[1]陈芳,周畅,陆艳霞,等.Hsa-miR-186在结肠癌细胞中的表达及作用[J].南方医科大学学报,2013,(05):654.
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Hsa-miR-186在结肠癌细胞中的表达及作用()
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《南方医科大学学报》[ISSN:/CN:]

卷:
期数:
2013年05期
页码:
654
栏目:
出版日期:
2013-05-15

文章信息/Info

Title:
Expression of hsa-miR-186 and its role in human colon carcinoma cells
作者:
陈芳周畅陆艳霞袁理彭璠莉郑林李学农
关键词:
结肠癌细胞hsa-miR-186慢病毒增殖迁移侵袭
Keywords:
colorectal cancer cell lines Has-mir-186 lentivirus proliferation migration invasion
摘要:
目的检测hsa-miR-186在结肠癌组织及细胞中的表达,探讨hsa-mir-186对结肠癌细胞生物学特性的影响及作用。方法
应用实时荧光定量PCR 检测了miR-186 在12 对配对的结肠癌组织、癌旁组织和5 株结肠癌细胞中的表达情况;扩增包含
miR-186前体序列在内的基因片段,并将其克隆至PLVTHM载体,将PLVTHM- miR186质粒转染SW620细胞,流式分选慢病
毒感染的SW620细胞;CCK-8法、划痕实验和Transwell检测细胞检测细胞的增殖、迁移和侵袭能力。Western Blot检测靶基因
YY1 蛋白水平的表达。结果与癌组织相比,12 例癌组织中miR-186 的平均表达量为0.0024±0.0027,显著低于癌旁组织的
0.066±0.068,P=0.008;miR-186 在高转移潜能的人结肠癌细胞SW620 和LOVO相对表达量分别为0.118±0.138 和0.157±
0.001,与在低转移潜能HT-29 细胞中表达量1.000 ± 0.00,差异具有统计学意义P<0.05;质粒双酶切及测序鉴定
PLVTHM-hsa-miR186重组质粒构建成功;荧光定量PCR表明稳定过表达miR-186的结肠癌细胞株SW620构建成功,且过表达
miR-186后降低了细胞的增殖、迁移及侵袭能力,差异均具有统计学意义P<0.05。稳定过表达miR-186的细胞中,miR-186的表
达明显高于对照组和未处理组,YY1蛋白质水平有所下降。结论hsa-miR-186在结肠癌组织以及在高转移潜能的人结肠癌细
胞SW620、LOVO中低表达,具有类似抑癌基因的作用;成功构建PLVTHM-miR186慢病毒重组质粒,并稳定转染结肠癌细胞
SW620,抑制了细胞的增殖、迁移和侵袭能力。miR-186调控YY1表达可能是抑制结直肠癌生物特性的重要机制之一。
Abstract:
Objective To explore the expression of hsa-mir-186 in colorectal cancer and study its role in regulating the biological
behaviors of human colorectal cancer SW620 cells in vitro. Methods The expression of hsa-miR-186 in colon cancer tissue and
the adjacent tissues as well as 5 colon carcinoma cells were analyzed using real-time quantitative RT-PCR. The precursor
sequence of miR-186 gene was amplified from the genomic DNA by PCR and cloned into the lentiviral vector PLVTHM labeled
with GFP. The colorectal cancer cell line SW620 was transfected with PLVTHM-miR186 vector and the lentivirus-infected cells
were sorted with flow cytometry. Cell counting kit-8 (CCK-8) assay was used to detect the proliferation of the cells. The
migration and invasion of SW620 cells were investigated using Transwell assay and scratch test. Western blotting was used to
detect the expression of YY1 protein in SW620 cell lines. Results The relative expression of miR-186 in the cancer tissues was
0.0024 ± 0.0027, significantly lower than that in the adjacent tissues (0.066 ± 0.068, P=0.008); the relative expression level of
hsa-miR-186 in SW620 and LoVo cells with a high metastatic potential was 0.118 ± 0.138 and 0.157 ± 0.001, respectively,
significantly lower than that in HT-29 cells with a low metastatic potential (1.000 ± 0.00, P<0.05). The recombinant lentiviral
vector PLVTHM-miR186, verified by enzyme digestion, sequencing and qPCR, caused significant inhibition of cell
proliferation, migration and invasion and suppressed the expression of YY1 protein in SW620 cells. Conclusion As a tumor
suppressor gene, Hsa-miR-186 is down-regulated in colon carcinoma tissues and in highly metastatic SW620 and LoVo cells.
Has-miR-186 can inhibit the cell proliferation, migration and invasion of colon carcinoma cells in vitro possibly by suppressing
YY1 expression.

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[1]黄宪章,赵学芹,李曼,等.Akt shRNA载体的构建及其对结肠癌细胞株Lovo Akt基因表达的抑制作用[J].南方医科大学学报,2011,(11):1914.

更新日期/Last Update: 1900-01-01