摘要
目的探讨高迁移率簇蛋白B1(high mobiligy group box-1,HMGB1)RNA干扰对人卵巢癌细胞株OV5生物学活性的影响。方法合成siRNA序列,构建HMGB1基因的慢病毒shRNA载体,转染卵巢癌OV5细胞。实验分为HMGB1-siRNA-1组、HMGB1-siRNA-2组、HMGB1-siRNA-3组、阴性对照组和空白组。采用Real-time RT-PCR和蛋白质印迹法等方法检测RNA干扰对HMGB1表达的影响;MTT法绘制生长曲线,检测细胞凋亡;Transwell小室检测卵巢癌细胞侵袭能力的变化。结果 HMGB1-siRNA-1组、HMGB1-siRNA-2组和HMGB1-siRNA-3组细胞中,HMGB1mRNA的表达量分别为1.261±0.131、1.109±0.074和0.483±0.059,明显低于阴性对照组的2.331±0.192和空白对照组的2.164±0.177,差异有统计学意义,P=0.005。实验组癌细胞生长明显受到抑制,与对照组相比,差异有统计学意义,P=0.007。HMGB1表达下调明显增加癌细胞的凋亡,HMGB1-siRNA-3组的凋亡率(16.89±1.13)%,明显高于阴性对照组的(5.87±0.47)%和空白对照组的(4.51±0.48)%,差异有统计学意义,P<0.01。Transwell小室法检测细胞侵袭力显示,HMGB1-siRNA-3组穿膜细胞数明显减少,与阴性对照组和空白组比较,差异有统计学意义,P<0.05。结论 HMGB1特异性siRNA序列可显著下调HMGB1的表达,抑制OV5细胞的增殖,促进细胞凋亡,也可明显降低OV5细胞的侵袭能力,HMGB1可能成为卵巢癌治疗的一个潜在靶点。
OBJECTIVE To study the effects of HMGB1 small interfering RNA on biological activity of OV5 human ovarian cancer cell lines.METHODS Synthesis of siRNA sequences and construction HMGB1 shRNA lentiviral vectors were transfected into OV5 ovarian cancer cell lines.There were four groups in the experient：HMGB1-siRNA-1group,HMGB1-siRNA-2group,HMGB1-siRNA-3group,negtive control group and blank control group.Real-time RT-PCR and Western blot were used to detect the effect of RNA interference suppression the expression of HMGB1.The cell proliferation was assayed by MTT to make the growth curve and apoptosis was assayed by FCM.Invasiveness of ovarian cancer cells was assessed by boyden chamber invasion assay.RESULTS Relative mRNA expression quantity of HMGB1 in the five groups were 1.261±0.131,1.109±0.074,0.483±0.059,2.331±0.192 and 2.164±0.177.The experiment groups were higher than control groups significantly（P=0.005）.Real-time RT-PCR and Western blot confirmed that the expressions HMGB1 in mRNA and protein levels were suppressed significantly comparing with control group（P=0.005）.Cancer cells growth of experimental group was significantly inhibited（P=0.007）.Apoptosis of OV5 cells in experiment group（16.89±1.13）% was significantly higher than that in the negtive（5.87±0.47）% and blank（4.51±0.48）% control group（P〈0.01）.Down-regulaton the expression of HMGB1 could inhibit the invasion of ovarian cancer cells significantly（P〈0.05）.CONCLUSIONS HMGB1-specific siRNA sequence can significantly inhibit the expression of HMGB1,and significantly induce apoptosis of OV5,also inhibit invasion of OV5 cells.HMGB1 may become a potential target for treatment of ovarian cancer.
出处
《中华肿瘤防治杂志》
CAS
北大核心
2015年第14期1109-1114,共6页
Chinese Journal of Cancer Prevention and Treatment
基金
山东省自然科学基金(ZR2013HL047)
