慢病毒介导下干涉酪氨酸激酶B表达对乳腺癌SK-BR-3细胞增殖的影响

摘要

Objective To explore the impact of lentivirus-mediated knockdown of tyrosine kinase B (TrkB) on the proliferation of breast cancer cell line SK-BR-3 and its the mechanisms.Methods TrkB expression in 4 breast cancer cell lines including MCF-7,BCAP-37,SK-BR-3 and MDA-MB231 were measured and the cell line with relatively high TrkB expression was selected for interference of TrkB using lentivirus infection.The Lv-sh-TrkB cell line with stable interference of TrkB was confirmed by Western blot assay detection; in addition,the Lv-scramble cell line was also built.The four methyl tetrazolium (MTT) method and flow cytometry assays detection were used to assess the proliferation and cell cycle changes.Results Expression level of TrkB in SK-BR-3 cell line was higher than that of other cell lines,and it was used to construct the Lv-sh-TrkB cell line with stable interference of TrkB.Western blot assay showed that TrkB of Lv-sh-TrkB was significantly reduced compared with that of control and Lv-scramble cell line.MTT assay showed that the cell proliferation of Lv-sh-TrkB cell line was significantly slower than that of control and Lv-scramble cell line (P ＜ 0.05).Flow cytometry assays showed that G1 phase arrest occurred in Lv-sh-TrkB cell line (Lv-sh-TrkB cell line G1 phase 78％,control cell line G1 phase 61％,Lv-scramble cell line G1 phase 62％).Conclusion The proliferation of breast cancer cell SK-BR-3 is significantly inhibited and the cell cycle G1 phase arrest is induced by interference of TrkB gene.%目的 探讨慢病毒介导下干涉酪氨酸激酶B(TrkB)表达对乳腺癌细胞SK-BR-3增殖的影响及其作用机制.方法 对MCF-7、BCAP-37、SK-BR-3和MDA-MB231乳腺癌细胞进行筛选,选取TrkB表达水平相对较高的细胞进行慢病毒感染以干涉TrkB表达,构建稳定干涉TrkB的乳腺癌细胞系Lv-sh-TrkB,通过蛋白质印迹(Western blot)实验检测TrkB干涉效果,同时构建乱序干涉细胞系Lv-scramble.采用噻唑蓝方法、流式细胞仪检测干涉TrkB后细胞的增殖情况以及细胞周期变化.结果 4株乳腺癌细胞中,SK-BR-3细胞TrkB表达水平相对较高,通过慢病毒感染SK-BR-3细胞后筛选稳定干涉TrkB表达的细胞株Lv-sh-TrkB,Western blot实验结果表明,Lv-sh-TrkB细胞中TrkB表达水平明显低于空白对照和Lv-scramble 细胞.细胞增殖检测表明Lv-sh-TrkB细胞增殖速度明显低于空白对照和Lv-scramble细胞;流式细胞仪检测细胞周期结果显示Lv-sh-TrkB细胞发生G1期阻滞,空白对照、Lv-scramble和Lv-sh-TrkB细胞G1期比例分别为61％、62％和78％.结论 干涉TrkB基因后乳腺癌细胞SK-BR-3增殖受到明显抑制,细胞周期发生G1期阻滞.