Martine Renard

Three months supplementation of hyperhomocysteinaemic patients with folic acid and vitamin B6 improves biological markers of endothelial dysfunction.

Hyperhomocysteinaemia is a risk factor for premature atherosclerosis and venous thromboembolic disease. Supplementation with folic acid and vitamin B6 has been shown to decrease plasma homocysteine but data fail to assess an effect on the progression of vascular disease. We measured plasma homocysteine and two markers of endothelial injury (plasma soluble thrombomodulin and von Willebrand factor) at baseline and after 3 months of treatment with folic acid and vitamin B6. After this treatment there was a significant decrease in fasting soluble thrombomodulin (−15 ng/ml, 95%CI 5–22.2). Von Willebrand factor was significantly raised after methionine load at baseline but did not significantly rise after supplementation.

Endothelial dysfunction during acute methionine load in hyperhomocysteinaemic patients

Hyperhomocysteinaemia has been associated with arterial and venous thrombosis possibly by causing damage to the endothelium. We hypothesised that an oral load in methionine, that increases plasma homocysteine, would also result in an increase in biological markers of endothelial or platelet dysfunction. Then we investigated two groups of patients with arterial or venous occlusive disease: 17 with hyperhomocysteinemia and 12 without hyperhomocysteinemia. We measured in both groups plasma soluble thrombomodulin, von Willebrand factor, P-selectin and tissue factor plasma inhibitor before and 6 hours after a load with 100 mg/kg oral methionine. Methionine load resulted in a significant increase in von Willebrand factor in both groups (P<0.02), suggesting that endothelial dysfunction occurs during the load.

Influence of pentoxifylline on membrane thrombomodulin levels in endothelial cells submitted to hypoxic conditions.

Thrombomodulin (TM) expression at the surface of endothelial cells is upregulated by cAMP analogues, whereas hypoxic conditions induce a decrease in this expression. Pentoxifylline (PTX) enhances the intra-cellular levels of cAMP in human umbilical vein endothelial cells (HUVECs). We therefore investigated the influence of this drug on TM expression in cells submitted to hypoxia. TM expression was quantified at the surface of HUVECs in the presence or absence of PTX during 18 h of incubation. Membrane TM was immunolabeled with a fluorescent monoclonal antibody and the expression was quantified by flow cytometric analysis. Our preliminary data revealed that PTX at 0.01 μM significantly increased TM expression levels compared with cells in hypoxia without the drug. Therefore, at this low dose PTX appears able to counterbalance the procoagulant effect of hypoxia.

Influence Of Hypoxia And Hypoxia-Reoxygenation On Endothelial P-Selectin Expression

P-selectin is an endothelial adhesion molecule involved in the initial step of the neutrophil recruitment. We investigated the effect of hypoxia (95% N2, 5% CO2) and of hypoxia-reoxygenation (95% air, 5% CO2) on the expression of P-selectin by human umbilical vein endothelial cells (HUVEC). P-selectin expression was detected by immunolabelling and quantified by flow cytometric analysis. Our data indicate that hypoxia induces an increase in P-selectin expression with a maximum reached after 90 minutes. A hypoxic exposure of 90 minutes results in a highly significant increase compared to normoxia (p < 0.001, n = 13). Furthermore, when a reoxygenation period follows 90 minutes of hypoxia, the initially elevated levels of P-selectin are dramatically enhanced with a maximum obtained after 60 minutes of reoxygenation.

Soluble P selectin in systemic sclerosis: relationship with von Willebrand factor, autoantibodies and diffuse or localised/limited disease

A.D. Blann*, J. Constans, P. Carpentier, M. Renard, B. Satger, V. Guerin, M.R. Boisseau, N. Neau-Cransac, C. Conri Haemostasis, Thrombosis and Vascular Biology Unit, University Department of Medicine, City Hospital, Dudley Road, Birmingham, B18 7QH, UK Service de Medecine Interne et Pathologie Vasculaire, Hopital St Andre, 1 rue Jean Burguet, 33075 Bordeaux, France Service de Medecine Interne et Vasculaire, Hopital de la Tronche, BP217, 38043 Grenoble, France Laboratoire d’Hematologie, Universite Victor Segalen-Bordeaux II, 33076 Bordeaux, France Laboratoire d’Immunologie, Hopital Pellegrin, 33076 Bordeaux, France

Effect of buflomedil on the neutrophil-endothelial cell interaction under inflammatory and hypoxia conditions.

In hypoxia/ischaemia and ischaemia/reperfusion, human neutrophils are likely to play an important role in the development of endothelial cell damage in the microcirculation. Buflomedil hypochloride improves the capillary perfusion in such related situations, evoking a possible effect upon neutrophils. Using in vitro models of cell adhesion, buflomedil decreased 100% of histamine related neutrophil adhesion (flow system) and partially inhibited adhesion after IL-1-4 hours (flow and stable systems). Hypoxia induced neutrophil adhesion (4 hours) was also reduced by buflomedil, which decreased the expression of P-selectin at the surface of endothelial cells. As adenosine (NECA) exhibited the same results in hypoxia and theophylline inhibited them, such results support an action of buflomedil presumably via the A2 receptor.