Dominance of cytokine- over FasL-induced impairment of the mitochondrial transmembrane potential (Deltapsim) in the pancreatic beta-cell line NIT-1.

Mitochondria of pancreatic beta-cells are potential targets of intrinsic and extrinsic apoptotic pathways in the autoimmune pathogenesis of type 1 diabetes. We aimed to investigate whether cytokine- and FasLigand (FasL)-induced apoptosis is associated with impaired mitochondrial transmembrane potential (Deltapsim) in the pancreatic beta-cell line NIT-1. NIT-1 cells were exposed to the interleukin-1beta/interferon-gamma (IL-1beta/IFN-gamma) cytokine combination to induce apoptosis in vitro. Low concentrations of cytokines resulted in Deltapsim impairment, and increasing concentrations had only a minor additional effect. Treatment with the inducible nitric oxide synthase (iNOS) inhibitor Nw-nitro-L-arginine methyl ester hydrochloride (L-NAME) prevented cytokine-mediated Deltapsim impairment, implying that cytokines affect Deltapsim via nitric oxide. The broad-spectrum caspase inhibitor Z-VAD(Ome)-FMK (ZVAD) revealed dichotomic actions. In the presence of ZVAD, cytokine-induced nitrite generation was increased but cell death and Deltapsim impairment were reduced. Deltapsim impairment was also reduced by inhibitors of caspases 1, 6 and 8. Induction of Fas by IL-1beta/IFN-gamma coupled with activation by Super-FasL augmented cytokine-induced cell death. We observed a clear dominance of cytokine- over FasL-induced effects on Deltapsim. Our findings show that IL-1beta/IFN-gamma cytokines have a strong effect to impair Deltaym and prime beta-cells for apoptosis via the intrinsic pathway mediated by iNOS and caspases. Furthermore, at least in NIT-1 cells, the extrinsic FasL/Fas pathway has only a minor additive effect on cytokine-induced Deltapsim impairment.

Interaction between factor V Leiden and serum LDL cholesterol increases the risk of atherosclerosis.

OBJECTIVES: We investigated the association between the factor V Leiden gene variant and carotid atherosclerosis in a cross-sectional study and explored possible associations between this gene variant and coronary artery disease (CAD) in a case-control study. METHODS: The presence (n=1696) or absence (n=703) of carotid atherosclerosis were sonographically assessed among participants of the population-based Study of Health in Pomerania (SHIP). The case-control study included 1021 patients with severe CAD and 2791 healthy SHIP participants. The factor V Leiden gene variant was determined by PCR and MnlI digestion. RESULTS: Multivariable analyses revealed no independent association between the factor V Leiden gene variant per se and carotid atherosclerosis or CAD. In the cross-sectional study, there was an interaction between the factor V Leiden gene variant and serum LDL cholesterol in non-diabetics with respect to the risk of carotid atherosclerosis. In the case-control study a similar interaction was found for CAD. In both studies the atherosclerotic risk increased with rising serum LDL cholesterol concentrations in carriers of the factor V Leiden gene variant. CONCLUSION: The co-existence between the factor V Leiden gene variant and high serum LDL cholesterol is independently associated with the risk of atherosclerosis.

Renin-angiotensin system and haemostasis gene polymorphisms and outcome after coronary artery bypass graft surgery.

BACKGROUND: Coronary artery bypass graft surgery is associated with a considerable 2-year mortality rate. Gene polymorphisms of the renin-angiotensin system may be associated with the risk of hypertension and cardiovascular disease. The angiotensin I-converting enzyme DD genotype has recently been identified as independent predictor of the outcome after coronary artery bypass graft surgery. Genetic factors of the clotting system may be related to the risk of myocardial infarction and restenosis after coronary interventions. The aims of the present study were to investigate whether gene polymorphisms of the renin-angiotensin system (angiotensinogen 235 M/T, angiotensin II type 1 receptor 1166 A/C) or the clotting system (glycoprotein IIIa PlA1/PlA2 and factor V Leiden 1691 G/A) are associated with the outcome after coronary artery bypass grafting. METHODS: A study population of 247 patients was followed-up 2 years after coronary artery bypass graft surgery. The primary end-point was total mortality. The secondary end-point was mortality from cardiac cause or the need for myocardial revascularization (percutaneous coronary interventions or recurrent surgery) during follow-up. Geno typing was performed by polymerase chain reaction- and restriction fragment length polymorphism-based techniques. RESULTS: An older age and the non-use of the internal mammary artery graft were identified as independent predictors of the primary end-point after coronary artery bypass grafting. A decreased left ventricular ejection fraction was an independent predictor for the secondary end-point. No association was found between any of the genetic factors and the outcomes after coronary artery bypass graft surgery in the main factor regression models. However, the angiotensin II type 1 receptor 1166 A/C gene polymorphism modulated the effects of age on the primary end-point, and the angiotensinogen 235 M/T gene polymorphism modulated the effects of age on the secondary end-point. CONCLUSION: We conclude that there are interactions between the angiotensin II type 1 receptor 1166 A/C as well as the angiotensinogen 235 M/T gene polymorphism and age with respect to the outcome after coronary artery bypass graft surgery. The glycoprotein IIIa PlA1/PlA2 and the factor V Leiden 1691 G/A gene polymorphisms were not associated with mid-term mortality or cardiac morbidity after coronary artery bypass grafting.

Preoperative plasma fibrinogen levels predict mortality after coronary artery bypass grafting.

This study was designed to investigate whether plasma fibrinogen levels as well as the beta-fibrinogen -455 G/A genotype are associated with outcome after coronary artery bypass graft (CABG) operation. We enrolled 249 consecutive CAD patients one day before they underwent a CABG operation. Data from 220 patients with available plasma fibrinogen levels were analyzed. The primary end-point was total mortality, the secondary end-point mortality from cardiac causes or the need for myocardial revascularization. The 2-year total mortality was 9.1% in the entire cohort. Multivariable analysis revealed an independent relationship between the primary end-point and preoperative plasma fibrinogen levels but not the beta-fibrinogen -455 G/A genotype. Neither preoperative plasma fibrinogen levels nor the beta-fibrinogen -455 G/A genotype could predict the secondary end-point. We conclude, that elevated preoperative plasma fibrinogen levels, but not the beta-fibrinogen -455 G/A genotype predict the total mortality after CABG operation.

Factor V Leiden and the risk of stillbirth in a German population.

An association between the factor V Leiden variant and an increased risk of pregnancy loss has been reported. Most previous studies were performed with clinically recruited patients and controls. This approach may cause selection bias. The present analysis was performed with the aim to investigate the association between the factor V Leiden mutation and the risk of stillbirth in a population-based sample. The Study of Health in Pomerania (SHIP) is a survey that was carried out in North East Germany. A random sample from the population aged 20 to 79 years was taken. The total SHIP population comprised 4,310 participants. The presence of the factor V Leiden variant was determined by PCR and Mnl I digestion. The presence of the factor V Leiden variant was neither associated with the number of pregnancies nor with the number of children per women. Data from 1,768 females who had at least one pregnancy with known outcome was available for the present analysis. Seventy-three women (4.1%) reported at least one stillbirth. Women with and without the factor V Leiden mutation did not differ with respect to the number of women with at least one stillbirth (OR for factor V Leiden variant 1.57; 95%-CI 0.76 - 3.25). Furthermore, the number of women with two or more stillbirths, the number of stillbirths per affected woman and the number of stillbirths per number of pregnancies per woman was similar between both genotype groups. In conclusion, there is no association between the factor V Leiden mutation and the risk of stillbirth in a representative population sample.

Neonatal sympathectomy reduces NADPH oxidase activity and vascular resistance in spontaneously hypertensive rat kidneys.

Neonatal sympathectomy reduces arterial pressure in spontaneously hypertensive rats (SHR). In SHR transplanted with a kidney from sympathectomized SHR, arterial pressure was lower and less Na+ sensitive than in SHR transplanted with a kidney from hydralazine-treated SHR. This study was performed to identify underlying renal mechanisms. Tests for differential renal mRNA expression of nine a priori selected genes revealed robust differences for renal medullary expression of the NADPH oxidase subunit p47phox. Therefore, we investigated the effects of neonatal sympathectomy on renal mRNA expression of NADPH oxidase subunits, NADPH oxidase activity, and renal function. In 10-wk-old sympathectomized SHR fed a 0.6% NaCl diet, medullary p47phox and gp91phox expression was 40% less than in hydralazine-treated SHR. Also, after a 1.8% NaCl diet, medullary p47phox mRNA expression was lower in sympathectomized than in hydralazine-treated SHR. We found lower cortical (-30%, P<0.01) and medullary (-30%, P<0.05) NADPH oxidase activities in sympathectomized than in hydralazine-treated or untreated SHR. Glomerular filtration rate, renal blood flow, medullary blood flow, and fractional Na+ excretion in kidney grafts from sympathectomized and hydralazine-treated donors (n=8 per group) were similar at baseline and in response to a 20-mmHg rise in renal perfusion pressure. Renal vascular resistance was lower in kidneys from sympathectomized than hydralazine-treated donors (25+/-2 vs. 32+/-4 mmHg.min.ml-1, P<0.05). The results indicate that the sympathetic nervous system contributes to the level of renal NADPH oxidase activity and to perinatal programming of alterations in renal vascular function that lead to elevated renal vascular resistance in SHR.

Candidate genetic markers and the risk of restenosis after coronary angioplasty.

The aim of the present study was to test for possible associations between candidate gene polymorphisms and the risk of restenosis and recurrent restenosis after percutaneous transluminal coronary angioplasty (PTCA) without stenting. We followed up 511 PTCA patients, and restenosis and recurrent restenosis were defined according to angiographical criteria. Genotyping of the beta-fibrinogen -455 G/A, glycoprotein (GP) IIIa PlA1/PlA2, plasminogen activator inhibitor-1 (PAI-1) 4G/5G, factor V Leiden 1691 G/A, tumour necrosis factor alpha (TNFalpha) -238 G/A, TNFalpha -308 G/A, interleukin (IL)-1alpha -889 C/T, IL-1beta -511 C/T, methylenetetrahydrofolate reductase (MTHFR) 677 C/T and endothelial nitric oxide synthase (eNOS) 4 b/a gene polymorphisms was performed by PCR and restriction-fragment-length-polymorphism-based techniques. One hundred and sixty patients (31.3%) developed restenosis and in 130 of these patients, of whom 123 were available for analysis, a second PTCA without stenting was performed. Of these patients, 35 (28.5%) developed recurrent restenosis. None of the investigated genotypes were associated with the risk of restenosis or recurrent restenosis after PTCA. The degree of stenosis before and immediately after PTCA and the severity of the lesion were independent predictors for restenosis after PTCA. In conclusion, there was no association between the beta-fibrinogen -455 G/A, GP IIIa PlA1/A2, PAI-1 4G/5G, factor V Leiden 1691 G/A, TNFalpha -238 G/A, TNFalpha -308 G/A, IL-1alpha -889 C/T, the IL-1beta -511 C/T, MTHFR 677 C/T and eNOS 4 b/a gene polymorphisms and the risk of restenosis after PTCA as well as recurrent restenosis after repeated PTCA.

Prevalence of eight molecular markers associated with thrombotic diseases in six Amerindian tribes and two African groups of Costa Rica.

Individuals belonging to six different Amerindian tribes and two African groups of Costa Rica were genotyped for factor V Leiden (FV), factor V haplotype HR2 (FV HR2), Factor II 20210G>A (FII), the methylenetetrahydrofolate reductase (MTHFR), factor VII polymorphisms (FVII IVS7, FVII R353Q), factor XIII (FXIII V34L), and the insertion/deletion (I/D) polymorphism of the gene of angiotensin converting enzyme (ACE). Clear differences in the prevalence were found and are first reported. The prevalence of some of the established genetic risk factors was low in Amerindians of Costa Rica (ACE) or even absent (FVL, FII), and others (MTHFR, FVHR2) had an extremely high prevalence. People of African origin carried very rare FVL or FII polymorphisms, but the DD genotype of ACE is the highest reported. Concerning the protective factors, the QQ genotype of FVII R353Q was absent in Amerindians, but the protective 7/7 genotype of FVII IVS7 frequently found. Novel alleles of FVII IVS7 (4, 8, and 9 monomers) were found. Intertribal heterogeneity was observed that may reflect the evolutionary history of these tribal groups and their admixture with other populations.

Prevalencia de la resistencia a la proteína C activada en poblaciones indígena y negra del occidente de Venezuela / Prevalance of the activated protein C resistence in indians and black populations from western Venezuela

La resistencia de la proteína C activa (RPCa) es el fenotipo común del factor V Leiden (Arg506G1n) reconocido como factor de riesgo trombótico. El objetivo del presente estudio fue determinar la prevalencia de RPCa y su asociación con el factor V Leiden en poblaciones indígena y negra del occidente de Venezuela, ya que hasta la presnte no hay publicaciones sobre esta asociación en grupos étnicos venezolanos. Se estudiaron 80 indígenas de la etnia Yukpa de la Sierra de Perijá y 91 individuos de raza negra habitantes de la región sureste del Lago de Maracaibo. La RPCa fue determinaad según el método de Dalhlback, modificado por Jorquera y col. y Trossaert y col y los resultados expresaron como n-PCa-SR (valor positivo menor e igual l0,75). Las muestras de sangre anticoagulada con EDTA fueron impregnadas en papel secante y procesada para la detección del factor V Leiden según técnicas estándar para PCR y de análisis de restricción, en el Institute of Human Genetics (Greisfswald, Alemania). No se encontró diferencia significativa entre la n-PCa-SR de los indígenas (X+-EE=1,13+-0,02; IC 95 por ciento=1,07-1,19) y el de los sujetos de raza negra (1,07+-0,02;IC 95 por ciento=1,03-1,12). La prevalencia de RPCa fue de 1,25 por ciento(1/80) en índigenas (el caso era heterocigoto para factor V Leiden) y 4,4 por ciento (4/91) en la raza negra( un individuo resultó ser heterocigoto para factor V Leiden). Ninguno de los pacientes con RPCa tenía antecedentes personales o familiares de trombosis. Esta investigación representa el primer informe sobre la asociación de RCPa y factor V leiden an aborígenes e individuos de raza negra venezolanos. Aunque entre los indígenas existe una alta endogamia no se puede descartar en ningunos de los grupos raciales la intervención de genes foráneos. El hallazgo de RPCa sin factor V Leiden (Arg506GIn) sugiere la presencia de una mutación diferente a la molécula del factor V. La determinación de la prevalencia de este fenotipo y su marcador molecular en diferentes grupos étnicos es importante para la interpretación de su papel como factores de riesgo para enfermedades trombóticas