Detailed kinetic data suggest that the direct transfer of plasmid DNA (YEp 351, 5.6 kbp, supercoiled, Mr approximately 3.5 x 10(6)) by membrane electroporation of yeast cells (Saccharomyces cerevisiae, strain AH 215) is mainly due to electrodiffusive processes. The rate-limiting step for the cell transformation, however, is a bimolecular DNA-binding interaction in the cell interior. Both the adsorption of DNA, directly measured with [32P]dCTP DNA, and the number of transformants are collinearly enhanced with increasing total concentrations [Dt] and [Cat] of DNA and of calcium, respectively. At [Cat] = 1 mM, the half-saturation or equilibrium constant is KD = 15 +/- 1 nM at 293 K (20 degrees C). The optimal transformation frequency is TFopt = 4.1 +/- 0.4 X 10(-5) if a single exponential pulse of initial field strength E0 = 4 kV cm-1 and decay time constant tauE = 45 ms is applied at [Dt] = 2.7 nM and 10(8) cells in 0.1 ml. The dependence of TF on [Cat] yields the equilibrium constants KCazero = 1.8 +/- 0.2 mM (in the absence of DNA) and K'Ca (at 2.7 nM DNA), comparable with and derived from electrophoresis data. In yeast cells, too, the appearance of a DNA molecule in its whole length in the cell interior is clearly an after-field event. At Eo = 4.0 kV cm-1 and T = 293 K, the flow coefficient of DNA through the porous membrane patches is Kto = 7.0 +/- 0.7 x 10(3)S-1 and the electrodiffusion of DNA is approximately 10 times more effective than simple diffusion: D/D0 approximately 10.3. The mean radius of these pores is rp = 0.39 +/- 0.05 nm, and the mean number of pores per cell (of size ø approximately 5.5 microns) is Np = 2.2 +/- 0.2 x 10(4). The maximal membrane area that is involved in the electrodiffusive penetration of adsorbed DNA into the outer surface of the electroporated cell membrane patches is only 0.023% of the total cell surface. The surface penetration is followed either by additional electrodiffusive or by passive (after-field) diffusive translocation of the inserted DNA into the cell interior. For practical purposes of optimal transformation efficiency, 1 mM calcium is necessary for sufficient DNA binding and the relatively long pulse duration of 20-40 ms is required to achieve efficient electrodiffusive transport across the cell wall and into the outer surface of electroporated cell membrane patches.

This review is directed to the redox-modulating properties and anticancer effect of vitamin K. The concept is focused on two aspects: (i) redox-cycle of vitamin K and its effect on the calcium homeostasis, “oncogenic” and “onco-suppressive” reactive oxygen species and the specific induction of oxidative stress in cancer; (ii) vitamin K plus C as a powerful redox-system, which forms a bypass between mitochondrial complexes II and III and thus prevents mitochondrial dysfunction, restores oxidative phosphorylation and aerobic glycolysis, modulates the redox-state of endogenous redox-pairs, eliminates the hypoxic environment of cancer cells and induces cell death. The analyzed data suggest that vitamin C&K can sensitize cancer cells to conventional chemotherapy, which allows achievement of a lower effective dose of the drug and minimizing the harmful side-effects. The review is intended for a wide audience of readers - from students to specialists in the field.

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A newly isolated indigenous strain BN10 identified as Pseudomonas aeruginosa was found to produce glycolipid (i.e., rhamnolipid-type) biosurfactants. Two representative rhamnolipidic fractions, RL-1 and RL-2, were separated on silica gel columns and their chemical structure was elucidated by a combination of nuclear magnetic resonance and mass spectroscopy. Subsequently, their cytotoxic effect on cancer cell lines HL-60, BV-173, SKW-3, and JMSU-1 was investigated. RL-1 was superior in terms of potency, causing 50 % inhibition of cellular viability at lower concentrations, as compared to RL-2. Furthermore, the results from fluorescent staining analysis demonstrated that RL-1 inhibited proliferation of BV-173 pre-B human leukemia cells by induction of apoptotic cell death. These findings suggest that RL-1 could be of potential for application in biomedicine as a new and promising therapeutic agent.

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The present study was designed to investigate whether poly-ion complex hollow vesicles (polymersomes), based on chemically modified chitosan, are appropriate for passive tumour targeting in the context of their application as drug carriers. The experiments were performed on colon cancer-grafted mice. The mice were subjected to anaesthesia and injected intravenously with water-soluble nanoparticles: (1) QD705-labelled polymersomes (average size ∼120 nm; size distribution ∼10%) or (2) native QD705. The optical imaging was carried out on Maestro EX 2.10 In Vivo Imaging System (excitation filter 435–480 nm; emission filter 700 nm, longpass). In the case of QD705, the fluorescence appeared in the tumour area within 1 min after injection and disappeared completely within 60 min. A strong fluorescent signal was detected in the liver on the 30th minute. The visualization of tumour using QD705 was based only on angiogenesis. In the case of QD705-labelled polymersomes, the fluorescence appeared in the tumour area immediately after injection with excellent visualization of blood vessels in the whole body. A strong fluorescent signal was detected in the tumour area within 16 hours. This indicated that QD705-labelled polymersomes were delivered predominantly into the tumour due to their long circulation in the bloodstream and enhanced permeability and retention effect. A very weak fluorescent signal was found in the liver area. The data suggest that size-controlled long-circulating polymersomes are very promising carriers for drug delivery in solid tumours, including delivery of small nanoparticles and contrast substances.

Electropermeabilization is a non-viral method that can be used to transfer plasmid DNA (pDNA) into cells and tissues. According the applications and considered tissues, this safe method can be less efficient than the viral approaches. Biophysical mechanisms of gene electrotransfer are not entirely known. Contrary to small molecules that have direct and fast access to the cytoplasm, pDNA is electrophoretically pushed towards the permeabilized membrane where it forms a complex before being transferred into the cytoplasm. In order to understand the biophysical mechanisms of gene electrotransfer and in this way to improve it, we investigated the dependence of the topology of pDNA i.e. linear versus supercoiled on both pDNA/membrane interaction and gene expression. Our results revealed that: i) even if pDNA/membrane interactions are only slightly affected by the topology of pDNA, ii) gene transfer and expression are strongly influenced by it. Indeed, the linearization of pDNA leads to a decrease in the transfection level.

In the present study it is shown that poloxamer 188, added before or immediately after an electrical pulse used for electroporation, decreases the number of dead cells and at the same time does not reduce the number of reversible electropores through which small molecules (cisplatin, bleomycin, or propidium iodide) can pass/diffuse. It was suggested that hydrophobic sections of poloxamer 188 molecules are incorporated into the edges of pores and that their hydrophilic parts act as brushy pore structures. The formation of brushy pores may reduce the expansion of pores and delay the irreversible electropermeability. Tumors were implanted subcutaneously in both flanks of nude mice using HeLa cells, transfected with genes for red fluorescent protein and luciferase. The volume of tumors stopped to grow after electrochemotherapy and the use of poloxamer 188 reduced the edema near the electrode and around the subcutaneously growing tumors.