The effect of exenatide (a GLP-1 analogue) and sitagliptin (a DPP-4 inhibitor) on asymmetric dimethylarginine (ADMA) metabolism and selected biomarkers of cardiac fibrosis in rats with fructose-induced metabolic syndrome.

Asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide (NO) synthesis, is a risk factor for endothelial dysfunction, a common pathophysiological denominator for both atherogenesis and cardiac fibrosis. We aimed to investigate whether the cardioprotective and antifibrotic effects of incretin drugs, exenatide and sitagliptin, may be associated with their ability to affect circulating and cardiac ADMA metabolism. Normal and fructose-fed rats were treated with sitagliptin (5.0/10 mg/kg) or exenatide (5/10 µg/kg) for 4 weeks. The following methods were used: LC-MS/MS, ELISA, Real-Time-PCR, colorimetry, IHC and H&E staining, PCA and OPLS-DA projections. Eight-week fructose feeding resulted in an increase in plasma ADMA and a decrease in NO concentration. Exenatide administration into fructose-fed rats reduced the plasma ADMA level and increased NO level. In the heart of these animals exenatide administration increased NO and PRMT1 level, reduced TGF-ß1, α-SMA levels and COL1A1 expression. In the exenatide treated rats renal DDAH activity positively correlated with plasma NO level and negatively with plasma ADMA level and cardiac α-SMA concentration. Sitagliptin treatment of fructose-fed rats increased plasma NO concentration, reduced circulating SDMA level, increased renal DDAH activity and reduced myocardial DDAH activity. Both drugs attenuated the myocardial immunoexpression of Smad2/3/P and perivascular fibrosis. In the metabolic syndrome condition both sitagliptin and exenatide positively modulated cardiac fibrotic remodeling and circulating level of endogenous NOS inhibitors but had no effects on ADMA levels in the myocardium.

Processes of plasma protein N-homocysteinylation in multiple sclerosis.

BACKGROUND: Homocysteine thiolactone (HTL) is a cyclic thioester of homocysteine (Hcy) contributing to the toxicity of this amino acid. HTL spontaneously reacts with protein lysine residues leading to altered properties of target proteins and induction of immune response. HTL is hydrolyzed to Hcy by plasma enzyme, paraoxonase 1 (PON1). Although both Hcy and PON1 may be involved in the pathogenesis of multiple sclerosis (MS), protein modification by HTL in this disease has not been studied so far. Purpose/Aim: The aim of this study was to assess the level of Hcy, HTL and autoantibodies against N-homocysteinylated proteins as well as PON1 activity in patients with MS. METHODS: The studies were performed in 61 MS patients with relapsing-remitting (RR group, n = 25) and secondary-progressive type of MS (SP group, n = 36), and in healthy people (C - control group, n = 44). RESULTS: Homocysteine level was significantly higher in MS patients comparing to control group (C vs. RR p < 0.01; C vs. SP p < 0.05). The level of HTL tended to be higher in RR-MS in comparison to control group, but it did not reach the level of significance. The level of antibodies against N-homocysteinylated proteins did not differ significantly between studied groups. PON1 activity was significantly lower in SP type of MS (SP vs. C p < 0.05; SP vs. RR p < 0.05). CONCLUSIONS: Although plasma Hcy concentration is higher in MS patients and PON1 activity is reduced in the SP form, MS is associated with minor or no changes in protein-attached HTL and anti-homocysteinylated protein immune response.

The effect of the angiotensin II receptor, type 1 receptor antagonists, losartan and telmisartan, on thioacetamide-induced liver fibrosis in rats.

It has been reported previously that the density of angiotensin II receptors is increased in the rat liver in experimentally-induced fibrosis. We hypothesized that pharmacological blockade of angiotensin receptors may produce beneficial effects in models of liver fibrosis. In this study, we used the widely used thioacetamide (TAA)-induced model of liver fibrosis (300 mg/L TAA ad libitum for 12 weeks). Rats received daily injections (i.p), lasting 4 weeks of the angiotensin II type 1 receptor antagonists, losartan 30 mg/kg (TAA + L) or telmisartan 10 mg/kg (TAA + T) and were compared to rat that received TAA alone. Chronic treatment with losartan and telmisartan was associated with a significant reduction in the activity of alkaline phosphatase, and decreased concentrations of tumor necrosis factor-alpha and transforming growth factor beta-1 compared to controls. We also found a significant reduction interleukin-6 in rats receiving telmisartan (P < 0.05) but not losartan. Both treatments increased the concentration of liver glutathione along with a concomitant decrease of GSSG compared to controls. In addition, increased paraoxonase 1 activity was observed in the serum of rats receiving telmisartan group compared to the TAA alone controls. Finally, histological evaluation of liver sections revealed losartan and telmisartan treatment was associated with reduced inflammation and liver fibrosis. Taken together, these results indicate that both telmisartan and losartan have anti-inflammatory and anti-oxidative properties in the TAA model of liver fibrosis. These finding add support to a growing body of literature indicating a potentially important role for the angiotensin system in liver fibrosis and indicate angiotensin antagonists may be useful agents for fibrosis treatment.

Protective effects of melatonin against thioacetamide-induced liver fibrosis in rats.

The aim of this study was to determine the effect of melatonin on thioacetamide (TAA) induced liver fibrosis in rats. The antifibrotic effects of melatonin were assessed by determining activity indirect markers of fibrosis: aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AP), and proinflammatory cytokines: interleukin 6 (IL-6), interleukin-1beta (IL-1ß), tumour necrosis factor alpha (TNF-α), transforming growth factor-beta (TGF-ß) and platelet-derived growth factor (PDGF). Parameters of oxidative stress: oxidised glutathione (GSSG), reduced glutathione (GSH) and presaged activity of paraoxonase 1 (PON-1), an antioxidative enzyme were determined. Inflammatory changes and fibrosis extent were evaluated histologically. Experiments were carried out in Wistar rats. Animals were divided into 4 groups: I - controls, water ad libitum for 12 weeks, group II - TAA, 300 mg/L ad libitum for 12 weeks, III - melatonin, 10 mg/kg b.w. intraperitoneally (i.p.) daily for 4 weeks, IV - TAA, 300 mg/L ad libitum for 12 weeks followed by melatonin, 10 mg/kg/b.w. i.p. daily for 4 weeks. Results of serum determinations demonstrated significantly lower activity of AST, ALT and AP in the group receiving TAA followed by melatonin compared to the group receiving only TAA. Immunoenzymatic findings on effect of melatonin on concentration of proinflammatory cytokines confirmed these data. Biochemical examinations in liver homogenates revealed statistically significant improvement (concentration of GSH increases and concentration of GSSG decreases) in animals with TAA-induced liver damage receiving melatonin. Moreover, the activity of PON-1 toward phenyl acetate and paraoxon was increased in liver homogenates and serum in the group receiving TAA followed by melatonin compared to the TAA group without melatonin treatment. Microscopic evaluation disclosed inhibitory effects of melatonin on inflammatory changes and extent of liver fibrosis.

Association of stromal-derived factor-1 alpha and endogenous sex hormones in men aged over 50 years with stable coronary artery disease.

PURPOSE: Many studies indicate an inverse relationship between stromal-derived factor-1 alpha (SDF-1 alpha), a chemokine, and coronary risk factors. Moreover, SDF-1 alpha is crucial in neoangiogenesis and in the mobilization and homing of endothelial progenitor cells to the ischemic coronary vessels. Numerous studies indicate that circulating sex hormones are associated with atherogenesis during male aging. The aim of this study was therefore to determine whether there exists a relationship between SDF-1 alpha and endogenous sex hormones in aging men with stable coronary artery disease (CAD). MATERIAL AND METHODS: Plasma concentrations of SDF-1 alpha, testosterone (T), estradiol (E2), and sex hormone binding globulin (SHBG) were measured and the E2/T ratio was calculated in a cross-sectional study of 82 men over 50 years of age with stable CAD. RESULTS: SDF-1 alpha was positively and significantly correlated with T (r = 0.233; p = 0.036) and with SHBG (r = 0.312; p = 0.004). There was a significant inverse correlation between SDF-1 alpha and the E2/T ratio (r = -0.463; p < 0.001). After adjustment for age, body mass index and smoking status, SHBG and E2/T ratio were the only factors associated with SDF-1 alpha. CONCLUSIONS: T and SHBG (directly) and the E2/T ratio (inversely) may be involved in the etiopathogenesis of CAD through their relationships to SDF-1 alpha.

Leptin decreases renal medullary Na(+), K(+)-ATPase activity through phosphatidylinositol 3-kinase dependent mechanism.

We examined the effect of leptin on renal function and renal Na(+),K(+)-ATPase and ouabain-sensitive H(+),K(+)-ATPase activities in the rat. Leptin was infused under general anaesthesia into the abdominal aorta proximally to the renal arteries. Leptin infused at doses of 1 and 10 microg/kg/min increased urine output by 40% and 140%, respectively. Urinary Na(+) excretion increased in rats receiving leptin at doses of 0.1, 1, and 10 microg/kg/min by 57.6%, 124.2% and 163.6%, respectively. Leptin had no effect on creatinine clearance, potassium excretion and phosphate excretion. Na(+),K(+)-ATPase activity in the renal medulla of rats treated with 1 and 10 microg/kg/min leptin was lower than in control animals by 25.5% and 33.2%, respectively. In contrast, cortical Na(+),K(+)-ATPase as well as either cortical or medullary ouabain-sensitive H(+),K(+)-ATPase activities did not differ between leptin-treated and control animals. The effect of leptin on Na(+),K(+)-ATPase activity was abolished by actin depolymerizing agents, cytochalazin D and latrunculin B, and by phosphatidylinositol 3-kinase (PI3K) inhibitors, wortmannin and LY294002. These results indicate that: 1). natriuretic effect of leptin is mediated, at least in part, by decrease in renal medullary Na(+),K(+)-ATPase activity, 2). inhibition of medullary Na(+),K(+)-ATPase by leptin is mediated by PI3K and requires integrity of actin cytoskeleton.

Nitric oxide decreases renal medullary Na+, K+-ATPase activity through cyclic GMP-protein kinase G dependent mechanism.

The aim of this study was to investigate the effect of nitric oxide on renal Na+,K(+)-ATPase and ouabain-sensitive H+,K(+)-ATPase activities. The study was performed in male Wistar rats. The investigated substances were infused under general anaesthesia into abdominal aorta proximally to the renal arteries. The activity of ATPases was assayed in isolated microsomal fraction. NO donor, S-nitroso-N-acetylpenicillamine (SNAP), infused at doses of 10(-7) and 10(-6)mol/kg/min decreased medullary Na+,K(+)-ATPase activity by 29.4% and 45.2%, respectively. Another NO donor, spermine NONOate, administered at the same doses reduced Na+,K(+)-ATPase activity in the renal medulla by 31.7% and 46.5%, respectively. Neither of NO releasers had any effect on Na+,K(+)-ATPase in the renal cortex and on either cortical or medullary ouabain-sensitive H+,K(+)-ATPase. Infusion of NO precursor, L-arginine (100 micromol/kg/min), decreased medullary Na+,K(+)-ATPase activity by 32.2%, whereas inhibitor of nitric oxide synthase, L-NAME (10 nmol/kg/min), increased this activity by 20.7%. The effect of synthetic NO donors was mimicked by 8-bromo-cGMP and blocked by inhibitors of soluble guanylate cyclase, ODQ or methylene blue, as well as by specific inhibitor of protein kinase G, KT5823. In addition, inhibitory effect of either SNAP or 8-bromo-cGMP on medullary Na+,K(+)-ATPase was abolished by 17-octadecynoic acid (17-ODYA), which inhibits cytochrome P450-dependent metabolism of arachidonic acid. These data suggest that NO decreases Na+,K(+)-ATPase activity in the renal medulla through the mechanism involving cGMP, protein kinase G, and cytochrome P450-dependent arachidonate metabolites. In contrast, NO has no effect on Na+,K(+)-ATPase in the renal cortex and on either cortical or medullary ouabain-sensitive H+,K(+)-ATPase.

The effect of peroxisome proliferator-activated receptors alpha (PPARalpha) agonist, fenofibrate, on lipid peroxidation, total antioxidant capacity, and plasma paraoxonase 1 (PON 1) activity.

The aim of this study was to investigate the effect of peroxisome proliferator activated receptors alpha agonist, fenofibrate, on the level of oxidative stress, total antioxidant capacity, and plasma paraoxonase 1 (PON 1) activity in the rat. The adult male Wistar rats received fenofibrate for 7 days. The drug was added to food at concentrations 0.005%, 0.05% and 0.5%, which corresponded to doses of 3, 30 and 300 mg/kg/day, respectively. Fenofibrate treatment dose-dependently reduced plasma concentration of malonyldialdehyde and 4-hydroxydialkenals. The level of these lipid peroxidation products in animals treated with 0.005%, 0.05% and 0.5% fenofibrate was lower than in control group by 52.8%, 62.7% and 87.1%, respectively. Lipid hydroperoxides in plasma decreased by 29.7%, 23.4% and 27.5% in these groups, respectively. The drug had no significant effect on total antioxidant capacity measured as ferric reducing ability of plasma (FRAP). Paraoxon-hydrolyzing activity (PON) of plasma paraoxonase was 81.5% lower in animals receiving 0.05% fenofibrate and 69.2% lower in rats treated with 0.5% fenofibrate than in control. Phenyl acetate hydrolyzing activity (arylesterase, AE) was reduced by 15.2%, 49.6% and 55.8% in rats receiving 0.005%, 0.05% and 0.5% fenofibrate, respectively. PON/AE ratio decreased following 0.05% and 0.5% fenofibrate by 64.9% and 30.4%, respectively. The drug had no significant effect on total plasma triglycerides and cholesterol concentrations. The results indicate that fenofibrate treatment favourably modulates oxidant-antioxidant balance and unfavourably affects plasma PON 1 activity in normolipidemic rats. These effects can contribute to the influence of PPARalpha agonists on pathological processes involved in atherogenesis.

The opposite effects of cyclic AMP-protein kinase a signal transduction pathway on renal cortical and medullary Na+,K+-ATPase activity.

Cyclic AMP-protein kinase A (PKA) pathway plays an important role in signal transduction in renal tubular cells, however, its role in transport regulation is not completely established. The aim of this study was to investigate in vivo the effect of PKA on renal Na, K-ATPase activity. The study was performed in male Wistar rats. The animals were anaesthetized with pentobarbital and investigated drugs were infused through the catheter inserted into the abdominal aorta. Na+,K+-ATPase activity was assayed in an isolated microsomal fraction of the renal cortex and medulla. Cell-permeable cAMP analogue, dibutyryl-cAMP (db-cAMP), dose-dependently stimulated Na+,K+-ATPase in the renal cortex and inhibited in the renal medulla. Maximal stimulation (+38.5%) and inhibition (-46.8%) were observed at a dose of 10(-6) mol/kg/min. Measurement of Na+,K+-ATPase activity at different Na' concentrations revealed that in the renal cortex db-cAMP increased Vmax of the enzyme without any effect on sodium affinity, whereas in the renal medulla decrease in Vmax was accompanied by decreased sodium affinity, evidenced by elevated K(0.5) for sodium. The effect of db-cAMP was mimicked by the infusion of either adenylate cyclase activator, forskolin, or inhibitor of phosphodiesterase, IBMX. Both stimulatory and inhibitory effects of db-cAMP were prevented by pretreatment with protein kinase A inhibitor, KT 5720 (10(-8) mol/kg/min) but not by inhibitor of protein kinase G, KT 5823. The inhibitory effect in the renal medulla was partially blocked by pretreatment with either ethoxyresorufin or 17-ODYA - two nonspecific inhibitors of cytochrome P450-dependent arachidonate metabolism, whereas an inhibitor of epoxygenase, miconazole, was not effective. Infusion of 20-hydroxyeicosatetraenoic acid (20-HETE) at a dose of 10(-10) mol/kg/min decreased medullary Na+,K+-ATPase activity by 24.2%. Exogenous protein phosphatases inhibitor, okadaic acid (OA, 10(-8) - 10(-7) mol/kg/min) caused dose-dependent decrease in renal medullary Na+,K+-ATPase activity, maximally by 31.9%, but had no effect in the renal cortex. The effects of OA and db-cAMP in the renal medulla were not additive. When OA administration (10(-7) mol/kg/min) was followed by 20-HETE (10(-10) mol/kg/min), medullary Na+,K-ATPase activity decreased by 48.6% and was similar as after db-cAMP. We conclude, that cAMP-PKA pathway activates Na+,K+-ATPase in the renal cortex and inhibits in the renal medulla. The inhibitory effect is partially mediated by cytochrome P450-dependent arachidonate metabolites and possibly also by PKA-dependent inhibition of protein phosphatases.

Selected markers of oxidative stress in rats with active Heymann nephritis.

BACKGROUND: Reactive oxygen species play an important role in the pathogenesis of different glomerulopathies. The purpose of this study was to examine selected markers of oxidative stress and antioxidant defense in active Heymann nephritis (AHN), which is a model of human membranous nephropathy. MATERIAL AND METHODS: AHN was induced in female Wistar rats by i.p. injection of proximal tubule brush border antigen (Fx1A). The control animals received an equivolume injection of saline. The animals were sacrificed at 6, 9, 15, 17, 18, 19, 22 and 23 weeks after Fx1A administration. We assayed the plasma concentration of thiobarbituric acid-reactive substances (TBARS) and nitric oxide metabolites (nitrites + nitrates, NOx) as well as total plasma antioxidant capacity (ferric reducing ability of plasma, FRAP) and serum paraoxonase (PON) activity. RESULTS: Fx1A-injected rats demonstrated a marked increase in proteinuria and impairment of renal excretory function evidenced by increased plasma creatinine and uric acid. Histologically, diffuse renal changes were seen, characterized by glomerular hypercellularity, increased glomerular size and narrowing of Bowman's space. Ultrastructural studies revealed diffuse fusion of foot processes and detachment of podocytes, subepithelial immune deposits in the glomerular capillary wall, thickening of the glomerular basement membrane, and local accumulation of lymphocytes and macrophages in glomeruli and interstitial cells. In the AHN group, TBARS increased significantly beginning in the 9th week, reaching maximum at the 23rd week (139.4% of time-matched control). The plasma concentration of NOx demonstrated a biphasic increase. The first peak (249.9% of control) was observed at the 9th week, followed by a decrease to normal between the 17th and 19th week. Then the NOx concentration increased to 211% of control at the 23rd week. FRAP began to increase in the 9th week and reached its maximum (134.9% of control) at the 15th week. After the 18th week FRAP returned progressively to control levels. PON activity was 17.0%, 19.1% and 21.3% lower in AHN than in the control group at the 19th, 22nd, and 23rd weeks, respectively. CONCLUSIONS: These results indicate that AHN is associated with oxidant-antioxidant imbalance, which may contribute to renal damage in this model of glomerulonephritis.

[Oxidative stress in glomerulonephritis]. / Stres oksydacyjny w zapaleniu klebuszków nerkowych.

The aim of this review is to focus on the possible role of reactive oxygen species (ROS) in the pathogenesis of glomerulonephritis (GN) and to review the potential mechanisms of this ROS-mediated renal injury. This paper examines the literature which demonstrates ROS as primary mediators in GN, responsible for modifications of glomerular permeability to proteins, development of morphologic lesions and alteration of glomerular haemodynamics (reduction in glomerular blood flow and glomerular filtration rate). In glomeruli, ROS are generated by both infiltrating cells (neutrophils, monocytes) and resident glomerular cells (mesangial and endothelial cells and podocytes). The participation of ROS in glomerular damage was proved in many experimental studies by detection of products of oxidant injury in renal tissue or urine and by demonstration of a protective effect of antioxidants in those lesions.

The effect of dietary-induced obesity on lipid peroxidation, antioxidant enzymes and total plasma antioxidant capacity.

The aim of this study was to examine the effect of dietary-induced obesity on some parameters of oxidative stress and antioxidant defence. The studies were performed in adult male Wistar rats. Control group received normal laboratory chow (62% calories as carbohydrates, 26% protein and 12% fat). High-calorie high-fat group (HCHF) was fed standard chow supplemented with lard (48% calories as carbohydrates, 20% as protein and 32% as fat) and high-calorie normal-fat group (HCNF) received standard chow and liquid diet containing sucrose, glucose, whole milk powder and soybean powder (60% carbohydrates, 26% protein, 14% fat). After 8 weeks body weight of HCHF and HCNF-fed rats was higher than body weight of controls by 9.3% and 15.2%, respectively. Plasma concentration of thiobarbituric acid-reactive substances (TBARS) increased in these groups by 43% and 52%, respectively. The activity of superoxide dismutase (SOD) decreased in HCHF group by 47.5% and in HCNF group by 21.1%. Glutathione peroxidase (GPx) activity in the blood tended to increase in both experimental groups but this was not significant. Plasma total antioxidant status (TAS) measuring the combined free radicals scavenging ability of nonenzymatic antioxidants was lower in HCHF and in HCNF group compared to control (-8.8% and -9%, respectively). The major observed lipid abnormalities were hypertriglyceridemia in HCHF group and decreased HDL-cholesterol in HCNF group. TBARS correlated negatively with SOD (r = -0.84, p < 0.001) and with TAS (r = -0.47, p < 0.05). These results indicate that obesity leads to oxidative stress which can contribute to obesity-associated diseases such as atherosclerosis, diabetes mellitus and arterial hypertension.

[Preliminary results of nasality tests in patients wtih cleft palate after operations performed with different surgical methods]. / Wstepne wyniki badan nosometrycznych u chorych z rozszczepem podniebienia operowanych róznymi metodami.

The study analyses results of tests carried out in a group of 82 patients with cleft palate defect after operations performed with different surgical methods. In the primary plastic group--the minimal degree of nasality was observed among patients after vomer flaps aged 2-3 years. The next group of patients after secondary surgical operations of phoniatric indication--better results were observed where Orticochea method was applied. The author expects to obtain more credible effects based on wider prospective and retrospective clinical observation.

[Dihydrotestosterone therapy of men with coronary artery disease and processes of peroxidation]. / Terapia dihydrotestosteronem mezczyzn z choroba wiencowa a procesy peroksydacyjne.

In 11 men after myocardial infarction (mean age 58.8 years) influence of 3 months transdermal dihydrotestosterone (DHT) treatment on malondialdehyde (MDA), total cholesterol, HDL cholesterol, triglycerides and platelets was estimated. Decrease in MDA concentration after DHT treatment only in men with low (< 13 nmol/l) testosterone (T) level (p = 0.06) was observed. MDA concentration after 3 months DHT treatment was significantly lower in hypotestosteronemic in comparison to normotestosteronemic men (p < 0.05). Plasma lipid levels and platelet count were not affected by DHT administration. Decrease serum MDA concentration in low testosterone men with coronary artery disease may suggest that DHT treatment in this group has beneficial effects on peroxidation processes.