miR-19c藉由調節Tgfβ1a影響淋巴管發育

Abstract

血管發育包含血管和淋巴管生成，是組成循環系統的基礎。之前在斑馬魚的研究發現微核糖核酸(microRNA)可藉由調控抑制血管生成的因子的表現，來促進血管生成。在此論文，我首先利用Q-PCR分析證實斑馬魚胚胎在受精後24小時到48小時之間，數種miRNA表現量具有顯著變化，其中一種miRNA - miR-19c的表現量會在受精後48小時有顯著增加，顯示miR-19c可能在後期血管發育上扮演很重要的角色。我們利用表現不同螢光蛋白於淋巴管及血管中之基因轉殖魚來觀察改變miR-19c表現量對血管發育的影響。增加miR-19c在斑馬魚中的表現量時，在受精五天後造成淋巴管過度分岔並產生水腫，不過對於血管並沒有看到明顯的影響。Q-PCR的分析顯示，miR-19c 在基因轉殖魚中過度表現會減少miR-19c可能的標的蛋白TGFβ1a的表現量。利用冷光蛋白分析試驗，我也證實了miR-19c可結合至tgfβ1a 端不轉譯區間。更重要的是，過量miR-19c所導致之水腫可被同時過量表現tgfβ1a所拯救。總結來說我的研究顯示，過量之miR-19c可藉由降解TGFβ1a之生成造成淋巴管過度增生及水腫表徵，因此證實miR-19c在淋巴管生成之重要性。

MicroRNAs (miRNA) are known regulators of gene expression in different cellular processes including development. Recently, different miRNAs were found to be enriched in endothelial cells (ECs) that suggests a role of miRNA in vascular development. miRNAs have been shown to mediate blood vessel development. However, their role in lymphatic vessel development is less well understood. Using zebrafish as a model, we analyzed miRNAs in ECs and found one miRNA, miR-19c, with increased expression between 24 to 48 hours post fertilization (hpf). We show that knocking down miR-19c by morpholino oligonucleotides has no effect on blood or lymphatic vasculature, while over expression by injecting miRNA mimic causes over-branching of lymphatic vessels. This suggests miR-19c as a positive regulator of lymphatic development. One probable target of miR-19c is transforming growth factor β 1a (tgfβ1a). Tgfβ1a may have the potential to prevent lymphangiogenesis and promote angiogenesis. Thus, we co-injected miR-19c mimic with tgfβ1a and they lessened the lymphatic defects observed. Moreover, we validated that tgfβ1a is a direct target of miR19c by Luciferase assay. Taken together, these results demonstrate that miR-19c is a positive gene regulator of lymphatic development and targets tgfβ1a to reduce anti-lymphatic signals during development.