Том 12, № 2 (2018)

Alpha-tocopheryl succinate (α-TS) is a vitamin-E derivative with potential antitumor properties. In this study, we have investigated the effect of α-TS on the morphology and motility of the A431 human epidermoid carcinoma cell line and HaCaT immortalized nontumorigenic human keratinocytes. Phase contrast and fluorescent microscopy, in vitro wound healing assay (scratch test), and scanning electron microscopy have been used. In both cell lines, alteration of cell shape, actin-cytoskeleton reorganization, cell-surface smoothing, disappearance of microvillae, and cell-motility inhibition have been shown. These effects are similar in both cell lines, but in the HaCaT cell line they occur after treatment with higher α-TS doses than for A431cells. Thus, we have demonstrated the selectivity of the effect of α-TS on tumor cells derived from human-skin stratified epithelia. The results of this work may be useful for further investigations focused on development of antitumor agents with selective action toward tumor cells.

Differences in the oxygen-dependent reactions of neutrophil granulocytes (NGs) depending on the nature of the agent affecting the cells were revealed. In vitro magnetite nanoparticles (MNPs) cause suppression of the NADPH–oxidase activity of NGs, which manifests itself in falling rates of reactions (NBT test) both with the effect of MNPs on NGs alone and a combined effect (MNPs and zymosan), as well as in the reduction of the index of activation (IA) and functional reserve of neutrophils (FRN). However, the introduction of MNPs dose-dependently stimulates the activity of myeloperoxidase (MPO). Gram-positive (S. aureus 2879 M) and gram-negative (E. coli 321) bacteria caused a respiratory burst of neutrophils, which manifested itself in a significant increase in the number of NBT-positive cells in single and combined influences (bacteria and zymosan). The lack of differences in the reaction of cells on opsonized and nonopsonized bacteria and the decrease in IA and FRN suggest that NGs are at the maximum level of functionality. Both strains of bacteria caused activation of the MPO.

CdSe and CdTe-based semiconductor fluorescent nanocrystals, also called quantum dots (QDs), attract the attention of biologists due to their wide range of emission in a visible light interval, high fluorescence quantum yield and photostability. However, their application is limited because of possible toxicity of cadmium. Indeed, there is a probability of metal leakage from QDs cores as a result of damage of both inorganic and organic layers of shells covering QDs. An alternative to cadmium QDs could be nanostructures having as a core, for example, non-toxical indium phosphide (InP), also emitting in the visible region of the spectrum. At present, there is few works on the use of these particles in biology. In this study, a comparative analysis of the spectral-luminescent properties of two InP/ZnS-QDs samples coated with PEG carrying- COOH or -NH₂ functional groups was performed. The obtained data were compared with the characteristics of CdSe/ZnS-QDs coated with PEG. The photophysical properties of all QDs in aqueous solution corresponded to the information claimed by manufacturers, but the fluorescence quantum yield of InP-based nanoparticles was found to be lower than that of CdSe-QDs. We also show that the photoluminescence of all types of QDs at pH 4.0 was lower than at pH 7.4, while the decrease in fluorescence intensity was minimal in the case of QDs-PEG-COOH. Studying the uptake of all three types of QDs by J774 macrophages, we found that the fluorescence spectra of internalized QDs do not change in comparison with those in solution. All three types of QDs after 24 hours of incubation were accumulated in the cells, but while QDs-NH₂ and QDs without reactive groups were detected mainly in vesicular-like discrete structures, the QDs-COOH were diffusely distributed throughout the cytoplasm. This fact indicates different mechanisms of interaction with cell membranes. In nonphagocytic HeLa cells all types of QDs behaved similarly, but the overall level of cells fluorescence was much lower. This may be due to both reduced nonspecific uptake and possible quenching of QDs fluorescence in acidic endolysosomes. Cytofluorimetric analysis of propidium iodide accumulation showed that after 24 hours incubation with all studied types of QDs as well as in control (no QDs), the proportion of dead HeLa cells did not exceed 10%. Thus, it has been demonstrated that non-toxic InP-based QDs can be used as an effective tool for biological research.

Mutations in the low density lipoprotein receptor gene (LDLR) frequently impair folding and intracellular traffic of the receptor protein, resulting in the development of a monogenic disorder, familial hypercholesterolemia (FH). Identification of novel LDLR mutations requires confirmation of their functional importance in distinguishing pathogenic mutations from neutral changes in the aminoacid sequence. To elaborate a system for evaluation of the effect of mutation on the folding and intracellular transport of the LDLR, as well as its ability to bind low density lipoprotein (LDL), we constructed a plasmid containing LDLR cDNA and the gene of enhanced green fluorescent protein (EGFP). Confocal microscopy has shown that, upon transient transfection of HEK293 cells with the plasmid, the recombinant fusion protein LDLR–EGFP is transported onto the cellular membrane and binds labeled LDL. This construct will be further modified by site-directed mutagenesis to reproduce the LDLR missense mutations most common in the population of northwest Russia so as to study the subcellular localization and function of the modified chimeric protein.

The aim of the present study was to compare structure of the nucleoli of ependymocytes, tanycytes, and secretory cells of the subcommissural organ using immunohistochemical staining for nucleolin and confocal laser microscopy. The study was performed in samples from the diencephalon of adult male Wistar rats (n = 6). The samples were fixed in zinc–ethanol–formaldehyde, a fixative providing a high level of preservation of antigen determinants. In the present study, we estimated diameters of nucleoli and their number in various types of cells lining the third ventricle. We compared for the first time the nucleoli of different subpopulations of tanycytes and report data on the distribution of nucleolin protein in the cells lining the ventricles. The content and location of nucleolin reflect the functional state of the cell. Our data will promote understanding of the interrelationships between the indices of the nucleolar apparatus and the functional state of the cell under various conditions, including stress, neoplastic transformation, and other pathological conditions.