[Apoptosis and necrosis of lymphocytes induced by UV-radiation in the presence of autological plasma].

The influence of UV-light (240-390 nm) in doses 151-3020 J/m2 on the nature of the death of lymphocytes cells of donor's blood (using markers of apoptotic and necrotic death of cells) and on the level of expression of the marker of apoptotic pre-preparation--CD95-receptor has been investigated. We have shown that UV-radiation increases expression of CD95-receptors which is caused mainly by de novo synthesis of the receptors. It has been revealed that during daily incubation of photo-modified lymphocytes (151 and 755 J/m2) without autological blood cell death occurs by receptor-involved apoptosis. Exposure to high doses of radiation (1510 and 3020 J/m2) causes massive necrotic death of immunocytes. The use of autologous blood plasma during incubation of photo-modified lymphocytes allows decreasing the number of both apoptotic and necrotic cells.

[Micronucleus test of human oral buccal epithelium: problems, progress and prospects].

The articles by russian and foreign authors for the period from 2000 to 2012, devoted to the problems of application, analysis and interpretation of the results of micronucleus test in human buccal epithelium has been analyzed in the review. Nuclear abnormality founding in the cells of the oral mucosa has been described. The paper summarizes works devoted to the analysis of the influence of the micronucleus test methods (painting, taking scrapings) to its results. Modern opinions about the factors of different etiology (sex, age, genotype, psycho-physiological characteristics, immune status, diseases of different etiology, man-made pollution, climatic and geographical conditions, ionizing and nonionizing radiation, chemical compounds (drugs, dietary supplements, androgenic steroids, etc.), dental fillings, occupational exposures, alcohol, using tobacco blends) inducing the estimation of nuclear aberration has been summarized as a scheme. The problems and unresolved issues related to the peculiarities of micronucleus test has been noted.

[Inulinases from various producers: the features of their permolecular organization].

The structural organization of inulinases from yeasts, fungi, and plants are researched. For studying their sizes, molecular weight, and permolecular organization, an approach consisting of a combination of atomic force microscopy with methods of dynamic light scattering, gel chromatography, and electrophoresis was used. It is shown that inulinases from Kluyveromyces marxianus and Aspergillus niger form geterodimers and inulinases from tubers of Helianthus tuberosus are present as both dimers and monomers. The role of various forms in the functional activity of inulinase molecules is discussed.

[Research on structural and functional characteristics (indicators) of apoptosis of human blood lymphocytes under the influence of carbon monoxide and ultraviolet (UV)-radiation].

The changes in the level of expression of membrane CD95 receptor, the structural state of DNA, parameters of biochemiluminescence of human blood lymphocytes under conditions of exposure to carbon monoxide (60-90 min) and UV-radiation (240-390 nm) at the doses of 151, 453 and 755 J/m2 have been investigated. The decrease in the level of expression of Fas-markers on the surface of immunocytes after incubation in the atmosphere at (60-90 min), the absence of changes in the DNA structure and the decreased intensity of lipid peroxidation processes in the cells were also found. It is established that UV-light (151-755 J/m2) exhibits a proapoptotic action, as evidenced by the increase in the expression of CD95 receptors on the surface of human blood lymphocytes and the decreased electrophoretic mobility of DNA in UV-irradiated cells. However, the process of a programmed cell death is not leaking ("apoptotic ladder" on electrophoregramme in the conditions of the experiment is missing). It is shown that the combined effects of carbon monoxide and UV-light in the cells of the human blood leads to a decrease in the sensitivity of the membrane CD95 receptors to the action of UV-radiation. It is revealed that carbon monoxide can contribute to blocking the processes of peroxide oxidation of lipids (POL) and, as a consequence, activate antioxidant properties of cells. It was concluded that the CO molecule in physiological concentrations is likely to have the antiapoptotic effect in relation to lymphocytes.

[Structural and functional properties of inulinases. Ways to regulate their activity].

The presented review is devoted to the analysis of structural and functional properties of inulinases from various producers. A special attention is given to the questions of molecular and permolecular organization of enzymes, description of their functional features depending upon their structural conditions and in various microenvironment. Perspectives of the development of biotechnological processes with the use of free and immobilized inulinases are discussed from a biophysical viewpoint.

[Effect of antibacterial agents of different nature on the osmotic resistance of human erythrocytes].

We have studied the structural properties of human erythrocytes under the action of antibacterial substances of different nature (macrolides, fluoroquinolones, tetracyclines, lincosamides). It has been established that antibiotics modify the cytoplasmic skeleton of erythrocytes and induce their hemolytic efficiency. The character of changes in the osmotic resistance of erythrocytes depends on the chemical structure of modifiers.

[Influence of UV-radiation on physical-chemical and functional properties of free immobilized glucoamylase from Aspergillus awamori].

The effect of UV light (240-390 nm) at the doses of 151-1510 J/m2 on the catalytic activity of free and im- mobilized glucoamylase from Aspergillus awamori was investigated. It was established that the loss of catalytic activity of the enzyme was associated with photochemical transformations of Trp-120, which is part of the active site of glucoamylase. It was shown that the use of collagen as a carrier for immobilization of glucoamylase reduces the constant photoinactivation of the enzyme. It was suggested that the protective effect of collagen in relation to the glucoamylase subjected to UV irradiation may be related to the acceptance of reactive oxygen species by collagen and the formation of the complex carrier-enzyme which is more photostable than the native enzyme.

[Influence of UV-light on the profile of receptors of lymphocytes of donated blood].

The influence of UV-light (240-390 nm) at the dozes of 151 and 755 J/m2 on the maintenance of membrane markers of lymphocytes: CD3, CD19, CD4, CD8, CD25, CD95 using the method of flow cytometry was investigated. It was demonstrated that UV-irradiation of lymphocytes induced the decrease in the number of CD3, CD4 and CD19 markers together with a simultaneous increase of the amount of CD8, CD25 receptors during the daily incubation of cells in a nutrient medium without plasma. When autologous plasma is used during incubation of both native and UV-irradiated lymphocytes, the percentage of cells with the specified markers is not different from the corresponding values in the cells immediately after their purification. This finding points at the important role of antioxidants of the blood plasma and growth factors for the maintenance of the sub-population structure of lymphocytes. However, the level of expression of CD95, one of the markers of apoptosis, increases in the photomodified cells incubated both in the presence and in the absence of plasma. This increase was revealed to be caused mainly by the synthesis of the protein de novo under the influence of apoptosis-inducing factors.

[Ways of apoptosis development in human lymphocytes, induced by UV-irradiation].

The level of DNA damage and cytochrome c content in human lymphocytes in the dynamics of apoptosis induced by UV-light (240-390 nm) at doses of 151, 1510 and 3020 J/m2 is studied. DNA fragmentation is revealed in 20 h after UV-irradiation of lymphocytes at doses mentioned above. It is shown that DNA damages (single strand breaks) appear immediately after UV-irradiation of lymphocytes at doses of 1510 and 3020 J/m2 (comets of C1 type) and reach their maximum 6 h after cell modification (comets of C2 and C3 types). It is concluded that p53-dependent and receptor caspase pathways are involved in apoptosis development in the human lymphocytes, modified after UV-irradiation.

[Spectral and functional properties of ceruloplasmin under UV irradiation].

During oxydase reaction spectral characteristics of ceruloplasmin at absorption of copper ions and protein part of the molecule are shown to change. It has been ascertained, that when irradiating ceruloplasmin by UV-light the functioning of intramolecular electron transport chain is broken, the degree of positive cooperativity (a Hill's constant) on substrate decreases. It is supposed, that these changes are caused by disturbance of interdomain interactions in a protein molecule.

[The effect of nuclear dyes on the frequency of aberrations in mucosal cells of humans].

The effect of staining with commonly used nuclear dyes (orsein, light green, methylene blue, azure-eosin by Romanovsky-Giemsa) on the frequency of occurrence of nuclear disorders (micronuclei, perinuclear vacuoles, notches, protrusions, such as "tongue" and "broken egg") in human buccal mucosa was determined. It is shown that Romanovsky-Giemsa dye is the best type of dye both in the quality of the micropreparations and the spectrum of detected abnormalities.

[The protective effect of autologous plasma against the development of oxidative stress in UV-irradiated lymphocytes of peripheral blood of donors].

The influence of UV-light (240-390 nm) at the dozes of 151 and 755 J/m2 on the intensity of processes of the lipid peroxidation, activity of lactate dehydrogenase (LDH), succinate dehydrogenase (SDH), cytochrome c oxidase and the level of the energy supply of donors' blood lymphocytes in the absence and presence of autologous plasma was investigated. It was shown that during the incubation of native and UV-irradiated lymphocytes, autologous plasma reduces the intensity of lipid peroxidation, thus protecting cells from oxidative stress. As a result, the endocellular level of ATP is restored in UV-irradiated lymphocytes (during the daily incubation), which reflects the intensification of the adaptive ability of cells in the presence of autologous plasma.

[Investigation of permolecular structure of lipase from Rhizopus niveus].

It has been shown by classical biophysical and biochemical methods in combination with atomic microscopy that lipase from Rhizopus niveus exists in a water solution as a dimer with a molecular weight of 96 kDa. The rate of splitting of triglycerides by a dimeric molecules is twice that of monomers. The heat stability of the monomeric form of lipase at temperatures of 20-60 degrees C is significantly higher than that of the native molecule.

[Ways of realizing apoptosis of human lymphocytes induced by UV-light and reactive oxygen species].

Changes of DNA structural condition, the level of membrane Fas-receptor expression, caspase-3 functional activity, concentrations of Ca2+, p53 and cytochrome c proteins of human lymphocytes in dynamics of apoptosis development induced by UV-light (240-390 nm) at doses 151, 1510, 3020 J/m2 and reactive oxygen species (superoxide anion-radical, hydroxyl radicals, hydrogen peroxide, singlet oxygen) have been studied. UV-light and reactive oxygen species have been established to induce fragmentation of lymphocyte DNA after 20 h incubation of the modified cells. It has been shown, that the increase in the expression level of membrane death Fas-receptors is observed during 1-5 h after exposure oflymphocytes to UV-light and ROS compared with intact cells. Also revealed is augmentation of lymphocyte caspase-3 functional activity 4 h after generation of singlet oxygen, hydroxyl radical and hydrogen peroxide addition, as well as 8 and 24 and 6 and 8 h after UV-irradiation of the cells at doses 151 and 1510 J/m2, correspondingly. Using DNA-comet method made it possible to tape that DNA damages (single-strand breaks) appear 15-20 min after lymphocyte UV-irradiation at doses 1510 and 3020 J/m and addition of hydrogen peroxide in concentration 10(-6) mol/l (C1 type comet) and reach their maximum 6 h after modification of the cells (C2 and C3 type comets). It has been observed, that 6 h after exposure oflymphocytes to hydrogen peroxide and UV-light at doses 1510 and 3020 J/m2, the p53 level of investigated cells raises. It has also been shown that the higher level of calcium in lymphocyte cytosol in conditions of UV-light exposure (1510 J/m2) and exogenous generation of reactive oxygen species is caused by Ca2+ exit from intracellular depots as a result of activating the components of the phosphoinositide mechanism for transferring information into a cell. Ideas about correlation between alterations of the calcium level and initiation of programmed cellular destruction of human lymphocytes after exposure to UV-irradiation and ROS is proposed. The authors come to the conclusion about the leading role of receptor-mediated (Fas-dependent) caspase- and p53-dependent ways of realizing apoptosis oflymphocytes induced by UV-light at doses 151 and 1510 J/m2 and active oxygen metabolites. The pattern of the possible intracellular events leading to apoptotic destruction of lymphocytes after their UV-irradiation is offered.

[Studies of physical and chemical properties of the vacuum UV-irradiated molecules of human hemoglobin and its components (heme and globin)].

The purpose of the present work is to study the vacuum ultraviolet radiation action on the optical and chromatographic characteristics of human hemoglobin molecules and their components - haem and globin. Using the methods of spectrophotometry and thin layer chromatography (TLC), we have investigated into the structural changes of molecules of human hemoglobin, haem and globin, induced by the influence of vacuum UV light (gamma = 118-134 nm, dose - 1.2 kJ/m2). It has been shown that vacuum ultraviolet radiation induces an infringement of the higher types of the spatial organization ofglobin molecules, thus leading to the changes in the structural state of the albuminous globule.

[Peculiarities of metabolism of UV-irradiated lymphocytes].

The influence of UV-light (240-390 nm) at a dose of 151 and 755 J/m2 on the functional properties of lymphocyte metabolism key enzymes from donors' human blood: lactate dehydrogenase (LDH), cytochrome c oxidase, succinate dehydrogenase (SDH), Ca2(+)-ATPase of plasma membranes has been investigated. It has been revealed that photoinactivation of enzymes immediately after UV-irradiation which leads to the decrease of the ATP content in lymphocytes is replaced by the increased activity of the enzymes under investigation during daily incubation of lymphocytes. As a result, the level of ATP in photo-modified lymphocytes does not differ from that in native cells before incubation. This indicates the normalization of biochemical processes in lymphocytes influenced by UV-light applied in autotransfusion of UV-irradiated blood.

[Level of receptor complex molecule expression of human blood T-lymphocytes under conditions of their UV-irradiation].

The influence of a wide range of doses of UV-light (240-390 nm) on the expression level of receptor complex molecules (CD3, CD4, CD8 markers) on the membrane surface of human blood T-lymphocytes has been studied using flow cytometry and enzyme linked immunosorbent assay. UV-light at small and medium doses (15 1, 453 and 906 J/m2) has been established to have a unidirectional (activating) effect on the expression level of receptor complex molecules, and at a high dose of 1359 J/m2 it can either increase (CD4 and CD8 markers) or reduce (CD3 complexes) the quantity of the analyzed molecules on the membrane surface of T-lymphocytes.

[Fragmentation of DNA lymphocytes the human in dynamics of development apoptosis, induced influence of UV-radiation and reactive oxygen species].

It is established that UV-light (240-390 nm) in doses of 151, 1510 and 3020 J/m2 and reactive oxygen species and singlet oxygen induce DNA fragmentation lymphocytes cells of the human 20 h after influence. Using a method of DNA-comets it is revealed that DNA damages (single strand breaks) are found out right after UV-irradiations of lymphocytes in doses of 1510 and 3020 J/m2 and additions hydrogen peroxide in concentration of 10-6 mol/l (a comet of type C1) and reach a maximum through 6 h after influence on of cells UV-light and ROS (comets of types C2 and C3). Assumption about the leading part of a p53-dependent way in realization apoptosis human lymphocytes in the conditions of influence of UV-light and reactive oxygen species is put forward.

[Thermal inactivation of free and immobilized inulinase].

Thermal inactivation of the Kluyveromyces marxianus inulinase in a free form and immobilized on VION KN-1 cation exchange fiber was studied. Atomic force microscopy demonstrated an oligomeric structure of this enzyme, composed of two subunits differing in their size. It was assumed that the intersubunit contacts were destroyed at 60 degrees C, and the inulinase molecule dissociated into two monomers located separately.

[Receptor caspase-dependent and caspase-independent ways of human lymphocytes apoptosis induced by UV-radiation].

Changes of a structural condition of DNA, expression level of the membrane Fas-receptor and of functional activity of the human lymphocytes caspase-3 in dynamics of apoptosis development, induced by influence of UV-light (240-390 nm) in doses 151, 1510 and 3020 J/m2 are studied. It is established, that the UV-irradiation of lymphocytes in doses of 151 and 1510 J/m2 causes realisation of receptor caspase-dependent apoptosis way. The assumption about switching possibility of receptor caspase-dependent way apoptosis on receptor caspase-independent way at increase of a dose of UV-light to 3020 J/m2 is made.