Effects Of Icariine On Numbers And Function Of Peripheral Blood-derived Endothelial Progenitor Cells

Objective:To investigate the effects of Icariine on numbers and function of endothelial progenitor cells (EPCs) derived from peripheral blood-derived.Methods:1 To prepare and preservation Icariine drug-liquidOne ten-thousandth scale take 2mg of freeze-dried powder, with the charger take 2mlDMSO (dimethyl Chiara maple) dissolved in 5ml centrifuge tube, Gently rocking the oscillation completely dissolved,use charger take 20ull solution for small beaker charger 20mlPBS dilution, using microporous membrane pressure filter sterilization (membrane pore diameter of 0.22um) after the system was I to liquid liquor 20ml, concentration lug/ml.-Packing 4℃cold standby.2 Culture and identification of human EPCs from peripheral bloodThe mononuclear cells (MNCs)were isolated from human peripheral blood by density gradient centrifugation. Cells were plated on culture dishes coated with human fibronectin and maintained in Medium 199.After 7d of culture,adherent cells were collected.EPCs were characterized as adherent cells double positive for Dil-LDL uptaking and FITC-lectin binding by laser confocal microscope,and further demonstrated by expressing CD34 and CD133 using flow cytometry.3 Effects of Icariine Drug on numbers,function of EPCsAfter cultured for 7d,EPCs were randomized into 4 groups:control group,Icariine low dosage group,medium dosage group,high dosage group. The cultured EPCs were incubated with different cultivate liquid (include different concentration drug-containing serum and control serum respectively)for different periods(24 h,48 h and 72 h).The morphology of the cells was observed under reverse microscope. Numbers of EPCs,adhesion and the ability of cell proliferation was assessed by cell counting, adhesion assay, and MTT assay.Results:1 Characteristics of EPCsAfter7days culture,the MNCs showed as spindles-shaped,endothelial cells-like morphology. EPCs were characterized as adherent cells with double positive for Dil-LDL-uptake and lectin binding with laser scanning confocal microscope. Further the cells were demonstrated by expressing CD34+(38.36±3.70)% and CD133+(13.91±1.47)% using flow cytometry. 2 Effects of Icariine Drug on numbers,function of EPCsCompared with control group,the numbers and function of proliferation and adhesion of endothelial progenitor cells were remarkably increased by Icariine low and medium dose group,showing a time-effect relation. The Icariine low dosage group had the most prominent effect on the number and function of EPCs,begined at 24h and reached peak at 72h(P<0.05). Effect of the Icariine medium dosage group on the number and function of EPCs begined at 48h and reached peak at 72h(P<0.05). The Icariine high dosage group increase the number and function of EPCs at 72h only(P<0.05).Besides,the effect to the number and function of EPCs shows a dose-effect ralation at 48h and 72h,which was most prominent of The Icariine medium dosage group (P＜0.05).Conclusion:1,Icariine increases the numbers and the ability of adhesion and proliferation,of EPCs,showing a time-effect and dose-effect relation. which was most prominent at the concentration of medium dosage Icariine for 72 hours.2,Icariine drug-containing serum can premote the numbers and function of EPCs derived from peripheral blood.The results may be parts of the mechanisms of Icariine to prevent and treat vascular endothelial injury/dysfunction-related disease.