself is divided into 4 different subclasses, IgG1-4. There is a high homology between the subclasses, but differences in the hinge region and the heavy chain influence and activate the immune response in different ways.
We have analyzed the distribution of the 4 GADA IgG subclasses using a modified version of a conventional GADA assay in patient’s serum before and after GAD65 treatment. 

Ion-Electron Transducing Electrodes with Electrochemically Active Material for Sample-friendly Lab-On-A-Chip Devices
Per Erlandsson

Improved integrated electrodes can increase the potential of many microfluidic technologies such as micro cell culture chambers and lab-on-a-chip (LOC) systems. Electrodes are one of the critical components in microfluidic devices, as they can move liquid between reactors, separate samples and even function as detectors. However, electrochemical reactions are required to maintain electric fields, often hydrolysis, which generates O2 and H2 bubbles and/or H+ and OH- at the electrodes. In large systems the impact of hydrolysis can be mitigated, but when scaling down to LOC sizes, these reactions can change the pH which is unacceptable for sensitive biomolecules. Using conductive and electrochemically switchable materials as electrodes gives nearly hydrolysis-free reactions without generating gas or changing the solution’s pH.

Immunological characterization of a type 1 diabetes high risk group
Linda Åkerman

The etiology and pathogenesis of type 1 diabetes is still not fully established. Autoantibody screening among 17000 Swedish children has identified a group of 22 individuals that have a high risk to get the disease, and by studying this group we want to get a better picture of what is really going on in their immune system during the period preceding disease onset. We may also have a chance to find new markers of risk, and to find ways of distinguishing risk individuals that eventually get the disease from those who don’t. This information could be very useful in prevention and intervention trials. There are numerous ways by which this group can be studied, but the first step will be to analyze gene expression in circulating immune cells from the risk individuals and compare it to that of healthy children and type 1 diabetics. This is done by multivariate qPCR, measuring the expression of 88 different genes simultaneously. Based on the results, more analyses and methods will be added.

December 1, 2010

Structural and Dynamic Analysis of the Human Glutaredoxin 3 using NMR spectroscopy
Cecilia Andrésen

NMR spectroscopy is a powerful technique in protein chemistry. The method can be used for structural determinations of protein molecules and for interaction analysis, dynamic studies and kinetics.
For better understanding of the Glutaredoxin pathway in the mammalian cell it is of importance to generate the structure of human Grx3. The protein is essential and multifunctional, for example involved in cardiac physiology and the response to oxidative stress. The protein consists of three domains, an N-terminal Thioredoxin and two Monothiol Glutaredoxin domains in the c-terminal. We are interested in the interaction and the dynamics between the different domains.
The structure of the Trx domain has been generated using CS-Rosetta and CS23D where the NMR chemical shifts are used for the calculations. The determinations of the other domains are in progress.
The dynamic properties of Grx3 are also important to investigate for analysis of the cooperation between the different domains and therefore relaxation experiments, T1, T2, NOE and CPMG, have been analyzed.
All together, the information of the structure, domain positions and motions will give more knowledge of the mechanism of the Glutaredoxin protein.

How and when to pick up the best signals from markers associated with T-regulatory cells?
Anna Kivling

Regulatory T (Treg) cells are important tools with the purpose to control and regulate the immune system. These cells use FOXP3, TGF-β, CTLA-4 and sCTLA-4 to regulate other T-cells. Since cryopreservation of PBMC is a convenient way to handle samples, we investigated whether these cells will change their mRNA expression of Treg associated markers after cryopreservation.
PBMC from healthy adults were stimulated either fresh (48/96 hours) or after cryopreservation (48 hours), with PHA, TT, βLG, OVA or in culture medium exclusively (spontaneous). The mRNA expression of FOXP3, TGF-β, CTLA-4 and sCTLA-4 were studied with multiplex real-time RT-PCR.
Cryopreserved PBMC were appropriate for detection of the Treg associated markers FOXP3, CTLA-4, sCTLA-4 and TGF-β expressed spontaneously. Also antigen-induced mRNA expression of CTLA-4, sCTLA-4 and TGF-β, with the exception of FOXP3, preserved a stable transcription activity after cryopreservation.
In conclusion, cryopreserved cells are in general suitable for studying Treg associated markers.

Adsorption of Poly-L-Lysine on Platinum Electrodes as a Function of Applied Potential
Sara Nilsson, Mats Fahlman, Fredrik Björefors, Nathaniel D. Robinson

The charged polyelectrolyte poly-l-lysine (PLL) has, due to its biocompatibility, been investigated extensively for use in surface modifications, e.g. to make electrodes suitable for in vitro and in vivo biological experiments. Since biological cells tend to produce proteins in order to insulate themselves from hard materials such as metal electrodes, the effectiveness of the electrode for electronic stimulation or monitoring of biological signals quickly diminishes. Coating such electrodes with materials like PLL helps to reduce their rejection by the biological system. We have chosen to study the adsorption of PLL onto platinum electrodes, since platinum is a commonly used electrode material.

Many studies concerning the adsorption of PLL on various substrates deal with variables such as pH and salt concentration of the electrolyte while fewer reports the effect of the underlying substrate potential. However, it has been demonstrated that the weakly basic cation PLL adsorbs continuously on the conducting substrate indium tin oxide (ITO) when applying a potential and using optical techniques.

We examine the rate-limiting step of the deposition of PLL on platinum electrodes by employing the electrochemical quartz crystal microbalance with dissipation monitoring (EQCM-D) together with cyclic voltammetry (CV). This combination is very powerful in the simultaneous measurements of the change in frequency, dissipation and current in response to the potential applied, hence emphasizing the value of applying a potential to the underlying substrate while adsorbing PLL.

The Orientation of a Phenyl Boronic Acid Analogue on Gold Substrates: a Near-
Edge X-ray Absorption Fine Structure Spectroscopy Study
Cecilia Vahlberg

The orientation of self-assembly monolayers of a phenyl boronic acid analogue on gold
surfaces is investigated using NEXAFS. Our aim is to design well defined and highly
ordered biofunctionalized surfaces with a high degree of reproducibility that can be used
as model systems to mimic molecular recognitions processes. NEXAFS is a surface
sensitive technique that is useful in characterization studies of molecular adsorbates as it
can be used to for example indentify different functional groups or to investigate the
molecular orientation. The NEXAFS measurements need to be done at a synchrotron
light facility. All our molecular structure studies have been done at MaxLab in Lund.

November 3, 2010

CHARACTERIZATION OF THE HUMAN VIROME
Fredrik Lysholm1,2, 2Anna Wetterbom, Tobias Allander3, Bengt Person1,2, Björn Andersson2
1 IFM Bioinformatics, Linköping University, Linköping, Sweden
2 Science for Life Laboratory, Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden
3 Institutet, Department of Microbiology, Tumor- and Cell Biology, Laboratory for Clinical Microbiology, Karolinska University Hospital, Stockholm, Sweden

Virus infections impose a huge disease burden on humanity. While the human genome has been fully characterized, we still have very incomplete knowledge of the viruses that infect humans. At the same time, the vast majority of known human virus species are pathogenic. Nevertheless, most studies of virus disease are limited to the investigation of already known viruses, while the discovery of an unknown virus and production of the first diagnostic reagent is very difficult and a rare occurrence. Therefore, a broad and unselective exploration of the human virus flora, analogous to the human genome project and the various ongoing metagenomics projects, would be the most productive development of modern virology.
We aim to fully investigate the virome in various milieus such as the respiratory tract, feces, blood and tissue samples in a wide range of patients with different clinical diagnoses. By the broad use of Roche 454 sequencing machines; an ever increasing amount of data per run is produced and to efficiently analyze this data, a wide range of bioinformatics tools is used. Some of these tools are common and widely used within bioinformatics while others were developed specifically for solving the huge data-processing problem.
We have developed a new pipeline for viral metagenomics aimed at automatically charactering the viral content of the sample and provide a general sample overview. Within this pipeline several steps are customized to more accurately and efficiently process this vast amount of data. Through our samples we have characterized nasopharyngeal aspirates from children with severe lower respiratory tract infections, serum from patients diagnosed with chronic fatigue syndrome, serum from immunesuppressed individuals (non-HIV patents), feces from children with severe diarrheal disease and samples related to several types of auto-immune diseases.

Electroactive 3D Fiberstructures of PEDOT to Control Cell Signaling
Maria Bolin

Traditionally, in vitro cell studies are performed using planar and rigid Petri dishes comprising the cell culture and the associated aqueous culture medium. This is far different from the in vivo situation where cells grow in a complex 3D environment called the extra cellular matrix. Electrospun nano-fibrous structures provide a suitable engineered 3D surface similar to the extracellular matrix, which is characterized by a wide range of pore diameters, high porosity, and high mechanical endurance. Adding an electroactive coating would result in enhanced functionality over the present, passive 3D-scaffolds. We have earlier shown that planar electronically active cell seeding surfaces based on conjugated polymers can be used to control the cell adhesion and proliferation of various cell systems. In addition, these materials have been successfully utilized to induce and record signaling in neurons. Here, electronically conductive and electrochemically active 3D-scaffolds based on electrospun poly(ethyleneterephthalate) (PET) nano-fibers with a conformal coating of PEDOT are presented. SH-SY5Y neuroblastoma cells were grown on the PEDOT coated fibers to study Ca2+ signaling.

Protein quantification using a combination of electrochemiluminescence detection and patterned arrays
Maria Jonson

It is known that many diseases, such as Alzhiemer’s disease, Parkinsson’s disease and Prion diseases are caused by an accumulation of amyloid forming proteins, leading to neurodegeneration and celldeath. Over the past decade Drosophila melanogaster has emerged as a promising model system for amyloid disease research.
In my work we use transgenic Drosophila models of Alzheimer’s disease (human Aβ and tau expressors) to analyze tissue/cell specific degeneration by variable expression of these amyloidogenic proteins. Amyloid associated proteins are expressed selectively in different tissues and the protein levels can be measured by MSD immunoassays.
This immunoassay uses electrochemiluminescence detection to detect binding events on patterned arrays and to determine the concentration of a specific protein in your sample. They also provide multi-array technologies which offer arrays within the well for increased throughput and efficiency.
The MSD immunoassay will be the main focus during my talk, where I will explain the method, give you some advantages and disadvantages and finally show you some results from my own studies.
 

October 6, 2010

Biophysical characterization of a new sequence variant in the human Thiopurine Smethyltransferase gene
Patricia Wennerstrand

Thiopurine S-methyltransferase (TPMT) is involved in treatment of thiopurines in leukemia and inflammatory bowel disease. Thiopurines such as 6-mercaptopurine, azathioprine and 6-thioguanine are converted to active metabolites which are incorporated into the DNA or inhibit purine de novo synthesis. TPMT is a polymorphic enzyme with at least 29 different allelic variants known today. These varaints cause low, intermediate or high activity of the TPMT enzyme. We have characterized a novel sequence variant, TPMT*28. Routine blood sample are taken of patient affected with inflammatory bowel disease taken prior to thiopurine threatment. This variant was discovered due to intermediate TPMT enzyme activity in such a sample. We have recombinantely expressed human TPMT*28 and have been able to explain the underlying mechanism causing intermediate activity.

Amyloid formation in Drosophila melanogaster: towards molecular mechanisms, or “A good picture is worth more than 1000 words”
Sebastian Schultz

Patients with type 2 diabetes have a marked reduced beta cell mass and fail to produce sufficient amounts of insulin required for regulation of glucose homeostasis. Recent research supports that intracellular aggregation of islet amyloid polypeptide (IAPP) leads to cell death making it a plausible cause for the beta cell reduction. Little is known about the mechanisms that precede amyloid formation or which cellular pathways are involved in this process. To gain better understanding we have established a Drosophila melanogaster model, where GAL4 drives expression of human proIAPP (the precursor of IAPP), human IAPP, and the non-amyloidogenic mouseIAPP (mIAPP) in a cell specific pattern. When expressed in 16 neurons of the fly brain we found that proIAPP and IAPP expression lead to reduction of these 16 cells over time. With genetical tools and the help of confocal microscopy we lately have started to dissect the pathways that are involved upon expression of our aggregating proteins. In my talk I will focus on explaining some principles of how confocal microscopy works, its advantages over other visualisation methods and will then give some examples of my own project in order to show how this method can be used to explain molecular mechanisms.

How and when to pick up the best signals from markers associated with T-regulatory cells?
Anna Kivling

Regulatory T (Treg) cells are important tools with the purpose to control and regulate the immune system. These cells use FOXP3, TGF-β, CTLA-4 and sCTLA-4 to regulate other T-cells. Since cryopreservation of PBMC is a convenient way to handle samples, we investigated whether these cells will change their mRNA expression of Treg associated markers after cryopreservation.
PBMC from healthy adults were stimulated either fresh (48/96 hours) or after cryopreservation (48 hours), with PHA, TT, βLG, OVA or in culture medium exclusively (spontaneous). The mRNA expression of FOXP3, TGF-β, CTLA-4 and sCTLA-4 were studied with multiplex real-time RT-PCR.
Cryopreserved PBMC were appropriate for detection of the Treg associated markers FOXP3, CTLA-4, sCTLA-4 and TGF-β expressed spontaneously. Also antigen-induced mRNA expression of CTLA-4, sCTLA-4 and TGF-β, with the exception of FOXP3, preserved a stable transcription activity after cryopreservation.
In conclusion, cryopreserved cells are in general suitable for studying Treg associated markers.

September 8, 2010

An introduction to Structural Biology: Questions, methods and possibilities
Sara Helander

Investigation of the structure and the biophysical properties of human proteins are of major importance and may lead to improved understanding of various diseases. Different questions related to structural biology will be discussed, together with various methods like Circular Dichroism, NMR and SPR. Results related to each method will be shown and explained.
In order to show one example where structural biology methods can function as a complement to cell biology studies, results from previous studies with c-myc95-158wt and two different mutants will be described.

Regulatory T cells four years after antigen treatment
Mikael Pihl

In trials of immunotherapy with the Type 1 Diabetes (T1D) associated autoantigen Glutamic Acid Decarboxylase (GAD) 65 (Diamyd), we have shown an altered response to GAD65 in treated children with T1D. The treatment is thought to act by inducing immunological tolerance toward GAD65. Treatment with GAD65 maintains insulin secretion in treated individuals. We hypothesize that treatment with GAD65 induces antigen-specific regulatory T cells (Treg) in children with T1D. We have monitored the expression of phenotypic markers for Treg in the participants throughout the trial. There are no differences in expression of markers for Treg in resting cells four years after injection of Diamyd. FOXP3 expression increased significantly after antigen stimulation in cells from treated individuals. The frequency of CD25hiCD127lo cells also increased upon antigen stimulation in patients receiving Diamyd. In addition, we are expanding cells to perform further analyses of Treg from these patients.

Nanostructured materials for high temperature gas sensing applications
Kristina Buchholt

Around the world emission laws and legislations are becoming stricter due to the increased knowledge of the adverse environmental, and health effects of emissions from cars and industry. To be able to fulfill these strict emission laws the car industry is in need of selective, long-term stable gas sensors able to withstand high temperatures and corrosive environments. My research is focused on developing materials for the sensors which can survive the harsh environment present inside an exhausts system. Investigating the properties of new materials involves using many material characterization techniques, one being Transmission Electron Microscopy (TEM). TEM is not only useful in materials science but also has applications in biology, for example in cancer research and virology.

Methods and materials for organic electronic/ionic devices
Erik Gabrielsson

There is a large interest in using drug delivery devices for stimulation of various biological systems. These devices are often based on micro-fluidics and thus incorporate valves and pumps. Unfortunately, such devices inherently produces interfering fluidic flow into the biological system.

In our group, we a have long experience in using organic electronics for precise delivery of various positively charged biomolecules without fluidic flow. This is applied into devices that, for example, can be used to interact with cultivated neural cells. In my work, I try to extend our current knowledge of controlling ion fluxes in micrometer-scale devices to include transport of anions. To reach this goal, new ion exchange materials is needed as well as detailed characterization of the resulting ion transporting micro-channels.

The final results will hopefully be useful for both delivery of anion biomolecules, such as retonic acid or ATP, and as part of more complex organic electronics/ionics systems.

May 19, 2010

Nanoparticle induced functionalization of designed polypeptides and the usage of pyrenyl-residues as fluorescent probes
Jutta Speda

The functionalization of de novo polypeptides gives rise to a wide application in industry and science. Special designed peptides are capable to form helical secondary structures upon addition of silica nanoparticles, even resulting in switch-like characteristics that refer to an attached catalytic side for ester hydrolysis.
In my studies, the basic peptide design is further varied to investigate the influence of rather bulky pyrene-labeled cysteine side chains onto the polypeptide secondary structures. Peptides exhibiting one or two labels in different positions are examined with respect to associated changes in silica nanoparticle induced secondary structures. Additionally the effects of secondary structure formation on fluorescent emission spectra are investigated as pyrene-labels tend to form excimers upon convergence.
The obtained results are important to understand the influence of polypeptide modifications on secondary structure for further optimizations.

Expansion and suppresive function of regulatory T cells from Type 1 diabetic- and high-risk individuals
Anna Rydén

Type 1 diabetes (T1D) is caused due to pancreatic beta-cell destruction in a process with autoimmune features. Hence, T1D is classified as an organ-specific autoimmune disease even though the pathological mechanisms leading to disease development are not fully understood. The process leading to beta-cell destruction may be initiated several years before T1D diagnosis. Therefore it’s important to study the diabetic and the prediabetic phase to increase our knowledge and pinpoint possible targets for future therapeutic models.
Previous data from us and others have shown imbalance between two prominent parts of the immunesystem, the T-helper 1 and -2 cells, during T1D and other autoimmune states. Regulatory T-cells (Tregs) are known to be important to maintain this balance, self-tolerance and immune homeostasis, and in this study we want to investigate the possibility to expand Tregs from patients in the pre- and diabetic phase for functional studies and eventual future immune intervention.

R1ρ methyl proton spin relaxation experiments for the accurate measurement of microsecond chemical exchange in proteins
Janosch Hennig
NMR spin relaxation in the rotating frame (R1ρ) is a method for atomic-resolution characterization of conformational exchange processes occurring in the microsecond time scale. Here, we add methyl proton relaxation to the repertoire of R1ρ relaxation experiments, to be able to measure the dynamics in side-chain protons of proteins during conformational exchange. This is of special interest for measuring dynamics of protein-protein complexes at their binding interface, where often hydrophobic side chains undergo conformational exchange. The pulse sequence is designed to reduce interference from cross-relaxation effects by applying inversion pulses for 13C and 2H during the 1H spin lock period and by maintaining a tilt angle of θ ≈ 35° throughout the series of measurements and NOESY and ROESY effects may be minimized. 13C-13C coupling during t1 is suppressed by using constant-time evolution. Unwanted proton background is eliminated by sample deuteration. The pulse sequence has been validated using an SH3 domain of yeast abp1p. Crosspeaks corresponding to methyl protons of non-deuterated (CH3), single-deuterated (CH2D), and double-deuterated (CHD2) methyl groups are sufficiently separated, and dispersion profiles in the absence of µs-ms time-scale dynamics are flat. Work is in progress to further validate the pulse sequence on other systems, including the E140Q mutant of the C-terminal domain of calmodulin, which has been shown to exchange between two conformations with a time constant of approximately 20 µs, and on the PDZ-2 domain of SAP97.

Difficulties in doing your Ph.D. - a family perspective
Janosch Hennig

In the second part I will talk about my life during my Ph.D. studies with the focus on later years and general problems, which might occur when you want to get finished and have family to think about.
 

The protein dynamics of the kinase domain of EphB2 studied by NMR spectroscopy
Alexandra Ahlner

Ephrin type-B receptor 2 (EphB2) is a member of the Eph receptor tyrosine protein kinase family that together with their ligands, the ephrins are best known for their role in cell migration and axon guidance during development. EphB2 have also been shown to be involved in intestinal cancer. We are studying the cytosolic kinase domain of EphB2, which seems to be activated by a mechanism different from the one proposed for all other kinases studied so far. Specifically, the crystal structure of the supposedly active form of the protein is reminiscent of the structure of inactive kinases. Interestingly, using nuclear magnetic resonance (NMR) spectroscopy we have identified a small population of another conformation and our working hypothesis is that this conformation is responsible for catalytic activity. By NMR relaxation experiments we hope to verify this and we will also compare the activity of the wild type protein with mutants designed to stabilize the active form of the protein.

April 21, 2010

Synthesis of molecular probes for detection of protein aggregates
Leif Johansson

Protein aggregation is an event related to numerous neurodegenerative diseases, such as Alzheimer’s disease and other prion diseases. Little is still known as to how and why these aggregates form. It has earlier been shown that molecular probes can distinguish different types of amyloid and selectively stain protein aggregates. During this project a library of different probes will be synthesised, and further on used to study the protein aggregation. The probes are thiophene based and emit light with different wave length depending of what they are binding to and therefore the interaction between probe and protein can be investigated by e.g. fluorescent microscopy. 

Searching for a difference between cancer cells and normal cells by using luminescent conjugated polythiophenes, LCPs
Karin Magnusson

Luminescent conjugated polythiophenes, LCPs, can be used for targeting of various components in a complex system of cultured cells. When bound to different entities, the polythiophene derivative probes emit light with different colors due to the unique spectral properties of these conformation sensitive probes. One of the probes can also be used for vital staining of live cells. This anionic probe stains some components in the cytoplasm. The targeting can not be demonstrated in transformed cells, indicating a difference in the localization, structure, accessibility, or quantity of the target in normal cells as compared with malignant cell lines. Determination of the chemical nature of this LCP-biomolecule complex indicates that the biomolecule is some kind of nucleic acid complex that does not derive from the nucleus.
The cells are studied with fluorescence microscopy. The fluorescent target is then purified, amplified and studied by ultracentrifugation, High Performance Liquid Chromatography (HPLC), cloning and DNA-sequencing.

How to prepare yourself for the dissertation
Tobias Ekblad

This presentation will be a bit different, not focusing so much on actual research as on the experience of obtaining a PhD, in particular the final processes. Having recently graduated, and now becoming a Transformer, I would like to try to help you prepare yourself for what is coming, using myself as an example. In short, the presentation will be a retrospective analysis of one PhD - mine. I will try to mainly address general issues which I think you will all encounter at some point.

Insulin signaling in human diabetic and non-diabetic fat cells
Cecilia Karlsson

When insulin binds to the insulin receptor at the cell surface it initiates a cascade of phosphorylation and de-phosphorylation of numerous of proteins through the cell that eventually leads to glucose uptake. These post translational modifications of the proteins can act either activating or deactivating. In type II diabetes when the cells are insulin resistant more insulin is needed to get the same effect as before. One possible explanation is that the phosphorylation pattern is changed, affecting the signal in an inhibiting way. By using immunoprecipitation and mass spectrometry I will isolate insulin receptor substrate 1 and analyze the phosphorylation pattern to see how it differs between diabetic and non-diabetic patients.

March 24, 2010

Synthetic receptors synthesized through SPPS for colorimetric protein sensing
Robert Selegård

Since introduced in the 1960’s solid phase peptide synthesis (SPPS) has become the standard chemical technique for synthesis of peptides and small proteins.
Apart from SPPS offering a strategy to synthesize de novo designed sequences, it allows for an alternative way to produce naturally occurring peptides which are difficult to express in bacteria. The versatility of the technique allows for incorporation of unnatural amino acids, D-amino acids and for alteration of the peptide backbone.
This presentation outlines the general principles of SPPS and its use in the synthesis of a versatile synthetic receptor for colorimetric protein sensing.

Expansion and suppresive function of regulatory T cells from Type 1 diabetic- and high-risk individuals
Anna Rydén

Type 1 diabetes (T1D) is caused due to pancreatic beta-cell destruction in a process with autoimmune features. Hence, T1D is classified as an organ-specific autoimmune disease even though the pathological mechanisms leading to disease development are not fully understood. The process leading to beta-cell destruction may be initiated several years before T1D diagnosis. Therefore it’s important to study the diabetic and the prediabetic phase to increase our knowledge and pinpoint possible targets for future therapeutic models.
Previous data from us and others have shown imbalance between two prominent parts of the immunesystem, the T-helper 1 and -2 cells, during T1D and other autoimmune states. Regulatory T-cells (Tregs) are known to be important to maintain this balance, self-tolerance and immune homeostasis, and in this study we want to investigate the possibility to expand Tregs from patients in the pre- and diabetic phase for functional studies and eventual future immune intervention. 

Nanoparticles for biomedical imaging; a combination of fluorescent probes and MRI contrast agents
Linnéa Axelsson

An important diagnostic and imaging tool in medicine is magnetic resonance imaging (MRI) based on the physical phenomena of nuclear magnetic resonance (NMR). MRI images the NMR signal from the hydrogen nuclei, which constitutes about 63% of the human body. The contrast between tissues can be enhanced by chemical compound called contrast agents that interact with the relaxing nuclei. Another imaging method in medicine is fluorescent labeling, which can be used as an imaging technique for diagnosis of for example cancerous cells. My objective in this project is to combine these two properties, designing a fluorescent MRI contrast agent. 

Effect of Water Fluxes through Aquaporin-9 on Cell Morphology and Motility
Thommie Karlsson

Water fluxes over the plasma membrane are mainly driven by pore-forming proteins known as aquaporins (AQPs). Until today, 13 different AQPs have been discovered and they are known to be expressed in most tissues of the body. AQP9 is known to be particularly abundant in highly motile human leucocytes. Incidentally, several AQPs have been shown to play pivotal roles in cell motility, but the mechanisms behind these findings remain unknown. Our aim is to address this issue by using various live-cell imaging techniques, such as total internal reflection microscopy, laser scanning confocal microscopy and grid confocal microscopy combined with time-lapse image acquisition to visualize potential effect s of AQP9 on cell motility and morphology at the single cell level. Our results indicate that there is enhanced migration, when over-expressing AQP9 in various cell types. Wesuggest, that distinct, dynamic re-distributions of this protein promote the formation of highly dynamic filopodia and lamellipodia. Here, we also present a hypothetical working model for such AQP9-induced membrane protrusions.

February 24, 2010 

Micro- and nanostructured materials for cell research
Anders Elfwing
In current neural stem cell research the standard tool for in-vitro studies are polystyrene petri dishes. Petri dished differs, however, significantly from the developing brain regarding stiffnes, surface chemistry and surface structure. The use of a surface also means the cells deprived of a three dimensional environment.
The aim of my project is to develop a three dimensional system for the study of neural stem cells with an ordered micro- and nano-structure with the possibility to add functionalities like electronic stimulation.
In this talk I will focus on the presentation of the problem.

A phase III clinical trial to investigate the impact of GAD₆₅ (Diamyd) in newly diagnosed type 1 diabetic children
Stina Axelsson
Type 1 diabetes (T1D) is caused by a deficiency of insulin as a result of an autoimmune destruction of the pancreatic beta cells. Glutamic acid decarboxylase (GAD) 65 is one of the main targets of the immune system in T1D. Clinical results from a Swedish phase II study have shown that vaccination with GAD₆₅ (Diamyd™) in children with T1D induces the preservation of the beta cell function. Recently a European clinical phase III study was initiated, including 148 children from 20 clinics in Sweden. The purpose of this study is to determine whether Diamyd (GAD65 formulated in alum) is effective in preserving the body's own insulin producing capacity in patients recently diagnosed with T1D. Our hypothesis is that administration with GAD₆₅ will induce tolerance by modulation of the GAD₆₅-specific immune response in the treated group compared to the non-treated group. My focus is to study the effect of vaccination on the immune system, using methods such as Luminex, PCR-array and 9-color flow cytometry.  

January 27, 2010

ELISA - A simple way to determine protein levels
Linda Andrén
It is already known that amyloid proteins aggregate and form plaques, tangles and other destructive structures. It is thought that this is due to an accumulation of protein over time, in my work with Lysozyme amyloidosis, determining the levels of Lysozyme at different times is very interesting in an attempt to investigate if this accumulation occurs continuously or intermittently.
A convenient method for determining the concentration of a specific protein in a solution is to use a sandwich ELISA, Enzyme-linked Immunosorbent Assay, where a pair of antibodies with an affinity for the analyte of interest is cleverly used to assess the protein levels.

Analysis of histone modifications in human fat cells
Åsa Svensson
Genetic studies of type 2 diabetes show that it is a very heterogeneous and polygenic disease. Recent studies have suggested that epigenetic factors, such as post-translational modifications of histones, may relate to insulin resistance.
To analyze modifications of histones in primary human adipocytes we isolated cells from subcutaneous abdominal fat tissue and developed an extraction method for basic proteins which gives samples containing 60-70 % histones.
By using mass spectrometry we can identify the proteins, identity and position of different modifications, as well as the combination of modifications on the histones. By proteolysis with ArgC-protease and subsequent LC/MS/MS analyses we identified all five histone classes (H1, H2A, H2B, H3 and H4) and several different histone isoforms. We also revealed many modifications of which the majority has not been annotated before.

Enhancement Strategies for Fluorescence-based Biosensors
Erik Martinsson
Fluorescence-based biosensors are becoming increasingly important in biomedical diagnostics, especially in point-of-care testing for rapid detection of disease-related biomarkers. These devices are usually based on an array of biorecognition elements which captures the analyte of interest and are detected through the use of fluorescent labels. Although, in order to meet the future requirements of biosensors with increased sensitivity and limit of detection, new ultra-sensitive devices must be developed and implemented.
In this presentation, I will describe some strategies for enhancing the analytical performance of fluorescence-based biosensors, focusing on a method known as plasmonic enhancement.

Title to be given later
Liza Ljungberg

November 25, 2009

Electronic Surface Switches Based on Conductive Polymers to Control and Regulate Cell Systems
Kristin Persson
Many different types of cells adhere to solid surfaces while in culture, and it is therefore of great interest to be able to control and regulate them by means of the surface they are attached to. We have worked with electroactive surfaces based on the conductive polymer PEDOT, which has been used in a number of applications involving cells and is known for its biocompatibility as well as for its electronic and ionic conductivity. We have previously shown that PEDOT can be used to control cell adhesion and proliferation and our ultimate goal is being able to regulate and control all events during cell growth; adhesion, spreading, proliferation, differentiation and release of cell systems. Thin films of PEDOT have been deposited by various means, such as electropolymerisation and vapour phase polymerisation, on surfaces dedicated for cell growth, and employed as electrodes during the cell culture period. The same kinds of surfaces have also been used to evaluate the effects on bacteria in culture, finding that the redox state of the polymer influences bacterial adhesion and subsequent biofilm formation.

Multi-scale top-down modelling of type 2 diabetes: extending a whole-body glucose homeostasis with adipocyte insulin signalling
Cecilia Brännmark
In type 2 diabetes (T2D) target tissues of insulin become resistant to insulin, and the concentration of glucose in the blood is increased. As this insulin-glucose interplay is highly complex, a model-based approach might be advantageous. To achieve the full effect of such modelling, it must be possible to combine detailed models for sub-system, into larger internally consistent whole-body models. We here present a combined top-down/bottom-up approach for the hierarchical development of such models.
We illustrate the approach on a pilot study where we have extended a physiologically-based whole-body model to include a zoomable module for the adipose tissue.
The resulting model translates mechanistically oriented simulations on the biochemical level, which is the level were drugs act, to the whole-body level, which is of clinical and diagnostic interest. Our approach and pilot study opens the door for future hierarchical modelling – and ultimately drug certification and treatment – of insulin resistance, glucose homeostasis and type 2 diabetes.

Colour Moiré Interferometry for Ubiquitous Physical Sensing
Stephen Macken
Digital cameras are easy to use and yet sophisticated equipment that sees improvements with each new generation. They are very versatile and in my work I use digital web cameras to perform physical measurements on different targets.
Presently I am working on using the web camera as a weighing scale. In my presentation I will bring you through the technique while giving you a brief explanation about how it all works.

The role of platelets in inflammation
Caroline Skoglund
Blood platelets have for long been recognized during haemostasis but are now also emerging as key actors during inflammation. Platelets also interact with bacteria and express innate immunity receptors, such as the toll-like receptors (TLRs). Toll-like receptor 2 (TLR2), recognize conserved microbial pathogen-associated molecular patterns and it has recently been shown that the TLR2/1 agonist Pam3CSK4 (a tri-acetylated lipopeptide) stimulates platelet activation. In the present study we have investigated the signaling events provoked in platelets upon stimulation with Pam3CSK4.
Platelet cytosolic Ca2+ levels were using the fluorescent ratiometric probe Fura-2. Briefly, platelets were loaded with Fura-2 by incubating platelet rich plasma with Fura-2-acetoxymethylester (AM) for 40 min at room temperature and gentle shaking. Platelets were then isolated and measurements of cytosolic Ca2+ were performed under stirring conditions (300 rpm) at 37 °C. Fluorescence emission was registered at 510 nm upon excitation at 340 nm and 380 nm. By addition of Triton X-100 followed by EGTA the maximal and minimal ratios were determined and the changes in intracellular Ca2+ was calculated.
The results showed that Pam3CSK4 treatment of platelets potently increased the [Ca2+]i within seconds after stimulation and that the increase was dose-dependent and comparable with the Ca2+ increase induced by thrombin, a well known platelet agonist. The [Ca2+]i increase was inhibited by an ATP-receptor antagonist (MRS 2159), a phospholipase C (PLC) inhibitor (U73122) and aspirin whereas ADP-receptor antagonists (MRS 2179 and cangrelor) had no effects. These results support the role of platelets in innate immunity.

October 28, 2009

Carbohydrate synthesis – The sweeter side of life
Timmy Fyrner
Areas in which carbohydrates can play a significant role are numerous, e.g., glycoconjugated vaccines, protein resistance surfaces or as ordinary sweeteners. However, due to the vast complexity of naturally occurring glycoconjugates, simplifications or appropriate model systems are required. The synthesis of orthogonally functionalized oligosaccharides allows for many possible conjugation methods and it requires persistence and high precision from the practitioner, using NMR as a powerful tool in product/structure elucidation. In our research a potentially protein resistant, highly defined, oligosaccharide-based linker molecule is being synthesized.

Immune modulation therapy using GAD65 to induce tolerance
Mikael Chéramy
Type 1 diabetes (T1D) is an autoimmune disease characterized by the destruction of insulin producing pancreatic β-cells. One T1D related autoantigen is Glutamic acid decarboxylase (GAD) 65, present in the cytoplasm of β- cells. Approximately 70-80% of newly diagnosed T1D patients have autoantibodies to GAD65 (GADA).
A number of strategies for treatment or prevention of T1D have been investigated or are currently ongoing. Immune modulation therapy using GAD65 to induce tolerance has been tested in a phase II trial conducted by our group.
An idiotope is the unique set of antigenic determinants (epitopes) of the variable portion of an antibody. If a separate antibody is produced that has specific binding capabilities to an idiotope, it is said to be an "anti-idiotypic antibody". The anti-idiotypic antibodies will be able to bind to the primary antibody (i.e GADA) and may inhibit the autoimmune response.
We will analyse anti-idiotypic antibodiesto GADA, using a new method, in serum from T1D patients treated with GAD65.

Controlled Protein Orientation by Chelation Assisted Photoimmobilization
Emma Ericsson
To enhance efficiency of biosensor surfaces a novel surface chemistry has been developed to render a proper orientation of the immobilized protein (ligand) and thereby facilitate the subsequent biomolecular interaction with the corresponding analyte. This is done in a two step process where the ligand is first oriented using chelate chemistry and then permanently locked in the proper orientation using photochemistry.
A protein modified with a hexahistidine (His) sequence can be coordinated to a chelate complex of a nickel ion (Ni2+) and NitriloTriacetic Acid (NTA). This is commonly used in protein chromatography but is used here as a means of orienting the protein by surface bound NTA. Protein immobilization to surfaces by photochemistry can be performed with the photolabile agent benzophenone (BP) which is activated by UV light. The use of BP alone results in a random orientation of the protein, but surface bound BP is used here to covalently bind the preoriented protein. We call this novel combination of surface bound NTA and BP “Chelation Assisted Photoimmobilization” (CAP).
Results from Surface Plasmon Resonance (SPR) measurements using a Biacore 3000 instrument indicate that a considerably larger analyte response is obtained with the CAP chemistry as compared to the response obtained with the random orientation achieved by photoimmobilization alone.

Improvement of metaphyseal bone formation by pharmacological treatment
Fredrik Agholme
Most fractures occur in osteoporotic cancellous bone. When an orthopaedic implant is inserted into bone, an injury is created which initiates bone repair. By influencing bone formation and repair it is possible to increase the healing rate, which leads to a shorter healing period for fractures and a better fixation of implants. The aim of this project is to evaluate the bone response to implant insertion trauma, in combination with pharmacological treatment. The prospect of increased healing rate will benefit all fracture patients. The goal is to expand the treatment options for the orthopaedic surgeon when treating bone disease and fractures. We have previously done much work on bisphosphonates and parathyroid hormone. However, as the importance of wnt signalling has emerged, most future work will concentrate on drugs that modulate this pathway. We study the effect on bone healing with in vivo models. By applying different screw coatings or treatments, the change in cancellous bone mechanics can be followed. Data is analysed using histology and mechanical testing.

September 30, 2009

TRIM21/Ro52, an E3 ligase involved in autoimmune diseases
Janosch Hennig
Several members of the tripartite motif (TRIM) protein family are involved in antiviral activity and immunity and have been linked to several diseases. TRIM21, the main object of this study, is involved in Sjögren syndrome (SS) and systemic lupus erythematosus (SLE), where patients often have autoantibodies against different epitopes on TRIM21. A role of TRIM21 in regulation of proinflammatory cytokines and autoimmunity emerged during recent years. The aim of this study is to provide a better understanding of the structure-function relationship of TRIM21. A wide range of biophysical methods were employed to establish a model of the protein domain arrangement of TRIM21, and functional implications were derived. By sequence comparisons, TRIM proteins are classified into three subgroups, sharing a conserved amphipathic helix in the region, linking the conserved N-terminal Zn$^{2+}$-binding domains RING and B-box, called the RING-B-box linker (RBL). A structural dependence of this region on the RING has been observed and a model of the RING-RBL was derived from bioinformatics and proteolysis data. Anti-RING-RBL antibodies inhibit the E3 ligase activity of TRIM21 in ubiquitination. Interferon regulatory factors (IRFs), the substrate for TRIM21-dependent ubiquitination could therefore retain their high cellular levels after stress-induced inflammation, increasing the susceptibility to SS and SLE. According to NMR data, the antibodies bind to the Zn$^{2+}$-binding loop regions of the RING, which usually bind to the E2 conjugating enzyme. Antibodies against the C-terminus of the RBL region do not inhibit the E3 ligase activity.

Interfacial Molecular Design and Characterization of Biospecific Surfaces
Hung-Hsun Lee
Supported lipid membranes (SLMs) are a powerful model system in biophysical research and a promising technological platform for applications in biosensors and membrane protein arrays. They consist of a lipid bilayer supported by solid surface via a thin organic molecular monolayer. This study is to develop a flexible and stable SAM system which can be applied to bio-mimetic interfaces for the assembly of SLMs. The SAMs were characterized by means of ellipsometry, contact angle goniometry, and infrared reflection-absorption spectroscopy (IRRAS). Vesicle fusion of 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC) on the SAM-coated solid surfaces was investigated employing QCM technique. QCM provides real-time measurement of both mass and structural properties of adsorbed molecular layers.

Organic Bioelectronics for Controlled Release of Ions and Biomolecules
Klas Tybrandt
The use of conducting polymers in biological applications has received significant attention lately. Desirable properties such as electronic and ionic conductivity, flexibility and biocompatibility make some polymers well suited for use in biological systems. We have previously demonstrated an organic electronic ion pump (OEIP) for controlled release of metal cations. By applying a voltage across the device, ions are transported from one container to another through the polymer film. The delivery rate is controlled by the applied voltage and the amount of delivered ions agrees with the integrated electric current. Here we report on a novel device architecture, which enables both spatially and temporally controlled release of charged biomolecules. To demonstrate the functionality of the device, the neurotransmitter acetylcholine is used to stimulate SH-SY5Y neuroblastoma cells in vitro.

Differences in the signalling cascades triggered by PAR1 and PAR4
Karin Vretenbrant Öberg
The platelet protease activated receptors (PAR) 1 and 4 have traditionally been considered to act through the same signalling cascades. To further elucidate this issue, we chose to study the action of PAR1 and PAR4 in combination. Our hypothesis was that there are differences in the signalling cascades triggered by PAR1 and PAR4, respectively. If the receptors were acting through the same cascade, simultaneous activation of them would result in an additive response. However, if they in any respect differ in action one would expect higher or lower response than additive. The response to combinations of the PAR1 activating peptide (AP) SFLLRN and PAR4-AP AYPGKF were determined in parallel with the concentration–response for each agonist alone with both flow cytometry and platelet aggregation. We detected a strong synergistic effect on platelet activation when PAR1 and PAR4 were co-stimulated with low concentrations of activating peptides. This strong synergy effect was also evident in aggregation of platelet rich plasma. Because when PAR1 and PAR4 were simultaneously activated the yielded response was much higher than additive indicating a strong synergistic action. This result leads us to conclude that the two thrombin receptors have differences in their signalling pathways.

September 2, 2009

The development of analysis methods based on luminescent conjugated oligothiophenes (LCOs) to study protein amyloid disease
Therése Klingstedt
The formation of highly ordered aggregates of intra- or extracellular proteins underlies a wide range of neurodegenerative conditions including prion, Parkinson¿s, Huntington¿s and Alzheimer¿s diseases. The main structural hallmark of protein aggregates is the cross -structure found in the highly ordered inner core and this is thought to be responsible for the tinctorial properties. We have introduced luminescent conjugated oligothiophenes (LCOs) as a novel class of conformation sensitive optical probes for selective staining of various types of protein aggregates. The property of LCOs to give rise to conformation-dependent emission spectra upon binding to protein aggregates has been utilized in several platforms such as cell systems and tissue samples. My aim is to develop methods based on LCOs to characterize protein aggregates found in different parts of the body such as the brain, liver and heart.

T-cell polarisation profiles in children with and without celiac disease (CD) and type 1 diabetes (T1D)
Anne Lahdenperä
Celiac disease (CD) and type 1 diabetes (T1D) are immunological diseases associated with inflammatory reactions in the target tissues. The reactions include T cell infiltration, cytokine production, mainly T helper(Th)1 cytokines, and antibody production. Both genetic and environmental factors are of importance in the disease development. The gut-intestinal immune system seems to be an important link. Circulation of autoreactive T cells in the body, due to abberant immunological responses in the gut-intestine, might induce inflammation, autoimmune reactions and β-cell death in pancreas. Little is known about the T-cell polarisation profiles in peripheral blood mononuclear cells (PBMC) and gut-intestinal biopsies from children with and without CD and T1D. The purpose of the study is to investigate T-cell expression profiles in PBMC and small intestinal biopsies from children with and without CD and T1D, and to study the effect of the gluten free diet (GFD) treatment. Real-time RT-PCR arrays (containing 84 genes) are used in the study, which are promising tools for gene expression studies. The aberrant T-cell responses in children with CD were generally normalized after GFD-treatment. However, the gene expression profile in blood was generally not reflecting the gene expression profile in the small intestine.

Rare earth nanoparticles as contrast agent in MRI :
Nanomaterial design and biofunctionalization
Maria Ahrén
Today, gadolinium ion complexes and superparamagnetic iron oxide nanoparticles are used routinely as magnetic resonance imaging (MRI) contrast agents. Nanoparticles of iron oxide 2-6 nm, capped with a passivating non-ionic material, have been shown to produce negative contrast in MRI, whereas gadolinium ion complexes produce positive contrast. Our aim is to design biofunctionalized Gd2O3 nanoparticles to achieve a contrast agent more efficient than the commercially available Gd chelates. Capping and biofunctionalization of the nanoctrystals are done to increase solubility, reduce the toxicity and increase specificity of the material. The chemical and physical properties of the nanomaterial are investigated with Fourier Transform Infrared Spectroscopy, X-ray Photoelectron Spectroscopy, Transmission Electron Microscopy and Dynamic Light Scattering. The proton relaxation times as a function of dialysis time and functionalization are measured at 1.5 T.

Title to be presented later
Hanna Björck

May 13, 2009

Biophysical Analysis of Proteins using MALDI-TOF-MS
Cecilia Andrésen
MALDI-TOF-MS (Matrix-Assisted-Laser-Desorption-Ionization-Time-of-Flight-Mass-Spectrometry) is frequently used for determination of the molecular weight of biomolecules but the technique is also suitable for a large number of biophysical analyses of proteins such as peptide mapping, disulphide bridge determination, protein labeling assays and folding properties. The technique involves soft ionization of molecules preventing fragmentation of sensitive proteins and low protein concentration can be used in physiological buffers.
When analyzing the oxidation state of Grx4 (Glutaredoxin from e-coli), MALDI was a useful tool together with enzymatic digestion. During the oxidized state, a disulfide bridge could also be determined showing the two Cysteins involved.

Drosophila melanogaster – a model system for protein aggregation
Sebastian Schultz
Recent research supports that aggregation of islet amyloid polypeptide (IAPP) leads to cell death and makes it a plausible cause for the halving of beta cell mass that occurs in patients with type 2 diabetes. IAPP and insulin are both produced as prohormones and processed into biological active hormones in the secretory granules by the convertases PC1/3 and PC2, prior to their stimulated co-secretion. In vitro, IAPP is one of the most amyloidogenic peptides known, while the presence of insulin can inhibit IAPP-fibril propagation. It is unclear if the intragranular equilibrium of prohormones and their putative processing metabolites affect amyloid formation. At present, I am in progress to establish a Drosophila melanogaster model that will allow studies of this. So far we were able to show the production of amyloid in Drosophila and now have the chance to utilize our newly established model system to dissect interactions involved in amyloid formation in vivo.

Force measurements for novel surfaces
Feng-I Tai
Atomic force microscopy (AFM) associated analytical methods have been introduced into multidisciplinary research fields and extensively developed for various studies in molecular scale. Hence information of morphology, magnetic/electric/chemical potential, force interactions, etc. can be obtained with possible atomic-scaled resolution.
Molecular self-assembly building blocks and nano-scaled functional materials, such as polypeptides and polyelectrolytes in this work, are potentially designed to for novel applications. In particular, for biological functionalities and surface modifications, the two systems introduced above have been intensively investigated.
Direct force measurements were meant to study both the structural and the intermolecular force properties of novel surfaces. The surface charge properties of polyelectrolyte gradients are demonstrated as an example.

Blood flow measurements at different depths using laser Doppler flowmetry and Photoplethysmography techniques
Sara Bergstrand
In the health care section there are special mattresses for prevention and treatment of pressure ulcer. Thus, there is no consensus of which parameters that are most suitable for evaluation of these products. We state that interface pressure, blood flow, temperature, humidity and shear force is important variables for evaluation. This project will develop a multi parametric system for evaluation of special mattresses for pressure ulcer. For the blood flow measurements, photoplethysmography (PPG) and laser Doppler flowmetry (LDF) are used. The LDF and PPG techniques reflect the blood flow at different levels. The main findings in the first study with healthy subjects are that the blood flow increases when the subject’s tissue is exposed to mechanical load. Reactive hyperaemia, a strong indicator of previous ischemia, seems to occur more frequent in the superficial layers of the tissue, as LDF following by green light at PPG most often detected the phenomenon.

April 15, 2009

Biofunctionalized Surfaces: The Molecular Orientation and Interaction Studies
Cecilia Vahlberg
Design of biologically inspired materials, is important in biosensor applications. Well organized self assembled monolayers, mimicking naturally occurring biomolecular structures, are useful as model systems. Our aim in this project is to design biofunctionalized surfaces to mimic e.g. biorecognition processes. We investigate self assembled monolayers of peptides, neurotransmitter derivatives and boronic acid derivatives. The molecular adsorbates are studied with surface sensitive techniques such as X-ray Photoelectron Spectroscopy (XPS), Infrared Reflection Absorption Spectroscopy (IRAS) and Near Edge X-ray Absorption Fine Structures (NEXAFS). Surface Plasmon Resonance (SPR) has been used to investigate biointeraction processes.

Tailoring nanoscale films
Sara Nilsson
Interactions between charged surfaces and various molecules regulate many industrial processes and the technique of continuous adsorption offers, in a single step, the possibility of tailoring nanoscale films. It has previously been shown that the adsorption of amine sidechain-containing polycations, upon a modest applied potential, is often asymptotically linear and weakly dependant on concentration for long duration measurements. This suggests that film growth is limited by a chemical process within the film itself, which has not been identified. My research this spring will involve investigations of the adsorption of charged poly-L-lysine under the influence of an applied electrical potential. I will use the EQCM-D technique, which enables label-free measurements of molecular adsorption on various surfaces in real-time. The dissipation parameter, available in QCM-D measurements offers information about the viscoelastic properties of the adsorbed layers. I expect that this data, when combined with previous measurements, will help elucidate the chemical mechanism within the film that regulates the deposition rate.

Modeling Transthyretin Amyloid Diseases in Drosophila Melanogaster
Ina Berg
Amyloid diseases arise from the misfolding and aggregation of a wide variety of nonhomologous extracellular proteins. Amyloid fibrils or oligomeric precursors thereof, appear very similar regardless on protein. Therefore, it is likely that increased understanding of amyloid diseases can be gained by studying and comparing a variety of different amyloid proteins and diseases. The Drosophila model system is increasingly being used for modelling human degenerative diseases. We generated transgenic Drosophila expressing single gene products of the human amyloidgenetic proteins Transthyretin under control of the Gal4/ UAS system (expressing system in Drosophila). We find that expression of Transthyretin in the central nervous system results in a reduced lifespan when compared to control flies, which was comparable to the effect of Aβ1-40 and Aβ1-42 expressing flies run in parallel. Since amyloisgenic proteins lose their normal conformation when building up amyloids, staining with antibodies are difficult. Since the expression of proteins in Drosophila are under the control of the Gal4/UAS system, methods on amyloid staining of different proteins can be tested out in Drosophila.

Title to be announced later
Pernilla T Eliasson
Text to be submitted later

March 18, 2009

Molecular probes for selective in vivo imaging
Andreas Åslund
Molecular probes for selective in vivo imaging of protein aggregates are important to advance our understanding of the molecular mechanisms underlying neurodegenerative cerebral amyloidoses. Here we report the chemical design of luminescent conjugated oligothiophenes (LCOs) which displayed selective and specific binding towards both recombinant Aβ fibrils and protein aggregates in human brain tissue samples with Alzheimer’s disease (AD) pathology. One of the LCOs, p-FTAA, showed conformation-dependent optical properties distinguishing i) the major hallmarks of AD brains, namely Ab aggregates versus neurofibrillary tangles and ii) prion deposits from two mouse-adapted prion strains. Furthermore, LCOs crossed the blood-brain barrier following intravenous administration and allowed real-time visualization of Aβ aggregates in a mouse model of AD by multiphoton microscopy. Overall, LCOs can be utilized as practical research tools for studying protein aggregation diseases and facilitate the study of amyloid origin, Ab-tau interactions and pathogenesis both ex vivo and in vivo.

Title will be added later
Mikael Pihl

Conjugated luminescent polymers
Jens Wigenius
Conjugated luminescent polymers (LCP) are soluble in water and interact none-covalently with biomolecules as DNA, this interaction has previously been used to study a number of bimolecular reactions as for example DNA hybridization. When interacting with DNA, the conformation and packing of the LCP is distorted, leading to changed luminescent properties hence a shift in the emission peek and intensity. This shift could enhance energy transfer from the LCP to a conventional chromophore attached to a DNA strand for detection of DNA hybridization. Combine with soft lithography we show an easy way to fabricate a simple DNA-chip.

What effect has Thrombospondin-1 on focal adhesion kinase phosphorylation and platelet-induced bronchial smooth muscle cell proliferation?
Ann-Charlotte Svensson Holm
Smooth muscle cell (SMC) proliferation is a process associated with many different disorders, e.g. the chronic inflammatory diseases asthma and atherosclerosis. Platelets may play a role in inflammatory diseases, since they possess both inflammatory and proliferative properties. We are, in the current project, going to investigate the effect of Trombospondin-1 on focal adhesion kinase (FAK) phosphorylation and what impact this could have on platelet-induced bronchial smooth muscle cell (HBSMC) proliferation. We are going to use several different methods, e.g. different assays to measure proliferation, Western blot analyses and fluorescent microscopy. We know from our previous studies that platelets bind to HBSMC and induce proliferation and our hypothesis is that platelets release Thrombospondin-1 upon binding to HBSMC and thereafter activate FAK, resulting in an increased cell proliferation.

February 18, 2009

Luminescent Conjugated Oligomers: Molecular probes
Rozalyn Simon
Conjugated polymers have been utilized in a variety of applications ranging from organic electronic devices to chemical sensors. Delocalization of electrons over the conjugated backbone allows for the emergence of a variety of useful features such as conductivity and luminescence. Polythiophene is one such polymer that has been exploited for just these features. Upon interaction with neighboring molecules, changes in the planarity of the thiophene rings result in differences in conjugation lengths over the polymer backbone and thus a spectral shift. Functionalization of the backbone can also be used to alter conjugation length. Utilizing functionalization of this type, one can tailor the spectral response upon interaction with specific molecules. While the potential applications for this technique are innumerable, our immediate aim is to synthesize a variety of oligomeric thiophenes tailored for the detection of misfolded protein aggregates. Via the synthesis of a small group of similar oligomers with slight variations, it is possible to study the importance of the functionalization at specific positions on the polymer and in this way, optimize for the intended applications in molecular detection.

A phase III clinical trial to investigate the impact of GAD65 (Diamyd) in newly diagnosed type 1 diabetic children
Stina Axelsson
Canceled because of sickness
Type 1 diabetes (T1D) is caused by a deficiency of insulin as a result of an autoimmune destruction of the pancreatic beta cells. Glutamic acid decarboxylase (GAD) 65 is one of the main targets of the immune system in T1D. Clinical results from a Swedish phase II study have shown that vaccination with GAD65 (Diamyd™) in children with T1D induces the preservation of the beta cell function. Recently a European clinical phase III study was initiated, including approximately 100 children from 20 clinics in Sweden. The purpose of this study is to determine whether Diamyd (GAD65 formulated in alum) is effective in preserving the body's own insulin producing capacity in patients recently diagnosed with T1D. Our hypothesis is that administration with GAD65 will induce tolerance by modulation of the GAD65-specific immune response of the treated group compared to the non-treated group. My focus is to study the effect of vaccination on the immune system, using methods such as Luminex and real-time PCR.

Title will be added later
Maria Bolin
Abstract will be added later

January 21, 2009

Title will be added later
Fredrik Lysholm
Abstract will be added later

Expression of mutant IRS-1 in human adipocytes using an adenovirus based system
Siri Fagerholm
Our group has earlier found a link between impaired insulin signaling in fat cells (adipocytes), and type 2 diabetes. Following insulin stimulation of an adipocyte, complex intracellular signaling pathways becomes initiated. One of these pathways engages the insulin receptor activated protein insulin receptor substrate 1 (IRS1). Following phosphorylation on multiple tyrosine sites, IRS1 becomes activated and thus further propagates the insulin signal downstream by activating a number of signal mediators. The phosphorylation of tyrosines is regulated by a serine residue (Ser307), phosphorylation of this site mediates a positive feedback loop that increase the steady-state level of tyrosine phosphorylation. Our group has found an impaired tyrosine and Ser307 phosphorylation in adipocytes from patients with type 2 diabetes.

Our aim is to ”rescue” adipocytes from patients with diabetes from their diabetic state through restoration of their IRS1 mediated insulin signaling pathway. Using adenoviruses we will overexpress a mutated form of IRS1 where Ser307 has been exchanged to an aspartic acid residue, mimicking a constantly phosphorylated IRS1. We will also induce overexpression of Ser307Ala IRS1 (to mimic an unphosphorylated state) and wt IRS1 (as a control). If needed, we will try to knock-down the expression of endogenous IRS1 using siRNA. Using this method we hope not only to be able to study the specific effect of serine307 phosphorylation on insulin signaling but also other IRS1 residues and proteins in the insulin signaling pathway.

Sensors for control of nitrogen oxides in exhaust gases
Kristina Buchholt
Fuel consists of hydrocarbons which break down into smaller molecules during combustion. For diesel engines some of the fuel ends up as particulate matter, one of the main pollutants in exhaust gases. Another main pollutant is nitrogen oxides. NOx react with hydrocarbons in sunlight and produce ozone and photochemical smog. Ozone can cause breathing difficulties and damage to plant life and NOx can increase respiratory illnesses and contributes to acid rain. Around the world emission laws and legislations are becoming stricter due to the increased knowledge of the adverse environmental, and health effects of emissions from cars and industry. To be able to fulfill these strict emission laws the car industry is in need of selective, long-term stable sensors able to withstand high temperatures and corrosive environments. My research is aimed at achieving sensors fulfilling these demands with focus on developing new sensing layers, and ohmic contacts, for SiC-based field effect sensors.

Food intake in a home-living elderly population
Yvonne Johansson
Most investigations of food intake among elderly people have been performed in nursing homes and hospitals. The aim of this study was to investigate home-living elderly people’s food intake, related to risk for malnutrition, symptoms of depression and demographical factors. Another aim was to describe changes of food intake during two years follow up. One hundred and fifteen 80-years old individuals were interviewed about their meal habits at baseline and after one and two years. A dietary assessment, 24-h recall, was performed once a year with the objective to obtain data of food consumption and a possible change of habits. At baseline 12% were at risk for malnutrition assessed with Mini Nutritional Assessment. Data from the 24 h recall showed, irrespective of nutritional status, that 40% of the participants reported an inadequate energy intake according to Nordic Nutrition Recommendations. Seventy-six percent reported inadequate levels of iron and 82% inadequate levels of D-vitamin.

November 26, 2008

Prenatal hypoxia programs an altered β1/β2 adrenoceptor subtype ratio in the heart of adult broiler chickens
Isa Lindgren
β-adrenoceptors (βARs) are essential for fetal development of the heart and regulation of cardiac activity. Adverse events during the development of the fetus may have deleterious effects on the adult phenotype and may predispose the latter for pathologies such as cardiac failure and hypertension.
Previous studies from our lab show that chronic prenatal hypoxia renders an adult animal with decreased sensitivity to cardiac βAR stimulation. In this study, we investigated whether low fetal oxygen conditions have a programming effect on the β1/β2AR ratio in the adult, causing the previously shown decrease in adult βAR sensitivity.
We studied total β-AR density (Bmax) and the ratio of β1/β2 subtypes in the broiler chicken heart (14 and 35 days post hatching, P14 and P35) prenatally developed in normoxia (21% O2, N) or chronic hypoxia (14% O2, H). To obtain total Bmax we incubated ventricular tissue with the tritiated β-antagonist [3H]CGP-12177. The β1/β2 ratio was obtained by specific inhibition of β1 by CGP-20712A and β2 by ICI-11855 in the presence of 2nM of [3H]CGP-12177.
Total Bmax did not differ between treatments in either P14 or P35, but βAR density decreased with age. There was significantly more β2 than β1 in P35H, while P35N showed a 50/50 ratio of the two subtypes.
Prenatal hypoxia programs an altered βAR subtype ratio in the adult in favor of β2 which may be the cause of the previously observed decrease in βAR sensitivity. Supported by FORMAS (Dnr. 2004-1331).

Analysis of histone post translational modifications in adipocytes
Åsa Svensson
Histones are small, basic, nuclear proteins that are responsible for the structure of DNA in eukaryotes. DNA wrap around a group of histones forming nucleosomes, which in turn are the basis of chromosomes. The histone N-terminal tails are pointing out of the nucleosomes and are heavily modified. These modifications are often called “the Histone Code” and the combination of modifications control gene transcription together with transcription factors.
The rise of a new, soft fragmentation technique for mass spectrometry, Electron Transfer Dissociation (ETD) allows us to study all posttranslational modifications (PTMs) sequentially intact on peptide fragments. Compared to older fragmentation techniques where many modifications were lost without a trace- this is a big advantage in the exploration of for example the Histone Code. My aim is to study the behaviour of the Histone Code in type2-diabetic and non-diabetic patients.

A Flow System for Urea and Glucose Measurement with a Self-Polishing Electronic Tongue
John Olsson
A temporary flow system connected to a self polishing pulse voltammetric electronic tongue was evaluated for prediction of small changes in urea and glucose concentrations in phosphate buffer. Two batches of samples standards were used; one for calibration and the other for validation. The ad-hoc system could easily predict changes at around 2mM for both urea and glucose. No significant difference between the two batches was seen.

Controlled drug delivery enhance the implant fixation on bone
Paula Linderbäck
To achieve long-time survival for bone implants and to avoid clinical failure, early fixation of an implant is of great importance. Different kind of coatings together with Immobilization of drugs has been used to develop a bioactive surface to enhance bone formation. Bisphosphonates are used for the treatment of osteoporosis and have been showed to have a positive effect also on bone implant. Recently developed drug strontium ranelate, is a new potential treatment against osteoporosis. The objective of my study is to investigate new method to immobilize medicines to the implant surface. For this purpose, bisphosphonates was bound to sol-gel derived titania coated screws through calcium crystals. The screws were implanted in rat for 4 weeks. The purpose of the second study was to investigate the effect of strontium on implant healing in rat.
 

October 29, 2008

Thiopurine methyltransferase activity effects chemotherapy
Patricia Wennerstrand
Thiopurines are frequently used drugs in order to treat childhood leukemia and autoimmune diseases. The key enzyme, in the metabolic pathway to convert Thiopurines into active metabolites, is Thiopurine methyltransferase (TPMT) where the natural substrate of TPMT is still unknown. TPMT is a polymorphic enzyme, to date 26 mutations are known causing low activity. Patients suffering from low activity are at greatly increased risk for potentially life threatening Thiopurine toxicity. To individualize treatment, patients are genotyped and phenotyped as standard routine to determine the TPMT activity. The theory of what causes the low activity is believed to be degraded protein but the theory has not yet been proved. By expressing recombinant TPMT variants and characterize them biophysically we are trying to reveal the mystery of the variants and explain what causes the low activity. I will present ongoing stability and activity data and focus on a method to determine the stability of proteins.

Mechanisms of initial transient signalling by insulin in adipocytes, a systems biology approach
Cecilia Johansson
Malfunction in the cellular response to insulin, insulin resistance, is a characteristic feature of type 2 diabetes, a disease that affects almost a quarter of a billion people. In diabetes the response to insulin is defect at an early step in signal transduction. Although the basic features of insulin signalling are well established, many important details such as feedbacks, time-scale, and intracellular location, remain largely uncharacterized. We have employed a systems biology approach to elucidate more details concerning these features. Experimentally we are examining primary human adipocytes and monitor the phosphorylation of key molecules in the insulin signalling network upon insulin stimuli. By using mathematical models of these signalling events we are able to evaluate a plethora of hypotheses according to the experimental data obtained. Our data show that the phosphorylation of insulin receptor substrate-1 by the occupied insulin receptor, displays an overshoot behaviour, with a transient peak reached within one minute and a stable steady-state response after the next 3-5 minutes. Such an overshoot behaviour can be explained by several mechanistic hypotheses, including feedback from a downstream signalling molecule and/or internalization and recycling of the insulin receptor to the cell membrane. Our mathematical analysis shows that this overshoot observation alone is sufficient to rule out the assumption that there are no feedbacks during the first minutes. To further elucidate features of these significantly active feedbacks, we perturbed the system with more complex input signals, e.g. containing several steps in the insulin concentration. The data obtained and observations of signalling in cellular sub-fractions, combined with mathematical analysis, give novel and important mechanistic information. With more detailed knowledge of the insulin signalling mechanisms it is possible to understand differences between a normal and an insulin resistant state and develop new strategies to treat type 2 diabetes.

Surface plasmon resonance characterization of the CD20 disease related epitope
Daniel Kanmert
This project, relevant to clinical rheumatology, aims at understanding interactions between Fc-parts of human IgG and the Fcγ receptor IIIa. The FcγRIIIa interacting with IgG-Fc influences the extent to which monocytes/macrophages release pro-inflammatory cytokines. The interaction between FcγRIIIa and human IgG is of low affinity and multivalent binding is needed for cell activation. We have mimicked a cell- or tissue surface opsonized by IgG class antibodies by immobilizing the mAb Rituximab in an orientation controlled manner using the specific interaction with a synthetic peptide derived from the membrane bound protein CD20. This model surface will be used as a tool to identify possible down-regulators of FcγR-initiated inflammatory response. This presentation will focus on the characterization of the mAb/peptide interaction by the use of surface plasmon resonance.

October 1, 2008

Surface functionalization of ZnO nanoparticles and thin films for sensor applications
Linnéa Selegård
A new procedure has been developed for functionalization of electrochemically produced ZnO nanoparticles. The core of the particles was characterized using TEM, PEEM and LEEM. Single crystal nanoparticles, with uniform spherical morphology with a size of approximately 50Å were obtained. The first aims of ZnO nanoparticle functionalization were to make a stabilizing molecular layer at the surface and to prepare for further linking possibilities, for use in different types of sensing applications. The functionalization process was investigated and the molecular layer was verified by XPS and FT-IR. Parallel studies on biofunctionalization of plane ZnO thin films were performed to obtain a suitable reference system.

The role of blood platelets in inflammation
Caroline Skoglund
Blood platelets have for long been recognized during haemostasis but are now also emerging as key actors during inflammation following implantation of a biomaterial, as well as in diseases with an inflammatory component such as atherosclerosis. A material in contact with blood will adsorb plasma proteins within milliseconds, e.g. immunoglobulin G (IgG) and albumin. Surprisingly, little is known regarding how protein adsorption regulates the platelet activation and their subsequent impact on other cells. Complement activation is known to occur on top of adsorbed plasma proteins on a foreign surface. Platelets express receptors for C1q, one of the first proteins in the classical complement cascade, although the physiological role of these receptors is still unclear.

Chaperonins and “Binding induced unfolding"
Satish Moparthi
We investigated, how Mreb protein (actin homologue) is affected when it is bound inside the cavity of a chaperonins like GroEL/ES (prokaryotic chaperone), TRiC (Eukaryotic chaperone), and also how the mechanism of the chaperonins subsequently affect the bound Mreb substrate. The fluorescence methods, fluorescence anisotropy, and homo-FRET were used to follow the conformational changes in the presence of different chaperonins, to compare the similarities and differences of interaction of prokaryotic and eukaryotic chaperones with substrate protein, and also understand the importance of the evolutionary changes of the structure and function of actin homologue Mreb. The present results shows that the Mreb structure is rearranged and expanded by “binding induced unfolding” mechanism in TRiC, GroEL, GroES chaperonins. We observed that TRiC binds to Mreb substrate as efficiently as GroEL/ES, moreover both GroEL, TRiC prevents the aggregation of Mreb efficiently during folding compare to spontaneous refolding. Interestingly GroES alone, also preventing Mreb aggregation in a little extent besides the structural rearrangements during refolding, shows that GroES not merely acts as a co-chaperone to the GroEL, which was believe from ages.

Immune modulation therapy using autoantigens
Camilla Skoglund
Immune modulation therapy using autoantigens to induce tolerance was tested with the 65kDa isoform of glutamic acid decarboxylase (GAD65), the major autoantigen in type 1 diabetes (T1D). Our objective was to investigate whether subcutaneous injections with 20 µg recombinant alum formulated human GAD65 (DiamydTM) given as a prime and boost injection affected the humoral response regarding GAD65 autoantibody (GADA) epitope pattern in children with T1D.
Samples of patients treated with GAD65 (n=32), or placebo (n=28) were collected at the initial visit and 1, 3, 9, and 15 months after the initial injection. These samples were tested for GADA titer and epitope specificities.
The GADA levels increased significantly in patients treated with GAD65 as compared to the placebo group. However, no longitudinal changes in GADA epitope specificities were observed when comparing placebo and treatment group.
We conclude that the administration of 20 µg GAD65 stimulates autoantibody production without inducing epitope spreading.

September 3, 2008

The effect of GAD65 treatment (Diamyd™) on autoantibody levels in type 1 diabetic (T1D) children
Mikael Chéramy
T1D is an autoimmune disease where the insulin producing pancreatic β-cells are destroyed. In recent years several vaccination trials has been performed with the aim to halt the disease. We investigated the effect of treatment with GAD65 on the antibody response in T1D children and its relation to C-peptide preservation and adverse events.
The effect of GAD65-treatment on the humoral response was antigen specific and induced a long lasting specific B cell memory, without adverse clinical events. C-peptide preservation was best in patients with short T1D duration. Our data constitute novel information in the understanding of the effect of treatment with GAD65 in children with T1D.

Marine biofouling
Tobias Ekblad
I will talk about marine biofouling and my efforts to create non-toxic surface chemistries that inhibit settlement and growth of fouling organisms. I will show a little bit of what has been done in the field previously, and mention different ideas suggesting how the goal might be achieved. The technique that I will discuss is free-radical grafting polymerization, which is a versatile method to create a thin polymer layer on virtually any type of surface. The properties of the grafted polymer can be tuned by choosing a suitable combination of monomers. We have used this method for marine biofouling applications as well as in projects aimed at biosensors or biomaterials.

Drug release systems for local treatment of bone
Fredrik Agholme
A bone fracture can occur due to trauma, cancer, metabolic diseases or osteoporosis. Bone is one of the few tissues that heal by regeneration. This provides a good but slow healing of the fracture. However, when foreign objects (implants) are inserted into bone, like when joints are replaced or complicated fractures are fixated, the healing process can be slow and incomplete. By covering the surface of an implant with matrixes that can release drugs that are beneficiary to bone growth we try to improve the bone healing around it. One matrix that we use is made by grafting fibrinogen layer by layer and then immobilizing the drug onto the fibrinogen by different bonding techniques. This method has successfully been used in earlier studies and we are now developing it further to be used with new types of drugs.

June 11, 2008

Synthesis of functionalized oligosaccharides
Timmy Fyrner
Glycoconjugates exist in nature in forms of glycoproteins, glycolipids and lipopolysaccharides.
The extent and function of these glycoconjugates can vary from a few carbohydrate units up to a polysaccharide containing network, called glycocalyx.
The synthesis of various functionalized oligosaccharides allows for the study of many different macromolecular interactions such as protein adhesion and/or biomolecular recognition. Using orthogonally functionalized oligosaccharides, different model systems such as self-assembly or incorporation into artificial cell membranes are available. Depending on the carbohydrate backbone, linear or a proposed helical conformation could be achieved. Earlier studies with surfaces containing saccharide-functionalities have shown protein resistant properties. In current work, these adhesion results are combined to synthesize protein resistant linker molecules.

How does the immune system react in children with type 1 diabetes, allergy and/or coeliac disease?
Anna Kivling
T-helper (Th) cells of type 1 and type 2 (Th2) are in balance with each other. A shift in this balance can cause different diseases. Type 1 diabetes (T1D) and coeliac disease (CD) have a deviation against Th1, while allergy has a deviation against Th2.
Do children with a Th1-skewed disease have an opposite immune response compared to children with a Th2-skewed disease? What happens when a Th1-associated disease is combined with a Th2-associated disease? Will the immune system be compensated and have a similar immune response as healthy children? Are regulatory T-cells involved in theses diseases? Our aim is to start solving these questions.
Our preliminary results show that when combining a Th1- and Th2-deviated disease the immune system show an increased immune response against a generally stimulating antigen, and that two Th1-deviated diseases in combination show decreased immune response against inhalation- and food allergens.

A CANONICAL CORRELATION APPROACH TO HEART BEAT DETECTION IN TEXTILE ECG MEASUREMENTS
Linda Rattfält
Research in textile sensors has lead to new ways to measure the electrocardiogram (ECG). However, additional disturbances from e.g. muscular noise and high skin-electrode impedances often result in poor signal quality. I will present a simple application of canonical correlation analysis (CCA) on multi channel ECG signals recorded with textile electrodes. Using CCA to solve the blind source separation (BSS) problem, we intend to separate the ECG signal from the various noise sources. The method (CCABSS) was compared to simple averaging of the ECG channels and to the independent component analysis method (ICA). A heart beat detector was used to evaluate the signal quality. Results show that the signal was completely lost while simulating various noise in 33%, 17% and 7% of the cases for averaging, ICA and CCA, respectively.

Haemocompatibility of biomaterials
Lars Faxälv
The haemostasis is a complex and delicate system that keeps the blood in balance between bleeding and thrombosis. This balance can be disrupted by a state of disease or when the blood is exposed to foreign surfaces inside or outside the body. Therefore it is of great importance that the mechanisms behind the interactions between foreign surfaces and blood are studied in order to minimize unnecessary reactions in terms of bleeding or thrombosis when blood and foreign surfaces meet. I have developed methods to test the interactions between coagulation and surface as well as platelets and surface. These methods have been used to test novel biomaterials intended for use as vascular grafts etc.

May 14, 2008

Design and synthesis of chelating and photo labile alkane thiols for surface modification
Lan Bui
A number of NTA- and benzophenone thiol derivates have been synthesized and immobilized onto gold surfaces. We combine the well-known NTA chelating chemistry with the photochemistry of benzophenone to attach biomolecules specific and covalently to a solid surface with well defined orientation of the recognition molecules without lost of functionality. This surface will contribute to the development of large-scale screening systems, micro arrays based on His-tagged labeled binders and antibodies.

Aberrance of soluble CTLA-4 regulation in Type 1 Diabetic children
Anna Rydén
Type 1 diabetes (T1D) is a serious disease associated with a defect regulation of the immune system resulting in destructive insulitis. The onset is abrupt and occurs only after about 80-90% of the insulin secreting β-cells has been destroyed. We hypothesize that dysfunction in T-regulatory (Treg) cells play a keyrole in the destructive autoimmune process. The aim of this study was therefore to investigate mRNA and protein of markers associated with Treg, and especially the spliced form of the cytotoxic lymphocyte antigen-4 (sCTLA-4), in children with T1D, high-risk individuals and healthy children. CTLA-4 is a molecule involved in the down regulation of T-cells activated towards self proteins.
We found an inverse relation between sCTLA-4 transcription and sCTLA-4 protein secretion in children with T1D suggesting a perturbed expression and RNA splicing of the CTLA-4 gene in T1D which might be involved in disease pathogenesis by affecting the regulation of autoimmune responses.

Metal-Enhanced Fluorescence using Metallic Nanoparticles
Erik Martinsson
Metallic nanoparticles and structured metallic surfaces significantly increase the emission of photons from nearby fluorescent molecules. By utilizing this effect called metal-enhanced fluorescence (MEF), a considerable enhancement of the fluorescence intensity can be obtained. The aim of this project is to study the interaction between fluorophores and metal nanoparticles and investigate how different parameters affect the enhancement capacity. The final goal is to incorporate metallic nanoparticles in a fluorescent-based biochip in order to get an amplified fluorescent intensity and thereby increase the sensitivity of the biochip.
The methods primary used in the project are UV-Vis spectroscopy and fluorescence spectroscopy. I will present some results achieved so far and show preliminary results of metal-enhanced fluorescence from small silver nanoparticles.

Platelets play an important role in the coagulation
Karin Vretenbrant Öberg
Platelets play an important role in the coagulation as they are first at the site of a vessel wall injury where they start to adhere and forming a plug to stop the bleeding.
Thrombin is an essential enzyme formed within the coagulation cascade and is also a strong platelet agonist. Both platelets and thrombin is important for the coagulation but in for example atherosclerosis, they are also involved in the formation of thrombosis. The research on platelet thrombin receptors are therefore of great importance to be able to understand and in the future; treating; thrombosis.
We have measured the number of platelet thrombin receptor PAR1 and correlated that to the response achieved when the receptors are activated. Now we want to measure the number of the other platelet thrombin receptor, PAR4, in the same way and possibly see if the number of PAR4 correlates to the number of PAR1.

April 16, 2008

The study of protein structures
Jonas Carlsson
Mutations and misfolding of proteins are the cause to numerous diseases such as cancer, diabetes and alzheimers disease. With the help of two molecular modeling techniques, monte carlo minimization and molecular dynamics, we are trying to investigate how the mutations affect the protein and how the proteins get misfolded.
The proteins we are investigating are the p53 cancer suppressor protein that is involved in about 50% of all cancers, CYP21 and CYP11B1 responsible for certain hormone activation, and IAPP that is involved in type 2 diabetes.
With the help of monte carlo minimization it is possible to get the stability of mutations in a protein. This together with other measurments can be used to evaluate the effect of a mutation. The molecular dynamics can used to study the dynamics of small peptides in different conformations and mutations on short time scales in order to get a better understanding of how a protein can be misfolded.

T-cell polarisation profiles in children with and without celiac disease (CD) and type 1 diabetes (T1D)
Anne Lahdenperä
Celiac disease (CD) and type 1 diabetes (T1D) are immunological diseases associated with inflammatory reactions in the target tissues. The reactions include T cell infiltration, cytokine production, mainly T helper(Th)1 cytokines, and antibody production. Both genetic and environmental factors are of importance in the disease development. The gut-intestinal immune system seems to be an important link. Circulation of autoreactive T cells in the body, due to abberant immunological responses in the gut-intestine, might induce inflammation, autoimmune reactions and b-cell death in pancreas. Little is known about T cell polarisation profiles in peripheral blood mononuclear cells (PBMC) and gut-intestinal biopsies from children with and without CD and T1D.
Th1-Th2-Th3 gene expression profiles were investigated in PBMC and small intestinal biopsies from children with and without CD and T1D, before and after 1 year of treatment with gluten free diet (GFD). RNA was isolated from cryopreserved samples and the gene expression signals were analysed on Real-time RT-PCR arrays, containing 84 Th1-Th2-Th3 associated genes. The gene expression profiles of the samples were analyzed, after housekeeping genes (HKG) normalization. Scatterplots, clustergrams, pathway analysis, K-means diagrams will be used for data analysis and visualisation.
The Real-time RT-PCR arrays are a promising tool for gene expression studies. Analysis of the T cell polarization profiles is on going… Some preliminary results…the Th1-Th2-Th3 gene expression profile in blood does not seem to reflect the gene expression profile in the small intestine. A Th1-polarized response (more Th1 & less Th2) was seen in children with CD, compared to children without CD. The T-cell polarization profile changed, the Th1-skewing was “normalized” (less Th1 & more Th2), after GFD-treatment.

Nonlinear Phonocardiographic Signal Processing
Christer Ahlström
The aim of this presentation is to present signal analysis methods for a computerized cardiac auscultation system. In particular, the work focuses on classification and interpretation of features derived from the phonocardiographic (PCG) signal by using advanced signal processing techniques.
The PCG signal is traditionally analyzed and characterized by morphological properties in the time domain, by spectral properties in the frequency domain or by nonstationary properties in a joint time-frequency domain. Here we will introduce nonlinear analysis techniques based on dynamical systems theory to extract more information from the PCG signal. Especially, Takens' delay embedding theorem will be used to reconstruct the underlying system's state space based on the measured PCG signal. This processing step provides a geometrical interpretation of the dynamics of the signal, whose structure can be utilized for both system characterization and classification as well as for signal processing tasks such as detection and prediction. In this presentation, sample entropy and mutual information will be described as means to assess the severity of aortic stenosis (AS). We will also have a look at very recent approaches based on dimension reduction using manifold learning techniques.

March 19, 2008

In vitro studies of fibril formation of the human Prion Protein
Sophie Nyström
Different model systems are employed to understand the mechanisms protein misfolding diseases as well as finding therapeutics and means of prevention. In our group we recombinantly express and purify proteins for in vitro studies of the pure protein.
The prion protein is responsible for several different diseases in humans including Creutzfeldt-Jakobs disease. The human genome contains a polymorphism in position 129, leading individuals homozygous 129M, homozygous 129V or heterozygous 129M/129V. This in turn gives rise to differences in symptoms of disease as well as disease onset and susceptibility to transmission of disease. I will present some preliminary data from my ongoing project concerning this position and also tell you how we fibrillate proteins and monitor fibril formation.

Monoclonal Antibodies as a tool for protein studies
Sebastian Schultz
A typical feature of type 2 diabetic patients is amyloid deposition, which is made up of beta cell derived islet amyloid polypeptide (IAPP). IAPP derives from the prohormone proIAPP which has to be processed by the prohormone convertases PC2 and PC1/3. Insulin which is produced simultaneously in the beta cells and co-secreted with IAPP likewise derives from a prohormone (proinsulin). Sequential processing of proinsulin to insulin and C-peptide by the same prohormone convertases PC2 and 1/3 occurs either via C-peptide-A-chain (CA) (90%) or via B-chain-C-petide (BC) (10%).
Addition of C-peptide triggers amyloid formation whereas equimolar concentrations of insulin abolish fibril formation. Even in C-peptide presence. The processing state of proIAPP and/or proinsulin and consequential interactions of the different processing intermediates play a major role in amyloid generation.
Monoclonal antibodies are designed to recognize only a single epitope and therefore offer a powerful tool to analyse protein processing and occurrence of different processing populations in vivo. Monoclonal antibodies can also be selected for different techniques, e.g. Western Blot, immunhistochemistry, immuno precipitation etc. Furthermore, monoclonal antibodies can be designed to be species specific, e.g human insulin B-chain vs. mouse insulin B-chain.

CSPT- Using Everyday Objects to make Scientific Measurements
Stephen Macken
The computer screen photo assisted technique (CSPT) is a technique whereby you use a computer screen as a light source and a web camera as a detector. By then inserting different types of sensors in between these two it is possible to make different analyses. My work involves the development of different types of sensing applications using this setup. To begin with I have developed SU-8 microstructures, using photo-lithographic methods, on microscope glass slides to enhance the detection of fluorescent substances. Also due to the fact that I use a web camera it is possible to differentiate between many different fluorescent substances simultaneously. The next phase will be to develop the setup so that it is possible to do SPR measurements using the same micro-pillars.

Hypoxia rescues primitive hematopoietic bone marrow cells from oxidative stress induced cell death and reduces their proliferative rate
Pernilla M Eliasson
Hematopoiesis, blood formation, is regulated by a dynamic balance between stem cell self-renewal and differentiation to mature blood cells. Oxidative stress reduces the life span of the hematopoietic stem cell (HSC). A family of proteins called, FOXO proteins, is thought to play an important role in protection of HSCs from oxidative stress and maintain the hematopoietic stem cell pool. Our studies show that hypoxia upregulates Foxo and rescue cells from oxidative stress-induced cell death. We believe that the hypoxic areas in the bone marrow provide HSCs with an appropriate environment for optimal survival and maintenance by inhibiting cell cycle, and that the metabolic resistance of HSCs to ROS depends on hypoxia and the transcription factor HIF-1α.

Feb 20, 2008

MDRs, HMMs and computational grids
Joel Hedlund
Medium-chain Dehydrogenases/Reductases (MDR) constitute a wide-spread superfamily of proteins whose roles are centered on shuffling hydrogens on and off substrates. Knowledge begets more knowledge, but to be able to take full advantage of the vast amount of prior knowledge available for these proteins, and to guide future research efforts, it is necessary to divide the family into functional and evolutionary subgroups. I have done this using bioinformatic methods, primarily profile Hidden Markov Models (HMM). For the calculations, I have been able to use the Nordic distributed computational facility Nordugrid. Before I started investigating the MDRs, only eight member families were characterized. Now, it is clear that the number of disjoint member families is at least 19.

The effect of HLA genotype or CTLA-4 polymorphism on cytokine response in healthy individuals
Jenny Walldén
Type 1 Diabetes (T1D) is considered as a T cell mediated autoimmune disease, affected by HLA class II alleles and CTLA-4 polymorphisms. HLA class II is involved in antigen presentation and CTLA-4 have the purpose of down-regulating an activated T cell. This suggests that both molecules are important and can contribute to pathogenesis in T1D.
We have tested whether these T1D related gene polymorphisms are associated with an aberrant regulation of cytokine response, which could contribute to the development of T1D.
We have done this after antigen stimulation in non-diabetic children and measured cytokine response with ELISA-technique.
We found that the cytokine responses to studied antigens were weakly modified by HLA class II risk haplotypes but all studied CTLA-4 polymorphisms were associated with enhanced IFN-, suggesting impaired function of these CTLA-4 polymorphisms.

Molecular gradients on bipolar electrodes
Christian Ulrich
If a conducting surface is placed in an electric field in a solution, electrochemical reactions can (under certain conditions) be induced on both ends of the non-contacted surface. The surface will then become a bipolar electrode, i.e. an electrode acting as both an anode and cathode. Since the potential difference between a point on the bipolar electrode and the solution will vary laterally along the surface, the rates of the reactions will also vary laterally. If the result of such reactions on either end of the bipolar electrode is the adsorption or desorption of a specific molecule, a gradient of that molecule can be created on the surface. One important example is monolayers of mixed ω-functionalized alkane thiols on gold, offering excellent control of surface properties. Other interesting applications include gradients of metals and electrodeposited polymers.

Health and Nutritionstatus among elderly people
Yvonne Johansson
Background: In the middle of 90th about 6 % among home living elderly people were malnourished and between 8%-38% when admission to hospital or a nursing home. Quality of life can be influenced negatively with an increased dependence from home care service and next of kin.
The aim of the project was to prospectively follow the development of nutritionstatus related to health among elderly people, with regard to demographical, social, medical and functional factors. Another aim was to find predictors for malnutrition.
Method: At baseline, 579 individuals (49%) participate, 75 and 80 years old. They were followed up once a year, 3-5 visits including baseline with different questionnaires, biochemical tests, anthropometrical and physical measurements. Multiple logistic regression analyses were used to find predictors comparing individuals at new risk and at no risk of malnutrition.
Results: An increased risk of developing malnutrition were found in following factors: increased age, decreased self-perceived health and increased number of symptoms of depression.

Jan 23, 2008

Biophysical Characterizations of Proteins using Circular Dichroism Spectroscopy
Cecilia Andrésen, Molecular Biotechnology
Because of the relation between several diseases and proteins it is important to learn more about the mechanisms behind protein structure and/or function. For structural determination of proteins a variety of techniques can be used such as Nuclear Magnetic Resonance (NMR), Fluorescence Spectroscopy and Circular Dichroism Spectroscopy.
To evaluate resistance mechanisms in MexR, a protein involved in antibiotic resistance in Pseudomonas Aeruginosa, on a molecular level, we have studied recombinant MexR wt- and mutant proteins with biophysical and biochemical methods. In particular, we choose to study mutations R21W, R70W, G58E and L13M in the MexR protein. MexR is a dimeric repressor protein that, when binding DNA, represses the efflux system, responsible for transportation of toxic molecules, in P. Aeruginosa. Wild-type protein has high affinity towards DNA but all mutants have no or poor DNA-binding ability.
Circular Dichroism Spectroscopy (CD) is a commonly used method for secondary and tertiary structure evaluations of proteins.
When analyzing the wild-type MexR and mutants by CD we could see that two of the mutants had reduced stability compared to the wild-type due to loss of electrostatic interactions within the protein dimer. Surprisingly however, the other two mutants showed much higher stability probably because of stronger interaction in the dimerization domain.
The results probably correspond with the general assumption that wt-MexR only binds to DNA in a specific conformation and that any change in the stability as a result of mutations gives none or poorer affinity leading to increased antibiotic resistance in the host organism P. Aeruginosa.

HLA-class II tetramers in flow cytometry
Mikael Pihl
In trials of immunotherapy with the Type 1 Diabetes (T1D) associated autoantigen Glutamic Acid Decarboxylase (GAD) 65 (Diamyd), we have shown an altered response to GAD65 in treated children with T1D. The treatment is thought to act by inducing immunological tolerance toward GAD65. Treatment with GAD65 has been shown to help maintain insulin secretion in treated individuals. We hypothesize that treatment with GAD65 induces antigen-specific regulatory T cells in children with T1D. To prove this hypothesis we intend to make use of GAD65 HLA class II tetramers. These phycoerythrin-conjugated tetramers will stain the scarce T-cells induced by the treatment with a T-cell receptor recognizing different fragments of GAD65. We aim to obtain T-cell clones specific for GAD65 using tetramers and single cell sorting flow cytometry. The clones’ response to GAD65-stimulation will be analyzed in terms of proliferation, cytokine secretion, gene transcription and in mixed lymphocyte cultures.

Folding induced assembly of nanoparticles and peptide fibers
Daniel Aili
Self-assembly has emerged as a promising and interesting alternative for nano-fabrication. The building blocks can range from simple organic molecules to “large” inorganic nanoparticles, and by choosing building blocks with suitable properties, it is possible to exert a fine control on the processes of self-assembly in order to obtain complex architectures and functional nanostructures. My project is focused on how synthetic de novo designed polypeptides can be utilized to control the assembly of gold nanoparticles and for self-assembly of fibrous materials and peptide based nanostructures. Gold nanoparticles have many interesting physical properties and are excellent building blocks in bottom-up nanofabrication due to the ease of production and convenient functionalization. The versatility of the synthetic polypeptides used here, in terms of folding and dimerization properties, allows for control of self-assembly of nanoparticles and peptides into structures and materials with applications in e.g. biosensing and plasmonics.

Health-related quality of life after osteoporostic fractures
Inger Hallberg
Osteoporosis is a skeletal disorder characterised by compromised bone strength predisposing a person to an increased risk of fracture. The global burden of osteoporosis includes considerable amounts of fractures, morbidity, mortality and expenses, mainly due to vertebral, hip and forearm fractures.
My research mostly concerns health-related quality of life (HRQOL) after osteoporotic fractures. The general aim of the present thesis is to describe and analyse bone mineral density (BMD) and HRQOL in women after osteoporotic fractures and follow-up. The methods in the project are questionnaires about risk factors, HRQOL-instruments (general and disease specific), BMD, X-ray of the spine, physical function tests and interviews with qualitative approach. In summary, osteoporosis is still a neglected health problem. Hip, vertebral or multiple fractures have a considerable larger impact on BMD and HRQOL than forearm and humerus fractures.

Nov 28

Mutations in the MexR gene are involved in multidrug resistance in bacteria
Anngelica Jarl
Multidrug resistant bacteria is of growing concern today, causing difficulties when treating bacterial infections. The bacteria Pseudomonas aeruginosa is one of the effected species and many patients with Cystic fibrosis die of untreatable Pseudomonas aeruginosa infections. MexR is the repressor for the MexAB-OprM Operon, which encodes a multidrug efflux system in Pseudomonas aeruginosa. Mutations in the MexR gene leads to increased resistance to antibiotics due to over expression of this efflux system. Studies of MexR- DNA interactions and how mutations in the MexR gene affect these interactions can lead to increased understanding about antibiotic resistance and the mechanism behind this phenomenon.

LPS induced cell signalling in relation to TLR4 polymorphism
Anna Lundberg
Lipopolysaccharide (LPS) is a part of the gram-negative bacterial cellwall. LPS is recognised by antigen presenting cells through the Toll-like receptor 4. The activation of TLR4 leads to intracellular signalling resulting in various cellular immune responses such as upregulation of co-stimulatory factors or secretion of cytokines. We have studied a polymorphism Asp299Gly in the gene coding for TRL4. We have investigated LPS signalling pathways in PBMC from individuals heterozygote for the polymorphism and in individuals without the polymorphism. This was done by analysing phosphorylation of several intracellular signalling proteins in PBMCs after LPS stimulation. Bead-based phosphoprotein assays were used for the detection of protein phosphorylation and were analysed with Luminex technique. We found that the protein IKBα was less phosphorylated after LPS stimulation in heterozygotes for the polymorphism compared to individuals with the wildtype.

Structural and intermolecular force study of polypeptides using atomic force microscopy
Feng-I Tai
During the last decade, atomic force microscopy (AFM) applications have been extensively developed for various studies in nano-scale. The elegant interaction between the tip and sample surface offers a non-destructive analysis process; one can obtain information of morphology, magnetic/electric/chemical potential, force interactions, etc. with atomic-scaled resolution. while AFM-associated analytical methods introduced into biological research, it is possible to visualize the structures of proteins and DNAs, and also study the intermolecular interactions, for instance, the folding force of proteins. More recently, there are publications which used fluidic cells in experimental, where allowed the proteins stay in liquid surroundings with controlled temperatures, and could be close to their natural free-standing states.
Thus, the polypeptides: JR2EC and JR2KC are examined in two perspectives. One is to monitor the structures of the hetrodimerized fibers formed by JR2EC and JR2KC at pH7.0 both in dry and liquid states. The other approach is to quantify the folding force of JR2EC in liquid state at pH4.5. By studying both in parallel, it is expected to provide essential knowledge including structural formation and folding dynamics of polypeptides.

Oct 31

Design of novel materials for bioassay applications based on synthetic polypeptides and gold nanoparticles
Irina Nesterenko
Synthetic polypeptides are highly interesting biomolecules for advanced surface design since they combine chemical and functional diversity with robustness and can be synthesized at moderate cost and effort by well-established methods. There are numerous strategies for covalent immobilization of polypeptides onto modified metal and other surfaces. The process of peptide fibres formation was also investigated. JR2EC and JR2KC have a Cys residue in the loop region that can be oxidized to link two monomers together into fibre-forming units, JR2EC2 and JR2KC2. When mixing the fibre-forming units, they rapidly assemble into fibres. Formation of fibres was monitored by DLS and TEM. According to DLS, the average size of fibres was about 100 nm. Aggregation of gold nanoparticles with immobilized JR2EC was observed by adding JR2KC2 at different concentrations. Presumably, aggregation was induced by the formation of peptide fibres. This was confirmed in a parallel experiment with the negative control D- alanine containing peptide JR2ECref, which is unable to form the helices necessary for fibre formation at any circumstances. In this case, aggregation was not observed. These experiments were evaluated by TEM. The average distances between gold particles were 5 nm and 20 nm for JR2EC and JR2ECref, respectively.

Insulin-like growth factor (IGF-I) and insulin in vascular smooth muscle and endothelial cells – from a diabetes perspective
Git Johansson
In patients with diabetes mellitus, micro- and macrovascular disease is the major cause of morbidity and mortality. The diabetic vascular disorders are associated with both metabolic and hormonal imbalances, and involves several kinds of cells, including vascular endothelial and smooth muscle cells. When discussing insulin action, one usually focuses on the more classically known insulin target tissues, such as sceletal muscle, liver and adipose tissue. But the fact is, several studies have shown insulin to be a important hemodynamic factor in blood vessels and has been described to have vasodilator actions such as stimulation of nitric oxide production. Also, in diabetes, not only the insulin concentration is altered, but the IGF-I system is disturbed as well. Insulin and IGF-I are similar hormones sharing homology to a large extent, and because of the similarity insulin and IGF-I can cross react with each others receptors although with a 100-1000 fold lower affinity than to their own receptors. In several in vitro studies showing insulin action, insulin has been used in supraphysiological concentrations. Hence, one could speculate that these studies which have shown substantial effects of insulin are perhaps not propagated by the insulin receptor, but instead due to activation of IGF-IRs and possibly also hybridRs, which are misinterpreted as insulin receptor effects.
Our work has been mainly focused on characterizing, at the receptor level, which action is attributable to which hormone acting through which receptor.

Field effect gas sensors and SLPT
Roger Klingvall
Field effect gas sensors are cost effective gas sensors that can be used in a variety of applications. The gas sensing properties are mainly dependent on the catalytic metal layer that can consist of various metals such as Pt, Pd or Ir. Our research focuses mainly on the gas sensing mechanisms and on important parameters in gas sensing, e.g. sensitivity, selectivity and stability. We are also developing methods and instruments that can help us to gain an increased knowledge of the sensors. One technique that we use is the Scanning Light Pulse Technique, SLPT, which can be used to investigate the gas sensing properties of various gas sensing surfaces. One example of a successful project is the UV-SLPT project where we have extended the use of SLPT to include wide band gap type of substrates, thus enabling the use of the technology for evaluation of high-temperature sensor surfaces. Another example is the combinatorial approach to field effect gas sensor research and development, where we used SLPT to evaluate a 2D array that had a sensor surface consisting of more than 100 different thickness combinations as the gas sensitive layer. Hopefully our research can lead not only to better understanding of field effect gas sensors but also to improved sensors that are suitable for various applications such as leak detection or process control.

Oct 3

Chemical Synthesis and Characterization of Core-shell Au-SiO2 Nanoparticles
Fatima Monteverde
Gold nanoparticles are versatile materials in biomedical applications, one of them being the use of the surface plasmons of the particles for enhancing the fluorescence properties of biosensing probes. Thus it is important to investigate new ways to produce gold nanoparticles with different sizes, shapes and coatings. Syntheses were performed by wet chemistry using tetrachloroauric acid, sodium citrate, sodium borohydride (NaBH4), tetraethylorthosilicate (TEOS) and other organic solvents. Molar ratios of gold, TEOS, sodium citrate, mercaptoacetic acid and NaBH4 (reducing and capping agent) were varied for the purpose of obtaining stable and uniform particles. Prepared materials were characterised with transmission electron microscopy (TEM) and UV-Visible spectroscopy. Increasing the concentration of TEOS in the reaction mixture appears to promote pure silica nanoparticles as well as of core-shell particles while the opposite hinders silica coating of gold nanoparticles instead of producing thinner shells as was intended.
Future work will focus on controlling size and increasing stability and uniformity of gold-silica core-shell particles by varying experimental parameters such as temperature, reaction time and reagent concentrations.

Does cryopreservation influence the expression of different markers in peripheral blood mononuclear cells?
Stina Axelsson
Functional analysis of peripheral blood mononuclear cells (PBMC) is an important tool to study immunological features of several diseases, and further to evaluate effects of clinical interventions. In clinical trials, PBMC are frequently studied over time, with blood samples drawn at several time points. Hence cryopreserved cells are often used in order to minimize inter- assay variations. To evaluate the effect of cryopreservation on different immunological markers we used multiplex fluorochrome technique (Luminex) to detect cytokine and chemokine secretion in both freshly handled and cryopreserved/thawed PBMC, cultured for 72 hours with or without stimuli (antigen/mitogen). The PBMC samples were collected from 22 type 1 diabetic children. We find that cryopreserved PBMC are, equally to fresh cells, able to respond to antigen and mitogen stimulation. Thus, cryopreserved PBMC are well suited to establish expression of different immunological markers, but should not be compared with results from freshly handled cells.

Detection of autoantibodies related to type 1 diabetes using Biacore and
A new concept for diabetes control based on a lectin panel
Jenny Carlsson
Type 1 diabetes is an autoimmune disease characterized by destruction of beta-cells in the pancreas leading to the need for insulin treatment therapy. Autoantibodies to beta-cell proteins are often generated and may be used for predicting disease in at-risk populations. The goal is to develop a sensitive and specific method based on the Biacore technique for detection of autoantibodies in serum samples from individuals with high risk of developing type 1 diabetes. Glycosylation changes of serum proteins in type 1 diabetic patients have been investigated based on the interaction of the saccharide moiety of serum proteins with different lectins (proteins that bind carbohydrates specifically). MVDA techniques were used to analyze data. The method is intended to be used in order to get a further understanding of the characteristics and development of the disease.

Cardiovascular effects of nicotine and tobacco use
Liza Ljungberg
Cardiovascular diseases (CVD) are the major cause of death in Western countries. Smoking tobacco is strongly associated with increased risk for CVD, but the cellular mechanism behind this association is unknown. Angiotensin-converting enzyme (ACE) and nitric oxide formation in the vascular endothelium are two important parameters for maintaining normal vascular tone. An increase in ACE expression and a decrease in nitric oxide formation can be seen in atherosclerotic plaque and appears to be important factors in the development of CVD. In this study, we examine how use of nicotine and different tobacco products affect ACE activity/expression and nitric oxide formation. The results showed that nicotine and liver metabolites of nicotine can increase ACE activity, but do not affect the nitric oxide formation. The findings from this study could explain one cellular mechanism behind increased risk of cardiovascular diseases in smokers.

Sep 5

Modeling Amyloid Diseases in Drosophila Melanogaster
Ina Berg
Amyloid diseases arise from the misfolding and aggregation of a wide variety of non-homologous extracellular proteins. The Drosophila model system is increasingly being used for modelling human degenerative diseases. To develop Drosophila models of Familial Amyloidotic Polyneuropathy (FAP) and Alzheimer’s disease we generated transgenic Drosophila expressing single gene products of the human amyloidgenetic proteins transthyretin (TTR) and A1-42, under control of the Gal4/UAS system. We find that expression of TTR and A1-42 in the nervous system results in a reduced lifespan when compared to normal flies. The effect of the TTR familial mutation V30M resulted in a more significant decrease in lifespan when compared to expression of the wild type protein. Expression of the same proteins in the eye led to progressive tissue degeneration that increases over time.

Separation of different cellular fractions using differential and density gradient centrifugation
Siri Fagerholm
In the cell biology field, it is of great interest to study subcellular processes such as the localization and phosphorylation state of proteins and their intracellular movement between different compartments of the cell. To be able to seperate different fractions of the cell, e.g. the plasma membrane from the nucleous, then becomes a prerequisite. Isolation of cellular fractions, in addition to the preparation of biological “particles” (proteins, organelles etc.), can effectively and rather rapidly be performed using centrifugation techniques.
Using different sequential centrifugation steps we have isolated three cellular fractions from primary rat adipocytes; a plasma membrane fraction, an endosomal fraction and a microsomal fraction. Both differential centrifugation and sucrose density gradient centrifugation were used to isolate the fractions depending on the particles respective properties in the solution.

Fabrication of Functional Protein Nanoarrays by Dip-Pen Nanolithgraphy
Tomas Rakickas
Dip-Pen Nanolithography (DPN) is quite new and promising technique for making organic and inorganic structures on solid surfaces. The advantage of this technique is possibility to make patterns of different compounds without intermediate steps. This opens the possibility to position the proteins to desired places by creation of well defined areas with terminal affinity tags. In our work DPN was used to make nanostructures of multivalent chelators terminated thiol compounds on gold surface. Later on these structures were used to immobilize the oligohistidine-tagged proteins. The functionality of proteins on nanostructures was tested by fluorescent microscopy and imaging surface plasmon resonance.

May 30

TiO2 sol-gel coating as a drug release matrix for orthopaedic implantations
Paula Linderbäck
Hip replacement survival is as low as 75 % in young patients after 10 years, and the failure rate for knee implants varies between 7 and 23 %. Good fixation of an orthopaedic implant to the surrounding tissue is thus of great importance for a successful treatment. Coatings, such as synthetic hydroxyapatite (HA) and sol-gel derived TiO2 coatings produced by different methods are then commonly used. Our group has previously shown positive results with implant bound bisphosphonate in rat tibia. However, in these studies the amount of released bisphosphonate was hard to control. The objective of the present study is to evaluate new method for bounding bisphosphonates to sol-gel derived TiO2 coatings.

LiTHe Doqtor, nätverk för doktorander
Johanna Wallén
(The presentation will be given in English. (The abstract below is taken from the home page.))
LiTHe Doqtor är ett nätverk av kvinnliga doktorander på Tekniska högskolan. Syftet med nätverket är flera: behov av kvinnliga kollegor, kunskapsbildning t.ex. föreläsningar, kontakt över institutionsgränser, att ge kvinnliga forskarstuderande ökad synlighet som grupp.
De flesta träffarna har hittills skett i form av ett gemensamt torsdagsfika. Temana för diskussionerna har tagit upp många ämnen: Ny doktorand, personligt nätverk, lärarrollen, undervisning för kvinnliga studenter, doktorandkurser, forskare & kvinna, ledarskap, handledare & forskningsfrågor, HSV:s utvärdering om teknikvetenskaplig forskarutbildning. Seminarier eller föredrag har tagit upp t.ex. kvinnliga pionjärer inom forskning, stresshantering och konsten att göra sin röst hörd som kvinna i den akademiska miljön (retorik).
Nätverket stöds av tekniska fakultetens forskarutbildningsnämnd.
Är du intresserad av nätverket LiTHe Doqtor?
Anmäl dig till en e-postlista genom att skicka ett mejl till Eva Lindblad, forskarutbildningsnämndens sekreterare.
Vill du veta mera om nätverket? Har du förslag på aktiviteter?
Kontakta Linnéa Rosenbaum, doktorand på ISY, Institutionen för systemteknik, eller Eva (se ovan)

May 16

Effects of hypoxia on maintenance of hematopoietic stem cells
Pernilla Eliasson
My preliminary project is to investigate the effects of hypoxia on hematopoietic stem cells (HSC). HSCs are located in special stem cell niches within the bone marrow in close contact with osteblasts and supportive stromal cells. This environment is thought to be hypoxic, low oxygen level. The aim of my work is to investigate how hypoxia affects maintenance, growth and self-renewal of HSCs. An important regulator for hypoxia-mediated effects on cell function is the transcription factor hypoxia-inducible factor 1 alpha (HIF-1α).
During the first months of this year we have constructed a retroviral vector with a mutated form of HIF-1α in which two prolins P402 and P564 in the oxygen dependent degradation domain (ODDD) have been changed to P402A/P564A, thereby disabling degradation of the protein when cells are exposed to oxygen.
We have also set up real time PCR assays for measuring gene expression of some key molecules associated with the HSC niche, for instance Hes-1, Tie-2, FoxO proteins, GLUT-1, and VEGF.
Recent studies in our group have shown that HSCs cultured in hypoxia survive better when exposed to reactive oxidative species (ROS).
Some questions we are trying to find answers for are: Is HIF-1α involved in the maintenance of the stem cells? How does hypoxia rescue HSCs from cell death when exposed to oxidative stress?

Protein adsorption on Carbon Nitrides and multilayer growth using in situ ellipsometry
Torun Berlind
There has been an extensive research on carbon nitrides and carbon based materials for tribological and electrical applications the last decades, but far fewer studies have been published regarding carbon nitrides and biointeractions. The objective with this study is to investigate the interaction of carbon and carbon nitride surfaces with proteins. The films were exposed to human serum albumin (HSA) and the adsorption was monitored in situ using dynamic ellipsometry and the protein layer index was described with a Cauchy model. In addition the films were incubated in plasma followed by anti-fibrinogen, anti-HMWK or anti-C3c hopefully giving information of the materials response to complement and contact activation. If time the first preliminary results from multilayer growth of fibrinogen will be presented as well.

May 2

A short introduction to Control of Cell Adhesion Using Conducting Polymers
Maria Bolin*; Kalle Svennersten*, Emeilien Saindon, Agneta Richter-Dahlfors and Magnus Berggren
What could be done if it would be possible to control the adhesion of cells to different surfaces? This is one of the inspiring questions that we try to answer with this on going project. Surfaces are developed to construct surface switches, based on conducting polymers that enable electronic control of the surface tension chemistry properties to be utilized to electronically control the adhesion, growth and release of living cells.
Petri dishes of polystyrene and plastic foils are coated with PEDOT. The polymer surface is then divided into two adjacent electrodes using over-oxidation, or sctatching, as the patterning method. This enables to exclusively reduce of one PEDOT-electrode while the other is oxidized. Then we study how cells adhere on the two electrodes while biased with an electric signal. The cells show a remarkable trend to adhere to the reduced side while hardly any cells were found on the oxidized electrode.

Segmental control of cell fate
Daniel Karlsson
In the developing Drosophila ventral nerve cord, about 100 neurons
express the LIM-HD gene apterous (ap). An easily distinguishable
subclass of these, the Ap cluster neurons, consists of clusters of 4
cells located specifically in the lateral thoracic segments. In the Ap
cluster, one cell, the Tv neuron, expresses the neuropeptide gene
FMRFa, while another, the Tvb, expresses the Nplp1 neuropeptide gene.
Studies from several labs have led to the elucidation of a highly
cell-specific regulatory cascade acting to ensure proper Ap cluster
specification, and to activate the cell-specific expression of FMRFa
and Nplp1.
Recent studies from our lab have determined that the origin of these
Ap cluster cells is the neuroblast 5-6 (NB 5-6). With this information
at hand, we are now asking the important question of which upstream
regulatory events act to ensure that only the thoracic NB 5-6, and no
other nerve cord neuroblast, activates the complex genetic cascade
leading to the formation of the Ap cluster.

April 18

Optical Biosensing with imaging SPR ­ Applications in Surface Patterning and Protein Micro-arrays
Olof Andersson
Using imaging surface plasmon resonance (SPR) and tailored surface chemistry it is possible to monitor multiple biochemical interactions on a surface in real-time. In this presentation I will give some examples of how imaging SPR can be used to study surface patterning and antigen-antibody interactions. More specifically, I will show two sensor systems; First, we employ a 3D surface consisting of circular spots of a functionalized polyethyleneglycol (PEG) matrix with different thicknesses as a template for protein arrays. Secondly, we have worked with a flat surface, patterned with four synthetic peptides with different affinities for a particular FAB.

Cavities in coiled-coil polypeptides as nanoreactor for the production of gold nanoparticles
Jagoba Iturri
Tetrabrachion, a protein complex found on the surface layer of the hyperthermophilic archaebacterium Staphylothermus Marinus, exhibits an extreme stability to heat denaturation far beyond its temperature of growth. Tetrabrachion protein complex consists of four identical subunits that form an α-helical coiled coil stalk anchored to the cell membrane at its C-terminal end and branches into four arms at its N-terminus.
A parallel right-handed coiled coil (RHCC) structure was proposed for the C-terminal end. This structure, in marked contrast to left-handed coiled coil tetramers, reveals large cavities placed at regular distances and connected to a continuous central channel.
The project regards the use of these well-defined cavities in the structure of the protein for the production of metal nanoparticles, as well as to verify both electrical and optical properties of such systems in order to exploit them for bio and chemical sensor applications.

April 4

Autoantibodies in sera from patients with Sjögrens syndrome can
penetrate living cells and inhibit the E3 ligase activity of Ro52.

Janosch Hennig
Ubiquitination of proteins is the most important quality control mechanism in living organism. Proteins which are targeted will be labeled with a chain of the small 8 kDa protein ubiquitin and recognized by the proteasome where it will be digested. Several proteins are involved in the ubiquitination pathway. The E3 ligase, which serves as substrate recognition module is one of them. Many E3 ligases have the ability to polyubiquitinate themselves as a self-destruct mechanism. One of them is Ro52/TRIM21, which is an autoantigen in autoimmune diseases such as Sjögren syndrome, lupus or congenital heart block.
E found that Ro52 autoantibodies specific for the RING domain inhibited Ro52 mediated polyubiquitination, but that anti-Ro52 antibodies specific for the leucine zipper or B30.2 domain of Ro52 did not inhibit Ro52 E3 ligase activity. Thus in patients with Sjögren syndrome and SLE, anti-Ro52 antibodies recognizing the RING domain may be actively involved in the pathogenesis by inhibiting Ro52 mediated polyubiquitination

March 21

Stretching capacity of GroEL is a debated characteristic by the chaperone from ages
Satish Moparthi
Abstract: The main idea behind this project is study the interactions between GroEL chaperone system and substrate proteins. We studied chaperone recognition of partially folded proteins by the use of site-directed labelling. Specifically ”analysis of interactions between the GroEL alone, GroES alone, GroEL/ES alone and GroEL/ES chaperone with fluorescence labelled Human Carbonic Anhydrase II mutants H64C, N67C, L79C, H10C, N253C and wild type” by using stopped flow fluorescence and steady state FRET measurements. We observed that GroEL alone is stretching the protein substrate as an early event in the protein folding process, when compared to spontaneous folding. GroEL and GroES interacts to the substrate protein independently as an early protein folding event. Interestingly, stretching or compressing of the protein substrate interaction with GroES depends up on the site of the protein substrate.

Molecular modelling of disease causing mutations
Jonas Carlsson
During my PhD I have had a few cooperations with KI and KS in Stockholm. During my presentation I will tell you about my part in two of these. The first one are already published work about mutational studies on a protein called CYP21, which then sort of evolved into the second where I am doing predictions on mutations in p53. All these studies are performed in the computer but large part of the data used are from "wetlab" experiments.

February 21

BioNanoIT! - A story about a research project from application to thesis
defence
Anna Herland
In the spring 2002 Olle Inganäs wrote an application to Vinnova's program BioNanoIT called "Biomolecule assisted assembly of molecular electronic systems". One year later he was able to hire two new PhD students, Anna Herland and Per Björk. Now 2007 both Anna and Per will defend their thesises.
In this presentation I want to tell you how me and Per have worked with our projects, comparing the application with the real lab work and publications.

A short introduction to the Control of Cell Adhesion Using Conducting Polymers
Maria Bolin*; Kalle Svennersten*, Emeilien Saindon, Agneta Richter-Dahlfors and Magnus Berggren
What could be done if it would be possible to control the adhesion of cells to different surfaces? This is one of the inspiring questions that we try to answer with this on going project. Surfaces are developed to construct surface switches, based on conducting polymers that enable electronic control of the surface tension chemistry properties to be utilized to electronically control the adhesion, growth and release of living cells.
Petri dishes of polystyrene and plastic foils are coated with PEDOT. The polymer surface is then divided into two adjacent electrodes using over-oxidation, or sctatching, as the patterning method. This enables to exclusively reduce of one PEDOT-electrode while the other is oxidized. Then we study how cells adhere on the two electrodes while biased with an electric signal. The cells show a remarkable trend to adhere to the reduced side while hardly any cells were found on the oxidized electrode.

Page responsible: jonathan.rakar@liu.se
Last updated: Mon May 13 10:55:13 CEST 2013