[1]马贺,梅鹏金,赵英玉,等.Dyskerin假尿苷合成酶1对胶质瘤细胞增殖、迁移和侵袭的影响研究[J].临床神经外科杂志,2020,17(03):292-297.[doi:DOI:10.3969/j.issn.1672-7770.2020.03.011]
 MA He,MEI Peng-jin,ZHAO Ying-yu,et al.Effect of DKC1 on cell proliferation, migration and invasion of human glioma U87 cells[J].Journal of Clinical Neurosurgery,2020,17(03):292-297.[doi:DOI:10.3969/j.issn.1672-7770.2020.03.011]
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Dyskerin假尿苷合成酶1对胶质瘤细胞增殖、迁移和侵袭的影响研究()
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《临床神经外科杂志》[ISSN:1672-7770/CN:32-1727/R]

卷:
17
期数:
2020年03期
页码:
292-297
栏目:
论著
出版日期:
2020-06-15

文章信息/Info

Title:
Effect of DKC1 on cell proliferation, migration and invasion of human glioma U87 cells
文章编号:
202003011
作者:
马贺梅鹏金赵英玉蒋昊张明明刘强范月超
221000 徐州,徐州医科大学附属医院(马贺,梅鹏金,赵英玉,蒋昊,范月超);徐州医科大学麻醉学院(张明明,刘强)
Author(s):
MA He MEI Peng-jin ZHAO Ying-yu et al.
The Affiliated Hospital of Xuzhou Medical University, Xuzhou 221000, China
关键词:
Dyskerin假尿苷合成酶1胶质瘤增殖迁移侵袭
Keywords:
dyskerin pseudouridine synthase 1 glioma proliferation migration invasion
分类号:
R739.41
DOI:
DOI:10.3969/j.issn.1672-7770.2020.03.011
文献标志码:
A
摘要:
目的探讨Dyskerin假尿苷合成酶1(Dyskerin pseudouridine synthase 1,DKC1)对人脑胶质瘤U87细胞增殖、迁移和侵袭的影响及其可能的作用机制。方法采用Control siRNA、DKC1 siRNA分别转染人脑胶质瘤U87细胞;Western blot检测DKC1蛋白表达水平及AKT、p-AKT、p70S6K、p-p70S6K蛋白表达水平的影响,基质金属蛋白酶(matrix metalloprotein,MMP)-2、MMP-9的表达量。细胞计数试剂盒(CCK-8)、细胞增殖实验检测DKC1 siRNA对人脑胶质瘤U87细胞增殖的影响;细胞划痕实验及transwell迁移、侵袭实验检测细胞的迁移、侵袭能力。结果 (1)通过CCK-8实验发现下调DKC1基因后,与Ctrl siRNA组比较,在24 h、48 h、72 h、96 h时间点DKC1组胶质瘤U87细胞OD值明显降低,差异有统计学意义(P<0.01-0.001)。(2)划痕实验显示:下调DKC1基因后,与Ctrl siRNA组比较,在0 h、24 h、48 h时间点DKC1组胶质瘤U87细胞迁移距离小,差异有统计学意义(P<0.01-0.001)。(3)transwell实验显示:下调DKC1基因后,与Ctrl siRNA组比较,DKC1组胶质瘤U87细胞迁移和侵袭细胞个数均明显减少,差异有统计学意义(均P< 0.01)。(4)Western blot实验显示:与Ctrl siRNA组相比,DKC1组的基因序列的蛋白表达明显降低(P<0.05)。下调DKC1基因后,DKC1组的AKT、pAKT蛋白表达较Ctrl siRNA组变化不明显,差异无统计学意义(均P>0.05),说明下调胶质瘤U87细胞中DKC1基因表达对于AKT、pAKT的表达无明显影响;DKC1组p70S6K、p-p70S6K蛋白表达较Ctrl siRNA组明显降低,差异有统计学意义(P<0.05- 0.01),表明在U87细胞中下调DKC1基因对p70S6K、p-p70S6K的表达水平有明显的抑制作用。DKC1组MMP2、MMP9蛋白表达较Ctrl siRNA组明显降低,差异有统计学意义(P<0.05-0.01),表明在U87细胞中下调DKC1基因对MMP2、MMP9的蛋白表达水平有明显的抑制作用。结论(1)DKC1对胶质瘤生物学行为的影响,提示DKC1可能参与了胶质瘤的发生和发展;(2)DKC1 siRNA靶向干扰了DKC1的表达,同时下调DKC1后AKT、p-AKT的表达水平无明显变化,p70S6K、p-p70S6K蛋白的表达水平明显降低;(3)DKC1组胶质瘤U87细胞MMP2、MMP9的表达水平较Ctrl siRNA组明显降低;(4)下调DKC1的表达水平能够抑制人脑胶质瘤U87细胞迁移、侵袭的能力。
Abstract:
Objective To investigate DKC1 on human glioma U87 cell proliferation, migration and invasion effect and its possible mechanism. Methods Control small interference RNA(siRNA) and DKC1 siRNA were used respectively to transfect human brain glioma U87 cells. The expression levels of DKC1 proteins in human brain glioma U87 cells was detected after DKC1 knockdown by using Western blotting. The expression levels of AKT、p-AKT、p70S6K、p-p70S6K、MMP2、MMP9 proteins in human brain glioma U87 cells were suppressed after DKC1 knockdown by using Western blotting. The influence of DKC1 knockdown by using Western blotting on the proliferation of human brain glioma cells was tested by using cell counting kit-8(CCK-8) assay. The migration and invasion ability of U87 cells were respectively measured by wound-healing assay and transwell migration and invasion assays. Results (1)According to CCK-8 assay, the loss of DKC1 can suppress the proliferation of U87 cells (P<0.01-0.001).(2)The result of wound-healing assay showed that the migration distance in the DKC1 group was significantly shorter than that in the Ctrl siRNA group (P<0.01-0.001). (3)The result of transwell detection showed that the number of migration and invasion cells in the DKC1 group was significantly lower than the Ctrl siRNA group (all P<0.01). (4) Western blot analysis showed that DKC1 gene expression in Ctrl siRNA group was significantly higher than that in DKC1 group. In DKC1 siRNA transfected cells, the expression of p70S6K, p-p70S6K, MMP2 and MMP9 was down-regulated(P<0.05-0.01), but the expression of AKT and p-AKT was without any changes(all P>0.05). Conclusion (1)DKC1 impacts on the biological behavior of glioma cells hint the DKC1 may participate in the development of glioma. (2)DKC1-targeted siRNA interferes with the expression levels of DKC1. The expression levels of p70S6K、p-p70S6K was down-regulated but the expression levels of AKT 、p-AKT without any change in DKC1 siRNA transfected cells (P<0.05). (3) As compared with Ctrl siRNA group, lower expression level of MMP2、MMP9 was detected in DKC1 siRNA group. (4)Downregulation of DKC1 expression can inhibit migration and invasion of human glioma U87 cells.

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更新日期/Last Update: 2020-06-17