[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.

Peculiarities of the free radical processes in rat liver mitochondria under toxic hepatitis on the background of alimentary protein deficiency.

The rate of superoxide anion radical, hydroxyl radical and hydrogen peroxide generation, the level of oxidative modification of mitochondrial proteins in the liver of rats with toxic hepatitis was investigated on the background of alimentary protein deficiency. We did not find significant increases of the intensity of free radical processes in liver mitochondria of rats maintained on the protein-deficient ration. The most significant intensification of free radical processes in liver mitochondria is observed under the conditions of toxic hepatitis, induced on the background of alimentary protein deprivation. Under these conditions the aggravation of all studied forms of reactive oxygen species generation was observed in liver mitochondria. The generation rates were increased as follows: O2 ­ by 1.7 times, Н2О2 ­ by 1.5 times, •ОН ­ practically double on the background of accumulation of oxidized mitochondria-derived proteins. The established changes in thiol groups' redox status of respiratory chain proteins insoluble in 0.05 M sodium-phosphate buffer (pH 11.5), and changes of their carbonyl derivatives content may be considered as one of the regulatory factors of mitochondrial energy-generating function.

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.

[NADH:ubiquinone reductase and succinate dehydrogenase activity in the liver of rats with acetaminophen-induced toxic hepatitis on the background of alimentary protein deficiency].

The ratio between the redox forms of the nicotinamide coenzymes and key enzymatic activity of the I and II respiratory chain complexes in the liver cells mitochondria of rats with acetaminophen-induced hepatitis under the conditions of alimentary deprivation of protein was studied. It was estimated, that under the conditions of acute acetaminophen-induced hepatitis of rats kept on a low-protein diet during 4 weeks a significant decrease of the NADH:ubiquinone reductase and succinate dehydrogenase activity with simultaneous increase of the ratio between redox forms of the nicotinamide coenzymes (NAD+/NADH) is observed compared to the same indices in the liver cells of animals with experimental hepatitis kept on the ration balanced by all nutrients. Results of research may become basic ones for the biochemical rationale for the approaches directed to the correction and elimination of the consequences of energy exchange in the toxic hepatitis, induced on the background of protein deficiency.

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.