| Objective:To study the expression and its relationship of Survivin gene and VEGF gene in the osteosarcoma cell(9607 cell line), as well as the cell proliferation after the interference of ASODN. So as to approach the interaction of Survivin gene and VEGF gene in osteosarcoma cell, which will provide some clues for the occurrence, development, metastasis of the osteosarcoma.Method:The osteosarcoma cells(9607 cell line)were cultured and divided into experimental group and control group. Survivin and VEGF phosphorothioate ASODN was transfected into 9607 cells by lipofectamine 2O00 respectively in experimental group, meanwhile 9607 cells in control group was treated by simple lipofectamine. The inhibition ratio of 9607 cells in different groups were determined by MTT, the expression of Survivin mRNA and VEGF mRNA were determined by RT-PCR, the expression of Survivin protein and VEGF protein were determined by immunocytochemistry.Results:(1)After the transfection of Survivin-ASODN ,the inhibition ratio of 9607 cells at 24 hours,48 hours and 72 hours was (31.400±0.691)%,(52.515±0.870)%,(78.473±1.025)% respectively. After the transfection of VEGF-ASODN ,the inhibition ratio of 9607 cells at 24 hours, 48 hours and 72 hours was(28.511±0.825)%,(49.292±0.900)%,(71.349±1.223)% respectively. The inhibition ratio of 9607 cells in Survivin-ASODN group and VEGF-ASODN group were higher than that in control group in different time points, P<0.05.(2)The expression of Survivin mRNA in 9607 cells of Survivin-ASODN group and VEGF-ASODN group was ( 1.5198±0.084 ) and ( 1.6261±0.091 )respectively, which were lower than that of control group(1.9610±0.094), P<0.05 . The expression of VEGF mRNA in 9607 cells of Survivin-ASODN group and VEGF-ASODN group was(1.7489±0.065)and(1.5429±0.064)respectively, which was lower than that of control group(2.0453±0. 082), P<0.05. The expression of Survivin mRNA and VEGF mRNA were both decreased in Survivin-ASODN group and VEGF-ASODN group. The inhibition ratio of the expression of Survivin and VEGF mRNA was positively correlated in Survivin-ASODN group(r=0.602) and VEGF-ASODN group(r=0.702)(.3)The appearance of cells in control group was justo,growth prosperity;the appearance of cells in ASODN group was deform in microscope. (4)Both the expression of Survivin protein and VEGF protein were decreased in experiment group compared to the control group, P<0.05. Conclusion: (1) The expression of Survivin gene and VEGF gene were found in the 9607 cells. (2)The proliferation of 9607 cells were inhibited after ASODN of Survivin gene and VEGF gene were transfected, which indicate the ASODN could block target gene with a time dependent form.(3)The expression of Survivin gene was decreased with the down-regulation of VEGF gene , meanwhile the expression of VEGF gene was decreased with the down-regulation of Survivin gene. So there may be a mutual effect between the expression of the two genes. Further study on the regulation mechanism of the two genes can provide some evidence for the growth of the osteosarcoma cells and the vascular formation of the osteosarcoma. |
