Magister biofizyki, specjalność biofizyka molekularna, Zakład Fizyki Makromolekularnej, Wydział Fizyki, Uniwersytet im. Adama Mickiewicza w Poznaniu, od 2015 roku. Tematem pracy badawczej jest wpływ wybranych surfaktantów na właściwości amyloidogenne peptydów A beta. Zainteresowania naukowe: badanie systemów na bazie surfaktantów do terapii genowej, badania strukturalne i spektroskopowe peptydów i kwasów nukleinowych, badania niskokątowym rozpraszaniem promieniowania rentgenowskiego (SAXS). Kilkuletni członek Biophysical Society oraz Polskiego Towarzystwa Synchrotronego.
Dobies M., Iżykowska J., Wilkowska M., Woźniak-Braszak A., Szutkowski K., Skrzypczak A., Jurga S., Kozak M.
Dispersion of water proton spin-lattice relaxation rates in aqueous solutions of multiwall carbon nanotubes (MWCNTs) stabilized via alkyloxymethylimidazolium surfactants Carbon nanotubes and a number of other carbon nanomaterials have a tendency to aggregate, which often resulted in difficulties of dispersion of these nanomaterials in aqueous solutions. The ability of dicationic (gemini) surfactants to disperse multiwall carbon nanotubes in water and the dynamic processes taking place at the water-MWCTs interface are studied. Stable dispersions of multi-wall carbon nanotubes with selected gemini surfactants (1,1?-(1,6-hexanediyl)bis(3-alkyloxymethylimidazolium) dichlorides) were prepared and characterized by nuclear magnetic relaxation dispersion (NMRD), NMR diffusometry, scanning and transmission electron microscopy, and Fourier transform infrared spectroscopy. The addition of multiwall carbon nanotubes to aqueous solutions of studied gemini surfactants leads to significant paramagnetic enhancement of the spin-lattice relaxation processes, which gets more pronounced with increasing concentration of well-dispersed MWNTs in water. The dominant role of outer sphere (OS) relaxation mechanism in total observed R1, governed by two-dimensional diffusion of water on the carbon nanotube surface in the vicinity of paramagnetic centers incorporated in the MWCNTs side-walls (mainly of iron origin), was assumed to explain NMRD data. The NMR diffusion experiments confirm the existence of restricted water diffusion in the studied supernatants. The NMR diffusion results are consistent with the FTIR and NMR proton spin-lattice relaxation dispersion in which the more effective R1 dispersion noticed for the sample with IMIC6C12 was ascribed to the better accessibility of water molecules to the surface of the MWCNTs.
The Journal of Physical Chemistry, 121(21), 11839-11850 (2017)
Andrzejewska W., Wilkowska M., Chrabąszczewska M., Kozak M.
The study of complexation between dicationic surfactants and the DNA duplex using structural and spectroscopic methods Dicationic (also known as gemini or dimeric) bis-alkylimidazolium surfactants belong to a group of non-viral transfection systems proposed for the successful introduction of different types of nucleic acids (i.e., siRNA, DNA oligomers, and plasmid DNA) into living cells. Our studies reveal the formation of complexes composed of dicationic (gemini) surfactants, 3,3?-[?,?-(dioxaalkane)]bis(1-dodecylimidazolium)chlorides, and 21 base pair deoxyribonucleic acid duplexes (dsDNA). The studied dsDNA and its complex formation process was analysed by small-angle X-ray scattering (SAXS), molecular modelling (dsDNA), agarose gel electrophoresis (AGE) and circular dichroism spectroscopy (CD). We observed the formation of stable complexes for charge ratio values of p/n > 2. Moreover, we noted conformational changes similar to those observed during the transition of B-DNA to C-DNA, X-DNA, and Z-DNA in several spatial structures (i.e., micellar, hexagonal and cubic) formed in mixtures. The surfactants used in this study were investigated for the influence of dioxaalkane spacer length and the presence of an imidazolium moiety on the complexation process. The complexes formed were stable, and the complexation process was reproducible and efficient. Toxicity tests done on HeLa cells allowed for the determination of non-toxic concentrations of studied surfactants. Transfection tests have confirmed that the studied surfactant systems are effective DNA carriers.
Moliński A., Zaręba J., Iżykowska J., Skupin M., Andrzejewska W., Jurga S., Kozak M.
C-60 fullerenes as contrast agents - structural, spectroscopic and nano-toxicity studies Recently, C60 fullerenes have been proposed as contrast agents for MRI method . Especially promising for in vitro and in vivo NMR imaging are their complexes with gadolinum (containing Gd3+ ions entrapped inside the fullerene cage) known also as gadofullerenes .
The general problem of carbon nanomaterials is their toxicity towards living cells . The aim of our study is to devise fullerene-based suspensions characterized with low toxicity, which could later be developed into metallofullerene contrast agents. We want to achieve that goal using C60 fullerenes, pluronics to lower the toxicity and phosphatidylcholine derivatives (DMPC or DPPC) to enhance biocompatibility of fullerenes and to stimulate the cellular absorption. In this work we would like to present results of toxicity studies of C60 fullerene systems pluronic modified C60 fullerene systems towards HeLa cells, along with results of structural analyses by AFM and SEM microscopies, FTIR and SAXS. This study was supported by Ministry of Science and Higher Education (Poland), within the project, Najlepsi z najlepszych!?
Skupin M., Iżykowska J., Andrzejewska W., Dobies M., Jurga S., Kozak M.
Interactions of carbon nanotubes stabilized by selected gemini surfactants with model biomembranes Thanks to the extraordinary mechanical strength and high electrical conductivity multiwalled carbon nanotubes are currently used in electronics, medicine (as biomedical sensors, transporters or drugs) as well as in the production of lightweight and durable construction. The aim of this study was to determine the possibility to use different cationic gemini surfactants with different spacer lengths or alkyl chain lengths in more efficient systems for dispersing nanostructures in aqueous solutions. The most important advantages of these systems are their non-immunogenic, biocompatible properties and generally low toxicity . Therefore nanotubes, surrounded by surfactants, have the potential to interact with biological membranes. For this purpose we studied the influence of dispersed CN solution on the phase behavior of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) - a phospholipid most often present in membranes of nerve cells. The microstructure of the stable suspension of carbon nanotubes was investigated using high-resolution Transmission Electron Microscopy and Atomic Force Microscopy. Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC) were used to analyze the influence of surfactants studied, used for CN dispersion, on the phase behavior of DMPC bilayers. A series of measurements of toxicity of these systems were performed in HeLa and fibroblast cell cultures. This work was supported by the Ministry of Science and Higher Education, within the project ?Najlepsi z najlepszych!? dec.
Andrzejewska W.J., Skupin M., Skrzypczak A., Kozak M.
Dicationic and tricationic surfactants as transgene carriers - comparison of their ability to dsdna and sirna binding In last decades, many different compounds have been tested as potential delivery systems of nucleic acids in gene therapy. Compounds subjected for delivery of nucleic acids should be characterized by low toxicity and high ability to form stable complexes with nucleic acids. In practice they should support efficient transfer of therapeutic material (transgene) to the pathological cells. Introduced transgene by interaction with the corresponding cellular genome induce a permanent curative effect. Our previous study indicated, that good transfection properties, have three-dimensional structures formed by self-organized compounds which are structurally similar to the natural lipids. Particularly, new oligomeric amphiphiles, dicationic and tricationic surfactants, seems to be quite promising. The physicochemical properties of these compounds promote their ability to create a stable, biocompatible complexes with dsDNA and siRNA. In this work we present results of structural studies of the complexes formed on the basis of selected novel dicationic or tricationic surfactants with short nucleic acid oligomers (dsDNA and siRNA, 21 bp). These systems were examined by the use of atomic force microscopy (AFM), small angle scattering of synchrotron radiation (SR-SAXS), circular dichroism (CD) and gel electrophoresis. We performed also toxicity tests on HeLa cells. Studies have shown formation of stable systems with the desired biological and chemical properties.
Iżykowska J., Skupin M., Andrzejewska W., Dobies M., Jurga S., Kozak M.
Carbon nanostructures of different spatial geometry: their dispersion and influence on model biological systems The discovery of carbon nanostructures (CN) in the last century has revolutionized science, opening new research opportunities in biophysics, material sciences, biomedicine and pharmacology . Unfortunately, carbon nanomaterials show a strong tendency towards aggregation and poor stability in solutions. Such properties, especially the suspension stability, are critical for bio-applications of carbon nanostructures . Surface functionalization of nanocarbons improves the dispersing properties in water solutions but it may also influence their physicochemical properties . For preparation of biocompatible CN samples we used novel dicationic surfactant (gemini) with imidazole head groups. Nanocarbon suspensions were investigated in the context of their anticancer activity, as well as their application as multimodal contrast/drug delivery agents. Different spatial geometries of CN may be of key importance for designing of the efficient anti-cancer system. Such systems were subjected to the cytotoxicity tests on HeLa cell cultures. The structural properties of selected, the most effective systems based on CN suspensions were characterized by the use of SEM, AFM and NMR methods. This study was supported by the Ministry of Science and Higher Education (Poland), within the project ?Najlepsi z najlepszych!?
Zaręba J., Iżykowska J., Skupin M., Moliński A., Dobies M., Jurga S., Kozak M.
The influence of surfactant structure and geometry on nanotoxicity and dispersion of carbon nanotubes Graphene, nanotubes and fullerenes, also known as carbon nanomaterials (CN), because of the their unique properties have potential applications not only in preparation of new composite materials or in electronics, but they can be also components of drug delivery systems. However, the applications of CN in life sciences or biomedicine require stable dispersions of these materials in water. The hydrophobic nature of CN causes serious problems with their separation in water solutions. Our study is aimed to find a way of dispersing such CN systems in water, by the use of non-covalent surface modification via different surfactants. In this work we focused on the use of trimeric, dimeric and monomeric surfactants fornon-covalent surface modification of selected carbon nanotubes. The toxicity of these systems (aqueous solutions of CN with surfactants and surfactants solutions) towards selected cell cultures (e. g. HeLa cells) was also tested. The surfactants studied can interact with carbon nanotubes in a number of different modes: hydrophobic interactions (surfactant chains with CN side walls) or ?-? interactions (aromatic rings of surfactants with CN surface). Infrared spectroscopy and atomic force microscopy were used for characterization of the systems studied. Finally, we focused on mechanical properties of cells exposed to surfactants and CN. This study was supported by the Ministry of Science and Higher Education (Poland), within the project ?
Andrzejewska W., Pietralik Z., Skupin M., Kozak M.
Structural studies of the formation of lipoplexes between siRNA and selected bis-imidazolium gemini surfactants Dicationic (gemini) surfactants are agents that can be used for the preparation of stable complexes of nucleic acids, particularly siRNA for therapeutic purposes. In this study, we demonstrated that bis-imidazolium gemini surfactants with variable lengths of dioxyalkyl linker groups (from dioxyethyl to dioxydodecyl) and dodecyl side chains are excellent for the complexation of siRNA. All of these compounds effectively complexed siRNA in a charge ratio range (p/n) of 1.5-10. The low resolution structure of siRNA oligomers was characterised by small angle scattering of synchrotron radiation (SR-SAXS) and ab initio modelling. The structures of the formed complexes were also analysed using SR-SAXS, circular dichroism studies and electrophoretic mobility tests. The most promising agents for complexation with siRNA were the surfactants that contained dioxyethyl and dioxyhexyl spacer groups.
Colloids and Surfaces B: Biointerfaces, 146, 598-606 (2016)
The system with zwitterionic lactose-based surfactant for complexation and delivery of small interfering ribonucleic acid-A structural and spectroscopic study Systems suitable for the effective preparation of complexes with siRNA (small interfering RNA) are at the center of interest in the area of research work on the delivery of the RNA-based drugs (RNA-therapeutics). This article presents results of a study on the structural effects associated with siRNA complexation by a surfactant comprising a lactose group (N-(3-propanesulfone)-N-dodecyl-amino-beta-D-lactose hydrochloride, LA12). The double stranded siRNA oligomer (21 base pairs) used in this study is responsible for silencing a gene that can be important in the therapy of myotonic dystrophy type 1. The obtained siRNA/LA12 lipoplexes were studied using the methods of small angle scattering of synchrotron radiation, circular dichroism spectroscopy, Fourier transform infrared spectroscopy, and electrophoretic mobility tests. Lipoplexes form in solution stable lamellar or cubic phases. The surfactant selected for the study shows much lower cytotoxicity and good complexation abilities of siRNA than dicationic or polycationic surfactants.
Study of molecular mechanisms of amyloid beta-peptid variants aggregation in presence of surfactants It is estimated that in 2050 at least 1.25 % of population might have Alzheimer's disease. It is supposed that the most critical changes relevant for the progress of this disease are related to development and deposition of amyloid plaque whose main components are amyloid ?-peptides (A?).
The aim of this study was characterization of the structure and conformational changes of ?-amyloid peptides in the presence of a wide group of surfactants. The kinetic study of the aggregation behavior of different peptides in solution was also undertaken. Four groups of surfactants (cationic, dicationic, tricationic and zwitterionic) and different ?-amyloid peptides were selected for the study. The surfactant concentration effect was tested on 1-42 A? peptide and its shorter variants (N-terminal with hydrophilic properties and hydrophobic C-terminal fragment).
Analysis of the secondary structure of A? peptide in the presence of different concentrations of surfactants as well as a function of temperature, was carried out using FTIR spectroscopy and circular dichroism method. The kinetics of aggregation processes of the peptides and formation of plaques was studied using fluorescence spectroscopy and Thioflavin T assay. The size distribution of aggregates of A? peptide was evaluated on the basis of gel electrophoresis.
Results of our study showed different impact of surfactants studied on the conformations of beta-amyloid peptides. Preliminary examination of the cytotoxicity of selected surfactants on HeLa cells was conducted in order to verify their suitability for therapeutic purposes.
Andrzejewska W.J., Skupin M., Skrzypczak A., Kozak M.
Structural characterization of dsDNA nanocomplexes based on ammonium gemini surfactants One of the most intensively studied groups of chemical compounds are gemini surfactants, because of their specific and attractive properties, especially the capability of stable binding of the nucleic acids, formation of spatial structures with high symmetry and relatively low cytotoxicity. They have found a broad range of applications in medicine, chemical technology, pharmaceutical industry. These complexes can be applied as nonviral transfection delivery systems in gene therapy, a novel and most promising method for the treatment of a broad range of genetic diseases. Dicationic surfactants allow introduction of a transgene without inducing natural immunological response, and release it undamaged inside the cell. However, the successful completion of this process is difficult, because of the need for many studies in order to determine all necessary parameters.
In our study we analysed nanocomplexes of cationic gemini surfactants(1,n-dodecanebis[(oxymethyl)alkyldimethylammonium]chlorides) with DNA oligomers using synchrotron radiation of small angle X-ray scattering, circular dichroism spectroscopy, agarose gel electrophoresis, polarization microscopy and transmission electron microscopy. We also analysed the cytotoxicity of our systems using MTT colorimetric assay on model cell
We observed the formation of complexes, their morphology, spatial nanostructure, conformation of nucleic acid inside them and influence on the cells. The systems studied were found to be stable and the process of their formation was reproducible.
Andrzejewska W.J., Skupin M., Murawska M., Skrzypczak A., Kozak M.
The study of complexation process between cationic gemini surfactants and dna using structural and spectroscopic methods Dicationic (gemini) surfactants are intensively studied group of chemical compounds, because of the broad range of applications in medicine, chemical technology or pharmaceutical industry. In solution they can form with nucleic acids the complex structures (lipoplexes), which can be used as drug delivery systems in nonviral transfection. Lipoplexes in gene therapy offer efficient introduction of a therapeutic material to the living cells. Gemini surfactants also allow introduction of a transgene without inducing natural immunological response and release it inside the cell. In our study, we analyzed the process of complexation of cationic gemini surfactants (3.3'- [1,6- (2,n-dioxyalcane)] bis(1-dodecyloxyimidazolium dichlorides)) with DNA, using small angle X-ray scattering, circular dichroism spectroscopy and gel electrophoresis. Surfactants which have been used had of variable length of the spacer group. We observed the formation of stable complexes in these systems and the process of complex formation was reproducible, efficient and immediate.
Nanosystem based on phospholipids and surfactants as innovative delivery system for gene therapy Amphiphilic dicationic surfactants, known as gemini surfactants, are currently studied for gene delivery purposes. The biggest advantages of these systems are that they are non-immunogenic and generally have low toxicity. One of the most important advantages of these systems is improved transfection efficiency. The aim of this study was to determine the possibility to use amphoteric surfactants (zwitterionic derivatives of sulfobetaine with carbohydrate moiety) and sulfobetaine/gemini surfactant mixtures as complexing agents for nucleic acids, with potential applications for gene delivery to reduce the toxicity and improved transfection. Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC) were used to a7nalyze influence of surfactants on the phase behaviour of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) bilayers with the presence of different DNA forms (small DNA oligomers, cDNA, low and high-molecular mass DNA). The influence of different concentrations of sulfobetaine and sulfobetaine/gemini surfactant mixtures with the presence of DNA on creating stable complexes was investigated using circular dichroism (CD) spectroscopy and electrophoresis. A series of measurements of toxicity and transfection of these lipoplexes were performed in HeLa cells. These compounds appear to be excellent for creating complexes with DNA. Thanks to their construction this DNA carrier molecules might be able to deliver genes to the cells of almost any DNA mo7lecular size, unattainable when using viral gene delivery systems.