E. B. Burlakova

Biomarkers of oxidative stress and smoking in cancer patients

CONTEXT Increased oxidative stress is a significant part of pathogenesis of smoking-related cancer. AIM The study aims to investigate changes in antioxidant status induced by chronic cigarette smoking in cancer patients and healthy subjects. SETTING AND DESIGN We examined the venous blood samples of 54 healthy subjects, both smokers (25) and non-smokers and of 50 patients with smoking-related cancer, both smokers (34) and non-smokers. MATERIALS AND METHODS We measured the activities of five antioxidant (AO) enzymes: glutathione peroxidase, glutathione transferase (GST), glutathione reductase, superoxide dismutase and catalase in the blood of 50 cancer patients and 54 healthy persons. Damage to cellular structures (level of malonic dialdehyde, micro viscosity of erythrocyte membranes, number of leukocyte DNA breaks) was determined. Statistical analysis of results obtained was performed using conventional and multi-factorial statistical methods. RESULTS Statistically significant increase in GST activity and DNA breaks, but decrease of membranes micro viscosity in cancer patients, compared with healthy subjects were obtained. In the cancer patients, no influence of smoking on studied parameters was found. Correlations of parameters within cancer patients and healthy subjects group did not coincide with each other. CONCLUSIONS Changes of AO status parameters and oxidative damages in cell structures are related to tumor processes indicating the augmentation of oxidative stress in human blood. This study demonstrated potential applicability of a statistical model based on the evaluated biomarkers of oxidative stress to determine a smoking-induced harm of cancer incidence in healthy subjects.

Effect of oregano essential oil on the engraftment and development of Lewis carcinoma in F1 DBA C57 black hybrid mice

The effect of a low uptake dose of oregano essential oil with drinking water for three months (Origanum vulgare L.) on the degree of Lewis carcinoma engraftment and some parameters of oxidative stress has been studied in vivo using F1 DBA C57 Black hybrid mice. Oregano essential oil has been established to possess an anticancer activity. The degree of tumor engraftment decreased by 1.8 times, its size decreased by 1.5 times, and the development of tumor was significantly suppressed in sick mice under the effect of oregano essential oil. It was found that the uptake of essential oil did not affect the intensity of lipid peroxidation in the brain of mice and resulted in a significantly (by 36%) decreased content of secondary lipid oxidation products in the liver as shown in a reaction with thiobarbituric acid as compared to control subjects. The activity of antioxidant enzymes was found to increase after three months of essential oil uptake (by 1.5–3 times) as compared to the control group. This effect of essential oil supports the presence of bioantioxidant properties in this essential oil.

[Comparison of the antiradical activity of ionol, components of fresh ginger, and its extracts].

The antiradical properties of three samples of ginger (Zingiber officinale R.)—juice from fresh rhizome, essential oil, and extracts (oleoresin)—were studied and compared with the properties of synthetic antioxidant ionol (butylatedhydroxy-toluene, BHT). Reaction antioxidants with stable free 2,2-diphenyl-1-picrylhydrozyl radicals were used as model systems. DPPH equivalents per gram of ginger sample, EC50, and antiradical efficiency (AE) were determined. The EC50 and AE values for ginger oleoresin and BHT were similar. They were the same as those of highly active natural antioxidants, and the values for essential oil and ginger juice were lower by two orders of magnitude. On the base of kinetic parameters, the ginger samples may belong to antiradical compounds with prolonged action.

Effect of antioxidant β-(4-hydroxy-3,5-ditertbutylphenyl)Propionic acid (phenosan) on the development of malignant neoplasms

The effect of different doses of synthetic antioxidant β-(4-hydroxy-3,5-ditertbutylphenyl)propionic acid (phenosan) on the development of spontaneous leukemia in AKR mice was studied. The drug efficiency was determined from the survival curves, animal life spans, and the incidence of leukemia. Phenosan exhibited a pronounced antitumor activity at therapeutic (10−4 mol/kg, 4 administrations) and ultra-low (10−14 mol/kg, 4 administrations) doses. The dose of 10−4 mol/kg proved most efficient to increase the life span of the short-lived subpopulation, while the dose of 10−14 mol/kg increased the life span of the long-lived subpopulation. The ultra-low dose of the drug seems promising as a prophylactic agent.

[Effects of low doses of essential oil on the antioxidant state of the erythrocytes, liver, and the brains of mice].

We studied the effects of essential oils from oregano and clove and a mixture of lemon essential oil and a ginger extract on the antioxidant state of organs in intact and three experimental groups of Balb/c mice. We found that in vivo essential oils were efficient bioantioxidants when mice were treated with it for 6 months even at very low doses, such as 300 ng/day. All studied essential oils inhibited lipid peroxidation (LPO) in the membranes of erythrocytes that resulted in increasing membrane resistance to spontaneous hemolysis, decreasing membrane microviscosity, maintenance of their integrity, and functional activity. The essential oil significantly decreased the LPO intensity in the liver and the brain of mice and increased the resistance of liver and brain lipids to oxidation and the activity of antioxidant enzymes in the liver. The most expressed bioantioxidant effect on erythrocytes was observed after clove oil treatment, whereas on the liver and brain, after treatment with a mixture of lemon essential oil and a ginger extract.

Inhibition of mouse aging by using an essential oil composition

167 Due to increasing life expectancy, the proportion of older people in the population increases, and the social importance of cognitive abilities is growing steadily. It is well known that, with age, the vast major ity of older people develop persistent pronounced cog nitive deficits that leads to disability and household independence and is defined as mild cognitive impair ment (MCI) [1]. This syndrome is a transitive condi tion between the physiological decline in cognitive function due to aging and the initial (preclinical) stage of Alzheimer’s disease (AD). The syndrome is charac terized by a borderline disorder of cognitive functions, including memory. In general, the MCI syndrome is extremely widespread. Since AD can be diagnosed in 40% of patients within three years after the diagnosis of MCI, an early diagnosis of the MCI syndrome and the prescription of appropriate drugs can delay the onset of dementia or even promote regression of cog nitive impairment characteristic of the early stages of AD [2].

Imbalance in the Enzymatic System of Production and Consumption of Active Oxygen Species in Liver of AKR Mice with Spontaneous Leucosis

Activities of protective antioxidant enzymes, the rate of superoxide formation (v) in microsomal membranes and submitochondrial particles (SMP), and the concentrations of reduced and oxidized glutathione in cytosol were studied in the liver of AKR mice during the development of spontaneous leucosis. It was found that in the latent period of leucosis (mice of 3-6 months of age) the glutathione reductase (GR) activity in cytosol and mitochondria decreased and v in SMP increased. The increase in v in SMP did not result in the induction of Mn-SOD. In this stage of leucosis, the activities of Cu,Zn-SOD, GSH-Px, and G-6-PDH in cytosol were unchanged; at the same time, the GR activity and the concentration of reduced glutathione smoothly decreased. In the stage of developed leucosis (mice of 7-9 months of age), non-synchronous changes in the antioxidant system resulting in the shift of metabolism towards the prooxidant state were found. Comparison of our findings and the literature data demonstrates that the observed decrease in the SOD/GSH-Px ratio, the decrease in GR activity, and the increase in the v/Mn-SOD activity ratio are typical for pre-neoplastic changes in cell metabolism.

The effect of volatile antioxidants of plant origin on leukemogenesis in mice

The effect of savory essential oil added with drinking water (150 ng/ml) or with feed (2.5 μg/g) on the lifetime of AKR mice and the parameters of oxidative stress in animal blood were investigated. It was found for the first time that long-term administration of an essential oil in low doses increased the average lifetime of mice by 20–35% and was accompanied by a decrease in the hemolysis level and the content of lipid peroxidation products in erythrocytes of mice, as well as alteration in the structural state of their membranes and stabilization of polyunsaturated fatty acids level in mice liver cells.

Alterations in fatty acid composition of mice brain and liver with aging and at savory essential oil administration

80 The ratio of saturated, monounsaturated, and polyunsaturated fatty acids in cell membrane lipids determines their common structural characteristics and functions. Changes in the composition of fatty acids affect the activity of receptors, transport of metabolites into and out of cells, as well as hormonal and other signal transduction processes [1, 2]. Upon aging, the content of polyunsaturated fatty acids in lipids (especially in the brain) decreases, which affects its function [1–3]. The results of some experiments in animals showed that the addition of essential oils to food of laboratory animals stabilized the levels of poly unsaturated fatty acids in brain cell membranes [4–6]. This is due to the fact that essential oils of many aro matic plants exhibit antioxidant activity and can be used for prevention and treatment of diseases caused by oxidative stress [7–9]. For example, it was found that some essential oils inhibited the growth of cancer cells [7, 8]. The essential oils that exhibit antioxidant properties are now considered as promising objects for experimental verification of their anticancer activity [7, 8]. We have shown earlier that daily administration of 0.3–0.5 μg of savory essential oil increased by 20% the lifespan of mice of the high cancer strain AKR, which develop spontaneous leukemia with age in nearly 100% of cases, as well as reduced the incidence of leu kemia by 35% [10]. It should be noted that murine spontaneous leukemias are most similar to human leu kemia in origin, clinical manifestations, and patholog ical and morphological features [11]. The purpose this work was to study the alterations occurring in the fatty acid composition in the liver and brain of mice of the high cancer strain AKR with age as well as to assess the effect of the essential oil of the summer savory (Satureja hortensis L.; Lionel Hitchen Ltd., United Kingdom), which was given to mice with drinking water, on the fatty acid composition of these organs. This study was performed with AKR mice (which develop spontaneous leukemia) that were three months of age at the beginning of the experiment. The control group of mice received normal drinking water; the experimental group, water supplemented with savory essential oil (0.15 mg per liter); i.e., each mouse received approximately 0.3 μg of savory essential oil per day. The experiment continued for ten months. The development of leukemia in animals was deter mined by an increased size of the thymus (30 mg) and spleen (greater than 150 mg). For biochemical studies, we used the brain and liver of mice at an age of four, six, eight, and ten months (the control group) and at an age of 4 and 6 months (the experimental group). The fatty acid composition of brain and liver cells was studied by gas chromatog raphy and gas chromatography–mass spectrometry. Lipids isolated from organs were subjected to acidic methanolysis. Fatty acid methyl esters (FAMEs) were analyzed using a Crystal 2000 M chromatograph (Russia) equipped with a flame ionization detector and a DB 1 quartz capillary column (50 m × 0.32 mm, 0.25 μm phase layer; Supelco, United States). The temperature of the column was programmed to change from 120 to 270°С at a rate of 4°C/min. The content of FAMEs in samples was calculated as the ratio of the relative peak area of a corresponding acid to the sum of peak areas of all fatty acids and expressed in percent. Data were processed and averaged using the ANOVA software at p < 0.05. Plots were con structed and processed using the Origin 7.5 program. Mice of the high cancer AKR strain suffered from leukemia upon ageing. This disease develops sponta neously with different rate in different animals. It is determined by the content of leukocytes and the weight of the thymus and spleen. Judging by these parameters, mice in out study became ill with leuke mia after eight months of age; this disease was pro Alterations in Fatty Acid Composition of Mice Brain and Liver with Aging and at Savory Essential Oil Administration

Study of the effect of melafen at low doses on animal malignant neoplasms in vivo and in vitro

76 Plant growth regulator melafen (melamine salt of bis (oxymethyl)phosphinic acid) [1, 2] effectively stimulates plant growth at low concentrations but sup presses it at high doses, i.e., has opposite effects on proliferating cells. With allowance for the fact that proliferation is an inherent characteristic of malignant tumor cells, it was of interest to study the effect of melafen on the development of experimental malig nant neoplasms in vivo and in vitro in animals. An ear lier study of the effect of melafen on the structural and functional characteristics of animal cells showed that melafen affects two targets on cell surface—PY2 puri nergic receptors and capacity entrance Ca2+ channels and significantly decreased their activity [3]. In Ehr lich ascites carcinoma (EAC), which was used as a mode, melafen suppressed the Ca2+ signaling system even at concentrations 10–11 to 10–10 M. It is known that the Ca2+ signaling system in cells affects S100 calcium binding proteins, which bind to the p53 reg ulatory protein and change its transcriptional activity, thereby ensuring apoptotic signal transduction. The S100B protein has also opposite effects (protective at nanomolar concentrations and proapoptotic at micro molar concentrations).