The present study was designed to elucidate the possible beneficial effects of naftidrofuryl on ischemia-induced endothelium damage. For this purpose, an in vitro model was developed wherein human endothelial cells isolated from umbilical vein were submitted to hypoxia. Long-term hypoxia incubation (6 h) induced cell mortality, and naftidrofuryl strongly protected endothelial cells against this mortality in a dose-dependent manner and at concentrations as low as 10(-9) M. 66% protection was still observed after 16 h of hypoxia. Naftidrofuryl had to be present during the hypoxia incubation to exert its action; preincubation up to 24 h in the presence of naftidrofuryl could not protect endothelial cells incubated under hypoxia without naftidrofuryl. Short-term hypoxia, which does not induce mortality, strongly activates the endothelial cells with an increase in the cytosolic calcium concentration, in the phospholipase A2 activity, and in the synthesis of prostaglandin and of platelet-activating factor. It also enhances the adherence of polymorphonuclear neutrophils. Naftidrofuryl was able to markedly inhibit this whole cascade of events in a dose-dependent manner. We also demonstrated that naftidrofuryl could block the decrease in ATP concentration that results from the hypoxic conditions. These results indicate that by preserving the energetic level of the cells, naftidrofuryl prevents the activation of endothelial cells and the cell mortality induced by hypoxia. By maintaining an intact endothelium in vivo during ischemia, naftidrofuryl could prevent the further damage induced by leukocyte recruitment and activation.
Recent discoveries have shown that venous diseases have a multifactorial etiology. One of the factors which is definitely involved in this pathologic process is the change in the concentration of oxygen. An increase in the concentration of oxygen, hyperoxia, or reoxygenation following hypoxia, damages the tissues by stepping up the production of free radicals. In addition, a reduction in oxygen concentration, or hypoxia, is also damaging, probably through a reduction in ATP synthesis. From a therapeutic standpoint, the veins, and more particularly the endothelium, must be protected against the impact on the tissue of these changes in oxygen concentration. In this study, the effects of Ginkor Fort were tested on cultured endothelial cells subjected to varying oxygen pressures. The results show that Ginkor Fort can provide good protection of endothelial cells against hyperoxia and hypoxia-reoxygenation. These beneficial effects are probably due to the presence of flavonoids in the Ginko biloba extract; these flavonoids have an anti-oxidant effect. In addition, this substance also protects the cells against hypoxia, possibly by increasing the availability of oxygen for ATP synthesis. This dual protective effect, which is produced by two different mechanisms, may account for the wide spectrum of Ginkor Fort in its use in venous diseases.
RNA‐mediated gene silencing (RNA interference) is a powerful way to knock down gene expression and has revolutionized the fields of cellular and molecular biology. Indeed, the transfection of cultured cells with small interfering RNAs (siRNAs) is currently considered to be the best and easiest approach to loss‐of‐function experiments. However, several recent studies underscore the off‐target and potential cytotoxic effects of siRNAs, which can lead to the silencing of unintended mRNAs. In this study, we used a low‐density microarray to assess gene expression modifications in response to five different siRNAs in various cell types and transfection conditions. We found major differences in off‐target signature according to: (a) siRNA sequence; (b) cell type; (c) duration of transfection; and (d) post‐transfection time before analysis. These results contribute to a better understanding of important parameters that could impact on siRNA side effects in knockdown experiments.
The protection of human diploid fibroblasts against high oxygen tension was investigated using various combinations of the three major antioxidant enzymes: superoxide dismutase, catalase and gluthathione peroxidase. α-Tocopherol, a well-known hydrophobic antioxidant, was also tested in combination with the different enzymes. Microinjection of solutions containing different combinations of the three enzymes was compared with the injection of each single enzyme. We observed that the protections given by catalase or superoxide dismutase on the one hand, and by glutathione peroxidase on the other hand, were additive. Surprisingly, the combinations of catalase and superoxide dismutase were less effective than catalase alone and was even toxic at low SOD concentrations. Addition of α-tocopherol following the injection of any of the three enzymes was highly beneficial, but the strongest synergistic effect was obtained with glutathione peroxidase. These results stress the importance of membrane protection by α-tocopherol and indirectly by glutathione peroxidase. They also showed that any injection leading to the decrease in the O2−. or H2 O 2 concentration combined with one of these two protectors is very beneficial for the cells probably by decreasing the OH concentration. This is also proven by the very good protective effect obtained with desferrioxamine.
