[1]付　锴　江普查　宫　睿　王　伟.表达VASH1基因的人脑胶质瘤U-87MG细胞对化疗药物敏感性的变化[J].中国临床神经外科杂志,2016,(01):34-37.[doi:10.13798/j.issn.1009-153X.2016.01.012]
　FU Kai,JIANG Pu-cha,GONG Rui,et al.Effect of lentiviral vector-mediated VASH1 gene on chemosensitivity of human glioma U-87MG cells[J].,2016,(01):34-37.[doi:10.13798/j.issn.1009-153X.2016.01.012]
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表达VASH1基因的人脑胶质瘤U-87MG细胞对化疗药物敏感性的变化()
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《中国临床神经外科杂志》[ISSN:1009-153X/CN:42-1603/TN]

卷:
期数:
2016年01期
页码:
34-37
栏目:
论著
出版日期:
2016-01-25

文章信息/Info

Title:
Effect of lentiviral vector-mediated VASH1 gene on chemosensitivity of human glioma U-87MG cells
文章编号:
1009-153X（2016）01-0034-04
作者:
付　锴　江普查　宫　睿　王　伟
430071 武汉，武汉大学中南医院神经外科
Author(s):
FU Kai;  JIANG Pu-cha;  GONG Rui;  WANG Wei.
Department of Neurosurgery, Zhongnan Hospital, Wuhan University, Wuhan 430071, China
关键词:
脑胶质瘤; U-87MG细胞; 血管生成抑制因子１; 慢病毒载体; 化疗; 药物敏感性
Keywords:
VASH1;  Lentiviral vector;  Glioma;  Chemosensitivity
分类号:
R 739.41; R 730.53
DOI:
10.13798/j.issn.1009-153X.2016.01.012
文献标志码:
A
摘要:
目的 探讨表达人血管生成抑制因子１（VASH1）的人脑胶质瘤U-87MG细胞对化疗药物的敏感性变化。方法 构建针对VASH1的慢病毒载体pGCL-GFP-VASH1，经测序鉴定后转染293T细胞，筛选出适合浓度的慢病毒转染人脑胶质瘤U-87MG细胞，荧光显微镜下检测转染效率；通过RT-PCR和Western blot分析U-87MG细胞VASH1 mRNA和蛋白表达水平；用CCK-8法检测U-87MG细胞在化疗药物顺铂和替莫唑胺作用下的存活率。流式细胞仪检测U-87MG细胞凋亡。结果 成功构建pGCL-GFP-VASH1慢病毒载体，并成功转染U-87MG细胞，转染率达70％以上；RT-PCR和Western blot结果证实转染VASH1慢病毒载体的U-87MG细胞表达VASH1 mRNA和蛋白。在顺铂或替莫唑胺作用下，表达VASH1的U-87MG细胞存活率均较未表达VASH1的U-87MG细胞明显降低（P<0.01），而且U-87MG细胞凋亡率明显增加（P<0.01）。结论 VASH1慢病毒载体转染U-87MG细胞可使其稳定表达VASH1，并提高人脑胶质瘤U-87MG细胞对化疗药物敏感性、增加细胞凋亡率。
Abstract:
Objective To construct the lentiviral vector over-expressing VASH1 and study the relationship between the expression of VASH1 and the chemosensitivity of human glioma U-87MG cells. Methods The lentiviral vector carrying human VASH1 gene (pGCL-GFP-VASH1), which was constructed and sequenced, was transinfected into 293 T cells. The transinfection efficiency was evaluated and then the lentiviral vector was transfected into the human glioma U-87MG cells. Transfection efficiency was determined by Fluorescence microscopy. The expression of VASH1 mRNA and protein in U87MG cells were detected respectively by RT-PCR and Western blot. The changes in the sensitivity of transfected U-87MG cells to the chemotherapeutic drugs including cispatin and temozolomide were determined by cell counting kit-8 (CCK-8). The apoptosis of U-87MG cells transfected with pGCL-GFP-VASH1 was determined by flow cytometry (FCM). Results The lentiviral vector-mediated VASH1 gene was successfully constructed and transfected into the U-87 cells. Immunofluorescence assay demonstrated that the transfection efficiency was above 70%. RT-PCR and Western blot analyses demonstrated that pGCL-GFP-VASH1 significantly increased the expressions of VASH1 mRNA and protein by 21.57% and 27.29% respectively in transfected U-87MG cells 96 hours after the transfection compared to the control group(P<0.01); CCK-8 Results showed that when exposed to cisplatin or temozolomide, the survival rate of pGCL-GFP-VASH1-transfected U-87 cells were significantly decreased to 23.96 and 17.24% respectively, which were significantly lower than those in the control groups. FCM Results showed that the apoptosis rate of U-87MG cells transfected with pGCL-GFP-VASH1 was significantly higher than that in the control groups (P<0.01). Conclusion The lentiviral vector-mediated VASH1, which can significantly increase the expression of VASH1 in U-87 MG cells, may enhance chemosensitivity and apoptosis of human glioma U-87MG cells.

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更新日期/Last Update: 2016-01-30