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1391~1396.锌指结构蛋白148 可变剪切体在结直肠癌发生发展中的作用[J].何平,高显华,许硕贵,丁杰,况飞.中国肿瘤生物治疗杂志,2017,24(12)
锌指结构蛋白148 可变剪切体在结直肠癌发生发展中的作用    点此下载全文    点此浏览HTML全文
何平  高显华  许硕贵  丁杰  况飞
1.南昌大学第三附属医院 普外科,江西 南昌 330008;2.上海长海医院急诊科,上海200433,1.南昌大学第三附属医院 普外科,江西 南昌 330008;2.上海长海医院急诊科,上海200433,1.南昌大学第三附属医院 普外科,江西 南昌 330008;2.上海长海医院急诊科,上海200433,1.南昌大学第三附属医院 普外科,江西 南昌 330008;2.上海长海医院急诊科,上海200433,1.南昌大学第三附属医院 普外科,江西 南昌 330008;2.上海长海医院急诊科,上海200433
基金项目:国家自然科学基金资助项目(No. 81572332)
DOI:10.3872/j.issn.1007-385X.2017.12.009
摘要:
      目的:探讨锌指结构蛋白148(zinc finger protein 148,ZNF 148)基因的两种可变剪接体对结肠癌细胞侵袭转移能力的影响及相关作用机制。方法:RT-PCR检测人结肠癌SW480 细胞中的ZNF148 两种剪接体的表达,构建ZNF148 干扰载体和过表达载体,将构建成功的ZNF148 干扰载体及过表达载体转染至人结肠癌SW480 细胞中,分为ZNF148FL-siRNA 组、ZNF148FL-Over express 组、ZNF148ΔN-siRNA 组、ZNF148ΔN-Over express 组、Control siRNA 组、Control Over express 组以及Normal control 组。RTPCR检测各组细胞mRNA表达水平,MTT、Transwell、划痕实验和流式细胞术检测人结肠癌SW480 细胞的增殖、体外细胞侵袭、迁移和凋亡情况。结果:RT-PCR 扩增获得全长分别为2 385 bp 以及2 004 bp 的ZNF148FL与ZNF148ΔN两种不同剪接体产物。ZNF148FL-siRNA组ZNF148FL的表达量明显下降,但ZNF148ΔN的表达水平上升;ZNF148FL-Over express 组ZNF148FL的表达量明显上升,而ZNF148ΔN的表达水平下降(均P<0.05)。ZNF148ΔN-siRNA组的ZNF148ΔN表达水平明显下降,但ZNF148FL的表达水平上升;ZNF148ΔN-Over express 组ZNF148ΔN表达水平升高,而ZNF148FL的表达水平下降(均P<0.05)。ZNF148FL-Over express 组及ZNF148ΔN-siRNA组SW480 细胞增殖能力增强,而ZNF148FL-siRNA组和ZNF148ΔN-Over express 组SW480 细胞增殖能力下降(均P<0.05)。ZNF148FL-siRNA 组和ZNF148ΔN-Over express 组穿膜SW480 细胞数量及迁移活性显著下降,而凋亡率显著上调;ZNF148FL-Over express 组和ZNF148ΔN-siRNA 组穿膜细胞数及迁移活性显著上调,而凋亡率显著下调(均P<0.05)。结论:ZNF148FL表现为增加结直肠癌细胞的增殖、侵袭和转移活力,而ZNFΔN作用与之相反,ZNF148 的ZNF148FL与ZNFΔN两种剪接体对于结直肠癌的恶性生物活性可能表现出了互相拮抗作用。
关键词:锌指结构蛋白148  可变剪接体  结直肠癌  SW480 细胞  侵袭  转移  凋亡
Roles of zinc finger protein 148 alternative splicing isoforms in occurrence and development of colorectal cancer    Download Fulltext
HE Ping  GAO Xianhua  XU Shuogui  DING Jie  KUANG Fei
1.Department of General Surgery, the 3rd Affiliated Hospital of Nanchang University, Nanchang 330008, Jiangxi, China;2. Department of Emergency, Changhai Hospital, Shanghai 200433, China,1.Department of General Surgery, the 3rd Affiliated Hospital of Nanchang University, Nanchang 330008, Jiangxi, China;2. Department of Emergency, Changhai Hospital, Shanghai 200433, China,1.Department of General Surgery, the 3rd Affiliated Hospital of Nanchang University, Nanchang 330008, Jiangxi, China;2. Department of Emergency, Changhai Hospital, Shanghai 200433, China,1.Department of General Surgery, the 3rd Affiliated Hospital of Nanchang University, Nanchang 330008, Jiangxi, China;2. Department of Emergency, Changhai Hospital, Shanghai 200433, China and 1.Department of General Surgery, the 3rd Affiliated Hospital of Nanchang University, Nanchang 330008, Jiangxi, China;2. Department of Emergency, Changhai Hospital, Shanghai 200433, China
Fund Project:Project supported by the National Natural Science Foundation of China(No. 81572332)
Abstract:
      Objective:To investigate the effect of two alternative splicing isoforms of zinc finger protein 148 (ZNF148) gene on the invasion and metastasis of colon cancer cells and their related mechanisms. Methods: RTPCR was used to determine the expressions of two ZNF148 alternative splicing isoforms in the human colorectal SW480 cells. ZNF148 interference vector and ZNF148 over-expression vector were constructed and transfected into the SW480 cells; them were divided into ZNF148FL-siRNA group, ZNF148FL-Over express group, ZNF148ΔN-siRNA group, ZNF148ΔN- Over express group, Control siRNA group, Control Over express group and Normal control group. The mRNA expressions in each group were examined by RT-PCR; the proliferation, invasion and migration in vitro as well as apoptosis of SW480 cells were detected by MTT, Transwell, scratch assay and flow cytometry, respectively.Results: Two splicing isoforms (ZNF148FL of 2 385 bp and ZNF148ΔN of 2 004 bp) were obtained by RTPCR.The expression level of ZNF148FL was significantly decreased while the expression of ZNF148ΔN was increased in the ZNF148FL-siRNA group; the expression of ZNF148FL was significantly increased while the expression of ZNF148ΔN was significantly decreased in the ZNF148FL- Over express group (all P<0.050. The expression of ZNF148ΔN was significantly decreased while the expression of ZNF148FL was increased in ZNF148ΔN-siRNA group;the expression of ZNF148ΔN was significantly increased while the expression of ZNF148FL was decreased in ZNF148ΔN-over express group (all P<0.05). The proliferation of the SW480 cells was increased in ZNF148FL-over express group and the ZNF148ΔN- siRNA group, while the proliferation of the SW480 cells was decreased in the ZNF148FL-siRNA group and the ZNF148ΔN-Over express group. The transmembrane cell number and migration ability of the SW480 cells in the ZNF148FL-siRNA group and the ZNF148ΔN-Over express group were significantly decreased,but the apoptotic rate was significantly increased; However, ZNF148FL-Over Express group and ZNF148ΔNsiRNA group showed the significantly increased transmembrane cell number and migration ability but decreased apoptosis rate (all P<0.05). Conclusion: ZNF148FL could increase proliferation, invasion and metastasis of colorectal cancer cells, while ZNFΔN showed opposite effect; the two splicing isoforms of ZNF148 may exert mutual antagonistic effect to each other on the malignant biological activities.
Keywords:zinc finger protein 148  variable splicing  colorectal cancer  SW480 cell  invasion  metastasis  apoptosis
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