[Zinc essentiality and toxicity. Biophysical aspects].

In this review the current conceptions concerning zinc biology, its metabolism and transport into the cells, its homeostasis, a role in the functioning of the human immune and endocrine systems, participation in cell signaling and its cytotoxicity, as well as the biophysical mechanisms of action of zinc ions action at the elevated concentrations on human blood cells were analyzed.

[Effect of cholesterol on the functional activity of proteins responsible for the resistance of human lymphocytes to xenobiotics].

The influence of agents modifying cholesterol in plasma membranes on the functional activity of transporter proteins (P-glycoprotein (P-gp) and the multidrug resistance protein 1 (MRP1)) in human lymphocytes has been studied. It was shown that changes in lateral distribution of cholesterol using the polyene antibiotic filipin, which disturb the structure and function of glycolipid microdomains in plasma membranes of lymphocytes lead to a decrease in the transport activity of both P-gp and MRP1. It was found that the treatment of human lymphocytes with the cyclic oligosaccharide methyl-beta-cyclodextrine, which leads to cholesterol depletion and reduction of lipid bilayer microviscosity in membranes of these cells, also decreases the functional activity of these proteins. It was concluded that the transport activity of P-gp and MRP1 depends on the modification of cholesterol in the membranes of human lymphocytes, i.e., is closely associated with the level of cholesterol and its lateral distribution.

Activity of membrane-bound NADH-methemoglobin reductase and physical state of lipids in erythrocyte membranes.

The activity of membrane-bound NADH-methemoglobin reductase in erythrocytes and the physical state of lipids in erythrocyte membranes under oxidative stress in cells were studied. A decrease of the activity of membrane-bound NADH-methemoglobin reductase and a change of physical state of the lipid bilayer of membranes under oxidative stress were found in erythrocytes in vivo and in vitro.

[Changes in the fluorescence parameters of lipophilic probes bound to the erythrocyte membrane in cancer patients]. / Ob izmenenii parametrov fluorestsentsii lipofil'nykh zondov, sviazannykh s membranami éritrotsitov krovi onkologicheskikh bol'nykh.

The fluorescence of intensivity of membrane-bound diphenylhexatriene (DFHT) of erythrocytes of the cancer patients is found to increase as compared to that of donors. No reliable differences are detected in a degree of fluorescence polarization of DFHT and pyren excimerization in erythrocyte membranes of cancer patients and donors. The data obtained show that under the development of malignant tumours the structure state of lipid bilayer of erythrocyte membrane does not undergo significant changes. However an increase of the fluorescence intensivity of the membrane-bound DFHT, evidently, is the result of changes in protein-lipid interactions in the membrane.

[Elevated agglutinability of erythrocytes in cancer patients]. / Povyshennaia aggliutinabel'nost' éritrotsitov krovi onkologicheskikh bol'nykh.

Mediated agglutination of erythrocytes was studied in the presence of concanavalin A (Con A). Agglutinability of erythrocytes from cancer patients was found to be higher than that of normal cells as a result of chemical factors present in the blood plasma which transform the cell surface, the number of Con A receptors remaining unchanged. It is concluded that increased agglutinability of erythrocytes from cancer patients is mainly due to a decrease in the negative charge of cells and the appearance of echinocytic forms.

[Various structural-functional characteristics of erythrocyte membranes in malignant growths]. / Nekotorye strukturno-funktsional'nye kharakteristiki membran éritrotsitov pri zlokachestvennom roste.

Changes in the acetylcholinesterase activity, thermal and osmotic stability of erythrocyte membranes of the cancer patients are found as compared with this activity of erythrocyte membranes of donors. No differences were detected in the rate of phosphate anion transfer through the erythrocyte membranes in the examined persons. The data obtained show that the development of malignant tumours is followed by the changes in the structural and functional state of surface components of erythrocyte membranes.

[Changes in lipid asymmetry and transport of glutathione conjugates under the influence of calcium ions in human erythrocytes]. / Izmenenie asimmetrii lipidov i transporta kon''iugatov glutationa v éritrotsitakh cheloveka pod vliianiem ionov kal'tsiia.

The influence of calcium ions on the distribution of the fluorescent analog of phosphatidylcholine 2-(6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoyl-1-hexadecanoyl-sn-glycero-3-phosphocholine (C6-NBD-PC) in membranes and transport of glutathione-S-conjugates in human erythrocytes was studied. It was supposed that both processes were performed by the multidrug resistance protein. It was found that the increase in intracellular calcium concentration tended to both the redistribution of about 35% of C6-NBD-PC in the inner layer of the membrane and a decrease in the exit of glutathione-S-conjugates from erythrocytes. In both cases, the intracellular calcium concentration varied in the physiological (nanomolar) range. The results testify that Ca2+ participates in the regulation of the activity of the multidrug resistance protein.

[Structural modification of erythrocyte membranes during oxidative stress and activity of membrane bound NADH-methemoglobin reductase]. / Strukturnaia modifikatsiia membran éritrotsitov pri okislitel'nom stresse i aktivnost' membranosviazannoi NADH-methemoglobinreduktazy.

The activity of NADH-methemoglobin reductase (metHb-reductase) in membranes isolated from human erythrocytes treated with phenylhydrazine at its sublytic concentration was studied. A decrease in the activity of membrane-bound metHb-reductase was shown to depend on the concentration of phenylhydrazine. Simultaneously, an increase in the level of membrane-bound methemoglobin and a change in the fluorescence parameters of membrane-bound 4,4'-diisothiocy-anatostilbene-2,2'-disulfonic acid were registered. In the case when Hb-free erythrocyte ghosts were treated with 0.2-2.0 mM phenylhydrazine, the activity of metHb-reductase did not change. The obtained results indicate that the inhibition of the activity of membrane-bound metHb-reductase by phenylhydrazine-induced oxidative stress in human erythrocytes is not caused by the direct action of the oxidant on the enzyme. The reason for this is the interaction of the products of hemoglobin oxidation with erythrocyte membrane (protein band 3) and structural changes in membrane proteins.

[Changes in the lipid physical state in human erythrocyte membranes subjected lead exposure in vitro]. / Izmenenie fizicheskogo sostoianiia lipidov v éritrotsitarnykh membranakh cheloveka pri deistvii svintsa in vitro.

The effect of lead acetate on the physical state of membrane lipids in human erythrocytes in vitro was studied using the lipophilic fluorescence probe 1,6-diphenyl-1,3,5-hexatriene and spin probes 16-doxyl-stearate and iminoxyl palmitic acid. It was shown that 2-10 microM lead acetate causes an increase in both intensity and polarization of fluorescence of 1,6-diphenyl-1,3,5-hexatriene, indicating changes in the microviscosity of the lipid bilayer of erythrocyte membranes. Judging from the parameters of EPR spectra of 16-doxyl stearate and iminoxyl palmitic acid incorporated into erythrocyte membranes, 2-10 microM lead acetate increases the heterogeneity of the lipid bilayer in surface and deep hydrophobic layers of the erythrocyte membrane.

[Parameters of hemolysis by sodium dodecyl sulfate as an indicator of the structural states of erythrocyte membranes]. / Parametry gemoliza dodetsilsul'fatom natriia kak indikator strukturnogo sostoianiia membran éritrotsitov.

The relationship between the parameters (percentage and rate) of hemolysis by sodium dodecyl sulfate and the time of incubation of human erythrocyte suspensions in glucose-free medium at 37 degrees C was studied. The polyphasic changes in the parameters were found, which depend on the mode of pretreatment: ATP depletion by iodoacetate, heat denaturation of spectrin, and treatment of cells by valinomycin. It was found that the percentage and rate of detergent hemolysis do not always change in parallel.

[Oxidation of membrane proteins and modification of erythrocyte surface characteristics]. / Okislenie membrannykh belkov i izmenenie poverkhnostnykh svoistv éritrotsitov.

It is shown that the oxidation of membrane protein SH-groups of human erythrocytes by diamide leads to the intensification of vesiculation and the formation on the cell surface of "senescent antigens". These processes were studied during prolonged incubation of erythrocytes at 37 degrees C in the absence of glucose. It was found that the formation of vesicules in ATP-depleted cells correlated with the appearance of "senescent antigens" (r = 0.9, p < 0.01). The oxidation of protein SH-groups takes place at the early stages of vesiculation and strengthening of autologous immunoglobulin binding, indicating that the oxidation of the other groups of membrane proteins plays the leading role in the modification of surface properties.

[Content of intracellular ATP and structural state of proteins in the erythrocyte membrane]. / Socerzhanie vnutrikletochnoi ATF i strukturnoe sostoianie belkov v éritrotsitarnoi membrane.

Effects of depletion and resynthesis of intracellular ATP on the properties of human erythrocytes (deformability, osmotic fragility) and on the structural parameters of their membranes (U. V. fluorescence spectra, protein aggregation) were studied. Incubation of the erythrocytes in the absence of glucose at 37 degrees C was shown to result in a number of consequent alterations: crenating and decrease in deformability of erythrocyte cells--a change in relative contents of membrane proteins--aggregation of membrane proteins and reversible shift of their fluorescence spectra. The evidence is considered in favour of the spectra shift due to dephosphorylation of membrane proteins.

[Change of state of erythrocyte membrane proteins during storage of preserved blood]. / Izmenenie sostoianiia membrannykh belkov éritrotsitov v protsesse khraneniia konservirovannoi krovi.

Erythrocyte vesiculation, kinetic parameters of tryptophane phosphorescence at room temperature and activity of erythrocyte membrane during storage of human blood at 4 +/- 2 C were studied. It is shown that before the erythrocyte vesiculation stage (10-15 days of storage) two types of changes in the structural state of the membrane proteins occur: 1) on the 1-5th days and 2) on the 5-10th days of blood storage.

[Vesiculation of erythrocytes during storage and connection of it with other processes in the cell]. / Vezikuliatsiia -eritrotsitov pri ikh khranenii i sviaz' ee s drugimi protsessami v kletke.

The activation energies of three processes (AT-depended vesiculation, release of potassium ions from erythrocytes and formation of methemoglobin) in erythrocytes incubated in phosphate buffer are measured in the range 4-37 degrees C. It is show that the Arrhenius plots for the first two processes have the same characteristics namely break at temperature around 18 degrees C, and Ea-58 kJ/mol at t > 18 degrees C. These data suggest that the velocity of ATP-depended vesiculation of erythrocytes is determined by the membrane permeability to monovalent cations. The appearance of methemoglobin in extracellular medium is characterized by a distinct value Ea(120 kJ/mol), although kinetics of this process is similar to vesiculation kinetics. Connection between considered processes and interrelation between ATP-depended vesiculation and spontaneous vesiculation are discussed.

[Changes in the structural-functional state of erythrocyte membranes treated with anion transport inhibitors]. / Izmenenie strukturno-funktsional'nogo sostoianiia membran éritrotsitov pod vozdeistviem ingibitorov transporta anionov.

Effect of 4,4-dyisotiocyanostilben-2,2-disulfonate (DIDS) and 1-ftor-2,4-dinitrobenzol (NDFB) on the rate of phosphate ion transport in erythrocytes, filtrability and thermal stability of erythrocytes and on the structural state of the erythrocyte membrane estimated by UV-fluorescence, PAAG--electrophoresis and measuring of the activity of membrane-bound acetylcholinesterase (AChE) has been studied. Unpenetrating anion transport inhibitor DIDS is shown to induce structural modifications of bands 3 of protein and AChE, while DNFB penetrating the membrane causes a significant reorganization of many membrane proteins (including spectrin) resulting in changes of transport and mechanical properties of erythrocytes.

[Damage to erythrocyte membranes induced by high-intensity ultrasound]. / Povrezhdenie membran éritrotsitov pri ul'trazvukovom vozdeistvii vysokoi intensivnosti.

The effect of high-intensity ultrasound (11.2-54.2 W/cm2, frequency 36 kHz) on the structural and functional state of erythrocytes was investigated. It was shown that, at short-term action of the ultrasound (up to 1 min), the dose-dependent hemolysis of erythrocytes occurs. It was found that the exposure to ultrasound of high intensity (54.2 W/cm2) leads to the disruption of the structural state of erythrocyte membranes, which manifests itself in a change of microviscosity of the lipid bilayer of membranes and inhibition of the activity of the lipid-dependent membrane-bound enzyme acetylcholinesterase.

[Effect of reduced and oxidized glutathione on physico-chemical properties of erythrocyte membranes]. / Vliianie vosstanovlennogo i okislennogo glutationa na fiziko-khimicheskoe sostoianie membran éritrotsitov.

The parameters describing the structural and functional state of membranes depending on the level of reduced glutathione in erythrocytes were studied. It was shown, that the decrease in the concentration of reduced intracellular glutathione in erythrocytes upon metabolic depletion (prolonged incubation of cells at 37 degrees C in the absence of glucose) or a rapid irreversible depletion of glutathione with 1-chloro-2,4-dinitrobenzene enhances lipid peroxidation processes in membranes, inhibits the membrane-bound NAD.H methemoglobin reductase activity and decreases the intensity of 1,6-diphenyl-1,3,5-hexatrien fluorescence. The data obtained suggest that the depletion of reduced intracellular glutathione causes changes in the physicochemical state of the erythrocyte membrane: the accumulation of lipid peroxidation products, changes in the physical state of lipid bilayer and the inhibition of membrane-bound NAD.H-methemoglobin reductase activity.

[Pyruvate and glucose transport through the erythrocyte membranes and the role of spectrin in these processes]. / Transport piruvata i gliukozy cherez éritrotsitarnye membrany i rol' spektrina v étikh protsessakh.

Effect of antibody to peripheral protein spectrin and antibody to integral protein of band 3 on kinetic parameters of pyruvate and glucose transport in the pink erythrocyte ghosts has been studied. It is shown that spectrin structure reorganization induced by the antibody to this protein has different effect on pyruvate and glucose transport parameters. Band 3 protein modification with the help of the antibody to this protein changes pyruvate transport parameters, while glucose transport is not changed. The data obtained show that facilitated diffusion of glucose and anions in the erythrocyte membrane is carried out by different carriers, the action of these carriers essentially depending on the structure state of spectrin.

[Temperature transitions of spectrin in solution and in erythrocyte membranes]. / Temperaturnye perekhody spektrina v rastvore i v éritrotsitarnykh membranakh.

Temperature transitions of spectrin in solution and in human erythrocyte membranes are recorded in the region t greater than 40 degrees C by irreversible changes in protein fluorescence spectra. Structural changes are completed 20 min after the sample incubation at an increased temperature. Both for isolated spectrin and for erythrocyte ghosts the temperature of half-transition is 46 +/- 1 degree C. There is no transition in the membranes after the removal of spectrin. Transitions in erythrocyte ghosts and in spectrin solution disappear at pH 5 when spectrin is in an aggregated state. Spectrin is suggested to be responsible for the transitions at 50 degrees C; its state in the cells areas more thermostable than in isolated membranes.

[Effect of diamide on protein oxidation and physico-chemical properties of lipids in erythrocyte membranes]. / Okislitel'naia modifikatsiia belkov i fiziko-khimicheskoe sostoianie lipidov v membranakh éritrotsitov pri deistvii diamina.

The effect of diamide on the physicochemical state of proteins and lipids of human erythrocyte membrane was studied. It was found that diamide at a concentration of 1 mM decreases the content of the SH-groups of membrane proteins by approximately 50%, resulting in enhanced vesiculation of erythrocytes upon metabolic exhaustion of cells. It was shown using fluorescein isothiocyanate-labeled concanavalin A and 4,4'-diisothiocyano-2,2'-stilbene disulfonate that diamide changes the structural state of the main integral protein of erythrocyte membranes, the band 3 protein. Changes in the microviscosity of the membrane lipid bilayer depending on diamide concentration were determined from the changes in the fluorescence parameters of the lipophilic probes (pyrene and 1,6-diphenyl-3,5-hexatriene). The level of lipid peroxidation products in membranes remained unchanged. It follows from these data that the SH-oxidizing agent diamide does not directly interact with the lipid bilayer of membrane and produces changes in the physicochemical state of lipids presumably by disrupting protein-lipid interactions that take place upon oxidation of the SH-groups and cross-linking of membrane proteins.