Low-Intensity Femtosecond Radiation Activates the Natural Defenses of Mice in vivo.

The possibility of induction of cytogenetic damage in the bone marrow, changes in the cellularity of lymphoid organs and blood composition in mice irradiated with low-intensity femtosecond laser radiation at a power flux density of 5.1, 10.4, and 52 mJ/cm2 (0.5 mW for 5, 10, and 50 s) in vivo was shown. Using the radiation adaptive response test (0.1 Gy + 1.5 Gy), it was found that, when mice were exposed to femtosecond laser radiation in high doses, the body's natural defenses were activated in the same narrow range of energy flux density (2-16 mJ/cm2) as in the case of X-ray irradiation in a dose of 0.1 Gy (4 mJ/cm2). The data obtained suggest a similar mechanism of activation of the body's natural defense upon exposure to low doses of both ionizing and non-ionizing radiation.

A Study of the Cytotoxic Effect of Microcapsules and Their Constituent Polymers on Macrophages and Tumor Cells.

We studied the effect of different concentrations of polyelectrolytes poly(allylamine hydrochloride) (PAH) and polystyrene sulfonate (PSS) as well as the effects of microcapsules coated with these polymers on survival of Ehrlich ascites carcinoma cells and mouse peritoneal macrophages and on ROS production by phagocytes. PAH reduced viability of Ehrlich ascites carcinoma in a concentration-dependent manner (LD50=12-15 µg/ml). This effect was presumably determined by its ability to bind phosphates, thereby depleting the culture medium. At the same time, PAH did not affect the viability of macrophages. PSS produced no cytotoxic effect on the examined cells. Polyelectrolyte capsules with the shell architectonics (PAH/PSS)3 and (PAH/PSS)3PAH in the examined concentration range had no effect on the viability of macrophages and tumor cells. PAH microcapsules with positively charged surface much more rapidly and more intensively activated macrophages. The chemiluminescence response directly depended on the amount of capsules in the solution.

Effects of Polyelectrolyte Microcapsules with Different Surface Charge on Erythrocyte Sedimentation Rate.

Relationship between changes in the erythrocyte sedimentation rate in rats and concentration and charge of polyelectrolyte microcapsules was studied by the Panchenkov method. Positively charged microcapsules reduced erythrocyte sedimentation rate in a concentrationdependent manner. This effect was related to a decrease in the content of high-molecularweight proteins in the plasma due to their adsorption in positively charged microcapsules with polyacrylamide surface layer.

[DYNAMICS OF NITROGEN OXIDE METABOLITES IN THE PLASMA AND ASCITES DURING ZAJDEL HEPATOMA GROWTH IN VIVO].

The dynamics of extracellular nitrogen oxide metabolites localized in the plasma and ascites during Zajdel ascites hepatoma growth in the abdominal cavity has been investigated. An increase in peroxynitrite concentration was found by the levels of nitrotyrosine (up to 10-11 nM) in blood plasma at the initial stage of tumor cell development. In the course of further tumor development, an oxidative stress developed, which might cause oxidation of protein components including tyrosine. All these processes may cause a decrease in the accessible amount of tyrosine for nitration and lead to a fall in nitrotyrosine level (to 3-6 nM) at the final stages of tumor growth. Nitrotyrosine dynamics in the region of tumor growth is essentially analogous to that in the plasma because proteins during tumor growth cames from the blood plasma of tumor bearer. In studying the dynamics of nitrosylation of sulfur-bearing protein groups, an increase in the concentration of S-nitrosothyols was found to occur in the blood plasma for up to 6 days of the experiment, subsequently their concentration decreased. In the ascites, where protein R-SNO arrives, the mean concentration of nitrosothyols upon tumor growth is lower compared to that of the plasma. In studying the dynamics of final stable nitrogen oxide decay products--nitrites/nitrates, it has been found that during tumor development the concentration of these metabolites in the plasma varies only moderately within some range and sharply increases at the final stage of the experiment. In the area of tumor growth, an analogous trend in the behavior of nitrites/nitriaes has been registered (noted, marked), but with a higher background level, which might be due to both the functioning of immunocompetent cells, microphages in particular, and a decreased rate of utilization of substances from the ascites. Based on the aforesaid, it has been concluded that the nitrosylating stress in the organism of the bearer of a tumor is being developed along with the oxidative stress.

[Effect of Zajdela ascite hepatoma growth on the extracellular antioxidant system of tumor bearer].

The influence of tumor development on the blood plasma antioxidant system in the area of tumor growth has been studied. Zajdela ascite hepatoma transplanted to the abdominal cavity of Vistar rats was used as a model of tumor growth. It hase been found that tumor development produced and imbalance between pro- and antioxidant systems in the organism of tumor bearer. Besides, a sharp decrease in tocopherol and uric acid concentrations (twice), as well as in the concentration of protein SH-groups (seven times) was noted. In the tumor growth area, along with the tocopherol level decrease, a 5-7 fold increase in the concentrations of uric acid and protein SH-groups was observed. As the concentration of low-molecular antioxidants decreases, the major part is played by protein components which bind or oxidize ions of variable valence. Thus, the level of transferrin (Tf, which is responsible for the transport of iron ions, is reduced in 2.5 to 3 times (from 5.0 to 1.6 mg/ml) in the blood plasma, whereas the Tf level in the ascitic fluid increased from 1.5 to 2.7 mg/ml. The concentration dynamics of the other protein functioning together with Tf, ceruloplasmin (Cp), had opposite (inverse) tendencies. Thus, the Cp concentration in the blood plasma increased 1.5-2 times (from 0.55 to 1.1 mg/ml) whereas it decreased from 0.55 mg/ml to 0.35 mg/ml in the ascites.

[Effects of a nitric oxide donor, the substratum of NO-synthase, and an inhibitor of NO-synthase on the ROS-generation activity of phagocytes in the course of ascites tumor growth].

Experiments in vitro were performed to investigate the effects of the nitric oxide donor (SNP), the substratum of NO-synthase (L-arginine), and the inhibitor of NO-synthase (nitroarginine) on the ROS-generating activity of blood plasma polymorphonuclear leucocytes and ascitic fluid macrophages isolated at different times of tumor (Zaidel hepatoma) growth in animal organism. It was found that in the initial period of tumor growth the nitric oxide donor at a concentration of 8 x 10(-5) M reduced the potential ROS-generating activity of macrophages by 38.5 +/- 9.0% and that of polymorphic-nuclear leucocytes of plasma by 27.6 +/- 7.0 %. However, the dynamics of this process during tumor growth was conservative: variations in the production of ROS by phagocytes were 10 +/- 3.0%. L-arginine induced a decrease in the ROS-generating activity of granulocytes and mononucleares by 25-30%. This fact points to an inducible inhibiting effect of NO-synthase on the ROS-generating activity of NADPH-oxidase in the course of tumor growth. The inhibitor of NO-synthase, nitroarginine, produced a monotonous increase in the ROS-generating activity of phagocytes isolated from the tumor at different periods of its growth. The use NO-synthase inhibitors for increasing ROS levels in the region of tumor growth may favor the suppression of tumor cell growth in vivo.

[Beneficial action of the liposomal form of dihydroquercetin on the process of skin regeneration after thermal burn].

It was found that the flavonoid of plant origin, dihydroquercetin, being localized in lecitin nanoparticles in the presence of amino acid, reduced inflammatory reactions in the wound zone after thermal burn. Application of the liposomal complex on burn trauma stabilized the endogenous antioxidant system and minimized the area of the secondary necrosis in burn wound. Also, intensification of skin regeneration and repair of hair follicles and sebaceous glands were observed.

[Variations in cell population size and reactive oxygen species level in the blood and the ascites liquid of tumor carrier].

Using Seidel ascites hepatoma as a model, we studied in detail changes in cell population size and in the level of reactive oxygen species in the tumor growth zone and in the blood plasma of tumor carrier. It was found that reduction-oxidation conditions in the blood plasma and in the tumor growth zone were different. Thus, because of hyperactivity and increase in the number of leukocytes, the blood plasma exhibited strong oxidative stress inducing damage to healthy cells, whereas the tumor growth zone showed the decrease in macrophage concentration, as well as in oxygen and ROS levels. These conditions favor intensive growth of tumor cells.

Effects of a nanocomplex containing antioxidant, lipid, and amino acid on thermal burn wound surface.

Dihydroquercetin (flavonoid of plant origin) immobilized with an amino acid in lecithin nanoparticles promotes reduction of inflammatory reactions in the wound after thermal burn. The use of a liposomal complex in burn injury stabilizes endogenous antioxidant system and limits the secondary necrotic zone in the wounds. The treatment was associated with intensification of skin regeneration processes and reparation of hairy follicles and sebaceous glands.

[Effects of weak magnetic fields on the production of reactive oxygen species in peritoneal neutrophils in mice].

The influence of weak magnetic fields of different types on the rate of the formation of reactive oxygen species in mouse peritoneal neutrophils has been studied. It was found that the exposure of neutrophils activated by phorbol 12-myristate 13-acetate to the magnetic field tuned to the parametric resonance for Ca2+ ions leads to a decrease in the rate of the reactive oxygen species (ROS) generation by 23%. Conversely, the generation of ROS in neutrophils exposed to the same field but stimulated by the bacterial peptide FMLP (N-formyl-L-methionyl-L-leucyl-L-phenylalanine) increased by about 21%. Pulsed magnetic fields also changed the rate of ROS generation in phorbol-stimulated neutrophils by about 20%, but the sign of the effects observed in this case was opposite to those induced by the magnetic field tuned to the parametric resonance for Ca2+ ions.

Effect of liposomes containing antioxidant, phospholipid, and amino acid on skin regeneration after chemical burn.

Liposomes containing natural flavonoid dihydroquercetin, phospholipid lecithin, and zwitterionic amino acid glycine were used for the therapy of glacial acetic acid-induced chemical burn. No inflammatory reaction was observed in the region of wound. The area of postburn injury did not increase in time. Planimetry and histological studies showed that improved skin regeneration was accompanied by repair of hair follicles and sebaceous glands.

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.

[A comparison of antioxidant properties of hypoxen and duroquinone by the method of chemiluminescence].

The antioxidant activity of duroquinone and hypoxen was compared with that of alpha-tocopherol and vitamin C in a model system (luminol-peroxidase-H2O2), and their influence on the level of reactive oxygen species in systems containing polymorphonuclear leukocytes of healthy and tumor-bearing animals was studied. It was shown that, in a model chemical system, the concentrations of antioxidants (the inverse of antioxidant activity) necessary to decrease twice the intensity of the chemiluminescence answer (C50%) are arranged in the following order: alpha-tocopherol > duroquinone > hypoxen > ascorbic acid. In this case, the concentrations of the hydrophobic antioxidants (C50% for alpha-tocopherol and duroquinone 10-30 mkM) should be 20-50 times higher than for hydrophilic antioxidants (C50% for vitamin C and hypoxen 0.5-0.6 mkM). It was revealed that the generation of reactive oxygen species by blood phagocytes of tumor-bearing animals is 2-2.5 times higher than by phagocytes of healthy animals. The antioxidant concentration necessary to decrease the chemiluminescence answer in the cellular system should be one order of magnitude higher than in the model chemical system. The distribution of a hydrophobic antioxidant between water/lipid phases promotes an increase in the concentration of the antioxidant necessary to decrease the level of reactive oxygen species twice. Thus, the major factor influencing the antioxidant activity is the constant of distribution of these compounds in a water/lipid system.

[Changes of the goldfish Mauthner neuron ultrastructure and function under the influence of 3,4-dihydro-2(1H)-pyrimidinethione].

The effects of pyrimidine derivative 3,4-dihydro-2(1H)-pyrimidinethione, (DPT) used as a test-system for detection of tumor growth, on the goldfish Mauthner neurons (MN) ultrastructure and function, as manifested in behavioral changes, were studied. The results of investigations demonstrated that an application of DPT on MN had the effects similar to those of dopamine application, as established earlier, causing the enhancement of MN resistance to fatigue stimulation, accompanied by an increase of the dimensions of the actin containing desmosome-like afferent admembranous synaptic contacts, and formation of the cytoplasmic bundles of actin stress-fibers. Similarity of morpho-functional changes of MN, induced by DPT, an artificial chemical substance, which has no receptors on the neuronal membrane, and by natural neurotransmitter dopamine, allows us to suggest possible trophic stabilizing and polymerizing effects of both substances on cytoskeletal actin due to their direct penetration into postsynaptic neuron.

[Cytotoxicity of amyloid fibrils of X-protein].

It is known that amyloid oligomers, protofibrils, and fibrils induce cell death, and antibiotic tetracycline inhibits the fibrillization of beta amyloid peptides and other amyloidogenic proteins and disassembles their pre-formed fibrils. Earlier we have demonstrated that sarcomeric cytoskeletal proteins of the titin family (X-, C-, and H-proteins) are capable to form in vitro amyloid fibrils, and tetracycline effectively destroys these fibrils. Here we show that the viability of polymorphonuclear leukocytes in the presence of X-protein amyloids depends on the concentration of amyloid fibrils of X-protein and the time of incubation. In addition to the disaggregation of X-protein fibrils, tetracycline eliminated the cytotoxic effect of the protein. The antibiotic itself did not show a toxic effect, and the cell viability in its presence even increased. Our results evidence the potential of this approach for evaluating the effectiveness of drugs preventing or treating amyloidoses.

[Cytotoxic action of polymorphonuclear leukocytes on tumor and normal cells during ascite tumor development in vitro and in vivo].

A vast number of studies, including the authors' own research, support the important role polymorphonuclear leukocytes (PMNL) in the development of ascite tumors. The method of luminol-dependent chemiluminescence (CL) was used to show the presence of two functionally different PMNL pools in a tumor-bearing organism: 1) "primed" PMNL, which circulate in the blood stream, and 2) "activated" PMNL, which are accumulated in the tumor zone and are capable of spontaneous CL. The purpose of the present investigation was to compare cytotoxic effects of primed and activated PMNL on tumor cells (ascite Ehrlich carcinoma (AEC), ascite Zajdel hepatoma) upon co-cultivation, as well as on normal cells of the organism, erythrocytes in vitro and in vivo. Upon stimulation with phorbol myristate acetate (PMA), PMNL effectively damaged AEC cells within the first 24 h until PMNL apoptosis occurred. Upon further co-cultivation, the tumor cells grew in number, which suggest the participation of PMNL in tumor protection. When stimulated with PMNL, pools suppressed tumor growth in vitro, since in this case the cytotoxicity was due to both reactive oxygen species and proteolytic enzymes. As it has been shown earlier by the authors, the functional potential of PMNL increases many times during tumor growth, and we suggested that not only tumor but also normal cells could be damaged. In this connection, we have studied the cytotoxic effect of primed and activated PMNL on rat erythrocytes in vitro on their co-cultivation. On stimulation with PMA, the rate of lysis of erythrocytes by primed PMNL increase many times compared to the norm. The fMLP-stimulated cytotoxity was 1.5-2.0 times higher than in the norm. Activated PMNL without stimulation are capable of producing only a partial lysis of erythrocytes (5-7 %). In order to assess the cytotoxic action of PMNL on erythrocytes in vivo, the hemoglobin content in erythrocytes and blood plasm of rats was measured in the course of tumor growth. The hemoglobin content in erythocytes during growth tumor decreased from 135 +/- 10 to 85 +/- 5 g/l, whereas in the blood plasm the hemoglobin content gradually increased by almost two times. The results enable us to suggest that one of death causes of tumor-bearing organisms may be the cytotoxic action of PMNL on normal cells of the organism caused by hyperproduction of ROS.

Combined vitamins Bl2b and C induce the glutathione depletion and the death of epidermoid human larynx carcinoma cells HEp-2.

The combination of hydroxocobalamin (vitamin B12b) and ascorbic acid (vitamin C) can cause the death of tumor cells at the concentrations of the components at which they are nontoxic when administered separately. This cytotoxic action on epidermoid human larynx carcinoma cells HEp-2 in vitro is shown to be due to the hydrogen peroxide generated by the combination of vitamins B12b and C. The drop in the glutathione level preceding cell death was found to be the result of combined action of the vitamins. It is supposed that the induction of cell death by combined action of vitamins B12b and C is connected to the damage of the cell redox system.

[Glycoproteins of liver cell membrane]. / Glikoproteiny kletochnoi membrany pecheni.

Plasmatic and mitochondrial membranes of rat liver cells were isolated by centrifugation in sucrose density gradient, enzyme analysis of the obtained fractions being performed. Membrane proteins and glycoproteins were examined using electrophoresis in polyacrylamide gel, and changes in glycoprotein composition of plasmatic and mitochondrial membranes being demonstrated. Quantities and molecular weights of the glycoproteins were estimated.

[Generation of reactive forms of oxygen by polymorphonuclear leukocytes during hepatoma growth in the peritoneal cavity of animals]. / Generatsiia reaktivnykh form kisloroda polimorfnoiadernymi leikotsiamiv protsesse razvitiia gepatomy v briushnoi polosti zhivotnykh.

In the present work, an attempt was made to analyse generation of reactive oxygen species (ROS) by polymorphonuclear leucocytes (PMN) in the course of tumour growth, using chemiluminescence (CL). A multiple increase in the capacity of polymorphonuclear leucocytes of generating active forms of oxygen in the course of tumor growth was discovered. Two causes of this process were found. 1) the increase in specific activity of leucocytes; 2) the increase in the total quantity of PMN circulating in the blood. Leucocytes were also found in the ascite liquid. PMN leucocytes were shown to participate in the antitumor defence of the organism.

[The effect of catecholamines on the luminol-dependent chemiluminescence of rat polymorphonuclear leukocytes]. / Vliianie katekholaminov na liuminolzavisimuiu khemiliuminestsentsiiu polimorfnoiadernykh leikotsitov krysy.

The effects of catecholamines on reactive oxygen species (ROS) generation by polymorphonuclear leucocytes were studied using chemiluminescence (CL). It has been found that catecholamines in 10(-9)-10(-10) M concentration range are able to increase reactive oxygen species generation. Within the micromolar concentration range (10(-6)-10(-5) M) cetacholamines (CA) strongly suppress CL intensity and ROS production. The efficiency of inhibitory action of CA: noradrenaline > adrenaline > dopamine > L-DOPA correspond to their redox properties. A conclusion is made that peroxidase is one of the main targets of CA effect.