siRNA 沉默 Her-2／neu 基因对胰腺癌 BXPC-3细胞COX-2表达及细胞生物学行为的影响

摘要

目的：观察siRNA沉默胰腺癌BXPC-3细胞中Her-2／neu基因对COX-2表达及细胞增殖、凋亡、侵袭力的影响，探讨干扰Her-2／neu抑制BXPC-3生物学行为的可能机制，为胰腺癌的临床治疗提供新的思路。方法采用实时荧光定量PCR法（ RT-QPCR）和Western blot检测Her-2／neu、COX-2 mRNA和蛋白在胰腺癌BXPC-3细胞及正常胰腺HPDE6C7细胞的表达。设计并构建Her-2／neu siRNA慢病毒表达载体转染BXPC-3细胞，检测Her-2／neu基因沉默对COX-2 mRNA和蛋白表达的影响。 CCK-8法检测Her-2／neu siRNA对细胞增殖的影响。流式细胞仪检测Her-2／neu siRNA对细胞凋亡的影响。 Transwell小室实验检测Her-2／neu siRNA对细胞侵袭力的影响。结果 Her-2／neu、COX-2 mRNA和蛋白在胰腺癌细胞中均高表达，而在正常胰腺细胞株中极少表达，差异有统计学意义（ P ＜0换．05）。Her-2／neu基因沉默能显著抑制COX-2 mRNA和蛋白表达，细胞增殖、侵袭能力显著减弱，细胞凋亡率显著增加（ P ＜0．05）。结论胰腺癌细胞高表达Her-2／neu、COX-2，Her-2／neu基因沉默能够显著抑制细胞增殖和侵袭能力，并促进细胞凋亡。%Objective To investigate the specific effect of silencing of Her-2eu gene by small interfering RNA ( siRNA) on COX-2 expression and the cell biological behaviors of human pancreatic cancer cell line BXPC-3 in order to find an new therapeutic method for pancreatic cancer.Methods The expression levels of Her-2eu, COX-2 mRNA and proteins were detected by RT-QPCR and Western Blot in human pancreatic cancer cell lines BXPC3 and normal pancreas cell line HPDE6C7.The specific Her-2eu siRNA was synthesized and transfected into BXPC3 cells by lentivirus, and the expression levels of Her-2eu and COX-2 were detected.The CCK-8 assay was used to analyze the proliferation of BXPC3 cells.Flow cytometry was used to determine the apoptosis of BXPC3 cells.The cell invasion was evaluated by transwell assay.Results The overexpressions of Her-2eu, COX-2 mRNA and proteins were observed in human pancreatic cancer cell lines BXPC3. However the expressions of Her-2eu, COX-2 mRNA and proteins were almost not detected in normal pancreas cell line HPDE6C7, there were significant differences between the two kinds of cells ( P <0.05).The Her-2eu siRNA could remarkedly inhibit the expressions of COX-2 mRNA and proteins.The proliferation and invasiveness of BXPC3 cells in Her-2eu siRNA group were decreased obviously.However the apoptosis of BXPC3 cells in Her-2n/eu siRNA group was significantly increased (P <0.05).Conclusion The overexpressions of Her-2eu and COX-2 are observed in the human pancreatic cancer cell lines BXPC3,moreover, Her-2eu gene silence can obviously inhibit proliferative and invasive abilities of the cells, furthermore, which can induce apoptosis of BXPC3 cells.