Effects Of SPK On The Injured Endothelial Cell

Objective Extract the biological activity of Spirulina kinase (SPK) from the fermented spirulina powder.On one hand, the endothrlial damage thrombosis model in vivo to study the impact of SPK on living rats’vascular endothelial cells anticoagulant fibrinolytic related factors. The other hand, by in vitro cultured human umbilical vein endothelial cells (HUVECs) to explore the influence of SPK on HUVECs fibrinolytic gene expression. From the antithrombotic function and fibrinolytic function of endothelial cells to explore the Spirulina kinase protection on the cardiovascular system.Methods Establish animal endothelial model:The rats were randomly divided into sham operation group (Sham), model group (Model), Spirulina kinase low-dose group+model (L-SPK+M), Spirulina kinase meduim-dose group+model (M-SPK+M), Spirulina kinase low-dose group+model (H-SPK+M). After different doses of SPK intervene, animals were anesthetized and FeC13induced cephalic artery endothelial injury, blood was collected from the abdominal aorta in rats, anticoagulation after, detecte plasminogen (Pig), antithrombin Ⅲ (AT-Ⅲ) secreted from endothelial cells with ELISA.Besides thromboxane B2(TXB2) and6-keto-prostaglandin F1a (6-Keto-PGF1a) were determined by radioimmunoassay.2. Culture vitro HUVECs, establish adrenaline (Adr) damage model with different concentrations of SPK. And control with the blank group, heparin (Hep) as a positive control, after12h,24h,48h, reverse transcription polymerase chain reaction (RT-PCR), PCR products were gel electrophoresis, the gel image analyzer photography and scanning, determination of the t-PAmRNA, PAI-1mRNA and reference gene glyceraldehyde phosphate aldehyde dehydrogenation enzyme GAPDHmRNA relative expression levels.Results1. In vivo experiments showed that thrombosis model was constructed successfully significantly higher AT-III,6-Keto-PGF1a, TXB2, in rat blood, with the sham group difference was statistically significant (P<0.05), while the Plg content had no significant effect (P>0.05). After the different concentrations SPK intervention,compare with model group, in the high dose group Plg significantly higher (P<0.01), differences between meduim-dose and high-dose group were not statistically significant (P>0.05); AT-Ⅲ were significantly increased, especially in the middle dose group (P<0.01); levels of TXB2decreased significantly (P<0.01), no significant difference between the low-dose group and high-dose group (P>0.05);6-Keto-PGF1a in content of high, medium and low dose groups were reduced (P<0.01), the most significant is in the low and middle dose group, and difference between two groups was not statistically significant (P>0.05)2. Cultured in vitro human umbilical vein endothelial cells, studying the expression of t-PAmRNA at different solubility SPK:SPK alone to HUVECs with the blank control group have no significant change (P>0.05). The t-PAmRNA expression as a downward trend in the model group at48h (P<0.01); The t-PAmRNA expression of the heparin group and SPK groups all increase within12h between the two groups it was not statistically significant (P>0.05); SPK high dose group in the24h and48h time periods increase the expression of t-PAmRNA was significantly superior to heparin group (P<0.05).3. Cultured in vitro human umbilical vein endothelial cells, studying the expression of PAI-1mRNA at different solubility SPK:SPK alone to HUVECs with the blank control group have no significant change (P>0.05). After adrenaline intervention expression of PAI-1mRNA significantly increase (P<0.05). Different concentrations of SPK group at each time periods showed the solubility dependence down at expression of PAI-1mRNA (P<0.05). The ratio of expression of t-PAmRNA and PAI-1mRNA was improved.Conclusion the SPK promote secretion of Plg, AT-Ⅲ, inhibit the release of TXB2,6-Keto-PGF1a, while improving the ratio of expression of t-PAmRNA and PAI-1mRNA in the anticoagulant thrombolytic effect, increase the fibrinolytic activity of endothelial cells. SPK. play its antithrombotic effect from anticoagulation fibrinolysis.