Huang Fei, Lin Chen, Shi Yong-Hua, Kuerban Gulinar
Department of Pathology, Basic Medical Sciences, Xinjiang Medical University, Urumqi, China E-mail :
Asian Pac J Cancer Prev. 2013;14(10):5915-20. doi: 10.7314/apjcp.2013.14.10.5915.
Although aberrant miRNA expression has been documented, altered miR-101 expression in cervical cancer and its carcinogenic effects and mechanisms remain unexplored. The aim of our study was to investigate the role of miR-101 alteration in cervical carcinogenesis.
Expression of miR-101 was examined by quantitative real-time reverse transcriptase PCR (qRT-PCR) in Hela cells. After modulating miR-101 expression using miR-101 mimics, cell growth, apoptosis and proliferation, and migration were tested separately by MTT or flow cytometry and cell wound healing assay and protein expression was detected by qRT-PCR. The expression of COX-2 in Hela cell was also examined by immunohistochemical staining and the correlation with miR-101 expression was analysed.
The miR-101 demonstrated significantly low expression in Hela cell. When we transfected miR-101 mimics into Hela cells, the modulation of miR-101 expression remarkably influenced cell proliferation, cycling and apoptosis: 1) The expression of microRNA-101 tended to increase after transfection; 2) Overexpression of miR-101 was able to promote cell apoptosis, the apoptosis rate being markedly higher (97.6%) than that seen pre-transfection (12.2%) (P <0.05); 3) The miR-101 negatively regulates cell migration and invasion, scratch results being lower (42.7um±2um) than that observed pre-transfection (181.4 um±2 um); 4) miRNA-101 inhibits the proliferation of Hela cells as well as the level of COX-2 protein, which was negatively correlated with miR-101 expression.
Overexpression of miR-101 has obvious inhibitory effects on cell proliferation, migration and invasion. Thus reduced miR-101 expression could participate in the development of cervical cancer at least partly through loss of inhibition of target gene COX-2, which probably occurs in a relative late phase of carcinogenesis. Our data suggest an important role of miR-101 in the molecular etiology of cancer and indicate potential application of miR-101 in cancer therapy.
尽管已有文献记载miRNA表达异常,但宫颈癌中miR-101表达的改变及其致癌作用和机制仍未得到探索。本研究的目的是探讨miR-101改变在宫颈癌发生中的作用。
采用定量实时逆转录聚合酶链反应(qRT-PCR)检测Hela细胞中miR-101的表达。使用miR-101模拟物调节miR-101表达后,分别通过MTT法、流式细胞术检测细胞生长、凋亡和增殖情况,通过细胞划痕愈合试验检测细胞迁移情况,通过qRT-PCR检测蛋白质表达。通过免疫组织化学染色检测Hela细胞中COX-2的表达,并分析其与miR-101表达的相关性。
miR-101在Hela细胞中表达明显较低。当我们将miR-101模拟物转染到Hela细胞中时,miR-101表达的调节显著影响细胞增殖、周期和凋亡:1)转染后microRNA-101的表达趋于增加;2)miR-101的过表达能够促进细胞凋亡,凋亡率明显高于转染前(97.6%比12.2%)(P<0.05);3)miR-101负向调节细胞迁移和侵袭,划痕结果低于转染前(42.7um±2um比181.4 um±2 um);4)miRNA-101抑制Hela细胞的增殖以及COX-2蛋白水平,COX-2蛋白水平与miR-101表达呈负相关。
miR-101的过表达对细胞增殖、迁移和侵袭具有明显的抑制作用。因此,miR-101表达降低可能至少部分通过失去对靶基因COX-2的抑制作用参与宫颈癌的发生发展,这可能发生在致癌过程的相对晚期。我们的数据表明miR-101在癌症分子病因学中具有重要作用,并提示miR-101在癌症治疗中的潜在应用。