Study On Effect Of BFGF Gene Transfected BMSCs On PMVECs

Objective: Chronic Obstructive Pulmonary Diseases(COPD) was a common chronic disease for respiratory tract,and the current treatment focus upon symptom controlling.This study tries upon the co-cultivation of the lung microvessel of endothelial cells and MSC transfected by exogenous basic fibroblast growth factor(b FGF) gene, and observed the role of transfected stem cells on the lung microvessel of endothelial cells,thus, the new idea and method are explored for the COPD treatment.Method: 4 weeks old SD rats were selected(both male and female). Their BMSCs were separated and cultivated by bone marrow adherent cell cultivation. The b FGF-pc DNA3.1 recombinant plasmids were purchased, and the plasmids with b FGF gene were transfected into BMSCs by lipofectin-mediated transfection.The transfection efficiencywas observed flurecenee microscope.Moreover, the Western-blot was utilized to make further verification upon the transfection of recombinant plasmid b FGF-pc DNA3.1 and the expression of the target protein. The transfected b FGF-pc DNA3.1-BMSCs would be co-cultivated with the pulmonary microvascular endothelial cells(PMVECs) under the transwell system.It has six experimental groups in this program and one blank control group. The six experimental groups were: Group of recombinant plasmids(Group A), Group of blank plasmids(Group B), Group of transfected recombinant plasmids(Group C), Group of blank transfected plasmids(Group D), Group of BMSCs(Group E), Group of PBS(Group F), Group of Blank control: simple group of PMVECs cultivation, Group B: co-cultivation of blank plasmid pc DNA3.1 and PMVECs, Group C: co-cultivation of b FGF- pc DNA3.1-BMSCs and PMVECs, Group F: co-cultivation of PBS liquid and VECs. And it should be observed respectively after 24 h,48h,72 h upon the growth rate of the PMVECs for each group.Result: BMSCs were successfully extracted and cultivated, and flow cytometry was used to make inspections and the CD29, CD 90 were positive, CD34 and CD11 B were negative.The result was the needed BMSCs; 2. When plasmids transfected BMSCs,green fluorescent protein could be seen under fluorescent microscope,thus, the early transfection was successful; 3. Western-blot was used to inspect the expression of the b FGF protein after the recombinant plasmids carrying the b FGF gene transfecting the BMSCs, and the result showed that recombinant plasmids expresses a large amount of b FGF protein after transfecting the BMSCs. In comparison with BMSCs transfected by blank plasmids pc DNA3.1 and untransfected DMSCs, the expressed amount of the b FGF protein increased on large scale,whose differences owned the statistical significance.That was to say that the successful transfection of the b FGF gene carried by recombinant plasimids into BMSCs could express a large amount of b FGF protein.4. Group of transfected recombinant plasmids ranks as the highest in the growth rate of PMVECs, P <0.05 when it counted with PBS group, whose differences owned statistic significance.Compared with the growth rate of Group of recombinant plasmids and blank plasmids, P <0.05,whose differences owned statistic significance. Compared with the growth rate of Group of blank transfected plasmids and Group of BMSCs,the P >0.05, whose differences owned no statistic significance,but the growth rate of PMVECs had the increasing tendency compared with group of blank transfected plasmids and group of BMSCs,and the tendency extended higher as the time of co-cultivation grows; When the group of blank transfected plasmids was compared with group FBS, the growth rate of PMVECs has been clearly higher, P<0.05,whose differences owned statistic significance; when the growth rate of the group of blank transfected plasmids and group of recombinant plasmids and blank plasmids, the P<0.05,whose differences owned statistic significance.In comparison with BMSCs, P>0.05, whose difference owned no statistic significance; when the growth rate of group of BMSCs compared with group FBS,P<0.05,whose differences owned statistic significance, when the growth rate of group of BMSCs compared with group of recombinant plasmids and blank plasmids were compared, P<0.05,whose differences owned statistic significance;Conclusion: 1. Using liposome transfection methods,the b FGF gene was successfully transfected BMSCs and overexpression of b FGF protein; 2. BMSCs could stimulate the proliferation of pulmonary microvascular endothelial cells;3.BMSCs transfectionb FGF gene could stimulate the proliferation of pulmonary microvascular endothelial cells and it had time dependent,compared with non transfectedb FGFgene,the stimulation of pulmonary microvascular endothelial cells was stronger effect,but no significant difference between the two groups of value-added rate statistics.