| Background & Objective Tumor angiogenesis is closely related to tumor growth and tumor metastasis. Vascular endothelial growth factor(VEGF) and its receptors (VEGFR) play an important role in the tumor angiogenesis, division, proliferation of the breast cancer cells, contribute to tumor growth and metastasis. At present, we have discovered three kinds of VEGFR:VEGFR1 (FLT-1),VEGFR2 (FLK-1,KDR) and FLT-4. KDR plays the most important role in the cellular growth and differentiation and is connected with the proliferation of the endothelial cells and angiogenesis. Blocking KDR expression is a good anti-tumor therapeutic strategy. The objective of this study was to further explore the apoptosis inducing effect of KDR-siRNA on MCF-7 cells and its molecular mechanism, and to evaluate the impact of KDR-siRNA to tumor suppressor gene methylation, which may provide theoretical and experimental bases for clinical application.Methods siRNA sequence targeting KDR gene was obtained by in vitro chemically synthesis and was transfected into MCF-7 cells by using Lipofectamine2000TM, the morphology of apoptosis were observed by Hoechst 33258 staining under fluorescence microscope; the level of caspase-3,survivin,NES1 and RUNX3 were detected by RT-PCR; the methylation level of NES1 and RUNX3 gene were analysized with MSP; the activity of caspase-3 was measured by colorimetry and the expression of survivin was detected by immuno- histochemistry, the survivin expression was analysed by the image instrument.Results Experiments in vitro showed that siRNA directed against KDR enhanced the apoptosis of MCF-7, up-regulated caspase-3 mRNA and the activity of caspase-3 (p<0.05), the degree of NES1 and RUNX3 methylation cut down and the expression of their mRNA have recovered, down-regulated survivin mRNA and protein expression (p<0.05).Conclusions KDR siRNA may effectively cause demethylation,induce the apoptosis of MCF-7 through regulating the expression of caspase-3,survivin,NES1 and RUNX3. |
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