[Features of free radical processes in the liver of rats with a nutrient imbalance]. / Intensivnost' svobodnoradikal'nogo okisleniia biomolekul mitokhondrii gepatotsitov pri nutrientnom disbalanse.

The activity of free radical processes in liver mitochondria was investigated in rats kept on high-sucrose and low protein/high-sucrose diets. Excess of dietary sucrose caused intensification of free radical processes in liver mitochondria as evidenced by increased hydroxyl radical generation, accumulation of primary (conjugated dienes, ketodienes) and secondary products (TBA-reactive products) of lipid peroxidation, increased cholesterol/phospholipids ratio and also accumulation of oxidative modification products of proteins (carbonyl derivatives). Additional nutritional protein deficiency (low protein/high-sucrose diet) enhanced destructive changes in liver mitochondria. This suggests a critical role of nutrient protein supplementation for maintaining the functional activity of mitochondria. The established changes can be considered as one of possible mechanisms of functional liver activity violation in conditions of nutrient imbalance.

[Biochemical markers of the functional state of liver in rats fed diets with different protein and sucrose content].

The study of mechanisms of the metabolic disorders in conditions of deficiency or excess of individual nutrients in the diet is a live issue. The influence of the simultaneous excess sucrose intake and protein deficiency in the diet on the functional state of the liver remains poorly understood. The aim of the research was to study the rate of generation of the superoxide radicals, the content of triglycerides and glycogen in the liver, as well as the activity of enzymatic markers of the liver state in rats fed diets with different protein and sucrose content. Material and methods. The studies were conducted over 28 days on 48 white non-linear rats, randomized into 4 groups: 1 - animals fed full-value semi-synthetic ration (14% protein); 2 - animals receiving low-protein ration (4.7% protein); 3 - animals receiving high-sucrose diet (40% sucrose), 4 - animals receiving low-protein high-sucrose diet. Serum sorbitol dehydrogenase activity was determined by the kinetic method in the reaction of NADH-dependent reduction of D-fructose to D-sorbitol. Serum alanine aminotransferase activity and aspartate aminotransferase was evaluated using a kit of reagents (Ukraine). Results and discussion. It was found that in rats fed low protein diet, no changes in the de Ritis coefficient were observed, while the activity of sorbitol dehydrogenase in blood serum increased 1.7 fold. However, no changes in the rate of superoxide radical formation, as well as glycogen and triglyceride level in the liver were observed. In animals fed highsugar diet, a rise in the de Ritis coefficient on the background of increased serum sorbitol dehydrogenase activity (more than 3.5 times) was revealed. At the same time, the rate of the superoxide radical formation in the liver mitochondria enhanced by 3 fold, with an increased accumulation of glycogen and triglycerides. The most pronounced changes in liver state were observed in animals fed low-protein/high-sugar diet: a marked increase in the de Ritis coefficient with a 5-fold increase in the activity of sorbitol dehydrogenase, and a 6-fold elevation in the intensity of the superoxide radical generation in liver mitochondria. The triglyceride content in the liver doubled, while the glycogen content remained at the level of control values. Conclusion. The data obtained represent disturbances of the functional liver state as a consequence of the relatively short-term excessive consumption of sucrose, especially in combination with a alimentary protein deficiency. It was found that the leading factor in the formation of destructive changes in the liver was excessive sucrose consumption, while the concomitant protein deficiency exacerbated the functional changes in hepatocytes.

[Activity of liver mitochondrial NAD+-dependent dehydrogenases of the krebs cycle in rats with acetaminophen-induced hepatitis developed under conditions of alimentary protein deficiency].

Activity of isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, malate dehydrogenase, and the NAD(+)/NADН ratio were studied in the liver mitochondrial fraction of rats with toxic hepatitis induced by acetaminophen under conditions of alimentary protein deprivation. Acetaminophen-induced hepatitis was characterized by a decrease of isocitrate dehydrogenase, α-ketoglutarate dehydrogenase and malate dehydrogenase activities, while the mitochondrial NAD(+)/NADН ratio remained at the control level. Modeling of acetaminophen-induced hepatitis in rats with alimentary protein caused a more pronounced decrease in the activity of NAD(+)-dependent dehydrogenases studied and a 2.2-fold increase of the mitochondrial NAD(+)/NADН ratio. This suggests that alimentary protein deprivation potentiated drug-induced liver damage.

AN ASSESSMENT OF THE FERROKINETIC INDICES IN RATS WITH TOXIC HEPATITIS UNDER THE CONDITIONS OF ALIMENTARY DEPRIVATION OF PROTEIN.

The ferrokinetic indices of blood serum of rats with acetaminophen-induced hepatitis under the conditions of alimentary deprivation of protein were assessed in the work. Research was conducted on 4 groups of animals: 1 - control; 2 - rats maintained on low-protein diet; 3 - rats with acute acetaminophen-induced hepatitis, maintained on full-value ration, 4 - rats with acute acetaminophen-induced hepatitis, maintained under the conditions of alimentary deprivation of protein. The serum iron content of and serum iron binding capacity were determined colorimetrically,% of the transferrin iron saturation - as a ratio of serum iron concentration to maximal iron binding capacity of serum transferrin. The presence of hemosiderin inclusions and the character of hemosiderosis were determined in the liver sections, stained by Perls method. Qualitative determination of C-reactive protein in blood serum was carried out by immunoenzymatic method. It is shown, that in protein-deficiency rats any significant changes of iron metabolism indices and hemosiderin accumu- lation weren't observed. At the same time in rats with acetaminophen-induced hepatitis the 5-fold increase of the serum iron content against the background non-significant increase of serum iron binding capacity and the 2-fold increase of the transferrin iron saturation is established. Simultaneously in the hepatocytes and reticuloendothelial system cells the accumulation of hemosiderin in the low dispersion form is observed, equating the second degree of hemosiderosis on the background emerging of C-reactive protein in serum. In protein-deficiency rats with toxic liver injury an abrupt increase of serum iron against the background reduction of the total serum iron binding capacity and maximal saturation of transferrin by iron ions is observed. It is established, that for animals from current group the third degree of mixed type hemosiderosis and the intensification of the inflammatory reaction in liver is characterstic. Conclusion was made, that alimentary deprivation of protein under the conditions of toxic liver injury is the critical factor for structural-functional state of liver, being accompanied by the iron metabolism disturbances, development of hemosiderosis and intensification of the inflammatory reaction in liver. Research results may be used for the biochemical rationale of the therapeutic approaches for the elimination and correction of the toxic liver injury consequences.

[Peculiarities of the Structural-Functional State of the Cytochrome Part of Liver Mitochondrial Respiratory Chain under Conditions of Acetaminophen-induced Hepatitis against the Background of Alimentary Deprivation of Protein].

Activity of the key enzyme of the cytochrome part of the respiratory chain--cytochrome oxidase, quantitative redistribution of mitochondrial cytochromes b, c1, c and aa3, activity of the key enzymes of cytochromes' heme metabolism--delta-aminolevulinate synthase and heme oxygenase under conditions of acetaminophen-induced hepatitis against the background of alimentary deprivation of protein were studied. It was found out, that under conditions of acetaminophen-induced hepatitis against the background of alimentary deprivation of protein, an inhibition of cytochrome oxidase activity and a decrease in the quantitative content of mitochondrial cytochromes against the background of the increase in the delta-aminolevulinate synthase and heme oxygenase activity are observed. In animals with toxic liver injury, maintained under conditions of alimentary deprivation of protein, a progressive decrease in the quantitative content of mitochondrial cytochromes b, c1, c and aa3 against the background. of the increase in heme oxygenase activity and preservation of delta-aminolevulinate synthase activity on the control level is identified. The conclusion was made, that alimentary deprivation of protein is a critical factor for the development of the disturbances of structural-functional integrity of the cytochromic part of the respiratory chain. The identified changes may be considered as one of the possible mechanisms of energy biotransformation system disturbances under conditions of alimentary deprivation of protein.

The ratio of ubiqiunon redox forms in the liver mitochondria under toxic hepatitis induced on the background of alimentary protein deficiency.

The level of the total ubiqiunon and redox forms CoQ in the rat liver mitochondria under the conditions of alimentary protein deficiency and toxic hepatitis, induced on the background protein deficiency has been investigated. Research has been carried out on 36 white non-linear rats, divided into 4 groups: 1 ­ rats, maintained on the complete semisynthetic ration; 2 ­ rats, fed low-protein ration; 3 ­ rats with acute acetaminophen-induced hepatitis, maintained on complete ration; 4 ­ rats with acetaminophen-induced hepatitis, maintained under the conditions of protein deficiency. The content of total and oxidized ubiqiunon was determined spectrophotometrically at λ=275 nm (molar extinction coefficient 12.25 Mm-1×sm-1). Reduced ubiqiunon content was determined by the difference between total and oxidized ubiqiunon content. The amount of tyrosine in the liver was measured in deproteinised by 6% sulfosalicylic acid extracts of liver tissue on an automated amino acid analyzer. The decrease of the total ubiqiunon content in liver mitochondria by 35% on the background of 2-fold decrease of oxidized ubiqiunon and preservation of reduced ubiqiunon amount has been estimated under the conditions of low-protein diet. Simultaneously the 5-fold decrease of liver content of tyrosine ­ the ubiqiunon precursor ­ has been observed. It has been shown, that under the conditions of acetaminophen-induced hepatitis the content of total ubiqiunon and its redox forms in the liver mitochondria of rats fed complete diet didn't change significantly comparing to control. A decrease of total ubiqiunon by 60% on the background of acute (18-fold) decrease of reduced ubiqiunon in liver mitochondria of rats with hepatitis, fed low-protein diet, has been observed. Established changes of the content of redox ubiquinone forms (a key component of the oxidative phosphorylation system in the liver mitochondria) can be considered as one of the mechanisms of malfunction of energy biotransformation system under the conditions of toxic liver injury in animals with protein deficiency.

[State of the energy-supply system of the liver mitochondria under the conditions of alimentary deficiency of protein].

The NADH-dehydrogenase and succinate dehydrogenase activity of the rats' liver mitochondria under the conditions of alimentary deprivation of protein has been studied. Research was carried out on 65 white non-linear rats divided according to the diet protein content into three groups: 1--rats fed a hypoprotein diet (7% of protein, 10% of fat u 83% of carbohydrates; n = 26); 2--rats fed a protein-free diet (n = 26); 3--rats fed a complete semi-synthetic ration (14% of protein, 10% of fat u 76% of carbohydrates; n = 13). The NADH-dehydrogenase activity was estimated by spectrophotometric method, succinate dehydrogenase activity--by the intensity of reduction of the potassium ferricyanide. It has been estimated that the decrease of NADH-dehydrogenase activity of mitochondria occurred on the 14th day of feeding rats with protein-free diet, and four-week feeding of rats under these conditions lead to the decrease of enzyme activity by 5,5 fold compared with the control group (0.506 +/- 0.040 nmol NADH/min/mg of protein) and by 3,0 fold compared with the previous stage of the experiment. At the same time a hypoprotein diet caused 2-fold decrease of NADH-dehydrogenase activity of liver mitochondria only on the 28th day. It has been shown that the succinate dehydrogenase activity didn't change significantly after two-week maintenance of rats on a protein-free diet as compared with control, while the four-week maintenance on both hypoprotein and protein-free diet lead to the decrease of the succinate dehydrogenase activity. Specifically, under the conditions of the hypoprotein diet succinate dehydrogenase activity of liver mitochondria decreased twofold and under the conditions of the protein free diet-- threefold. Probably, the disorders at the level of Complex I of respiratory chain underlie the realization of the changes in the system of energy biotransformation in mitochondria under the conditions of alimentary protein deficiency.

[The state of the mitochondrial energy-supplying system of blood leukocytes in the dynamics of Guerin's carcinoma growth under the low-level irradiation conditions].

Mitochondrial NADH-dehydrogenase, succinate dehydrogenase and cytochrome oxidase activities of peripheral blood leukocytes of rats with the grafted Guerin's carcinoma were studied in the dynamics of oncogenesis under the conditions of the preliminary low-level irradiation. Tumor growth was accompanied by a decrease in NADH-dehydrogenase activity, an increase of succinate dehydrogenase activity. Cytochrome oxidase activity of leucocytes remained at the control level up to the terminal stages of tumor growth. Preliminary low-level irradiation of the tumor bearing animals caused a tendency to the decrease of enzymatic activities studied. This tendency was observed from the initial stages of oncogenesis.

[Activity of the marker liver enzymes under the conditions of toxic hepatitis and alimentary deprivation of protein].

The activity of the sorbitoldehydrogenase (SDH), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) in the blood serum of rats with acetaminophen-induced hepatitis under the conditions of alimentary deprivation of protein was studied. The animals were divided into 3 groups: 1--rats with acute acetaminophen-induced hepatitis, maintained on the full ration; 2--rats with acute acetaminophen-induced hepatitis, maintained under the conditions of alimentary deprivation of protein; 3--control. The activity of the sorbitol dehydrogenase in blood serum was determined by the kinetic method, activity of the alanine aminotransferase and alkaline phosphatase - photometrically. It is shown, that in animals with the model hepatitis the activity of sorbitol dehydrogenase in blood serum increases 20-fold, wherein statistical significance between animals with hepatitis maintained under the conditions of full ration and those of low-protein diet is not established. In the group of animals with acetaminophen-induced hepatitis the preservation on the control level of the alkaline phosphatase activity on the base of the increase of alanine aminotransferase by 2.2 times and ratio ALT/ALP>5 testifies about hepatocellular liver injury. In the group of animals with drug-induced hepatitis and alimentary deprivation of protein, the increase of the alkaline phosphatase and alanine aminotransferase activity is observed, herewith the ratio ALT/ALP ranges from 2 to 5 and testifies about mixed liver injury. The conclusion was made, that alimentary deprivation of protein is the critical factor for the development of the disturbances of functional and structural liver integrity, and the therapeutic approaches to the correction of the drug-induced liver injury should be different depending on the value of protein ration in the anamnesis, taking into account the different types of liver injury.

Effects of low-dose radiation on glutamate dehydrogenase activity in tissues of rats with transplanted Guerin's carcinoma.

Glutamate dehydrogenase activity in mitochondrial fraction of the liver and muscle tissue of tumor-bearing rats was studied in the dynamics of Guerin's carcinoma growth and after preliminary low-dose irradiation. At the initial stages of Guerin's carcinoma growth, maximum activity of glutamate dehydrogenase was observed in the liver, while in the muscle tissue, catabolic transformations of amino acids was activated at the logarithmic phase of carcinogenesis and tended to inhibition at the terminal stages. Low-dose irradiation was followed by activation glutamate dehydrogenase in mitochondrial fraction of the liver and inhibition of this enzyme in mitochondrial fraction of the muscle tissue.

[The state of the cytochrome part of respiratory chain in tumor carrier rats' liver in the conditions of preliminary low-level irradiation].

The effect of low-level irradiation on the structural and functional organization of the cytochrome part of the respiratory chain in tumor carrier rats' liver is studied. The preliminary low-level irradiation leading to the mitochondrial cytochrome a, b and c content reduction at the latent stage of Guerin's carcinoma is shown. At the same time, the maximal reduction of the content of all liver cytochromes is observed at the terminal stages of oncogenesis. The content of cytochome c undergoes the most significant changes in the liver mitochondrial fracture. The possible mechanism of mitochondrial haem-containing cytochromes content reduction may be associated with the disorder of their formation caused by the heam synthesis inhibition found in our study. Simultaneously, the cytochrome oxydase (key enzyme of the cytochrome part) activity inhibition is observed to be caused by preliminary low-level irradiation at the latent growth stage of Guerin's carcinoma. The determined differences between irradiated and non-irradiated tumor carrier groups allow us to come to the conclusion that low-level irradiation has an impact only at the initial stages of the aftereffect. At the following stages, the state of the cytochrome part of the respiratory chain is defined by growth conditions of tumor.

[The change in the structural and functional organization of the Guerin's carcinoma cytochrome part of respiratory chain in tumor carriers in the conditions of preliminary low-level irradiation].

The effect of low-level irradiation of tumor-bearing rats on the structural and functional organization of the cytochrome part of respiratory chain of mitochondria isolated from Guerin's carcinoma has been investigated. The maximal reduction in the mitochondrial cytochromes a, b and c content was observed at the terminal stage of Guerin's carcinoma. A low-level irradiation during initial stages of oncogenesis produced opposite changes in the mitochondrial cytochrome content. The possible mechanism of mitochondrial haem-containing cytochromes content reduction may be attributed to impairment in their formation caused by inhibition of the key enzyme of haem synthesis, 5-aminolevulinate synthase. The determined changes of the mitochondrial cytochromes quantitative content were accompanied by decreased activity of cytochrome oxidase. The preliminary low-level irradiation of the tumor-bearing animals produced further reduction in the cytochrome oxidase activity observed in all experimental periods.