Experimental Study Of Proliferation Of EPCs Effected By 3-MA In Rat's Chronic Deep Vein Thrombosis

Objective To study the proliferation of endothelial progenitor cells(EPCs) effected by 3-MA(3-Methyladenine) in rat's chronic deep vein thrombosis .Methods 1.Rat bone marrow was obtained by rinsing bone marrow cavity,then bone marrow mononuclearcells were isolated by gradient centrifugation using Ficoll centrifugate. mononuclearcells were then cultivated with EGM-2MV medium cultivation. MTT assay were used to draw growth curve of cells.Cultvated cells werc identified by immunofluorescence and immunohistochemisty. 2. Experimental rat models(about 300g)of chronic deep vein thrombosis were obtained by partial ligation of inferior vena cava.The rats were randomly divided into four groups,and 15 animals in each group. 3. Transplantation is through the femoral vein.A group:control group,1ml EGM-2MV medium transplanted;B group:EPCs group,1ml 105 EPCs transplanted ;C group:3-MA group,1ml 3-MA(5mmol/L) transplanted;D group:EPCs effected by 3-MA group, 1ml EPCs have cultivated with 3-MA for 6 hours before injection.4.Collecting the thrombus in inferior vena cava at different time after transplantation.Then,western blotting of inferior vena cava thrombosis was used to detect LC3Ⅱ,VEGF,bFGF protein expression changes.5.SPSS 1 7.0 software Was used for analysis.Results 1.then rat bone marrow EPCs were isolated successfully by Ficoll centrifugate in vitro.2.The method of rat model of deep venous thrombosis was successfully set up,and it is feasible and reliable.3.The result of Western blotting:on the protein expression of LC3Ⅱ,the group C and D were lower than group A and B(P<0.05);On the protein expression of VEGF and bFGF,the group D and B were significantly higher than group A and C(P<0.05) . Conclusion After transplantation of EPCs effected by 3-MA ,the proliferation ability of EPCs is improved , and it is significantly up-regulate compared with transplantation EPCs only.