| Objective:By the recombinant lentirvirus vector(LV) of bFGF transfacting the rBM-MSCs , to observe the differentiation of rBM-MSCs modified by bFGF gene in vitro, to research the possibility and benefit of differentiation of rBM-MSCs modified by bFGF.Methods:Separating,cultivating and identificating the rBM-MSCs,then transfacting the rBM-MSCs with compeleted LV of bFGF, grouping the rBM-MSCs into three as the general group (the general rBM-MSCs group), the black control group (LV of GFP modifying rBM-MSCs group), the objective group (LV of bFGF gene modifying rBM-MSCs group); observing the expression of bFGF with culture solution by ELSIA and RT-PCR; and detecting the vigor of 3 groups by MTT assay ; cultivating the 3 groups of rBM-MSCs by VEGF in culture solution for 28 days in vitro, to observ the appereance of rBM-MSCs and the expression of CD31,CD34 and the FⅧby IHC then observing the phagocytosis phenomenon of Dil-AC-LDL ect.Results:1. rBM-MSCs at the passage 3 were positive for CD44 and CD90 (97.8%±0.9%, 96.9%±1.5%, respectively), but negative for CD11b/c and CD34 (14.2%±0.7% and 1.3%±0.5%, respectively).2. The rBM-MSCs transfected LV with GFP can express GFP highly ,and never disappear or weaken after passaging.3. The rBM-MSCs transfected LV with bFGF can inherit and secrete bFGF stably by RT-PCR and ELSIA, the objective group is (5.500±0.124)μg/ml, the black control group is (2.650±0.402)μg/ml, the general group is (2.763±0.253)μg/ml, the bFGF gene modified group is different from the GFP modified group and the general group (P=0.000<0.01 ), there is significance difference.4.There is no difference of the vigor among 3 groups (P =0.000>0.05), The objective group is 0.398±0.186, the general group is 0.392±0.189, the black control group is 0.394±0.243.5. Adjusting from morphological, The primary rBM-MSCs are likely trabs, after culturing the rBM-MSCs with bFGF for 28 days they are likely cobble-stone, and conjuncting closely-up, but there is no difference among the 3 groups .6. The objective group can get Dil-AC-LDL and express fluorescent , but little or none of other two groups After induction ( P=0.867<0.05),the objective group is 0.518±0.227, the general group is 0.047±0.027, the black control group is 0.071±0.019.7. After induction the objective group is high than other two group among CD31,CD34 and FⅧ(P<0.01), CD31 ,the objective group is (22.73±5.42)%, the general group is (2.57±1.21)%, the black control group is (7.85±1.83)%;CD34, the objective group is (26.32±3.95)%, the general group is (2.41±0.97)%, the black control group is (8.93±1.37)%; FⅧ,the objective group is (30.34±6.62)%, the general group is (1.97±0.52)%, the black control group is (8.93±17.37)% .Conclusions:Getting rBM-MSCs by the way of density gradient centrifugation;rBM-MSCs modified with LV of bFGF can secrete bFGF ; there is no difference of the vigor after being transfected by LV;the rBM-MSCs modified by bFGF are better to differentiate into cells as VECs than other two groups. |
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