The overall haemostatic potential (OHP) assay records fibrin polymerisation in plasma by areas un... more The overall haemostatic potential (OHP) assay records fibrin polymerisation in plasma by areas under the opacity curve. The authors reported high area values in pregnancy, and still higher values in preeclampsia. We wanted to see if the assay detects thrombophilia, and particularly, if the area values were high in congenital combined protein S (PS) and the FV R506Q Leiden mutation, since low protein S and increased activated protein C (APC) resistance occur in pregnancy and preeclampsia. The original overall haemostatic potential assay and our modified version with Protac and pentasaccharide to enhance inhibition by activation of protein C and by antithrombin (AT) were performed in plasma from 18 persons with thrombophilia. Seven had combined protein S deficiency and heterozygous FV Leiden mutation. In the original assay, median area values in controls and thrombophilic plasma samples were similar. In the modified assay, area values tended to be lower in controls than in the thrombophilia group (P=0.035). The enhanced inhibition reduced area values more in control plasmas (median reduction 34.7%) than in thrombophilic plasmas (median reduction 2.2%) (P=0.0017). The responses to activation were also low in warfarin-treated patients with thrombophilia. High area values in pregnancy with the original assay are probably not caused by insufficient inhibition. The response to activation of protein C and antithrombin, as calculated by the difference between the original and a modified assay, was subnormal in thrombophilic plasma, but there was an overlap with the results in controls.
Bone marrow angiogenesis is increased in non-Hodgkin lymphomas (NHL). Compounds affecting extrace... more Bone marrow angiogenesis is increased in non-Hodgkin lymphomas (NHL). Compounds affecting extracellular matrix (ECM) may modify angiogenesis. Here we investigated ECM regulators in 48 unselected NHL patients compared with 35 controls. Untreated patients had elevated (P < 0.05) serum matrix metalloproteinase (MMP) 9 and tissue inhibitor of metalloproteinase (TIMP) 1, while MMP-2, TIMP-2 and syndecan-1 were not significantly different from controls. MMP-9 mRNA was significantly up-regulated in blood mononuclear cells, while mRNA expressions of the other ECM regulators were unaltered. We found strong correlations between mRNA expressions of both vascular endothelial growth factor and fibroblast growth factor 2, and MMP-9, TIMP-1 and TIMP-2. After therapy, serum MMP-2 increased while MMP-9 decreased (P < 0.05), the others being unchanged. Several compounds affecting ECM may be involved in angiogenic activity in NHL.
Neovascularization and hemorrhaging are evident in advanced atherosclerotic plaques due to hypoxi... more Neovascularization and hemorrhaging are evident in advanced atherosclerotic plaques due to hypoxic conditions, and mediate the accumulation of metabolic substrates, inflammatory cells, lipids, and other blood born factors inside the plaque. Tissue factor (TF) pathway inhibitor (TFPI) is mainly expressed by endothelial cells and is the endogenous inhibitor of the coagulation activator TF, which together with the downstream product thrombin can drive plaque progression and atherogenesis. We aimed to investigate the effect of hypoxic conditions on endothelial cell expression and activity of TFPI and TF that are important in coagulation initiation. Hypoxia was induced in primary human umbilical vein endothelial cells using chemicals or 1% oxygen tension, and mRNA and protein expressions were measured using qRT-PCR, ELISA, and Western blot analysis. Microscopy of fluorescence-labeled cells was used to visualize cell-associated TFPI. Cell-surface factor Xa (FXa) activity was measured usin...
There is now circumstantial evidence that tissue factor pathway inhibitor (TFPI) is not only a ma... more There is now circumstantial evidence that tissue factor pathway inhibitor (TFPI) is not only a major anticoagulant, but also has proapoptotic properties. The current study was designed to address the role of TFPI on signalling pathways and apoptosis. The non-TFPI expressing cell line CHO-K1 was stably transfected with pcDNA3.1/V5-His-TOPO-TFPI and control cells were established by transfecting the CHO-K1 cells with pcDNA3.1/V5-His-TOPO. Sodium butyrate (NaBut) has been shown to induce the expression of recombinant proteins. Here we have used NaBut to increase the expression of TFPI as assessed by qRT-PCR and ELISA. Compared to the control cells, TFPI induced apoptosis in a concentration dependent manner as measured by a cell death detection assay. Independent of caspase-3 activation an increased cleavage of PARP was detected in the TFPI expressing cells. This was accompanied by downregulation of Bcl-XL, elevated levels of Bax, and increased translocation of the apoptosis initiating factor. Increased DNA binding activity of NF-κB was revealed by electrophoretic mobility shift assay when the TFPI level was elevated by NaBut together with an increased translocation of the NF-κB subunit p65. The results indicate that TFPI affected the apoptotic activity through a process independent of caspase-3, and was also able to increase the activation of the NF- κB pathway.
The overall haemostatic potential (OHP) assay records fibrin polymerisation in plasma by areas un... more The overall haemostatic potential (OHP) assay records fibrin polymerisation in plasma by areas under the opacity curve. The authors reported high area values in pregnancy, and still higher values in preeclampsia. We wanted to see if the assay detects thrombophilia, and particularly, if the area values were high in congenital combined protein S (PS) and the FV R506Q Leiden mutation, since low protein S and increased activated protein C (APC) resistance occur in pregnancy and preeclampsia. The original overall haemostatic potential assay and our modified version with Protac and pentasaccharide to enhance inhibition by activation of protein C and by antithrombin (AT) were performed in plasma from 18 persons with thrombophilia. Seven had combined protein S deficiency and heterozygous FV Leiden mutation. In the original assay, median area values in controls and thrombophilic plasma samples were similar. In the modified assay, area values tended to be lower in controls than in the thrombophilia group (P=0.035). The enhanced inhibition reduced area values more in control plasmas (median reduction 34.7%) than in thrombophilic plasmas (median reduction 2.2%) (P=0.0017). The responses to activation were also low in warfarin-treated patients with thrombophilia. High area values in pregnancy with the original assay are probably not caused by insufficient inhibition. The response to activation of protein C and antithrombin, as calculated by the difference between the original and a modified assay, was subnormal in thrombophilic plasma, but there was an overlap with the results in controls.
Bone marrow angiogenesis is increased in non-Hodgkin lymphomas (NHL). Compounds affecting extrace... more Bone marrow angiogenesis is increased in non-Hodgkin lymphomas (NHL). Compounds affecting extracellular matrix (ECM) may modify angiogenesis. Here we investigated ECM regulators in 48 unselected NHL patients compared with 35 controls. Untreated patients had elevated (P < 0.05) serum matrix metalloproteinase (MMP) 9 and tissue inhibitor of metalloproteinase (TIMP) 1, while MMP-2, TIMP-2 and syndecan-1 were not significantly different from controls. MMP-9 mRNA was significantly up-regulated in blood mononuclear cells, while mRNA expressions of the other ECM regulators were unaltered. We found strong correlations between mRNA expressions of both vascular endothelial growth factor and fibroblast growth factor 2, and MMP-9, TIMP-1 and TIMP-2. After therapy, serum MMP-2 increased while MMP-9 decreased (P < 0.05), the others being unchanged. Several compounds affecting ECM may be involved in angiogenic activity in NHL.
Neovascularization and hemorrhaging are evident in advanced atherosclerotic plaques due to hypoxi... more Neovascularization and hemorrhaging are evident in advanced atherosclerotic plaques due to hypoxic conditions, and mediate the accumulation of metabolic substrates, inflammatory cells, lipids, and other blood born factors inside the plaque. Tissue factor (TF) pathway inhibitor (TFPI) is mainly expressed by endothelial cells and is the endogenous inhibitor of the coagulation activator TF, which together with the downstream product thrombin can drive plaque progression and atherogenesis. We aimed to investigate the effect of hypoxic conditions on endothelial cell expression and activity of TFPI and TF that are important in coagulation initiation. Hypoxia was induced in primary human umbilical vein endothelial cells using chemicals or 1% oxygen tension, and mRNA and protein expressions were measured using qRT-PCR, ELISA, and Western blot analysis. Microscopy of fluorescence-labeled cells was used to visualize cell-associated TFPI. Cell-surface factor Xa (FXa) activity was measured usin...
There is now circumstantial evidence that tissue factor pathway inhibitor (TFPI) is not only a ma... more There is now circumstantial evidence that tissue factor pathway inhibitor (TFPI) is not only a major anticoagulant, but also has proapoptotic properties. The current study was designed to address the role of TFPI on signalling pathways and apoptosis. The non-TFPI expressing cell line CHO-K1 was stably transfected with pcDNA3.1/V5-His-TOPO-TFPI and control cells were established by transfecting the CHO-K1 cells with pcDNA3.1/V5-His-TOPO. Sodium butyrate (NaBut) has been shown to induce the expression of recombinant proteins. Here we have used NaBut to increase the expression of TFPI as assessed by qRT-PCR and ELISA. Compared to the control cells, TFPI induced apoptosis in a concentration dependent manner as measured by a cell death detection assay. Independent of caspase-3 activation an increased cleavage of PARP was detected in the TFPI expressing cells. This was accompanied by downregulation of Bcl-XL, elevated levels of Bax, and increased translocation of the apoptosis initiating factor. Increased DNA binding activity of NF-κB was revealed by electrophoretic mobility shift assay when the TFPI level was elevated by NaBut together with an increased translocation of the NF-κB subunit p65. The results indicate that TFPI affected the apoptotic activity through a process independent of caspase-3, and was also able to increase the activation of the NF- κB pathway.
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