Down Regulation Of Survivin And VEGF Double Gene Expression Via SiRNA Liposome Is Capable Of Promoting Apoptosis Of U251 Cells

Objective:To investigate if survivin and vascular endothelial growth factor (VEGF) genes targeting siRNA transfection gets biological response in human U251 cells.Methods:The mRNA sequences targeting survivin and VEGF were designed and their siRNAs were synthesized in vitro. The human gliomas cells (U251) were cultured, targeting survivin-siRNA and/or VEGF-siRNA entrapped in liposome was transfected. There were five groups:group S:transfected with survivin-siRNA, group V: transfected with VEGF-siRNA, group SV:transfected with survivin-siRNA and VEGF-siRNA, negative control(NC):transfected with scrambled-siRNA, blank control(BC):transfected with blank liposome. The mRNA and protein of survivin and VEGF were detected by real-time quantitative PCR and Western Blot respectively. The proliferation and apoptosis of U251 were detected by CCK-8 assay and flow cytometry respectively.Results:1. Real-time qPCR and Western Blot analysis showed that compared to groups NC and BC the expression of survivin mRNA and protein in groups S and SV were significantly down-regulated 48 hours after transfection with siRNA(P<0.01)while without any significant difference between these two groups;the expression of VEGF mRNA and protein were down-regulated in groups S, V and SV, especially in groups V and SV, based on group S at mRNA levels (P<0.05).2. CCK-8 assay showed that compared to groups NC and BC, the proliferation of U251 cells was significantly inhibited in groups S, V and SV 24,48 and 72 hours after transfection (P<0.01) and dual genes transfection (group SV) got stronger inhibition than single gene transfection (groups S, V) 72 hours after transfection (P<0.01).3.The apoptosis of U251 cells detected by flow cytometry assay was significantly increased in groups S, V and SV compared to the groups NC and BC (P<0.01), more, there was much more significant apoptosis in group SV compared to groups S and V (P<0.01).Conclusion:1.The siRNA of survivin or VEGF entrapped in liposome could decrease the expression of survivin or VEGF at mRNA and protein levels, and significantly inhibited proliferation and induced apoptosis in U251 cells.2. Dual genes transfection (group SV) got stronger inhibition than single gene transfection in the proliferation and apoptosis in U251 cells.3.The expression of VEGF mRNA and protein were down-regulated by siRNA with survivin, but the opposite was not.Therefore, survivin may be the upstream target of VEGF.