The present study was performed to evaluate the role of neuronal nitric oxide synthase (nNOS)-derived nitric oxide (NO) during the developmental phase of hypertension in transgenic rats harboring the mouse Ren-2 renin gene (TGR). The first aim of the present study was to examine nNOS mRNA expression in the renal cortex and to assess the renal functional responses to intrarenal nNOS inhibition by S-methyl-L-thiocitrulline (L-SMTC) in heterozygous TGR and in age-matched transgene-negative Hannover Sprague-Dawley rats (HanSD). The second aim was to evaluate the role of the renal sympathetic nerves in mediating the renal functional responses to intrarenal nNOS inhibition. Thus, we also evaluated the effects of intrarenal L-SMTC administration in acutely denervated TGR and HanSD. Expression of nNOS mRNA in the renal cortex was significantly increased in TGR compared with HanSD. Intrarenal administration of L-SMTC decreased the glomerular filtration rate (GFR), renal plasma flow (RPF) and sodium excretion and increased renal vascular resistance (RVR) in HanSD. In contrast, intrarenal inhibition of nNOS by L-SMTC did not alter GFR, RPF or RVR and elicited a marked increase in sodium excretion in TGR. This effect of intrarenal L-SMTC was not observed in acutely denervated TGR. These results suggest that during the developmental phase of hypertension TGR exhibit an impaired renal vascular responsiveness to nNOS derived NO or an impaired ability to release NO by nNOS despite enhanced expression of nNOS mRNA in the renal cortex. In addition, the data indicate that nNOS-derived NO increases tubular sodium reabsorption in TGR and that the renal nerves play an important modulatory role in this process., L. Červenka, H. J. Kramer, J. Malý, I. Vaněčková, A. Bäcker, D. Bokemeyer, M. Bader, D. Ganten, K. D. Mitchell., and Obsahuje bibliografii
It has been suggested that increase in acute nitric oxide (NO) or cyclic guanosine monophosphate production may be involved in cardioprotection induced by chronic hypoxia (CH). We studied the effect of NO donor molsi domine and phosphodiesterase type 5 inhibitor sildenafil on myocardial ischemia/reperfusion (I/R) injury in rats adapted to CH. Male Wistar rats were exposed to continuous hypoxia in a normobaric chamber (10 % O 2 , 4 weeks). Rats received either saline, mol sidomine (10 mg/kg body weight, i.v.) or sildenafil (0.7 mg/kg body weight, i.v.) 30 min before ischemia. Control rats were kept under normoxia and treated in a corresponding manner. Adaptation to CH increased the myocardial ischemic tolerance. Acute treat ment with either molsidomine or sildenafil significantly reduced infarct size in normoxic rats and further enhanced cardioprotection induced by CH. However, the cardioprotective effect of CH on I/R injury was not additive to the cardioprotection provided b y the drugs., P. Alánová, F. Kolář, B. Ošťádal, J. Neckář., and Obsahuje bibliografii
Remodeled pulmonary arteries return to normal structural conditions after the increase in pulmonary artery flow resistance is reversed. We studied whether proteolysis of extracellular matrix proteins and apoptosis occur during reversal of remodeling produced by chronic hypoxia in the rat. Main pulmonary arteries were removed at different times during a 10-day period of exposure to 10% O2 and 14 days after return to air. Content and rates of degradation of collagen and elastin as well as immunoreactive collagenase in tissue and isolated mast cells were measured. Immunoblots for collagenase and tissue inhibitor of metalloproteinases (TIMP) were performed. Apoptosis was assessed by cleavage of DNA and TUNEL assay. Excess collagen and elastin present at 10 days of hypoxia decreased to near normal levels after 3-5 days of air. Transient increases in collagenolytic and elastolytic enzyme activities accompanied the rapid decrease in matrix proteins. Mast cells containing collagenase accumulated in remodeled pulmonary arteries, and the active form of collagenase appeared at the time of peak proteolytic activity. TIMP increased during remodeling. Apoptosis was maximal 3 days after return to air. Our results suggest that activation of enzymes, which degrade matrix proteins, and apoptosis play a role in resolution of vascular remodeling., D. J. Riley, S. Thakker-Varia, F. J. Wilson, G. J. Poiani, C. A. Tozzi., and Obsahuje bibliografii
Fertilization process is a very clever and unique process comprising some essential steps resulting in formation of zygote. Tetraspanin CD9 is considered to be a serious candidate molecule participating in these events. The importance of CD9 has been discussed in relation to acrosome reaction, sperm-binding, sperm-penetration, sperm-egg fusion and eventually, egg activation. The abundant expression of CD9 oocyte plasma membrane and the presence of CD9-containing vesicles in the perivitelline space of intact oocytes have been confirmed. Despite the fact that majority of authors analyzed CD9 expressed on oocytes, several studies considered the function of sperm CD9, too. To understand CD9 involvement, various conditions of in vitro fertilization (IVF) assays using polyclonal as well as monoclonal antibodies or knockout mice were carried out. However, ambiguous data have been obtained about the importance of CD9 in sperm-egg binding or fusion. Although the current findings did not prove any hypothesis, the indispensable role of CD9 in fertilization process was not excluded and the precise role of CD9 remains unexplained., J. Jankovičová, M. Simon, J. Antalíková, P. Cupperová, K. Michalková., and Obsahuje bibliografii
Mammalian P2X receptors contain 10 conserved cysteine residues in their ectodomains, which form five disulfide bonds (SS1-5). Here, we analyzed the relevance of these disulfide pairs in rat P2X4 receptor function by replacing one or both cysteines with alanine or threonine, expressing receptors in HEK293 cells and studying their responsiveness to ATP in the absence and presence of ivermectin, an allostenic modulator of these channels. Response to ATP was not altered when both cysteines forming the SS3 bond (C132-C159) were replaced with threonines. Replacem ent of SS1 (C116-C165), SS2 (C126-C149) and SS4 (C217-C227), but not SS5 (C261-C270), cysteine pairs with threonines resulted in de creased sensitivity to ATP and faster deactivation times. The maximum current amplitude was reduced in SS2, SS4 and SS5 double mutants and could be partially rescued by ivermectin in SS2 and SS5 double mutants. This response pattern was also observed in numerous single residue mutants, but receptor function was not affected when the 217 cysteine was replaced with threonine or arginine or when the 261 cysteine was replaced with alanine. These results suggest that the SS1, SS2 and SS4 bonds contribute substantially to the structure of the ligand binding pocket, while the SS5 bond located towards the transmembrane domain contributes to receptor gating., M. B. Rokic, V. Tvrdoňová, V. Vávra, M. Jindřichová, T. Obšil, S. S. Stojilkovic, H. Zemková., and Obsahuje bibliografii
Steroid sulfatase (EC 3.1.6.2) is an important enzyme involved in steroid hormone metabolism. It catalyzes the hydrolysis of steroid sulfates into their unconjugated forms. This action rapidly changes their physiological and biochemical properties, especially in brain and neural tissue. As a result, any imbalance in steroid sulfatase activity may remarkably influence physiological levels of active steroid hormones with serious consequences. Despite that the structure of the enzyme has been completely resolved there is still not enough information about the regulation of its expression and action in various tissues. In the past few years research into the enzyme prope ties and regulations has been strongly driven by the discovery of its putative role in the indirect stimulation of the growth of hormone-dependent tumors of the breast and prostate., L. Kříž, M. Bičíková, R. Hampl., and Obsahuje bibliografii a bibliografické odkazy
Enhanced production of superoxide radicals by nicotinamideadenine dinucleotide phosphate (NADPH) oxidase in the brain and/or kidney of salt hypertensive Dahl rats has been proposed to participate in the pathogenesis of this form of experimental hypertension. Most information was obtained in young Dahl saltsensitive (DS) rats subjected to high salt intake prior to sexual maturation. Therefore, the aim of our study was to investigate whether salt hypertension induced in adult DS rats is also accompanied with a more pronounced oxidative stress in the brain or kidney as compared to Dahl salt-resistant (DR) controls. NADPH oxidase activity as well as the content of thiobarbituric acid-reactive substances (TBARS) and conjugated dienes (oxidative index), which indicate a degree of lipid peroxidation, were evaluated in two brain regions (containing either hypothalamic paraventricular nucleus or rostral ventrolateral medulla) as well as in renal medulla and cortex. High salt intake induced hypertension in DS rats but did not modify blood pressure in DR rats. DS and DR rats did not differ in NADPH oxidase-dependent production of ROS, TBARS content or oxidative index in either part of the brain. In addition, high-salt diet did not change significantly any of these brain parameters. In contrast, the enhanced NADPH oxidase-mediated ROS production (without significant signs of increased lipid peroxidation) was detected in the renal medulla of salt hypertensive DS rats. Our findings suggest that there are no signs of enhanced oxidative stress in the brain of adult Dahl rats with salt hypertension induced in adulthood., M. Vokurková, H. Rauchová, L. Řezáčová, I. Vaněčková, J. Zicha., and Obsahuje bibliografii
Two mechanisms contribute in the development of pulmonary hypertension in pulmonary embolism (PE) - obstruction of pulmonary blood vessels and vasoconstriction. We hypothesize that hypoxia, increased shear stress and/or activation of gathered leukocytes in the PE may cause a release of reactive oxygen species (ROS). Therefore our aim was to determine the influence of the ROS scavenger Tempol on pulmonary hypertension and to d escribe NO synthase activity and production of NO oxidative products (NOx) after PE. In general anesthesia sephadex microspheres suspended in PSS were applied in right jugular vein as the pulmonary microembolism. Than we measured in isolated salt solution -perfused lungs the changes in perfusion pressure, activity of NO synthase and NOx plasma concentration in 7 groups of rats: C: control group (n=5), CN: C + sodium nitroprusside (SN) (n=5), EN: PE + SN (n=5), ETN: Tempol + PE + SN (n=5), CL : C + L -NAME (n=5 ), EL: PE + L-NAME (n=5), ETL: Tempol + PE + L -NAME (n=5). Tempol was applied intraperitoneally before PE. Animals that received Tempol (groups TN, TL) had significantly lower basal perfusion pressure than those which did not rec eive Tempol (EN, EL). Overa ll we measured a higher decrease of perfusion pressure than in the control group (C) after applica tion of SN. Administration of L-NAME after PE (EL) increased the pressure more than in the control group (NL). NOx concentration was higher after PE. We found that preventive administration of Tempol decreases the increase in perfusion pressure after PE. PE increased NO release and concentration of NOx., R. Mizera, D. Hodyc, J. Herget., and Obsahuje bibliografii
Diabetes mellitus is associated with increased inflammatory response, which may contribute to atherosclerosis progression. Experimental results demonstrated anti-inflammatory activity of glitazones; their effect on leukocyte adhesion molecules has not been studied to date. We therefore studied the effect of rosiglitazone treatment on leukocyte surface expression of adhesion molecules in patients with type 2 diabetes mellitus and compared our results with findings in healthy subjects. 33 subjects with type 2 diabetes and 32 healthy controls were included; patients were examined at baseline and after 5 months of rosiglitazone treatment (4 mg /d). Leukocyte expression of adhesion molecules LFA-1, CD 18 and ICAM-1 was quantified using flow cytometry; in addition, CD14 (lipopolysaccharide receptor) expression was analyzed as a marker of nonspecific immunity. The expression of examined molecules at baseline was higher in patients compared to controls. Despite only mild decrease in blood glucose, ro siglitazone treatment induced substantial decrease of CD18 and CD14 expression and borderline decrease of LFA-1 and ICAM-1 expression (on monocytes only). We thus observed improvement in the expression of leukocyte inflammatory markers after rosiglitazone treatment. This effect is supposed to be mediated by direct effect of rosiglitazone on PPAR- γ receptors on leukocytes., T. Štulc, H. Svobodová, Z. Krupičková, R. Doležalová, I. Marinov, R. Češka., and Obsahuje bibliografii
Recently, we derived “humanized” spontaneously hypertensive rats (SHR-CRP) in which transgenic expression of human CRP induces inflammation, oxidative stress, several features of metabolic syndrome and target organ injury. In addition, we found that rosuvastatin treatment of SHR-CRP transgenic rats can protect against pro-inflammatory effects of human CRP and also reduce cardiac inflammation and oxidative damage. In the current study, we tested the effects of rosuvastatin (5 mg/kg) on kidney injury in SHR-CRP males versus untreated SHR-CRP and SHR controls. All rats were fed a high sucrose diet. In SHR-CRP transgenic rats, treatment with rosuvastatin for 10 weeks, compared to untreated transgenic rats and SHR controls, was associated with significantly reduced systemic inflammation which was accompanied with activation of antioxidative enzymes in the kidney, lower renal fat accumulation, and with amelioration of histopathological changes in the kidney. These findings provide evidence that, in the presence of high CRP levels, rosuvastatin exhibits significant anti-inflammatory, anti-oxidative, and renoprotective effects., J. Šilhavý, V. Zídek, V. Landa, M. Šimáková, P. Mlejnek, O. Oliyarnyk, H. Malínská, L. Kazdová, M. Mancini, M. Pravenec., and Obsahuje bibliografii