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標題: | 微核醣核酸(miR-19c)在斑馬魚血管發育扮演角色之探討 The role of miR-19c in zebrafish vessel development |
作者: | Ying-Hsien Liao 廖穎嫻 |
指導教授: | 李士傑(Shyh-Jye Lee) |
關鍵字: | 斑馬魚,血管新生發育,淋巴管新生發育,微核醣核酸,miR-19c, Angiogenesis,Lymphangiogenesis,Zebrafish,miRNAs,miR-19c, |
出版年 : | 2015 |
學位: | 碩士 |
摘要: | 血管及淋巴管是循環系統的基本構造,但其在脊椎動物之研究常因皮膚及其他阻隔而增加其困難度。斑馬魚由於其胚胎透明、並有幾種血管螢光魚品系易於研究,近年來已成為血管新生(angiogenesis) 及淋巴管新生(lymphangiogenesis) 的主流模式動物之一。微核醣核酸(miRNAs)在血管新生及淋巴管新生中扮演重要角色,前人研究發現幾種miRNAs在斑馬魚血管內皮細胞高量表現,並發現其中之一miR-221在血管新生成有其功能,但其他miRNAs所扮演角色則仍未知。在本論文中,我重複前人試驗發現在利用流式細胞儀分選出的內皮細胞中,一些miRNAs的表現量於產卵後22-48小時有顯著的變化。其中miR-19c被發現在22-48小時有顯著的上升情形,由已知的研究中知道這個時間點與血管和淋巴管發育皆有所相關,所以我大膽假設miR-19c可能參與血管發育或淋巴管發育。為了釐清這個問題,我以血管螢光轉基因斑馬魚為材料,利用morpholino/miRNA抑制劑與miRNA模擬物(mimic)分別抑制或增加miR-19c的表現來探討miR-19c在斑馬魚發育過程中所扮演的角色。我發現增加miR-19c的表現會造成淋巴血管系統缺陷,但不管是抑制或增加miR-19c的表現皆不明顯影響血管系統。利用微核醣核酸目標預測資料庫(Target Scan)與定量即時聚合酶鏈鎖反應(real-time PCR),我發現transforming growth factor beta 1a (tgfβ1a)為miR-19c的目標基因之一,且增加miR-19c表現可降低tgfβ1a表現量,但tgfβ1a在淋巴管新生成之功能則尚待釐清。總體而言,本研究結果顯示,miR-19c可能經由tgfβ1a來影響斑馬魚淋巴管的發育。 Vessel formation, including angiogenesis and lymphangiogenesis, paves the structural foundation of circulation. The study of vessel formation in vertebrates often hinders by the invisibility, but the availability of transparent zebrafish embryos greatly accelerates our understanding in the field. With the addition of several vessel reporter lines, zebrafish has become one of the popular models to explore the regulatory mechanisms of angiogenesis and lymphangiogenesis. microRNAs (miRNAs) have been reported to be involved in regulating angiogenesis and lymphangiogenesis. In particularly, several miRNAs have been shown to be enriched in blood endothelial cells. However, except miR-221, the functional roles of other miRNAs remain unknown. Here, I first validated the enrichment of those miRNAs in endothelial cells from embryos at 22-48 h post fertilization. Interestingly, I found one of miRNAs, miR-19c, increases notably during 22-48 hpf. This time is associated with blood vessel and lymphatic vessel formation, I hypothesized that miR-19c may regulate blood or lymphatic vessel formation. I performed loss and gain of function analyses by applying antisense morpholino/miRNA inhibitor and miRNA mimic against miR-19c to different transgenic zebrafish lines expressing GFP. I observed that overexpression of miR-19c impaired lymphatic development, but no significant change in blood vascular system was observed in embryos. Transforming growth factor beta1a (tgfβ1a) is a potential target of miR-19c and known lymphangiogenesis regulator. I found significant reduction of the expression of tgfβ1a in embryos injected with miR-19c mimic. The functional link between miR-19c and tgfβ1a still needs to be investigated. Collectively, I conclude that miR-19c is indispensable to lymphatic development in zebrafish possibly via tgfβ1a. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/7999 |
全文授權: | 同意授權(全球公開) |
電子全文公開日期: | 2025-06-24 |
顯示於系所單位: | 生命科學系 |
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ntu-104-1.pdf 此日期後於網路公開 2025-06-24 | 3.58 MB | Adobe PDF |
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