The CelCom research group focuses on studies of metabolism in prokaryotic and eukaryotic cells. Special emphasis has been on the sugar metabolism in lactobacteriae in neutrophils (white blood cells) and in the yeast Saccharomyces cerevisiae. The research encompasses synthesis of small nanobiosensors to be inserted into cells to perform time-resolved measurements of metabolites in intact cells and mathematical modelling of cell metabolism. These two approaches work in synergy: the models are based on what we do know about metabolism and what has been measured; the nanobiosensors are being developed to measure what we do not know in cell metabolism, but what can be predicted from the models. Recently we have started a new project in the Lundbeck NanoCAN consortium to synthesize biodegradable nanoparticles to be used for targeting and delivery of drugs to cancer stem cells.
Head of research: associate professor Lars Folke Olsen
Researchers and research group:
Laboratory technician Anita Lunding
Lars Folke Olsen is involved in the following national research networks:
Lundbeck NanoCAN: www.nanocan.org
The European Science Foundation collaborative network : http://funcdyn.org
and the EU 7FP Initial Training Network (ITN): www.isolate.se
Our specialties in research:
1. Development of aptamer-based optical nanosensors
Aptamers are small nucleotide sequences with ligand-binding properties. An aptamer with specificity for a particular target molecule and with a predefined affinity can be selected in vitro. Furthermore the aptamers can be chemically modified in various ways after they have been selected to be transformed into signalling molecules. An example is shown below for a socalled ATP aptamer switch probe (ASP). The aptamer (44 nucleotides) has been modified with a fluorophophore at the 5’-end and a complementary sequence to the 3’-end which is linked to the aptamer through a polyethylene glycol linker. A quencher has been attached to the complementary sequence. In the absence of the target (ATP) the complementary sequence is bound to the aptamer and the quencher quenches the fluorescence from the fluorophore. However, when the aptamer binds ATP it undergoes an intramolecular rearrangement so the quencher is separated from the fluorophore again leading to a a signal from the ASP, the size of which depends on the amount of ATP present.
ATP binding aptamer switch probe (ASP)
In order to be able to use the aptamer switch probe in a cellular environment we have encapsulated it in a polyacrylamide particle with a diameter of approximately 30 nm.
ASP encapsulated in a polyacrylamide particle
The particles are porous so small molecules can easily diffuse in and out of the particle, while larger molecules are prevented from entering. Also the aptamer switch probe is retained within the particle matrix. The sensors are biocompatible, i.e. the material does not interfere with any process within the cell. Furthermore, the amount of metabolite bound to the sensors is negligible compared to to amount present in the cell. We have used these sensors to measure glucose and ATP in various eukaryotic cells.
2. Mathematical modelling of cell metabolism
Mathematical models are extremely useful when analyzing experimental model. In the CelCom group mathematical models are used to explain the effect of perturbations of cell metabolism and to predict the effect of drugs on metabolism and cell signalling and ultimately on cell viability.
3. Synthesis of biodegradable solid nanospheres targeted to cancer stem cells
The unifying feature of nano-scaled drug delivery devices (NDDDs) in cancer treatment is that they display an improved passive targeting at the site of sufficiently large and vascularized tumors, referred to as the enhanced permeability and retention (EPR) effect. Due to enhanced vascularization and endothelial permeability on the one hand and impaired lymphatic drainage of macromolecules on the other hand, nano-scaled particles selectively enrich in solid cancer tissues. Numerous approaches include utilization of other physical properties of NDDDs, such as magnetic enrichment or hyperthermia-induced drug release from heat-sensitive liposomes. Heat-sensitive liposomes are presently in phase III clinical trials for cancer treatment. Cancer stem cells are suspected to be able to hide at sites distinct from the primary tumor, and one of the most pertinent problems are undetectable metastases that may already be present at the time point of the first diagnosis. Thus, NDDDs suitable for systemic administration will represent the focus of the core activities in NanoCAN. In general, artificial liposomes and biodegradable solid nanospheres are the best studied NDDDs for systemic applications. Comprehensive experiences have accumulated in the field of liposomal NDDDs in cancer treatment. Enhanced selectivity can be achieved by using liposomal NDDDs that are specifically degraded in the vicinity of cancer cells due to high levels of cancer cell-derived Phospholipase A2. In fact, the liposomal shell itself can be converted to a cytotoxic agent via this mechanism.
Solid nanospheres are commonly used as porous carriers for drug encapsulation . Drug release can be achieved through either biodegradation or swelling and consecutive drug diffusion out of the nanoparticles. Poly(D,L-lactic-co-glycolic acid) (PLGA), and polylactide acid (PLA) have, for example, been used for drug encapsulation for about two decades and are approved by the FDA. In the CelCom group we aim to synthesize such solid nanospheres and coat them with aptamers selected against cancer stem cells
Selected publications from the latest 5 years
Poulsen, A.K., Petersen, M.Ø. and Olsen L.F. (2007) Single-cell studies and simulation of cell-cell interactions using oscillating glycolysis in yeast cells. Biophys. Chem. 125, 275-280.
Poulsen, A.K. Scharff-Poulsen, A.M. and Olsen, L.F. (2007) Horseradish peroxidase embedded in polyacrylamide nanoparticles enables optical detection of reactive oxygen species. Analytical Biochemistry 366, 29-36.
Andersen, A.Z., Poulsen, A.K., Brasen, J.C. and Olsen, L.F. (2007) On-line measurements of oscillating mitochondrial membrane potential in glucose fermenting saccharomyces cerevisiae. Yeast 24, 731-739.
Poulsen, A.K., Andersen, A.Z., Brasen, J.C., Scharff-Poulsen, A.M. and Olsen, L.F. (2008) Probing glycolytic and membrane potential oscillations in Saccharomyces cerevisiae Biochemistry 47, 7477-7484.
Andersen, A.Z., Carvalho, A.L., Neves, A.R., Santos, H., Kummer, U. and Olsen, L.F. (2009) The metabolic pH response in lactococcus lactis: an integrative experimental and modelling approach. Comp. Biol. Chem. 33, 71-83.
Olsen, L.F., Andersen, A.Z., Lunding, A. , Brasen. J.C. and Poulsen, A.K.(2009) Regulation of glycolytic oscillations by mitochondrial and plasma membrane H+-ATPases, Biophys. J. 96, 3850-3861.
Brasen, J.C., Olsen, L.F. and Hallett, M.B. (2010) Cell surface topology creates high Ca2+ signalling microdomains, Cell Calcium 47, 339–349
Brasen, J.C., Barington, T. and Olsen, L.F. (2010) On the mechanism of oscillations in neutrophils. Biophys. Chem. 148, 82-92.
Nielsen, L.J., Olsen, L.F. and Özalp, V.C. (2010) Aptamers Embedded in Polyacrylamide Nanoparticles: A Tool for In Vivo Metabolite Sensing. ACS Nano 4, 4361-4370.
Özalp, V.C., Pedersen, T.R. Nielsen, L.J. and Olsen, L.F. (2010) Time-resolved measurements of intracellular ATP in the yeast saccharomyces cerevisiae using a new type of nanobiosensor. J. Biol. Chem. 285 (48) 37579-37588
Özalp, V.C., Nielsen, L.J. and Olsen, L.F. (2010) An aptamer-based nanobiosensor for real-time measurements of ATP dynamics. ChemBioChem 11 (18), 2538-2541
Brasen, J.C., Olsen, L.F. and Hallett, M.B. (2011) Extracellular ATP induces oscillations in cytosolic free Ca2+ but not in NADPH oxidase activity in neutrophils. Biocheim. Biophys. Acta (Mol. Cell. Res.) 1813, 1446-1452.
Ytting, C.K., Fuglsang, A.T., Hiltunen, J.K., Kastaniotis, A.J., Özalp, V.C., Nielsen, L.J. and Olsen, L.F. (2012) Measurements of intracellular ATP provide new insight into the regulation of glycolysis in the yeast Saccharomyces cerevisiae Integr. Biol. 4, 99 – 107
Kloster, A. and Olsen, L.F. (2012) Oscillations in glycolysis in Saccharomyces cerevisiae: The role of autocatalysis and intracellular ATPase activity. Biophys. Chem. 165-166, 39-47.
Stock, R.P., Brewer, J., Wagner, K., Ramos-Cerrillo, B., Duelund, L., Jernshøj, K.D., Olsen, L.F. and Bagatolli, L.A. Sphingomyelinase D activity in model membranes: structural effects of in situ generation of ceramide-1-phosphate. PLoS ONE 7(4) e36003.
Poulsen, A.K. and Olsen, L.F. (2008) Optical nanosensor for the detection of reactive oxygen species WO 2008/044138 A1
Olsen, L.F., Özalp, C. and Nielsen, L.J. (2011) Nucleic acid biosensors (SDU 647-123) Patent pending. WO 2011/003424 A1
Popular science papers:
Brasen, J.C. og Olsen, L.F. (2010) Langt fra ligevægt. Aktuel Naturvidenskab 2010(2), 31-34.