- 學位論文
以細胞模式探討miR-622抑制肺癌細胞侵襲轉移之機制
Inhibitory effects of miR-622 on tumor invasion and migration in lung cancer cells in vitro
摘要
MicroRNA(miRNA)是一段長度約23個鹼基對的微型核醣核酸，其透過結合在mRNA的3端不轉譯區域上，抑制基因轉譯達成調節基因功能的目的。近年來，許多文獻也證實miRNA與腫瘤的生長、癌化、藥物感受性、轉移以及侵襲有密切的相關性，miRNA在腫瘤發展過程中扮演一個極為重要角色。我們利用生物資訊技術在TargetScan資料庫中推測miR-622是缺氧誘導因子 Hypoxia-inducible factor 1-alpha(HIF-1α)的調控者，HIF1-α在多種癌症中都有被報導出與腫瘤的生長，轉移以及侵襲有關，因癌細胞快速生長造成局部缺氧，而缺氧反應會誘導HIF-1α表現量增加，形成一種正回饋調控，使癌細胞在缺氧環境下快速生長，甚至轉移到正常組織。但MicroRNA與HIF-1α相關的研究並不多，有鑑於此我們想要知道MicroRNA-622與HIF-1α兩者關係為何。 首先，利用生物冷光(luciferase)報導基因實驗，證實hsa-mir-622確實會結合到HIF-1α 3'UTR上而降低冷光表現，利用miR-622 mimic轉染到肺癌細胞株A549中大量表現miR-622，藉由Western Blot觀察 HIF-1α蛋白表現量，也有降低的情形，最後也探討了miR-622是否具有影響細胞的轉移和侵襲能力，結果顯示當大量表現miR-622時，細胞的轉移和侵襲能力明顯被抑制了60%，用miR-622 inhibitor抑制miR-622表現，細胞的轉移和侵襲能力回復。本篇研究，證實在肺癌細胞株中hsa-mir-622會直接結合在 HIF-1α 3'UTR上，抑制 HIF-1α表現量，使細胞的轉移侵襲能力降低。
並列摘要
Hypoxia-inducible factor-1α (HIF-1α) has been found to involves in tumor progression, angiogenesis, metastasis, and invasion. HIF-1α have also been shown to induce EMT and metastasis through the repression of E-cadherin. MicroRNAs (miRNAs) are a class of small noncoding RNAs that control gene expression by targeting mRNA 3’untranslated region. Recently, there are growing evidence showed that miRNAs can affect the genesis and development of tumor and play a kind of tumor suppressor or oncogenic function. In the present study, we identified that Hypoxia-inducible factor-1α (HIF-1α) is a putative target of hsa-miR-622.Using luciferase reporter assay we confirmed that miR-622 could reduce luciferase expression by binding HIF-1α 3’UTR. Overexpression of miR-622 in the lung cancer cell lines, A549 or H1299, HIF-1α protein level is downregulated by miR-622. Further, study in vitro has been shown that both capabilities of tumor invasiveness and motility were significantly inhibited by overexpression of miR-622 in lung cancer cells. In addition, we found that the levels of vimentin were drastically decreased, but levels of E-cadherin were induced in miR622-transfected cells. Our data indicate that miR-622 plays a key role in controlling the expression of HIF-1α may mediate the tumor suppressor effects of miR-622.