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作者(中文):王丹延
作者(外文):Wang, Dan-Yen
論文名稱(中文):在飲食限制或高油脂飲食的條件下纖維母細胞生長因子第一型在小鼠腦內的表現
論文名稱(外文):Spatial expression of FGF1 in the mouse brain under the condition of food restriction or high-fat diet
指導教授(中文):陳令儀
邱英明
指導教授(外文):Chen, Linyi
Chiu, Ing-Ming
口試委員(中文):紀雅惠
許益超
口試委員(外文):Chi, Ya-Hui
Hsu, Yi-Chao
學位類別:碩士
校院名稱:國立清華大學
系所名稱:分子醫學研究所
學號:100080605
出版年(民國):102
畢業學年度:102
語文別:英文
論文頁數:95
中文關鍵詞:纖維母細胞生長因子第一型(FGF1)腦室旁神經核(PVN)oxytocinnesfatin-1飲食調控
外文關鍵詞:FGF1PVNoxytocinnesfatin-1diet regulation
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摘要
纖維母細胞生長因子(Fibroblast growth factors, FGFs)在胚胎的發育以及成年時期的細胞增生控制、分化和遷移都扮演著重要的角色。纖維母細胞生長因子第一型(FGF1)是FGFs的其中一員,可以促進細胞生長、增生和神經的新生。為了研究在小鼠腦內內源性FGF1的效應,我們製造了三隻FGF1抗體且分別命名為FGF1#14、FGF1#15和FGF1#16,這三隻FGF1抗體是由打入特定的老鼠FGF1抗原至兔子體內所產生的。使用西方墨點法和石蠟切片免疫組織化學染色法分析,顯示出FGF1#14、FGF1#15和FGF1#16A這三隻抗體都能辨認出FGF1的重組蛋白以及小鼠腦內內源性FGF1。特別是FGF1#16A這隻抗體可以應用在西方墨點法、石蠟切片和震盪式切片的免疫組織化學法染色等方面。此外免疫吸附法分析的結果指出FGF1#16A可以專一性地偵測到FGF1。在免疫組織化學染色的結果中,我們發現有FGF1#16A免疫反應的細胞會表現在下視丘的腦室旁神經核(paraventricular nucleus, PVN),這個位置在腦中是調控能量動態平衡的重要區域。有趣的是,我們也發現到FGF1#16A免疫反應的細胞會與先前研究已指出抑制食慾的標記蛋白oxytocin和nesfatin-1表現在同一顆細胞中。因此我們準備了處於規律飲食、禁食、禁食再餵食和餵食高油脂飼料誘導肥胖等老鼠的腦片進行FGF1#16A/oxytocin和FGF1#16A/nesfatin-1的免疫組織化學染色來測定FGF1對於飲食調控的效應。我們的結果顯示禁食老鼠的FGF1#16A免疫反應細胞數量會在前囟門(bregma) -0.22 mm和-0.46 mm處增加,另外餵食高油脂飼料的肥胖老鼠FGF1#16A免疫反應細胞量在bregma -0.46 mm也有增加的現象。在之前發表的研究已有指出藉由顱內注射FGF1可以調控飲食的行為。另外也有研究說明經由高油脂飲食會使脂肪組織中的FGF1表現量提高。我們的實驗結果藉由計算特定bregma的FGF1#16A免疫反應的細胞數量顯示出在禁食老鼠和高油脂餵食的肥胖老鼠表現量有增加的現象。所以我們的實驗結果和FGF1在飲食調控上可能有關的主張是一致的。

關鍵詞:纖維母細胞生長因子第一型(FGF1)、腦室旁神經核(PVN)、oxytocin、nesfatin-1和飲食調控。
Abstract
Fibroblast growth factors (FGFs) play important roles in embryonic development and in adult life by controlling cell proliferation, differentiation and migration. FGF1 is one member of the FGF family, which promotes cell growth, proliferation and neurogenesis. To study the effect of endogenous FGF1 in the mouse brain, we produced FGF1 antibodies, named FGF1#14, FGF1#15 and FGF1#16, that are raised in the rabbits against specific mouse FGF1 polypeptides. We showed that FGF1#14, FGF1#15 and FGF1#16A recognized recombinant mouse FGF1 protein using Western blotting analysis. These antibodies could also recognize endogenous FGF1 in the mouse brain using immunohistochemistry-paraffin (IHC-P) analysis. Particularly, FGF1#16A could be used to apply to Western blot, IHC-P and immunohistochemistry (IHC). In addition, immunoabsorption assay results indicated that FGF1#16 were specific to detect FGF1. Among these results, we found that FGF1#16A immunoreactive cells were present in the paraventricular nucleus (PVN) of the hypothalamus, which is an important brain region in regulating energy homeostasis. Interestingly, we also found FGF1#16A immunoreactive cells could express both oxytocin and nesfatin-1. Both proteins are anorexigenic markers in several studies. We prepared mice that were maintained under regular diet, fasted, refed or high-fat diet (HFD) for double labeling with FGF1#16A/oxytocin or FGF1#16A/nesfatin-1 to determine the effect of FGF1 in diet regulation. Our results showed that FGF1#16A immunoreactive cells were higher in fasted mice at bregma -0.22 mm and -0.46 mm and in HFD mice at bregma -0.46 mm. Previous studies have indicated that FGF1 regulates feeding behavior through intracerebroventricular injection. It has also been reported that FGF1 is highly induced in adipose tissue in response to HFD. In conclusion, our results revealed that FGF1 is increased in the fasted mice and in HFD mice at specific bregma regions by counting immunoreactive cells. Thus, our results are consistent with the notion that FGF1 may have an effect in diet regulation.

Key words: FGF1, PVN, oxytocin, nesfatin-1 and diet regulation



Table of Contents
摘要.................................................................................................................................i
Abstract......................................................................................................iii
誌謝............................................................................................................................v
Table of contents...........................................................................................................vi
Abbreviations..............................................................................................vii
Introduction...................................................................................................1
Material and Methods...................................................................................10
Results.........................................................................................................................18
Discussion....................................................................................................................26
References....................................................................................................................34
Tables............................................................................................................................44
Figures..........................................................................................................................50

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