臺灣博碩士論文加值系統

微核醣核酸 (microRNA) 主要是透過與標的基因的結合，造成標的基因的表現下降而影響癌細胞的轉移能力。甲狀腺素 (3,3',5-triiodo-L-thyronine; T3)會與甲狀腺素受體 (TR, thyroid hormone receptor) 結合，改變基因的表現，進而影響細胞的分化和生長。目前微核醣核酸如何參與甲狀腺素和甲狀腺素受體促使提升癌細胞爬行能力的機制尚未被釐清。根據本篇的研究，在細胞和動物的實驗模式中，甲狀腺素使miR-17的表現量下降，啟動子活性測試和染色體免疫沉澱實驗中得到甲狀腺素受體會結合在miR-17啟動子-2234/-2000的位置。在細胞和動物模式中得到驗證，發現大量表現miR-17會抑制細胞的爬行和侵犯的能力，主要是藉由抑制下游標的基因matrix metalloproteinases (MMP)-3。另外，抑制miR-17的表現促使磷酸化的AKT表現上升，進而增加細胞的侵犯能力，同時此現象會被LY294002抑制劑所抑制。此外在高度侵犯能力的細胞中miR-17是低表現的現象。甲狀腺素和甲狀腺素受體促使細胞爬行的能力可以被大量表現miR-17所部分抑制。進一步分析甲狀腺素受體、miR-17和MMP3在肝癌病人當中的表現，結果顯示在肝癌病人中，甲狀腺素受體是大量表現並且使病人存活率呈現下降的情況。同時也發現miR-17的表現與甲狀腺素受體或是MMP3的表現是呈現有意義的負相關。根據我們的實驗結果，發現甲狀腺素和甲狀腺素受體使miR-17表現量下降，此時MMP3表現量上升，進而導致細胞爬行能力的上升。

MicroRNAs (miRNAs) are thought to control tumor metastasis through direct interactions with target genes. Thyroid hormone (T3) and its receptor (TR) are involved in cell growth and cancer progression. However, the issue of whether miRNAs participate in T3/TR-mediated tumor migration is yet to be established. In the current study, we demonstrated that T3/TR negatively regulates mature miR-17 transcript expression, both in vitro and in vivo. Luciferase reporter and chromatin immunoprecipitation (ChIP) assays localized the regions responding to TR-mediated repression to positions -2234/-2000 of the miR-17 promoter sequence. Overexpression of miR-17 markedly inhibited cell migration and invasion in vitro and in vivo, mediated via suppression of matrix metalloproteinases (MMP)-3. Moreover, phosphorylated AKT expression was increased in miR-17 knockdown cells that led to enhanced cell invasion, which was blocked by LY294002. Notably, low miR-17 expression was evident in highly metastatic cells. The cell migration ability was increased by T3, but partially reduced upon miR-17 overexpression. Notably, TRα1 was frequently upregulated in hepatocellular carcinoma (HCC) samples and associated with shortened overall survival (P = 0.023). miR-17 expression was significantly negatively associated with TRα1 (P = 0.033) and MMP3 (P = 0.043) in HCC specimens. Data from our study suggest that T3/TR, miR-17, p-AKT and MMP3 activities are interlinked in the regulation of cancer cell metastasis.

TABEL OF CONTENTS
Part I: Thyroid hormone receptor represses miR-17 expression to enhance tumor metastasis in human hepatoma cells
Chapter I Introduction......................................................................1
1.1 Background...................................................................................1
1.2 Objective of the study...................................................................6
Chapter II Materials and Methods...................................................7
Chapter III Results...........................................................................15
3.1 miR-17 is downregulated by the thyroid hormone...............15
3.2 T3-dependent repression of the miR-17 promoter via interactions with TR...............................................................16
3.3 miR-17 controls cell migration and invasion via modulation of p-AKT activity....................................................................16
3.4 MMP3 is a miR-17 target gene upregulated by T3/TR........17
3.5 miR-17 is downregulated in a highly metastatic cell line.....19
3.6 Overexpression of miR-17 blocks MMP3-mediated T3 promotion of cell motility.........................................................20
3.7 miR-17 is downregulated by T3 in hyperthyroidal rats and negatively correlated with TRα1 and MMP3 in a subset of human HCC............................................................................21
Chapter IV Discussion......................................................................23
Chapter V Conclusion......................................................................28
Chapter VI Figures and Figure Legends........................................29
Reference.................................................................................................52
Part II: Thyroid hormone suppresses cell proliferation through endoglin-mediated promotion of p21 stability……………………….61
Part III: Thyroid hormone regulation of miR-21 enhances migration and invasion of hepatoma………………………………......................73

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