Biomedical subjects

Agneta Siegbahn

Publications and source records attributed to Agneta Siegbahn.

27 records · Page 2Linked to original sources

PubMed2003

A quantitative real-time PCR method for tissue factor mRNA.

BACKGROUND: Tissue factor (TF) is primarily known for its function to initiate blood coagulation. The range of in vivo expression of TF is wide and requires a dynamic assay for monitoring. A general method for TF mRNA quantitation that is dynamic, sensitive and applicable to a variety of experimental systems or clinical situations is therefore desirable. OBJECTIVES: To develop a method for sensitive and dynamic quantitation of TF mRNA in human blood cells. METHODS: TF mRNA expression was analysed and evaluated in monocyte isolations, in whole blood (healthy volunteers and patients scheduled for percutaneous coronary intervention, PCI) and in a panel of human cell lines. RNA was extracted, reverse transcribed and subjected to real-time PCR amplification, according to the TaqMan technology. A TF plasmid was constructed as calibrator of the assay. Two housekeeping genes used as endogenous controls for cDNA quality and integrity were evaluated. RESULTS: The assay was linear by seven orders of magnitude and detected down to 10(2) copies of the TF plasmid. The coefficient of variation was 4% intra-assay and 28% between the assays when using beta2MG as endogenous control. The beta-actin gene expression was induced by treatment with lipopolysaccharide (LPS) in blood leukocytes and could not be used as an endogenous control. However, beta2MG showed only minor variations upon treatment with LPS. The TF mRNA and antigen expression, measured in a Western blot, correlated well (R(2)=0.903) in a panel of 11 human cell lines. CONCLUSIONS: We have established a method for sensitive and dynamic quantitation of TF mRNA in experimental systems and for clinical situations.

Base Sequence↗

PubMed2003

Low molecular weight heparin (dalteparin) compared to unfractionated heparin as an adjunct to rt-PA (alteplase) for improvement of coronary artery patency in acute myocardial infarction-the ASSENT Plus study.

BACKGROUND: Current thrombolytic-antithrombotic regimens in acute myocardialinfarction (AMI) are limited by incomplete early coronary reperfusion and by reocclusion and reinfarction. We compared the effects of low molecular weight heparin (LMWH) versus unfractionated heparin (UFH) as an adjunct to recombinant tissue-plasminogen activator (alteplase) on coronary artery patency and clinical outcomes in AMI. METHODS: Patients with AMI treated with alteplase (n=439) were randomised to either subcutaneous dalteparin (120 IU/kg every 12h) for 4-7 days or intravenous infusion of UFH for 48 h. Coronary angiography was performed between day 4 and hospital discharge. Clinical events and safety were evaluated until day 30. RESULTS: Overall there were higher thrombolysis in myocardial infarction (TIMI) flows in the infarct related coronary artery in the dalteparin group (p=0.016). The predefined primary end-point, TIMI grade 3 flow, did not reach statistical significance (dalteparin 69.3% versus heparin 62.5%; p=0.163). However, TIMI 0-1 flow (13.4 versus 24.4%; p=0.006) and its combination with intraluminal thrombus (27.9 versus 42.0%; p=0.003) were less common in the dalteparin group. During the period of randomised treatment there were less myocardial reinfarctions in the dalteparin group(p=0.010) but after cessation of dalteparin there were more reinfarctions resulting in no difference in death or MI at 30 days. There were no significant differences in major bleeding or stroke after 30 days. CONCLUSIONS: In alteplase treated AMI adjunctive dalteparin for 4-7 days seems to reduce the risk of early coronary artery occlusion and reinfarction. However, early after cessation of treatment there is a raised risk of events, which might eliminate any long-term gains.

Abciximab↗

PubMed2003

PML/RARalpha plays a role for basal activity and retinoid-induced repression of the tissue factor promoter in acute promyelocytic leukemia cells.

Constitutive expression of tissue factor (TF) by acute promyelocytic leukemia (APL) cells may contribute to thrombotic complications. In this study we examined the transcriptional mechanisms of all-trans retinoic acid (ATRA)-induced down-regulation of TF in the APL cell line NB4, by analysis of stable clones expressing the luciferase gene under the control of 5' flanking regions of the TF gene. We show that the TF promoter is constitutively active in NB4 cells, and that ATRA induces rapid suppression of the promoter. Basal activity and ATRA-induced suppression of TF promoter is determined by the proximal -383 to +121 bp of the promoter. Electrophoretic mobility shift assays demonstrate the binding of Fos/Jun complexes to two TF promoter AP-1 sites in this region. Both complexes were suppressed by ATRA treatment. The ectopic expression of the APL-specific PML/RARalpha oncoprotein in U-937 cells results in induction of TF mRNA and promoter activity. Interestingly, this PML/RARalpha-mediated increase in TF promoter activity is sensitive to ATRA treatment. These data indicate that TF expression in APL cells is exacerbated by the presence of the PML/RARalpha fusion protein, and implicates the loss of Fos/Jun binding to the TF promoter in ATRA-induced suppression of TF.

Cell Line, Tumor↗

PubMed2002

Tissue factor regulation and cytokine expression in monocyte-endothelial cell co-cultures: effects of a statin, an ACE-inhibitor and a low-molecular-weight heparin.

INTRODUCTION: Mounting evidence implies beneficial properties of statins and angiotensin converting enzyme (ACE)-inhibitors beyond those of their original indications in the treatment of coronary artery disease (CAD). Less is known of the mechanisms by which low-molecular-weight (LMW) heparin, also used in unstable CAD, affects the cellular micro-environment. The effects of these drugs in monocyte-endothelial cell co-culture systems have so far been sparsely investigated. MATERIALS AND METHODS: We studied the expression of tissue factor (TF) and the cytokines tumour necrosis factor (TNF)-alpha, interleukin (IL)-6 and IL-10 in a co-culture model with monocytic, vitamin D(3)(vitD(3))-differentiated U-937 cells and human coronary artery endothelial cells (HCAEC), and the effects of the above-mentioned drugs in this system. Cells were co-cultured for 18 h, with or without pre-stimulation of the HCAEC with interferon (IFN)-gamma, and in the presence or absence of simvastatin, enalapril or dalteparin. Analyses of surface tissue factor and intracellular cytokines were done by flow cytometry. RESULTS: Co-culture with activated HCAEC induced tissue factor expression in U-937 cells but not in the endothelial cells. All three drugs significantly reduced tissue factor up-regulation (p<0.001 for each). Co-culture also induced IL-6 expression in U-937 cells and an increase in IL-10 production by HCAEC, none of which was affected by drugs. When cultured separately, both cell types expressed TNF-alpha. This was attenuated in U-937 cells by all three drugs (p<0.001 for each), whereas only enalapril reduced the TNF-alpha content of activated HCAEC (p=0.02). Enalapril also down-regulated the basal expression of IL-6 (p=0.01) and IL-10 (p<0.01) in HCAEC, which simvastatin and dalteparin failed to do. CONCLUSIONS: In this study, we demonstrated for the first time that a statin, an ACE-inhibitor and an LMW-heparin all suppress tissue factor up-regulation in monocyte-endothelial cell co-cultures, thus adding new information regarding the cellular effects of these drugs that may be of importance in the treatment of CAD.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed2002

[Prophylaxis of postoperative thromboembolism. New alternatives to low-molecular-weight heparin].

For somewhat more than a decade low molecular weight heparins have dominated in the pharmacological prevention of postoperative venous thromboembolism. At present there are some new methods of potential interest both as prophylactic substances but also to better understand the pathophysiology of deep vein thrombosis. These are inhibition of factor VII a/tissue factor complex (NAP, Nematode Anticoagulant Protein), inhibition of activated factor X (the synthetic pentasaccharide fondaparinux) and thrombin inhibition (melagatran and its oral prodrug ximelagatran). They have been shown to be effective in high risk orthopaedic surgery. They have to show their place in the prophylactic arsenal in comparison with low molecular weight heparins (effect, safety, mode of administration, cost-effectiveness).

Anticoagulants↗

PubMed2002

Influence on coagulation activity by subcutaneous LMW heparin as an adjuvant treatment to fibrinolysis in acute myocardial infarction.

In this study, which includes 101 patients with acute ST segment-elevated myocardial infarction, we investigated the influence on the increased coagulation activity after streptokinase treatment by adding low-molecular-weight (LMW) heparin or placebo and the relation between the coagulation activity and ischemic episodes, coronary patency, and mortality. The expected increase of prothrombin fragment 1+2 (F1+2), thrombin-antithrombin (TAT), and D-dimer were significantly attenuated at 2, 6, and 18 h (D-dimer only at 18 h) in the dalteparin group compared to placebo. Ischemic episodes during the first 24 h appeared significantly more often in patients with F1+2 levels above the median at 18 h. There was a tendency to a lower frequency of Thrombolysis In Myocardial Infarction Trial (TIMI) grade 3 flow in the infarct-related artery in patients with TAT and D-dimer levels above the median at 18 h. F1+2, TAT, and D-dimer were significantly higher after 18, 6, and 18 h, respectively, in the deceased compared to surviving patients. Also, the lack of reduction of the levels of F1+2 between 6 and 18 h was related to a raised mortality. In conclusion, adjuvant treatment with LMW heparin to streptokinase attenuates increased coagulation activity. This might be of importance as remaining high coagulation activity is associated with signs of early reocclusion and raised mortality.

Acute Disease↗

PubMed2002

Coagulation, fibrinolysis, and cell activation in patients and shed mediastinal blood during coronary artery bypass grafting with a new heparin-coated surface.

OBJECTIVES: Heparin coating of the cardiopulmonary bypass circuit is shown to improve the biocompatibility of the surface. We have studied a new heparin surface, the Corline Heparin Surface, applied to a complete set of an extracorporeal device used during coronary artery bypass grafting in terms of activation of inflammation, coagulation, and fibrinolysis in patients and in shed mediastinal blood. METHODS: Sixty patients scheduled for coronary artery bypass grafting were randomized to one of 3 groups with heparin-coated devices receiving either a standard, high, or low dose of systemic heparin or to an uncoated but otherwise identical circuit receiving a standard dose of systemic heparin. Samples were drawn before, during, and after the operation from the pericardial cavity and in shed mediastinal blood. No autotransfusion of shed mediastinal blood was performed. RESULTS: The Corline Heparin Surface significantly reduced the activation of coagulation, fibrinolysis, platelets, and inflammation compared with that seen with the uncoated surface in combination with a standard dose of systemic heparin during cardiac surgery with cardiopulmonary bypass. Both a decrease and an increase of systemic heparin in combination with the coated heparin surface resulted in higher activation of these processes. A significantly higher expression of all studied parameters was found in the shed mediastinal blood compared with in systemic blood at the same time. CONCLUSIONS: The Corline Heparin Surface used in cardiopulmonary bypass proved to be more biocompatible than an uncoated surface when using a standard systemic heparin dose. The low dose of systemic heparin might not be sufficient to maintain the antithrombotic activity, and the high dose resulted in direct cell activation rather than a further anti-inflammatory and anticoagulatory effect.

Aged↗

PubMedDate not supplied

CRP and IL-6 concentrations are associated with poor glycemic control despite preserved beta-cell function during the first year after diagnosis of type 1 diabetes.

BACKGROUND: The role of non-specific inflammation in beta-cell loss in type 1 diabetes is unclear. In the present study, inflammatory markers were determined in patients with newly diagnosed disease and related to beta-cell function, glycemic control and autoimmunity. METHODS: Ninety-seven adult patients with type 1 diabetes mellitus (80% islet antibody positives, ab(+)) were examined at diagnosis and 3, 6, 9 and 12 months after the start of insulin treatment. Plasma C-reactive protein (CRP), interleukin-6 (IL-6), C-peptide, islet autoantibodies, insulin requirement and HbA(1c) were assessed. RESULTS: The concentrations of CRP were high-normal at diagnosis and did not change during the study period. A positive correlation between CRP at diagnosis and BMI was observed in ab(+) as well as in ab(-) cases. Detectable concentrations of IL-6 were found in 32% (157/485) of the samples and did not change during the study. Ab(-) patients had higher values of CRP at diagnosis and throughout the study compared to the ab(+). Among the ab(+) patients, CRP concentrations during the study were positively correlated to C-peptide at 12 months and an increase in HbA(1c) levels between 6 and 12 months. No associations between the presence or levels of islet autoantibodies and CRP were noted. CONCLUSIONS: In type 1 diabetes, the islet destructive process and the development of beta-cell remission are not associated with changes in CRP or IL-6. Instead, elevated CRP concentrations are prevalent and seem to reflect insulin resistance, as positive associations to BMI, C-peptide and deterioration of glycemic control were observed.

Adolescent↗

PubMedDate not supplied

Expression of fibrinolytic and coagulation factors in cocultured human endothelial and smooth muscle cells.

Interactions between endothelial cells and smooth muscle cells are interesting from a tissue-engineering point of view. We have developed a coculture system that allows direct contact between these two cell types. The fibrinolytic factors PAI-1, tPA, and uPA and the coagulation factor TF, were studied at the gene level by RT-PCR and at the protein level by ELISA. Significant changes of all studied factors were seen at the gene level in cocultured endothelial cells. tPA and TF were upregulated 4- and 7-fold, respectively, and PAI-1 and uPA were downregulated 4- and 1.5-fold, respectively, compared with single-cultured controls. In cocultured smooth muscle cells alterations of PAI-1 and TF were significant, with a 1.5-fold upregulation of PAI-1 and a 2.5-fold downregulation of TF. Results at the protein level mirrored the gene expression results. These findings indicate that cocultured endothelial cells are rendered both hypercoagulative and hyperfibrinolytic.

Blood Coagulation Factors↗