Plasma osteopontin levels, but not its myocardial expression, reflect heart failure severity in recently diagnosed dilated cardiomyopathy.

BACKGROUND: Elevated levels of the extracellular matrix glycoprotein osteopontin (OPN) may be detected in both myocardium and plasma under various pathological conditions affecting the heart. Several studies demonstrated increased plasma OPN levels in patients with heart failure due to dilated cardiomyopathy (DCM), while other studies showed high OPN expression levels in the myocardium of such patients. However, very little is known about OPN levels in both plasma and myocardium of the same individual with DCM. Therefore, we aimed to compare plasma OPN levels and levels of myocardial OPN expression in patients with recent-onset DCM (Ro-DCM). METHODS: We examined plasma OPN as well as creatinine, C­reactive protein (CRP), brain natriuretic peptide (BNP), and troponin I levels in 25 patients with Ro-DCM. Furthermore, all subjects underwent transthoracic echocardiography, selective coronary angiography, and endomyocardial biopsy (EMB) for the assessment of myocardial OPN expression. RESULTS: No significant correlation between myocardial OPN expression and clinical, biochemical, or echocardiographic parameters was found. In log transformation analysis, plasma OPN levels correlated significantly with BNP levels (r = 0.46, p = 0.031), with CRP levels (r = 0.52, p = 0.015), and with early diastolic mitral annular velocity (r = -0.57, p = 0.009). There was a borderline association between the plasma OPN log value and New York Heart Association class (p = 0.053). CONCLUSION: Plasma OPN levels reflect heart failure severity in patients with Ro-DCM. Myocardial OPN expression is not associated with either plasma OPN levels or markers of heart failure in these individuals.

Polymorphisms rs2167444 and rs508384 in the SCD1 Gene Are Linked with High ApoB-48 Levels and Adverse Profile of Cardiometabolic Risk Factors.

Elevated plasma concentration of apolipoprotein B-48 (apoB-48) is an independent risk factor of cardiovascular disease. Stearoyl-CoA desaturase-1 (SCD1) is a rate-limiting lipogenic enzyme and a key regulator of fuel metabolism. The aim of this study was to analyse associations between clinical, biochemical, and genetic factors and different apoB-48 levels in subjects at increased cardiometabolic risk. We examined 220 subjects exhibiting at least one metabolic syndrome (MetS) component. In conjunction with basic clinical, anthropometric and laboratory measurements, we analysed various polymorphisms of stearoyl-CoA desaturase-1 (SCD1). Subjects were divided into two groups according to the median apoB-48 level: (1) high apoB-48 (≥ 7.9 mg/l, N = 112) and (2) low apoB-48 (< 7.9 mg/l, N = 108). Neither group differed significantly in anthropometric measures. High plasma apoB-48 levels were associated with increased systolic blood pressure (+3 %; P < 0.05), MetS prevalence (59.8 vs. 32.4 %; P < 0.001), small-dense LDL frequency (46.4 vs. 20.4 %; P < 0.001), triglycerides (+97 %; P < 0.001), non-HDLcholesterol (+27 %; P < 0.001), and lower concentrations of HDL-cholesterol (-11 %; P < 0.01). This group was further characterized by a higher HOMA-IR index (+54 %; P < 0.001) and increased concentrations of conjugated dienes (+11 %; P < 0.001) and oxidatively modified LDL (+ 38 %; P < 0.05). Lower frequencies of SCD1 minor genotypes (rs2167444, rs508384, P < 0.05) were observed in subjects with elevated plasma concentrations of apoB-48. Elevated plasma concentrations of apoB-48 are associated with an adverse lipid profile, higher systolic blood pressure, insulin resistance, and oxidative stress. Lower proportions of minor SCD1 genotypes (rs2167444, rs508384) implicate the role of genetic factors in the pathogenesis of elevated levels of apoB-48.

Plasma osteopontin levels in patients with dilated and hypertrophic cardiomyopathy.

BACKGROUND: Osteopontin (OPN) is an extracellular matrix glycoprotein that plays a role in a variety of cellular activities associated with inflammatory and fibrotic responses. Increased OPN levels in myocardium and plasma have been demonstrated in patients with dilated cardiomyopathy (DCM). However, nothing is known about OPN levels in patients with hypertrophic cardiomyopathy (HCM). Therefore, the aim of our study was to compare plasma OPN levels in patients with these two most common cardiomyopathies. PATIENTS AND METHODS: We examined plasma OPN as well as creatinine, C­reactive protein (CRP), brain-type natriuretic peptide (BNP), and troponin I levels in 64 patients with DCM, 43 patients with HCM, and 75 control subjects. Transthoracic echocardiography was also performed on all cardiomyopathy patients. RESULTS: Plasma OPN levels were significantly elevated in patients with DCM compared with HCM patients (95 ± 43 vs. 57 ± 21 ng/ml; p < 0.001) and control subjects (54 ± 19 ng/ml; p < 0.001); however, there was no difference between HCM patients and control subjects. New York Heart Association (NYHA) class III or IV disease was more frequently present in DCM patients than in HCM subjects (44 % vs. 2 %, p < 0.0001). In multivariate analysis, BNP and CRP levels together with NYHA class were found to be significant predictors of plasma OPN levels in DCM patients (p = 0.002, p = 0.029, and p < 0.001 for BNP, CRP, and NYHA, respectively). CONCLUSION: Plasma OPN levels were associated with overall heart failure severity rather than with specific cardiomyopathy subtype in patients suffering from DCM or HCM, respectively.

Fructosamine 3-kinase and glyoxalase I polymorphisms and their association with soluble RAGE and adhesion molecules in diabetes.

Advanced glycation end-products (AGEs) are key players in pathogenesis of long-term vascular diabetes complications. Several enzymes such as fructosamine 3-kinase (FN3K) and glyoxalase I (GLO I) are crucial in preventing glycation processes. The aim of our study was to evaluate an association of FN3K (rs1056534, rs3848403) and GLO1 rs4746 polymorphisms with parameters of endothelial dysfunction and soluble receptor for AGEs (sRAGE) in 595 diabetic and non-diabetic subjects. Genotypic and allelic frequencies of mentioned polymorphisms did not differ between subgroups. In diabetic patients significant differences were observed in sRAGE concentrations according to their rs1056534 and rs3848403 genotype. While GG and CG genotypes of rs1056534 with mutated G allele were associated with significant decrease of sRAGE (GG: 1055+/-458 and CG: 983+/-363 vs. CC: 1796+/-987 ng/l, p<0.0001), in rs3848403 polymorphism TT genotype with mutated T allele was related with significant sRAGE increase (TT: 1365+/-852 vs. CT: 1016+/-401 and CC: 1087+/-508 ng/l, p=0.05). Significant differences in adhesion molecules were observed in genotype subgroups of GLO1 rs4746 polymorphism. In conclusion, this is the first study describing significant relationship of FN3K (rs1056534) and (rs3848403) polymorphisms with concentration of sRAGE in patients with diabetes.

Genetic variation screening of TNNT2 gene in a cohort of patients with hypertrophic and dilated cardiomyopathy.

Mutations in troponin T (TNNT2) gene represent the important part of currently identified disease-causing mutations in hypertrophic (HCM) and dilated (DCM) cardiomyopathy. The aim of this study was to analyze TNNT2 gene exons in patients with HCM and DCM diagnosis to improve diagnostic and genetic consultancy in affected families. All 15 exons and their flanking regions of the TNNT2 gene were analyzed by DNA sequence analysis in 174 patients with HCM and DCM diagnosis. We identified genetic variations in TNNT2 exon regions in 56 patients and genetic variations in TNNT2 intron regions in 164 patients. Two patients were found to carry unique mutations in the TNNT2 gene. Limited genetic screening analysis is not suitable for routine testing of disease-causing mutations in patients with HCM and DCM as only individual mutation-positive cases may be identified. Therefore, this approach cannot be recommended for daily clinical practice even though, due to financial constraints, it currently represents the only available strategy in a majority of cardio-centers.

Paraoxonase-1 (PON1) status in pancreatic cancer: relation to clinical parameters.

Human paraoxonase 1 (PON1) has been shown to decrease the level of systemic oxidative stress, which is thought to contribute to cancer development. The aim of this study was to examine the interrelationships between PON1 status and some clinical characteristics in patients with pancreatic cancer (PC). A group of 73 consecutive patients with PC (stage II-IV) and 73 control subjects were examined. Laboratory studies included five polymorphisms of the PON1 gene (L55M, Q192R, -108C/T, -126C/T, and -162A/G), PON1 arylesterase (PON1-A) and lactonase (PON1-L) activities, as well as some markers of protein metabolism, insulin resistance, and oxidative stress. In comparison with the control group, no difference in the distribution of the PON1 polymorphisms was found in cancer patients, both arylesterase and lactonase activities being significantly lower (-33, -47 %, respectively, both P < 0.001). There was neither statistically significant association of PON1 polymorphisms with tumour stages nor with diabetes mellitus connected with PC. The genotype distribution of L55M and ­108C/T differed only in a subgroup of patients presenting clinically relevant malnutrition (χ² = 6.50, 6.25, respectively, both P < 0.05). In the PC group, PON1-A and PON1-L activities correlated with Nutritional Risk Index (r = 0.351, 0.409, respectively, both P < 0.01), PON1-L with mid-arm muscle circumference (r = 0.328, P < 0.05), and PON1-A and PON1-L with serum albumin (r = 0.352, 0.391 respectively, both < 0.01). Our results suggest that PON1 plays an important role in PC, especially in cancer-associated malnutrition.

Pregnancy-associated plasma protein A polymorphisms in patients with risk pregnancies.

Pregnant women are often threatened by hypertension, symptoms of preterm labour, hepatopathy, and other. These complications might be the consequence of genetic factors together with involvement of environmental factors. We were searching for three polymorphisms Arg654Lys, Ala678Pro and Thr686Ala in exon 5, and two polymorphisms Phe802Leu, Ser827Ser/Leu in exon 7, and for the new mutations in exons 5 and 7 of the pregnancy-associated plasma protein A gene in the studied group consisting of 203 women - 79 pregnant women in time of preterm labour, 24 pregnant women suffering from preeclampsia, and 100 healthy pregnant and non-pregnant women serving as controls. We did not find any divergence from wild-type form of these polymorphisms in any of the studied groups, which led us to the hypothesis that these polymorphisms are not associated with our studied group of Caucasian origin. However, further studies with a larger group of subjects are needed to confirm our results.

Leptin and adiponectin in pancreatic cancer: connection with diabetes mellitus.

UNLABELLED: The aim of this study was to analyze the relationship of serum leptin as well as adiponectin and the manifestation of pancreatic cancer (PC). Serum leptin, adiponectin, glucose homeostasis and insulin resistance (expressed as HOMA-IR) were investigated in 64 patients with newly diagnosed PC and compared with 64 healthy controls (CON group) and 75 patients with type 2 diabetes (DM2). Seventy percent of newly diagnosed PC patients had DM2. The levels of leptin were lower, whilst adiponectin/leptin ratio was higher in PC patients (both with and without DM2), in comparison with CON and DM2 groups (P < 0.001) independently of age, BMI and waist circumference. Newly diagnosed PC is characterized with lower leptin concentrations and higher adiponectin/leptin ratio in comparison with CON or DM2 individuals. Analysis of these parameters could help in the screening of persons in high risk for PC, especially in those with DM2. KEYWORDS: adiponectin, leptin, pancreatic cancer, type 2 diabetes mellitus.

Polymorphisms of genes for brain-derived neurotrophic factor, methylenetetrahydrofolate reductase, tyrosine hydroxylase, and endothelial nitric oxide synthase in depression and metabolic syndrome.

The prevalence of metabolic syndrome as well as the occurrence of depressive disorder, which are both connected with increased risk of diabetes mellitus type 2 and cardiovascular diseases, is continually increasing worldwide. These disorders are interconnected at various levels; the genetic one seems to be promising. Contribution of genetic factors to the aetiopathogenesis of depressive disorder weighs within the range 40-50 %, whereas the genetic background for the manifestation of metabolic syndrome is more complicated. In this pilot study, we investigated the incidence of polymorphisms in several genes supposed to play a role in the development of both depressive disorder and metabolic syndrome such as brain-derived neurotrophic factor, methylenetetrahydrofolate reductase, tyrosine hydroxylase, and endothelial nitric oxide synthase. The entire group consisted of 42 patients with depressive disorder, 57 probands with metabolic syndrome and 41 control individuals. We found that genotype Met/Met of the Val66Met polymorphism of the brain-derived neurotrophic factor gene was positively associated with depressive disorder (P < 0.05), but we were not able to find any significant associations of both the depressive disorder and metabolic syndrome with the remaining polymorphisms studied (methylenetetrahydrofolate reductase 677CT, methylenetet rahydrofolate reductase 1298AC, endothelial nitric oxide synthase Glu298Asp, and tyrosine hydroxylase).

[Features of metabolic syndrome in patients with depressive disorder]. / Rysy metabolického syndromu u nemocných s depresivní poruchou.

BACKGROUND: Depressive disorder is a serious illness with a high incidence, proxime accessit after anxiety disorders among the psychiatric diseases. It is accompanied by an increased risk of development of type 2 diabetes mellitus, cardiovascular disease, and by increased all-cause mortality. Recently published data have suggested that factors connected with the insulin resistance are at the background of this association. METHODS AND RESULTS: In this pilot study we have investigated parameters of lipid metabolism and glucose homeostasis in consecutively admitted patients suffering from depressive disorder (DD) (group of 42 people), in 57 patients with the metabolic syndrome (MetS) and in a control group of 49 apparently healthy persons (CON). Depressive patients did not differ from the control group by age or body mass index (BMI) value, but they had statistically significantly higher concentrations of serum insulin, C-peptide, glucose, triglycerides (TG), conjugated dienes in LDL particles (CD-LDL), higher value of microalbuminuria and of insulin resistance (HOMA-IR) index. They simultaneously had significantly lower value of the insulin sensitivity (QUICKI) index. In comparison with the MetS group the depressive patients were characterized by significantly lower both systolic and diastolic blood pressure, BMI , serum TG, apolipoprotein B, uric acid, C-peptide and by higher concentrations of apolipoprotein A-I and HDL-cholesterol. On the contrary, we have not found statistically significant differences between the DD and MetS groups in the concentrations of serum insulin, glucose, HOMA and QUICKI indices, in CD-LDL and MAU. CONCLUSIONS: In this pilot study, we have found in patients with depressive disorder certain features of metabolic syndrome, especially insulin resistance and oxidative stress.

Polymorphism -23HPhI in the promoter of insulin gene and pancreatic cancer: a pilot study.

Pancreatic cancer (PC) is one of the most frequent gastrointestinal malignancies with extremely poor prognosis. In spite of a relative low incidence of PC, in comparison with other cancers, PC is the fourth leading cause of cancer death in USA in both sexes. The available data clearly suggest that diabetes mellitus (DM) can be both a long-standing cause of PC and an early manifestation of the disease. Besides of DM, insulin resistance and high insulin levels are linked as well with increased cancer risk, including PC. The variable number of tandem repeats (VNTR) locus upstream of the insulin gene (INS) regulates insulin expression and has been associated with susceptibility to many diseases including DM. It is known that there is nearly complete linkage disequilibrium of the insulin variable tandem of repeats (INS-VNTR) alleles I/III with neighboring -23 HphI A/T single nucleotide polymorphism (SNP) in Caucasians. Therefore, we have studied the association between SNP of -23HphI in promoter of INS with PC, DM Type 2 (2TDM) and healthy controls. In this study we investigated 153 subjects (86 M/67 F); 51 patients with newly-diagnosed PC (31 M/20 F), 45 patients with 2TDM (29 M/16 F) and 57 healthy control subjects (26 M/31 F). The polymorphism of -23HphI (A/T) in the promoter of INS was determined by the combination of polymerase chain reaction (PCR) with the restriction fragment length polymorphism (RFLP) methods. The results obtained by the PCR-RFLP analyses of SNP -23HphI were confirmed by a direct studied locus sequencing of the genomic DNA. The frequency of abnormal glucose metabolism (both DM and impaired fasting glucose) was 88 % (45/51) in PC group. The AA genotype in SNP -23HphI was more prevalent (67 % vs. 47 %; P<0.05) among PC patients compared to controls. Additionally, statistically significant differences were found in frequencies (%) of genotypes AA/AT/TT in groups with PC (67/27/6), 2TDM (53/40/7) compared to healthy controls (37/46/17) (P<0.05). Moreover, a statistically significant effect of -23HphI A/T polymorphism on tumor staging was found (P< 0.05). Polymorphism of -23HphI (A/T) in the promoter of INS may play a role in the pathogenesis of PC and could contribute to tumor staging. Key words: pancreatic cancer; insulin gene regulation; polymorphism of -23HphI; diabetes mellitus; disorders of glucoregulation.

Association of the -344T/C aldosterone synthase gene variant with essential hypertension.

The aldosterone synthase gene (CYP11B2) is an important candidate gene region in essential hypertension. We therefore studied the association of -344T/C polymorphism of the CYP11B2 gene with the presence and severity of hypertension in a case-control study. We studied 369 individuals, of whom 213 were hypertensive patients (139 controlled hypertensive, 74 resistant hypertensive) and 156 were healthy normotensive subjects. The -344T/C polymorphism of the CYP11B2 gene was determined using polymerase chain reaction - restriction fragment length polymorphism analysis. The distribution of genotypes in normotensive controls and hypertensive subjects were: TT 25.6 vs. 31.9 %, TC 51.9 vs. 57.3 % and CC 22.4 vs. 10.8 %. The -344T/C variant was associated with hypertension. Subjects carrying the -344T allele had a greater risk of hypertension compared to those having C allele (chi(2)=5.89, p<0.05). The frequency of CC genotype was significantly lower in hypertensive patients than in normotensive controls (chi(2)=9.44, p<0.01). A stepwise logistic regression analysis confirmed these findings. We did not find an association of -344T/C variant with the resistance of hypertensive patients to combination therapy, but we observed an association of -344T/C polymorphism of aldosterone synthase gene with increased risk of hypertension. These results support a potential role of -344T/C CYP11B2 gene polymorphism in genetic predisposition to develop hypertension.

[Metabolic syndrome and depression--clinical relations]. / Metabolický syndrom a deprese--klinické vztahy.

The occurrence of both obesity and type 2 diabetes mellitus is rapidly increasing; according to WHO data, this can be considered as a worldwide epidemic. The obesity is one of the components of metabolic syndrome, the cluster of several risk factors of atherosclerosis such as dyslipidemia, hypertension, impaired glucose homeostasis, pro-thrombotic state and subclinical inflammation. The importance of the metabolic syndrome is confirmed by findings of the several times increased risk of both the type 2 diabetes mellitus and cardiovascular disease. Similarly, as in the case of obesity and diabetes, the incidence and prevalence of depressive disorder are still increasing and depressive disorder belongs to the most important causes of disability. The interrelations between depressive disorder and diabetes are known for a long time. Diabetics very often suffer from depression and vice versa, the depressive disorder is a significant risk factor of type 2 diabetes mellitus development and worsens the survival of diabetics. Those relationships have been recently intensively studied. Our paper reviews genetic, nutritional, metabolic and hormonal factors, contributing to the above mentioned syndrome.

[Composition of the nonesterified fatty acids and lipid peroxidation in metabolic syndrome]. / Slození esterifikovaných mastných kyselin a lipoperoxidace u metabolického syndromu.

BACKGROUND: Composition of the nonesterified fatty acids in plasma in metabolic syndrome patients and in other syndromes of insulin resistance is altered. Fatty acid profile in plasma is related to the composition of dietary fat and to the metabolic changes of fatty acids, e.g. to de novo lipogenesis, beta-oxidation and conversion accompanying the oxidative stress. The aim of the work was to study the fatty acid composition in the major plasma lipid classes in relation to the insulin resistance, to some polymorphisms of candidate genes with activity related to insulin resistance, and to the lipoprotein composition and parameters of lipid peroxidation. METHODS AND RESULTS: 95 patients with metabolic syndrome (56 M/39 F) and 195 healthy persons (99 M/96 F) were included into the cohort. Basic clinical data, parameters of glucose homeostasis, lipid concentration in plasma and conjugated diens in LDL were determined. Fatty acids were detected by capillary gas chromatography. Polymorphisms of apolipoprotein E, intestinal isoforms of fatty acid binding protein (Ala54Thr) and y-2 isoforms of peroxisomal activated receptor (Alal2Pro) were analyzed using combination of polymerase chain reaction methods and by the detection of polymorphisms of the restriction fragment length. Persons with metabolic syndrome had higher concentrations of CRP and conjugated diens in LDL. In all lipid classes we proved a decreased concentration of n-6 polyunsaturated fatty acids and an increase of unsaturated fatty acids. From all the acids, the only significant was the decrease of linolic acid concentration and the increase of palmitic and palmitoyl acids. Results showed an increase of delta 9 palmitic acid desaturase activity, delta 6 linolic acid desaturase and elongase activity. Concentration of conjugated diens in LDL inversely correlated with linolic acid. Clinical or laboratory parameters and homozygotic combination of polymorphism studied were not mutually related. CONCLUSIONS: Changes in the profile of fatty acids during the metabolic syndrome results from the elevated lipogenesis and from the higher level of oxidative stress.

[Factors modulating parameters of cholesterol homeostasis in the metabolic syndrome]. / Faktory ovlivnující ukazatele homeostázy cholesterolu u metabolického syndromu.

BACKGROUND: Newly described component of the metabolic syndrome is the elevated synthesis of cholesterol accompanied with its decreased intestinal absorption. The aim of our study was to ascertain the incidence of genotypes and alleles of several candidate genes, which modulate insulin resistance and metabolism of lipids and to find their role in lipid, lipoprotein and cholesterol homeostasis. The concentrations of cholesterol precursors (lathosterol, desmosterol, respectively their rations to cholesterol) are related to the synthesis of cholesterol; concentrations of fytosterols (kampesterol, sitosterol, respectively their rations to cholesterol) are related to the intestinal absorption of cholesterol. METHODS AND RESULTS: 95 patients with metabolic syndrome (56 M/39 F) and 195 healthy persons (99 M/96 F) were included into the study. Beside the basic clinical and anthropometric data, parameters of glucose homeostasis, plasma concentration of lipids, ultracentrifugation separated lipoproteins, and conjugated diens in LDL were determined. Non-cholesterol sterols were estimated by capillary gas chromatography. Polymorphisms of apolipoprotein E, intestinal isoforms of fatty acids binding protein (Ala54Thr), microsomal transfer protein (-493G/T), and gamma-2 isoforms of peroxisomal proliferator activated receptor (Ala12Pro) were analysed by combination of methods of polymerase chain reaction and by determination of polymorphism of the length of restriction fragments. After adjustation to the age, patients with metabolic syndrome had higher BMI, body fat and lean body weight (all P < 0.001), waist circumference, systolic and diastolic blood pressure (all P < 0.01). At the same time they had higher levels of glucose, insulin (P < 0.001), C-peptide, CRP (P < 0.05), uric acid, conjugated diens in LDL and HOMA insulin resistance index (P < 0.001). After adjustation to the age, higher concentration of triglycerides (P < 0.001), apo B (P < 0.01), cholesterol and triglycerides in VLDL (both P < 0.001), triglycerides in LDL (P < 0.01) were found. Incidence of alleles and genotypes of studied polymorphisms did not differ in both groups. Cholesterol synthesis is modulated by the presence of metabolic syndrome and by sex; cholesterol resorption is modulated only by the presence of metabolic syndrome. CONCLUSIONS: In patients with metabolic syndrome we found higher synthesis and lower intestinal absorption of cholesterol. We did not confirm relation between alleles of studied polymorphisms and clinical and anthropometric parameters, neither relation of these alleles to lipid or lipoprotein levels, oxidation stress, inflammation, or parameters of synthesis and absorption of cholesterol.

[Nitric oxide synthase, typical flavohemoproteins and their complicated enzymology]. / Syntázy oxidu dusnatého, typické hemoflavoproteiny a jejich komplikovaná enzymologie.

Nitric oxide is a diatomic gaseous molecule with unpaired electron in the molecule. Physical properties such as solubility, diffusibility and half-life decide the chemical reactivity of nitric oxide. Nitric oxide is the unstable free radical in vessels, immune system and central nervous system. The reactivity of nitric oxide under physiological and pathological conditions depends upon its concentration and site of production. Nitric oxide is thought to play a role in many pathological situations: septic shock, cardiovascular diseases, arthritis, diabetes, multiple sclerosis, asthma, and hypertension. Nitric oxide synthase is a self-sufficient flavohemoprotein capable of producing nitric oxide from L-arginine by two successive monooxygenation steps. Although the N-terminal heme domain functionally resembles cytochromes P450, no structural similarities exist between cytochrome P450 and nitric oxide synthases heme domains. The C-terminal domain of nitric oxide synthases containing flavin adenine dinucleotide and flavin mononucleotide as cofactors exhibits a high degree of sequence similarity with NADPH-cytocrome P450 reductase. The reductase domains serve as an intermediary for the transfer of electrons from NADPH for the catalytic reaction. The connecting domain between the oxygenase and the reductase domains of nitric oxide synthase isoforms binds calmodulin in the presence of calcium. The binding of calmodulin to all nitric oxide synthase isoforms is obligatory for the production of nitric oxide. At the same time, the presence of one or more phosphorylation sites in nitric oxide synthase puts them among the kinase-mediated signaling pathways. This also means that nitric oxide synthases are regulated indirectly by the events that regulate kinases. This field of research of nitric oxide synthase regulation has become one of the most actively pursued and much has been learned from basic biochemical mechanisms to physiological processes and to medical applications, but many more questions still remain to be answered.

Recruitment of governing elements for electron transfer in the nitric oxide synthase family.

At least three building blocks are responsible for the molecular basis of the modulation of electron transfer in nitric oxide synthase (NOS) isoforms: the calmodulin-binding sequence, the C-terminal extension, and the autoregulatory loop in the reductase domain. We have attempted to impart the control conferred by the C termini of NOS to cytochrome P450 oxidoreductase (CYPOR), which contains none of these regulatory elements. The effect of these C termini on the properties of CYPOR sheds light on the possible evolutionary origin of NOS and addresses the recruitment of new peptides on the development of new functions for CYPOR. The C termini of NOSs modulate flavoprotein-mediated electron transfer to various electron acceptors. The reduction of the artificial electron acceptors cytochrome c, 2,6-dichlorophenolindophenol, and ferricyanide was inhibited by the addition of any of these C termini to CYPOR, whereas the reduction of molecular O(2) was increased. This suggests a shift in the rate-limiting step, indicating that the NOS C termini interrupt electron flux between flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) and/or the electron acceptors. The modulation of CYPOR by the addition of the NOS C termini is also supported by flavin reoxidation and fluorescence-quenching studies and antibody recognition of the C-terminal extension. These experiments support the origin of the NOS enzymes from modules consisting of a heme domain and CYPOR or ferredoxin-NADP(+) reductase- and flavodoxin-like subdomains that constitute CYPOR, followed by further recruitment of smaller modulating elements into the flavin-binding domains.

Clinical correlations of potential activity markers in systemic sclerosis.

The objectives of the project were the following: (1) to establish a group of patients with a confirmed diagnosis of systemic sclerosis (Ssc), (2) to perform a detailed entrance examination of each patient, (3) to determine concentrations of potential activity markers, and (4) to make a comprehensive examination of each patient 1 year after inclusion into the study. A total of 49 patients were examined, 36 with a limited form of SSc, 9 with diffuse SSc, and 4 with other forms of SSc. We determined plasma or serum levels of the N-terminal propeptide of procollagen type III (NPIIIP), interleukin-6 (IL-6), soluble receptor for interleukin-2 (sIL-2r), soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular adhesion molecule-1 (sVCAM-1), von Willebrand factor antigen (vWFAg), and big endothelin-1 (BET-1) using commercial kits, and urinary excretion of pyridinoline (PYR) and deoxypyridinoline (D-PYR) using high-performance liquid chromatography. Correlations of these markers with selected clinical data were calculated. The mean levels of all potential activity markers were increased compared with normal values, but differences were not significant. The levels of NPIIIP, D-PYR, and IL-6 were normal. The measured values after 1 year did not differ from the entry values. At entry, NPIIIP concentrations correlated with the finger-to-palm distance, and D-PYR corresponded with findings on a simplified health assessment questionnaire (FQ). IL-6 levels correlated with the leukocyte count, sIL-2r with the FQ, and ET-1 with the diffuse lung capacity for carbon monoxide. In general, we found only a few clinical correlates of potential activity markers. Our data confirmed the correlations of collagen metabolism markers with skin involvement and FQ, as was reported previously. Larger studies in this field are needed.

Role of endothelin and nitric oxide in the pathogenesis of arterial hypertension in autosomal dominant polycystic kidney disease.

The pathogenesis of arterial hypertension in autosomal dominant polycystic kidney disease (ADPKD) is complex and likely dependent on interaction of hemodynamic, endocrine and neurogenic factors. We decided to evaluate the role of endothelin (ET1) and nitric oxide (NO) in the regulation of arterial blood pressure (BP) and to determine plasma levels of ET1 and NO in the group of patients with ADPKD. The ADPKD group (18 patients, 6 men + 12 women, mean age 44.6+/-11.7 years, with creatinine clearancecorrig > 1.1 ml/s) was compared with a control group of 27 healthy volunteers of comparable age. Plasma levels of ET1 assessed by direct RIA determination in the group of ADPKD patients (11.03+/-1.8 fmol/ml) were significantly increased (p<0.001) in comparison with the control group (2.66+/-0.58 fmol/ml), while no significant differences were observed between normotensive and hypertensive patients in the ADPKD group. Serum levels of NO were evaluated according to the determination of serum levels of their metabolites - nitrites/nitrates. Serum levels of NO in the group of ADPKD patients (39.85+/-.38 micro mol/l) were significantly higher (p<0.05) in comparison with the control group (22.7+/-1.20 micro mol/l), whereas in the ADPKD group no significant differences were observed between normotensive and hypertensive patients. Thus, our study supports the concept of complex alteration of both vasoconstrictor and vasodilator systems in the pathogenesis of arterial hypertension in ADPKD.

[Is mild essential hypertension without obvious organ complications and risk factors associated with increased levels of circulating markers of endothelial dysfunction? Effect of ACE inhibitor therapy]. / Je mírná esenciální hypertenze bez zrejmých orgánových komplikací a rizikových faktoru provázena zvýsenými hladinami cirkulujících markeru endoteliální dysfunkce? Vliv lécby ACE-inhibitory.

The objective of the investigation was to assess whether circulating adhesion molecules, von Willebrand factor (vWf) and endothelin-1 are elevated in patients with mild uncomplicated essential hypertension without further risk factors of atherosclerosis and whether they could serve as indicators of endothelial dysfunction in this form of hypertension. Furthermore the authors investigated the effect of ACE inhibitor treatment (ACEI), quinapril, on the level of these markers of endothelial dysfunction. The level of adhesion molecules [intercellular cytoadhesion molecule-1 (ICAM-1), E-selectin, P-selectin], von Willebrand s factor (vWf) and endothelin-1 were assessed in patients with mild essential hypertension without further cardiovascular risk factors or clinical manifestations of atherosclerosis before and after quinapril treatment (n = 25) and compared with normotensive controls (n = 29). The results of the examinations provided evidence that contrary to controls the hypertensive subjects had significantly higher ICAM-1 levels (237.8 vs. 207.8 ng/ml, P = 0.02) vWf (118 vs. 106 IU/dl, p < 0.05) and endothelin-1 (5.81 vs. 5.15 fmol/ml, p < 0.05). Three-month treatment of hypertensive patients with ACEI led to a significant drop of endothelin-1 levels (5.81 vs. 5.26 fmol/ml, p = 0.01). The authors proved also an unequivocal declining trend of other cytoadhesion molecules and vWf after ACEI treatment, the changes however were not statistically significant. From the investigation it may be concluded that also patients with uncomplicated essential hypertension without other cardiovascular risk factors or clinical manifestations of atherosclerosis have significantly elevated plasma levels of ICAM-1, vWf and endothelin-1. Higher concentrations of these factors suggest endothelial dysfunction already in mild forms of essential hypertension without further risk factors or cardiovascular complications. A significant drop of endothelin-1 and declining trend of the other investigated indicators suggest that ACEI treatment can favourably influence endothelial dysfunction in hypertensive patients also independently on reduction of the BP.