2011-01-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/649457摘要：我們將在今年和國衛院楊良棟博士合作，利用他的Tgfb3-Cre小鼠來製造條件式破壞肺臟上皮Notch訊息的基因剔除鼠。我們先前發現若在肺臟發育的初期(E9.5)利用Shh-Cre的小鼠將肺臟初始上皮細胞的Notch訊息給破壞掉，那呼吸道上皮的Clara cell就無法形成，相對的整個呼吸道上皮就佈滿了纖毛細胞，這代表在肺臟發育過程中，Notch訊息是用來調控Clara cell 和纖毛細胞之間的平衡(Tsao et al. Development 2009; 136, 2297-2307)。可是，出生後肺臟上皮的Notch訊息到底扮演什麼角色則不清楚。最近我們利用Rosalacz小鼠來偵測Tgfb3-Cre的組織獨特性，我們發現Tgfb3-cre 的Cre蛋白表現在胚胎早期(E14.5,此時 Clara cell及纖毛細胞的分化正在進行)的肺臟上皮是呈現mosaic的型式，只有少數的肺臟上皮有表現cre，但是在成鼠大部份的呼道上皮都有表現cre。因此，我們可以利用Tgfb3-cre小鼠製造出肺臟呼吸道上皮(含Clare cell)形成之才去破壞Notch 訊息的基因剔除鼠。我們目前已成功的製造出條件型Pofutl基因剔除鼠(PofutlcTb3) ，這種突變鼠可以存活，但是它呼吸道的Clara cell數目會減少，同時呼吸道的Goblet細胞會過度增生。這種分泌黏液細胞過度增生(goblet cell metaplasia)的現象，十分類似慢性阻塞性肺疾病及氣喘病人的病理變化。國衛院已經同意我們變更原先之計畫，改為研究此基因突變鼠。這個有趣的發現，不只可以提供一個新的類似慢性阻塞性肺疾病及氣喘的動物模式，更可已利用研究此條件型 Pofut1 基因剔除鼠，來了解其致病機轉，並進而尋求新的治療方式，此部份預計會在這一年度會完成。<br> Abstract: we will cooperate with Dr. Liang-Tung Yang (NHRI) by using his Tgfb3-Cre deleter mouse line to inactive Notch signaling in airway epithelium. Previously, by using Shh-Cre to delete Pofut1 in very early lung progenitors before lung buds formation (E9), we successfully created a conditional Notch knockout mouse which had overpopulated ciliated cells and no Clara cells in the airways. This indicates that Notch signaling is required for the balance of ciliated and secretory cell fate in the developing airways (Tsao et al. Development 2009; 136, 2297-2307). However, the role of Notch signaling in postnatal airway epithelium is unknown. In our preliminary data, we found the Tgfbcre Rosalacz reporter showed a mosaic pattern at early embryonic stage (E14.5, the timing which airway epithelium starts to differentiate into Clara cell and ciliated cell), however, in the adult, signals were seen in the majority of the airway epithelial cells. This suggested that using tgfb3-cre deleter we could to inactive Notch signaling at later stages, presumably after Clara cell formation. We have created Pofut1 conditional knockout mouse (Pofut1cTb3) by crossing the Tgfb3-Cre deleter line and Pofut1 folx/flox mouse line successfully. The Pofut1cTb3 mutant mice were viable, but decreased Clara cell number and dramatic goblet cell metaplasia. This phenotype mimics human asthma or chronic obstruction pulmonary disease (COPD). Due to this striking phenotype, we have modified this year project to analyze this Pofut1cTb3 mouse carefully to elucidate the possible mechanism on the next year. This modified project has been accepted by NHRI already. Hope this mutant can be used as a novel animal model of airway goblet cell metaplasia which is a hall marker of asthma or COPD in the future.肺臟幹細胞呼吸道上皮lungstem cellairway epithelium