V. S. Shubina

Cytoskeletal regulation of the cellular function by dopamine

The interaction of dopamine with model membranes, isolated G-actin, and living cells, such as Mauthner neurons and fibroblast-like BHK-21 cells has been studied. It was found that in vitro dopamine passes through the phospholipid membrane and directly polymerizes G-actin due to incorporation into threads as their integral part. In in vivo conditions, it penetrates inside the cell and induces the appearance of a network of actin filaments in loci rich in globular actin. The data suggest that there exists a mechanism of dopamine interaction with living cells, which is based on direct polymerization of cytosolic G-actin as its cellular target. The reorganization of the actin cytoskeleton leads to changes in the morphofunctional status of cells.

Effect of Dopamine on Viability of BHK-21 Cells

We studied the effects of dopamine added to culture medium on survival of floating or adherent BHK-21 cells differing by organization of actin cytoskeleton. The viability of floating cells more drastically decreased with increasing dopamine concentration and duration of exposure than that of adherent cells. The cells worse adhered to the substrate and formed a monolayer. The formed monolayer degrades, cell borders become blurred, cells, polygonal in the control, are rounded. Preliminary blockade of dopamine receptors with haloperidol, inessential for cell survival and morphology, does not prevent the destructive effect of dopamine on the cells. Ultrastructural study revealed increased density of filamentous actin threads in deep compartments of cell cytoplasm after dopamine treatment, this increase being more pronounced in cells grown in suspension. Bearing in mind the polymerizing effect of dopamine on globular actin in vitro and the fact that the content of this protein in floating cells is higher than in adherent cells, we can conclude that the decrease in viability of BHK-21 cells is caused by interaction of dopamine with cytoplasmic globular actin.

Partitioning of taxifolin-iron ions complexes in octanol-water system

The composition of taxifolin-iron ions complexes in an octanol-water biphasic system was studied using the method of absorption spectrophotometry. It was found that at pH 5.0 in an aqueous biphasic system the complex of [Tf · Fe2(OH)k(H2O)8 − k] is present, but at pH 7.0 and 9.0 the complexes of [Tf2 · Fe(OH)k(H2O)2 − k] and [Tf · Fe(OH)k(H2O)4 − k] are predominantly observed. The formation of a stable [Tf3 · Fe] complex occurred in octanol phase. The charged iron ion of this complex is surrounded by taxifolin molecules, which shield the iron ion from lipophilic solvent. During transition from water to octanol phase the changes of the composition of complexes are accompanied by reciprocal changes in portion of taxifolin and iron ions in these phases. It was shown that the portion of taxifolin in aqueous solution in the presence of iron ions is increased at high pH values, and the portion of iron ions is minimal at pH 7.0. In addition, the parameters of solubility limits of taxifoliniron ions complexes in an aqueous solution were determined. The data obtained gain a better understanding of the role of complexation of polyphenol with metal of variable valency in passive transport of flavonoids and metal ions across lipid membranes.

The effect of the liposomal form of flavonoid-metal complexes on skin regeneration after chemical burn

The effect of the liposomal form formed by taxifolin and metals of variable valency was investigated. It was shown that application of preparations based on the free flavonoid and its complexes with Fe(II/III) and Cu(II) ions after chemical burns results in more effective skin regeneration and repair of hair follicles and sebaceous glands. A tendency toward more effective wound healing after the application of taxifolin-Cu(II) and taxifolin-Fe(III) liposomal complexes versus control was observed. It was assumed that the mechanism of action of these preparations is based on oxidative polymerization and conjugation of the flavonoid, which results in utilization of toxic metabolites and lipid peroxidation products.

Skin Regeneration after Chemical Burn under the Effect of Taxifolin-Based Preparations

We studied the effect of liposome preparation containing taxifolin oligomers and taxifolin conjugates with carbonyl compounds on skin regeneration after chemical burn. The preparations containing fl avonoid conjugates intensifi ed regeneration processes and reparation of hair follicles and sebaceous glands after chemical burn. The preparation based on taxifolin conjugate with acetaldehyde was most effective; its activity was comparable with that of the wound-healing preparation Olasol. Taxifolin conjugates with carbonyl compounds can be used for the creation of combined wound- and burn-healing preparations.

Cytochemical and Ultrastructural Characteristics of BHK-21 Cells Exposed to Dopamine

BHK-21 cells were incubated in medium containing dopamine (DA) and catecholamine contents were then measured using the Falck cytochemical method. As compared with controls, significant increases were seen in the fluorescence of cells and these were proportional to the concentration and duration of exposure to DA and more marked in cells in suspension than in attached cells. Parallel electron microscopic studies showed that the increased fluorescence intensity of the cytoplasm correlated with the presence of dense networks of fibrils which were identified on the basis of morphological characteristics as microfilaments consisting of F-actin. Prior blockade of dopaminergic receptors with haloperidol did not alter the subsequent effects of DA on fluorescence intensity or cell ultrastructure. These data suggest that, in conditions of chronic exposure, DA can penetrate into the cytoplasm, inducing actin polymerization and becoming bound to the newly formed actin cytoskeleton. Structurally, this can be apparent as hypertrophy of the cytoskeleton and its derivatives, with significant influences on the overall structure of the cell.

A new material based on collagen and taxifolin: Preparation and properties

The possibility of producing a material based on collagen and a biologically active polyphenol, viz., taxifolin, was investigated, and the properties of the material were studied. The data on the release dynamics and metal-reducing activity of polyphenol that is chemically linked to collagen are represented. The effects of taxifolin, taxifolin glutarate, and a polyphenol-containing gel on the production of reactive oxygen species by neutrophils that are stimulated with phorbol myristate acetate were examined. Polyphenol that was released from the gel was shown to have antioxidant and metal-reducing properties. This suggests that unoxidized polyphenol is effectively included in the collagen gel.

Ultrastructure of BHK-21 cells treated with dopamine

The ultrastructure of a suspension or of adhered BHK-21 cells exposed to dopamine has been studied. It was found that ultrastructural modifications were observed mainly in cellular regions enriched in actin, including in intercellular desmosome-like contacts, microvilli, and under the membrane cortical layer. Desmosome-like contacts are hypertrophied with increased electron density and fibrillar bridges that appear in specialized contacts. Microvilli fused with each other and with plasma membrane of neighboring cells or were split up and penetrated by fibrils. The cytoplasm is filled with randomly arranged F-actin filaments and has a blotchy appearance. The effect of dopamine is more pronounced in nonattached cells with undifferentiated actin cytoskeletons than in spread cells with stress fibers. The blockage of the D2 receptor with haloperidol does not affect the ultrastructure of BHK-21 or dopamine induced modifications. The data obtained suggest direct interactions between dopamine and the actin cytoskeleton.

Changes in the ultrastructure and function of goldfish Mauthner neurons in the presence of 3,4-dihydro-2(1H)-pyrimidinethione.

The behavioral effects of 3,4-dihydro-2(1H)-pyrimidinethione (DPT, a pyrimidine derivative), which is used as a test system for detecting tumor growth, on the ultrastructure and function of Mauthner neurons (MN), were studied in goldfish. Application of DPT to MN was found to lead to increased resistance of neurons to exhaustive stimulation, which was accompanied by increases in the sizes of actin-containing membrane desmosome-like contacts, along with the formation of bundles of actin stress fibers; these effects are similar to those previously reported with dopamine. The similarity of the morphofunctional changes in MN on exposure to an artificial chemical substance for which there are no membrane receptors and dopamine itself suggests that they have trophic effects on the stabilization and polymerization of cytoskeletal actin due to direct penetration into postsynaptic neurons.

Study of the cytotoxicity of protein X amyloid fibrils

Amyloid oligomers, protofibrils, and fibrils of various amyloidogenic proteins are known to induce cell death. Tetracycline prevents the formation of fibrils of Aβ peptide and other amyloidogenic proteins and decomposes mature fibrils. It was previously shown that sarcomeric cytoskeletal proteins of the titin family (protein X, protein C, and protein H) in vitro form amyloid fibrils and tetracycline decomposes them. In this work, the concentration and time dependence of the survival of polymorphonuclear leukocytes in the presence of protein X amyloid fibrils is demonstrated. It is also shown that the survival rate increases as fibrils are decomposed by tetracycline. The antibiotic itself is found to be nontoxic. The results obtained show that this approach can be used to evaluate the efficiency of drugs that prevent or rectify amyloidoses.