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By study on the effect of anisodamine on lipopolysaccharide- induced expression of tissue factor(TF) in vascular endothelial cells(EC) ,the mechanism of anisodam ine antithrom bosis,as well as in the treatm ent of bacteraemic shock was investigated.Human umbilical vein endothelial cells (HUVECs) were cultured by trypsin digestion m ethod.TF activity was measured in the lysates of HUVEC by using a single step clotting assay.Specific m RNA expression was detected by Northern blotting.In order to evaluate a possible contribution of the nuclear factor (NF) -κB pathway on the effects observed,electrophoretic mobility shift assays(EMSA) were performed using nuclear extracts from HU VECs and NF- κB- binding oligonucleotides.The results showed that treatment of HUVEC with L PS resulted in a significant increase in TF activity.Anisodamine dose- dependently inhibited L PS- induced upregulation of TF.These effects was also confirm ed on the level of specific TF m RNA expression by Northern blotting.Furtherm ore,EMSA showed that anisodamine com pletely abolished L PS- induced NF-κB DNA binding activity in nuclear ex- tracts from HUVECs treated with L PS together with anisodamine.The results suggest thataniso- damine counteracts endothelial cell activation by inhibiting L PS- induced TF expression in these cells.Its interference with the NF- κB pathway might- at least in part- contribute to this effect. The ability of anisodamine to counteract L PS effect on endothelial cells m ight be one underlying m echanism explaining its antithrombosis and efficacy in the treatment of bacteraemic shock.
By studying on the effect of anisodamine on lipopolysaccharide-induced expression of tissue factor (TF) in vascular endothelial cells (EC), the mechanism of anisodam ine antithrombosis, as well as in the treatm ent of bacteraemic shock was investigated. Human umbilical vein endothelial cells (HUVECs) were cultured by trypsin digestion m ethod.TF activity was measured in the lysates of HUVEC by using a single step clotting assay. Specific m RNA expression was detected by Northern blotting. Order to evaluate a possible contribution of the nuclear factor (NF) -κB pathway on the effects observed, electrophoretic mobility shift assays (EMSA) were performed using nuclear extracts from HU VECs and NF- κB-binding oligonucleotides. The results showed that treatment of HUVEC with L PS resulted in a significant increase in TF activity. Annisodamine dose-dependently inhibited L PS-induced upregulation of TF. These effects were also confirmed ed on the level of specific TF m RNA expression by Nor thern blotting.Furtherm ore, EMSA showed that anisodamine com pletely abolished LPS-induced NF-κB DNA binding activity in nuclear ex-tracts from HUVECs treated with L PS together with anisodamine. The results suggest that iso- damine counteracts endothelial cell activation by inhibiting L PS-induced TF expression in these cells. Its interference with the NF- κB pathway might- at least in part- contribute to this effect. The ability of anisodamine to counteract L PS effect on endothelial cells m ight be one underlying m echanism explaining its antithrombosis and efficacy in the treatment of bacteraemic shock.