Effect of Liposome-Encapsulated Dextrazide on Local Regulation of Extracellular Matrix Metabolism in Liver and Lungs of Mice with during Chronic BCG-Induced Granulomatosis.

The liposomal form of isonicotinic acid hydrazide conjugate with oxidized dextran (liposomeencapsulated dextrazide, LEDZ) injected intraperitoneally for 3 months to BALB/c mice with chronic BCG-induced granulomatosis (6 month after infection) down-regulated activities of protease (MMP) and antiprotease (TIMP-1, TIMP-2, α2-macroglobulin) components of the regulation system, which indicates a drop of destructive and inflammatory potential of BCGinduced granulomatosis. The persistent enhanced hyaluronidase activity in the liver and its reduced (normalized to control level) activity in the lungs after administration of LEDZ indicates a greater hydrolysis of hyaluronan in the liver than in the lungs. LEDZ-induced changes in MMP/TIMP system in mouse liver and lungs are characterized with relative elevation of protease activity over that of antiprotease one.

Content of the Major Extracellular Matrix Components of the Liver and Lungs in Mice with Chronic BCG-Granulomatosis Treated with Liposome-Encapsulated Dextrazide.

Administration of the liposome-encapsulated dextrazide (LEDZ) to mice 6 months after infection with mycobacterium BCG vaccine for 3 months modulated metabolism of collagens and the intensity of fibrosis of internal organs. In the liver, we observed a redistribution of glycosaminoglycans towards sulfated forms, a decrease in the content of hyaluronan, a change in the ratio of hydroxyproline fractions indicating a decrease in fibrosis via two mechanisms: suppression of synthesis and increased degradation of collagens. In the lungs, administration of LEDZ did not affect the content of sulfated glycosaminoglycans, but significantly reduced the content of hyaluronan, stimulated degradation of collagen, and reduced its synthesis, but these processes were insufficient for significant reduction of fibrotic complications in the lungs. In animals treated with LEDZ, the decrease in collagen synthesis in the liver was 2-fold more pronounced than in the lungs.

[Age-related features of the relationship between the content of vascular endothelial growth factor and indicators of lipid metabolism and extracellular matrix metabolism in men in the European part of the Russian Arctic.]

The content of vascular endothelial growth factor-A (VEGF-A) in blood plasma and its relationship with lipid and extracellular matrix metabolism in working-aged men (19-69 years), living and working in the European part of the Arctic zone of the Russian Federation (Russian Arctic), were studied. No age dependence of the plasma VEGF-A content was found. The correlation analysis, performed in different age groups, revealed significant associations of VEGF-A level with lipid parameters (CS, LDL-C, Apo B, atherogenicity coefficient, Apo B /Apo A1 ratio) and extracellular matrix metabolism (blood TIMP-4, MMP-2, MMP-3, MMP-9, hyaluronan, total and peptide-bound hydroxyproline, glycosaminoglycans). The established correlations indicate the formation of relationships between angiogenesis, atherogenesis and fibrosis at a specific period of life of northerners in the Russian Arctic.

Antifibrotics Effect of Liposome-Encapsulated Composition of Oxidized Dextran and Isonicotinic Acid Hydrazide in Mice with BCG-Induced Granulomatosis Depends on Administration Route.

We studied the response of the extracellular matrix of the lungs and liver in mice with BCGinduced granulomatosis (3 months) after inhalation and intraperitoneal administration of liposome-encapsulated dextrazide (LEDZ): a conjugate of oxidized dextran (40 kDa) and isonicotinic acid hydrazide (INH). LEDZ inhalation proved to be more effective in reducing fibrosis severity, both in the lungs and liver. However, the mechanisms of the antifibrotic effect were different: increased degradation and reduced collagen synthesis in the lungs and reduced collagen synthesis and collagen degradation in the liver. This suggest that drug administration routes and delivery to the target organs are crucially important in the therapy of tuberculosis. The antifibrotic effect depended on LEDZ administration route and was more potent after LEDZ inhalation.

Peculiarities of Collagen Turnover in Aging BALB/c Mice.

Examination of collagen turnover in the liver, lungs, and spleen of BALB/c mice revealed the age-related differences in the levels of hydroxyproline and its fractions. The highest level of total hydroxyproline was observed in the lungs and spleen of young (2-month-old) mice. In the liver, this level attained maximum at the age of 6 months at the expense of elevation of protein-bound hydroxyproline relatively its level in 2-month-old mice. At the age of 12 months, the levels of total hydroxyproline in the liver and spleen were lower than in 6-month-old mice. The decrease in the collagen turnover rate in the liver of 12-month-old mice reflected lower levels of hydroxyproline fractions in comparison with the corresponding values in 6-month-old mice. The rates of collagen turnover in organs differed in mice of different ages: it was maximum in the lungs and spleen of young animals and in the liver of middle-aged mice.

Effect of Liposomal Delivery of Oxidized Dextran with Isonicotinic Acid Hydrazide on Component Profile of Pulmonary Extracellular Matrix in Mice with BCG-Induced Granulomatosis.

Intraperitoneal injections of isonicotinic acid hydrazide (INH), dextrazide (oxidized dextran+INH), or liposomes loaded with dextrazide (INH dose of 14 mg/kg) over 2 months to mice with BCG-induced granulomatosis started from postinfection day 90 induced qualitative and quantitative changes in composition of pulmonary extracellular matrix. Both dextrazide and its liposomal form decreased the levels of sulfated glycosaminoglycans and uronic acids. In contrast to INH, both preparations did not decrease the levels of total glycosaminoglycans, proteins, and galactose. This difference is explained by the fact both free and liposomal dextrazide activated MMP, but did not increase the content of TIMP-1 and TIMP-2, whereas injection of INH was followed by an increase in TIMP-2 content and a decrease in the level of free hydroxyproline, which attested to down-regulation of collagen degradation and maintenance of the conditions for pulmonary fibrosis in mice of this group.

Remodeling of the Extracellular Matrix of the Liver in Mice with BCG-Induced Granulomatosis after Administration of Liposome-Encapsulated Composition of Oxidized Dextran and Isonicotinic Acid Hydrazide.

Remodeling of the extracellular matrix of the liver in mice with BCG-induced granulomatosis after 2-month course of intraperitoneal injections of isonicotinic acid hydrazide, oxidized dextran conjugated with isonicotinic acid hydrazide, dextrazide, and its liposome-encapsulated form manifested in modification of the proteoglycan composition and hydroxyproline fractions associated with changes in hyaluronidase and MMP activities. The antifibrotic effect of dextrazide and its liposome-encapsulated form manifested in reduction of hydroxyproline fractions that reflected the process of collagen synthesis. Administration of isonicotinic acid hydrazide was followed by a decrease in hydroxyproline fractions reflecting collagen synthesis (antifibrotics effect) and degradation (profibrotic effect), which can be explained by its hepatotoxic activity.

[Apolipoprotein A-I stimulates secretion of insulin and matrix metalloproteinases by islets of Langerhans]. / Apolipoprotein A-I stimuliruet sekretsiiu insulina i matriksnykh metalloproteinaz ostrovkami Langergansa podzheludochnoi zhelezy.

The development of type 2 diabetes mellitus (DM2) is accompanied by disturbances in lipid metabolism. These include the increase in serum levels of atherogenic fractions of very low-density (VLDL) and low-density lipoproteins (LDL), total cholesterol, triglycerides and apo B. In contrast, the level of antiatherogenic high density lipoproteins (HDL) and the content of apolipoprotein A-I (apoA-I) decreased. To study the effect of the observed metabolic changes on insulin secretion in vitro, we used the islets of Langerhans isolated from the rat pancreas. It has been found that incubation of the islets in the presence of serum of the obese patients and patients with decompensated DM2 leads to a decrease in insulin secretion by 2.4 and 5.0 times, respectively. On the contrary, the addition of HDL to the incubation medium increased the insulin secretion by 3.4 times. A similar effect was observed in the presence of apoA-I, the main protein component of HDL. In the presence of apoA-I, the extracellular activity of matrix metalloproteinases (MMPs) demonstrated a 10-fold increase. The addition of LDL and VLDL to the islets did not change the secretion of insulin and activity of MMP. Our results testify to the important role of HDL and apoA-I in regulation of the insulin secretion by b-cells and the activity of MMPs in the islets of Langerhans.

Changes in Activity of Matrix Metalloproteinases and Serum Concentrations of Proinsulin and C-Peptide Depending on the Compensation Stage of Type 2 Diabetes Mellitus.

In patients with type 2 diabetes mellitus, serum activities of MMP-2 and MMP-7 were substantially decreased in comparison with apparently healthy individuals. At the decompensation stage, along with the increased content of glucose and glycated hemoglobin, a pronounced (3-fold) increase in proinsulin concentration was observed. On the contrary, MMP activity and C-peptide concentration decreased at this stage. The ratio of proinsulin concentration to MMP activity at the stages of diabetes mellitus compensation and subcompensation was approximately 1:50, while at the stage of decompensation it was 1:12. Thus, the ratio of these blood serum parameters can be used as an additional diagnostic marker of diabetes decompensation and severity of its complications.

[Interrelation between the melatonin level and indicators of aging and fibrosis in men in the European part of the Arctic zone of Russian Federation.]

The results of the correlation analysis between the metabolite melatonin - 6-sulfatoxymelatonin (6-SMT) in the urine and the indicators of aging, interstitial fibrosis, the system of antioxidant protection in men in the Arctic are presented. Negative correlation of medium strength between 6-SMT with calendar age, biological age and molecular marker of aging p16INK4a in leukocytes was noted. A relationship of 6-SMT to fibrosis indices was found. It manifested itself in a negative correlation with sulfated glycosaminoglycans in the urine, with collagen markers: free hydroxyproline, peptide-linked hydroxyproline, and total hydroxyproline. Age-related decrease in the level of 6-SMT may be associated with an age-related increase in interstitial fibrosis in northerners. A positive correlation of medium strength between 6-SMT and hormones: free testosterone level (r=0,53, p=0,0002), total testosterone (r=0,43, p=0,003) and cortisol (r=0,33, p=0,007) was shown. Negative correlations of 6-SMT with the content of ceruloplasmin (r=-0,63, p=0,001) and SCORE scale (r=-0,52, p=0,001) indicate the relationship between melatonin and the antioxidant defense system and the risk of cardiovascular pathology.The results of the correlation analysis between the metabolite melatonin - 6-sulfatoxymelatonin (6-SMT) in the urine and the indicators of aging, interstitial fibrosis, the system of antioxidant protection in men in the Arctic are presented. Negative correlation of medium strength between 6-SMT with calendar age, biological age and molecular marker of aging p16INK4a in leukocytes was noted. A relationship of 6-SMT to fibrosis indices was found. It manifested itself in a negative correlation with sulfated glycosaminoglycans in the urine, with collagen markers: free hydroxyproline, peptide-linked hydroxyproline, and total hydroxyproline. Age-related decrease in the level of 6-SMT may be associated with an age-related increase in interstitial fibrosis in northerners. A positive correlation of medium strength between 6-SMT and hormones: free testosterone level (r=0,53, p=0,0002), total testosterone (r=0,43, p=0,003) and cortisol (r=0,33, p=0,007) was shown. Negative correlations of 6-SMT with the content of ceruloplasmin (r=-0,63, p=0,001) and SCORE scale (r=-0,52, p=0,001) indicate the relationship between melatonin and the antioxidant defense system and the risk of cardiovascular pathology.

Cystatin C as a Marker of Progressing Cardiovascular Events during Coronary Heart Disease.

The role of cystatin C, an inhibitor of cysteine proteases, as an alternative and potent predictor of acute cardiovascular events in coronary heart disease (CHD) patients was examined and compared to that of other markers of cardiorenal abnormalities. The patients with CHD demonstrated elevated serum cystatin C, especially in cases with serious risk of cardiovascular complications. In comparison with other indicators of cardiorenal dysfunction, cystatin C can be viewed as an alternative predictor of cardiovascular complications, although its sensitivity is inferior to that of high-sensitivity C-reactive protein and natriuretic peptide.

[Proinsulin as a diagnostic biochemical marker of decompensated diabetes mellitus type II.]

The proinsulin is one of indices reflecting functional activity of pancreas. Under insulin-independent diabetes ration proinsulin/insulin increases. The study was carried out to evaluate content of proinsulin and other biochemical indices of blood serum depending on stage of compensation of diabetes mellitus type II. The content of proinsulin in blood serum was detected using "sandwich"-technique enzyme-linked immunosorbent assay (BioVender, Czechia). The concentration of proinsulin in blood serum of control group comprised 2,56 ± 0,23 nmol/l. No gender and age differences were established concerning content of proinsulin in blood serum of patients. The concentration of proinsulin was twice higher than control level in group of patients. Two main groups were singled out: one with high content (concentration thrice exceeded average value in control group) and another with low content of proinsulin (within standard value). The patients with stages of compensated and sub-compensated diabetes (n=35) retained low concentration of proinsulin. At the stage of decompensation of diabetes (n=40) concentration of proinsulin was thrice higher than standard value. This group was characterized by increasing of concentration of glucose, fructosamine and decreasing of concentration of C-peptide. At the stage of decompensation concentrations of triglycerides, total cholesterol and atherogenicity index increased reliably. The concentration of proinsulin at the stage of decompensation positively correlated with level of glucose and concentration of triglycerides. Therefore, measurement of concentration of proinsulin can be used as an important diagnostic criterion permitting to judge about degree of decompensation of diabetes and development of its complications.

[Quality of life for men of different ages in the russian European North and its relationship with self-reported health and hormonal status].

Assessment of the quality of life for male inhabitants of the Russian European North demonstrated decreased indices of physical health component (the scale of role functioning determined by physical condition) and mental health component (the scales of social functioning and role functioning determined by emotional state and mental health) compared to the inhabitants of Siberia as a comparison group. Men aged up to 29 had the highest values at all scales. The values decreased with age. The most prominent decrease was observed for the scales of general well-being and pain intensity. There was a moderate decrease for the scales of physical functioning, role functioning determined by emotional state, and role functioning determined by physical condition. Quality of life was closely related to self-reported health of northerners, pathological index, molecular marker of aging, and biological age. Negative correlation between these values implies the deterioration of physical and mental health with biological age. The quality of life for northerners also correlated with hormonal status. There was a relationship between testosterone and three scales of physical health components; between cortisol and one scale of physical component; and between both dehydroepiandrosterone sulfate and insulin and the scales of physical and mental components of health. The effect of various hormones on different age groups of northerners was demonstrated.

Content of circulating extracellular DNA, plasma activities of matrix metalloproteinases, and ultrastructure of the myocardium in hypothyroid rats with hypercholesterolemia.

Free circulating extracellular DNA, plasma activities of matrix metalloproteinases in hypothyroid rats, and ultrastructural changes in the myocardium were studied under conditions of experimental hypercholesterolemia. For suppression of thyroid function, the animals received antithyroid drug mercazolyl under conditions of cholesterol diet. Hypercholesterolemia in hypothyroid rats (thyroxine concentration 2-fold below the normal) was paralleled by a pronounced increase of the concentration of free circulating extracellular DNA and total matrix metalloproteinases 2 and 7 activity. These changes were associated with lytic and destructive changes in cardiomyocytes and blood capillary endotheliocytes. Changes in the cardiomyocyte and endotheliocyte ultrastructure were more pronounced in hypothyroid rats.

Structural changes in the myocardium and serum lipid spectrum in experimental hypercholesterolemia and hypothyroidism.

We studied the peculiarities of lipid spectrum of the blood and structural reorganization of the myocardium in experimental hypercholesterolemia with and without hypothyroidism. It was found that alimentary hypercholesterolemia accompanied by elevated total cholesterol, LDL, HDL, and triglyceride concentrations led to a decrease in body weight, heart weight, number of cardiomyocytes in the heart and induced pronounced lytic changes in cardiomyocytes, circulation disorders (sludge syndrome, echinocytosis of erythrocytes, lymphostasis), diffuse fibrosis of the stroma, and appearance of foam cells among diffuse mononuclear infiltrate cells. The combination of hypercholesterolemia with hypothyroid status caused more pronounced changes in the lipid spectrum and atherogenic index and more pronounced lytic and necrobiotic changes in cardiomyocytes. These findings suggest that elevated cholesterol concentrations in the blood, especially against the background of suppressed thyroid function, can directly induce considerable damage to cardiomyocytes, intramural vessels, and erythrocytes without the development of myocardial ischemia and in the absence of atherosclerotic plaques.

Activity of matrix metalloproteinases during antimycobacterial therapy in mice with simulated tuberculous inflammation.

Matrix metalloproteinases are shown to be involved in the pathogenesis of tuberculosis inflammation. In the early stages of BCG-granuloma formation in mouse liver and lungs, the serum levels of matrix metalloproteinases 2 and 7 increased by 4.5 times and remained unchanged while the pathology developed. Antimycobacterial therapy with isoniazid reduced enzyme activity almost to the level of intact control. The decrease in activity of matrix metalloproteinases 2 and 7 that play the most prominent role in the development of destructive forms of tuberculosis is of great therapeutic importance.

Effect of polysaccharides and human plasma lipoproteins on the secretion of cystatin C by peritoneal macrophages from normal and tumor bearing mice.

Intact peritoneal macrophages in vitro secreted the cysteine proteinase inhibitor cystatin C. Polysaccharides stimulated cystatin C secretion: lipopolysaccharide < carboxymethylated beta-D-glucan < sulfoethylated beta-D-glucan. Human plasma low-density- (LDL) and high-density lipoproteins (HDL) are still more potent inducers of cystatin C secretion by macrophages. Peritoneal macrophages from mice with experimental HA-1 hepatoma compared to those from intact mice secreted more cystatin C with maximum polysaccharide-stimulated secretion after 30 min of incubation. LDL and HDL induced cystatin C secretion by tumor macrophages also.

Detection of apolipoprotein A-I, B, and E immunoreactivity in the nuclei of various rat tissue cells.

Proteins with apolipoprotein A-I immunoreactivity were detected in the fraction of non-histone acidic proteins isolated from nuclei of various rat tissue cells. These proteins were detected in the brain, liver, kidney, lung, heart, skeletal muscle, testis, spleen, and bone marrow. In the same fraction from liver nuclei, proteins with apoB and apoE immunoreactivity were also detected. The composition of these proteins was studied by immunoblotting. ApoA-I immunoreactivity in the liver nuclei is due to two proteins. One 28-kD protein corresponds to the mature form of the plasma pool of apoA-I and another 14-kD protein is the product of limited proteolysis of apoA-I. The highest content of apoA-I immunoreactivity was detected in transcriptionally active chromatin and nuclear matrix. ApoB immunoreactivity is due to six proteins with molecular weights from 15 to 100 kD. ApoE immunoreactivity is due to a single protein corresponding to the 35-kD form of plasma apoE. Proteins with apoA-I, apoB, and apoE immunoreactivity may be involved in the regulation of transcriptional activity of chromatin.

Distribution of tocopherol and apolipoprotein A-I immunoreactivity in rat liver chromatin.

[125I]Labelled lipoproteins (HDL, LDL, and VLDL) are actively taken up by the rat liver. The uptake rate in nonparenchymal liver cells was significantly higher than that in hepatocytes. Analysis of the distribution of the labelled lipoproteins in liver subcellular structures showed the presence of these compounds in the nuclei. The presence of apoA-I, apoB and apoE immunoreactivity in liver cell nuclei was demonstrated by immunochemical methods (ELISA and dot immunoassay). The nuclear matrix and transcriptionally active chromatin displayed the highest apoA-I levels, which considerably exceeded those in the total and transcriptionally inactive chromatins. A similar distribution pattern was observed for alpha-tocopherol. It is concluded that blood serum lipoproteins, primarily HDL, may transport tocopherol and other lipids into cell nuclei.