2016-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/642613摘要：腫瘤幹細胞是癌細胞中一群具有自我更新、多能分化並可以不斷新生成腫瘤的細胞。將腫瘤 幹細胞培養在含有bFGF及EGF的培養基中，細胞增生則會形成具有神經幹細胞球體，進一步 可加以誘導分化成神經細胞。研究顯示微小核醣核酸(microRNAs, miRNAs)可藉由與其標的基因 mRNA之3’-UTR序列互補結合，抑制轉錄後基因的表達，進而達到基因調控的目的。除了在腫 瘤幹細胞中扮演調控的角色外，部分miRNAs也與神經發育及腫瘤形成有關。然而進一步深入探 討miRNAs對於腫瘤幹細胞其自我更新及多能分化的影響相關研究則非常有限。在本研究中我們將探討在腫瘤幹細胞中miRNAs在自我更新及多能分化上如何進行調控。在 第一年計晝中，我們將培養大鼠C6 glioma細胞株及小鼠P19胚胎瘤細胞株形成球體，並且鑑定 球體細胞的自我更新、多能分化及致癌能力。同時也建立H9人類胚胎幹細胞的球體細胞及神經 分化模式。以miRNA microarray作為平台，檢測球體細胞與貼附細胞中miRNAs的表現，建立 C6、H9及P19細胞株經由幹細胞—球體形成—神經分化等一系列過程之miRNA表達譜分析。 在球體細胞中高度表現的miRNA如miRNA-30c與其標的基因也將進行探討。除延續第一年的研 究，第二年我們將建立動物模式檢測高度表現miRNAs的細胞在活體的調控機制並加以探討。第 三年我們將探討NCI-60癌細胞株miRNAs的表現，以及miRNAs在“tumor suppressor”及“EMT” 相關之訊息傳遞機制，包括球體癌細胞中miRNAs其相互調控的關係及所牽涉的訊息傳遞路徑。 本計晝的研究結果將有助於釐清miRNAs在腫瘤幹細胞中所扮演的角色及其與標的基因間的相 互關係。並瞭解miRNAs在正常細胞分化的角色亦或是腫瘤幹細胞中所參與的調控機制，將有助 於進一步瞭解miRNAs對於細胞行為的影響。<br> Abstract: Cancer stem cells (CSCs) are a subpopulation of cells in the tumor that have self-renewal property, multipotency and can give rise to heterogeneous cancer cells that constitute the tumor. CSCs were cultured in vitro as spheres using a serum-free medium containing basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Upon induction, CSCs could differentiate to neural cells and showed remarkable similarities to normal neural stem cells (NSCs). MicroRNA (miRNAs) play a regulatory role in CSCs including in glioblastoma multiforme (GBM) by imperfect base-pairing binding to complementary sequences in the 3’UTR of target mRNA transcripts. MiRNAs are also involved in neural development and tumor formation. However, the mechanism of miRNA-regulated self-renewal ability and multipotency is poorly understood.We aim to investigate how miRNAs regulate self-renewal ability and multipotent differentiation in CSCs. In the first year, we will examine the CSCs identified from rat glioma C6 and mouse embryonic carcinoma P19 cells under the “sphere forming” culture condition. The miRNA expression profiles of C6 cells, P19 cells and human embryonic stem cells (H9 cells) will be established. MiRNA microarray will be used as a platform to identify the miRNA contributes in the processes of sphere formation and neural differentiation in the cell models of C6, P19 and human embryonic stem cells (H9). The relationship of the highly expressed miRNAs, such as miRNA-30c, and their target genes in sphere cells will be explored. In the second year, in vivo tumorigenicity experiments and the miRNA regulation of the cells in SCID mouse model will be conducted. In the third year, we will study the miRNA expression patterns of NCI-60 cancer cell panel. The relationship of miRNAs and the involved signaling pathway including “tumor suppressor” and EMT in NCI-60 cancer cells will be investigated. The results of this study will contribute to clarifying the relationship of miRNAs and its target genes in CSCs. It also can help us to understand the influence of miRNAs in cell behaviors that not only the roles of miRNAs in normal cell differentiation buy also the mechanisms of regulation in CSCs.腫瘤幹細胞微小核糖核酸自我更新多能分化球體形成cancer stem cellmicroRNAself-renewalmultipotencysphere formation