摘要
目的探讨沉默GTPBP4基因对结肠癌细胞(HT29)生物学行为的影响及机制。方法将培养好的HT29细胞随机分为转染组与阴性对照组,转染组加入适量GTPBP4-RNAi慢病毒,阴性对照组转染含有非特异性干扰序列的阴性病毒。采用CCK8法检测细胞增殖能力,划痕实验检测细胞迁移能力,Transwell实验检测细胞侵袭能力,流式细胞术检测细胞周期,AnnexinⅤ-APC单染法检测细胞凋亡率,细胞克隆形成实验检测细胞克隆形成情况,Western blotting检测细胞内p53、Survivin蛋白表达,细胞免疫荧光法检测细胞内蛋白p53、Survivin定位。结果与阴性对照组比较,转染组细胞增殖能力低(P <0. 05),迁移距离短(P <0. 05),穿膜细胞数少(P <0. 05),G1期细胞比例高(P均<0. 05)、S期细胞比例低(P <0. 05),凋亡率高(P <0. 05),细胞克隆形成的细胞集落数目少(P <0. 05),p53蛋白相对表达量高(P <0. 05),Survivin蛋白相对表达量低(P <0. 05)。两组p53、Survivin蛋白均主要表达于细胞核,未发生表达位置改变。结论沉默GTPBP4基因可抑制结肠癌细胞HT29的恶性生物学行为,该作用可能与调节p53、Survivin蛋白表达有关。
Objective To investigate the effect and mechanism of silencing GTPBP4 gene on the biological behavior of colon cancer cells (HT29). Methods The cultured HT29 cells were randomly divided into the transfection group and negative control group. The appropriate amount of GTPBP4-RNAi lentivirus was added into the transfection group,and the negative control group was transfected with negative virus containing non-specific interference sequence. CCK8 was used to detect cell proliferation,scratch test was used to detect cell migration,Transwell test was used to detect cell invasion,the cell cycle was detected by flow cytometry,the apoptosis rate was detected by AnnexinV-APC single staining method,and the cell clone was detected by cell clone formation assay. The expression of P53 and Survivin of cells was detected by West-ern blotting. The immunofluorescen-ce assay was used to detect the localization of P53 and Survivin. Results Compared with the negative control group,the transfection group had lower proliferation ability (P < 0. 01),fewer transmembrane cells (P <0. 01),higher proportion of cells in the G1 phase (P < 0. 05),lower proportion of cells in the S phase (P < 0. 05),higher apoptotic rate (P <0. 05),and fewer colonies formed by cell cloning (P <0. 05). The relative expression of p53 protein was higher in the transfection group (P <0. 05),while the Survivin protein was lower (P <0. 05). The p53 and Survivin proteins were mainly expressed in the nucleus of the cells in both groups,and no change of expression location was observed. Conclusion Silencing GTPBP4 gene can inhibit the malignant biological behavior of colon cancer cells HT29,which may be related to regulating the expression of p53 and Survivin protein.
作者
唐丹
刘勋
杨志惠
TANG Dan;LIU Xun;YANG Zhihui(Southwest Medical College,Luzhou 646000,China)
出处
《山东医药》
CAS
2019年第1期34-37,共4页
Shandong Medical Journal
基金
泸州市科技局科技计划项目(2017LZXNYD-T04
2013LZLY-J44)
