Effects Of Thymosin Beta4 On Activity Of Bone Marrow-Derived Endothelial Progenitor Cells

Objective: To observe the biological behaviours of rat bone marrow derived endothelial progenitor cells(EPCs) by gradient concentration Thymosin beta 4(Tβ4), to improve the effectiveness of the EPCs transplantation trentment of the central nervous system damage and to provide a broader thinking about promoting the EPCs transplantation as the main body of the treatment in the clinical application.Methods: 90 ~ 100 g SPF SD rats, male and female unlimited, using density gradient centrifugation to collect mononuclear cells isolated from rat bone marrow cells, inoculating in 24 hole culture plate which packaged for 24 hours in advance with mice fiber link protein(FN), culturing with EGM- 2 complete medium which containing 10% fetal bovine serum(FBS), 7 days later cell morphology and immunocytochemistry double fluorescent antibody test(CD133 and VEGFR- 2) were used to test the cells andlaser confocal microscope was used to observe and record. At the same time Dil marked the acetylation of low-density lipoprotein(LDL) and fluorescein isothiocyanate whin lectin- 1(Dil- ac- LDL and FITC- UEA- 1) were also been used to further identify the cells to endothelial progenitor cells by phagocytic experiment, and also laser confocal microscope was used to observe and record. Adherent cells were stimulated with Tβ4 at gradient concentrations(1 000 ng/ml, 100 ng/ml, 10 ng/ml, 1 ng/ml) or vehicle control, EPCs proliferation, migration and adhesion were assated with CCK-8 assay, transwell migration assay and adhesion assay respectively. Meantime gradient time(24 h, 48 h, 72 h) with Tβ4(1 000 ng/ml) were also been arranged,respectively, in the same way to observe the proliferation、adhesion ability of EPCs′ of change.Results: Tβ4 can significantly enhance the EPCs′ ability of proliferation, migration and adhesion in vitro, and present a dose-effect relationship, when cultured in 1 000 ng/ml condition medium the proliferation, migration and adhesion ability were significantly enhanced(compared with control group, P < 0.05). When cultured in 1 000 ng/ml condition medium intervention gradually with time goes on, the proliferation capacity of EPCs at each time point were significantly increased and better than that of the blank control group(P < 0.05); Adhesion ability at 48 h achieve the maximum effect(compared with control group, P < 0.05).Conclusion: Tβ4 can significantly enhance the in vitro cultured rat bone marrow origin EPCs′ biological activities, and presents a certain dose- effect relationship, 1 000 ng/ml Tβ4 under the conditions of 48 h can obtain the most optimal biological effect.