臺灣博碩士論文加值系統

人體內主要有兩種類型的脂肪細胞，分別為白色脂肪細胞及棕色脂肪細胞，前者主司將能量以三酸甘油酯的形式儲存在體內；後者則是將能量轉換成熱能釋出。近來研究指出，增加白色脂肪組織中類棕色脂肪細胞(brite cells)可促進生熱對抗肥胖。Fibroblast growth factor 21 (FGF21)表現受Peroxisome proliferator-activated receptor alpha (PPARα)調控，為hepatokines之一，已知參與啟動生熱基因表現和白色脂肪褐化。本實驗室先前研究在懷孕母鼠餵食氧化炸油活化PPARα，發現子代成年後雄鼠不易胖，伴隨白色脂肪生熱基因表現增加，追溯胚胎期(E18天)可見肝臟FGF21 mRNA量增加，因此我們提出假說: 懷孕期間給予PPARα活化刺激可藉由提高胚胎肝臟FGF21分泌而影響日後脂肪組織生熱能力及肥胖敏感性。將懷孕C57BL/6J mice分成兩組，自懷孕第0天到第18天(E0-E18)分別給予control diet (Ctrl組)或含0.5% clofibrate diet (CF組)，仔鼠出生後母親統一給予control diet，斷乳仔鼠餵食chow diet 至7週齡，轉高脂飲食5週以誘發肥胖，觀察:(1) 母鼠孕期給予 Clofibrate 是否活化胚胎肝臟PPARα，並促使胚胎表達FGF21 (2) 胚胎肝臟大量表現FGF21對新生仔鼠脂肪組織發育影響 (3) 胚胎肝臟大量表現FGF21對成年後脂肪組織生熱及Diet-induced obesity的敏感性。結果顯示，懷孕全期給予clofibrate可藉由母親及胚胎肝臟PPARα活化，成功誘導E18胚胎肝臟FGF21表現及母體血液FGF21濃度，但新生小鼠出生後7天量測肝臟與血液FGF21已回復與Ctrl相當。成年後歷經DIO，由脂肪組織重量得知CF組雄性子代不易胖，而雌性子代易胖。與各自Ctrl littermate相較，CF組雄性子代鼠蹊白色脂肪中有較高的Uncoupling protein 1 (UCP-1)mRNA與蛋白質量及brite cell的標誌基因表現，相對CF組雌性子代brite cell的標誌基因表現降低。結論: 懷孕期間給予母鼠clofibrate，可大幅提高胚胎肝臟FGF21表現，可能影響成年後白色脂肪組織的生熱或褐化能力，但有性別差異。

There are two types of adipose tissue in mammals, white and brown. White adipose tissue stores excess energy in the form of triglycerides. Conversely, brown adipose tissue dissipates chemical energy in the form of heat through the action of mitochondrial uncoupling protein 1 (UCP-1). Recent study showed, there’s one kind of adipocyte emerge within white adipose tissue, which is called brown-fat like adipocyte (brite adipocyte) or beige adipocyte. Brite adipocytes display multilocular morphology and express brown-adipocyte-specific UCP-1. The development of these brite cells in WAT is dramatically enhanced during adaptation to cold or in response to treatment with β3-selective adrenergic agonists. According to Badman’s study, FGF21 is a target gene of Peroxisome proliferator-activated receptor alpha (PPAR-α) and FGF21 might play an important role for thermogenesis in adipocytes. Our previous study found that when oxidized frying oil (OFO), a dietary PPARα agonist, was fed to pregnant mothers, their male offsprings were resistance to diet-induced obesity (DIO), accompanied with an increased expression of thermogenic genes in adipose tissue. Therefore, we proposed a hypothesis: PPARα activation during pregnancy, through upregulated FGF21, might increase thermogenic activity of adipose tissue of offsprings and reduce their propensity to DIO. In our experiment, dams were divided into two groups to receive a control diet (Ctrl group) or 0.5% clofibrate diet (CF group) during the whole gestational period. The aims of this study were to investigate whether: 1) dams receiving CF diet during gestational period could activate PPARα, thus increasing FGF21 expression in fetus liver. 2) Impact of upregulated FGF21 expression in fetus liver on the development of newly form adipose tissue. 3) Impact of upregulated FGF21 expression in fetus liver on thermogenic ability of adipose tissue and the propensity of DIO at adulthood. Our results showed that the mRNA levels of PPARα target genes were significantly increased in livers of dams and their fetus of the CF group. Clofibrate administration successfully increased mRNA levels of FGF21 in liver of fetus and its blood levels in dams at pregnancy d18. However, the hepatic mRNA levels and blood levels of FGF21 recovered at postnatal d7 neonates. After challenging with a high-fat diet at adulthood, the fat mass showed the male offsprings from the CF group resisted to DIO, while the females offsprings from the CF group were prone to be obese. Compared with their Ctrl littermates, the CF male offsprings had significantly higher mRNA and protein levels of uncoupling protein-1 (UCP-1) and mRNA levels of brite cell markers (CD137、Tmem26 and Tbx1) in inguinal WAT. In contrast, the female offsprings of CF group showed the expression levels of brite cell markers were significantly lowered. We conclude that uteral PPARα activation caused by maternal clofibrate administration leads to a greater expression level of FGF21 in fetus liver, which might determine the browning capacity of WAT during adulthood, an effect with gender difference.

目錄 i
圖目錄 iii
表目錄 iv
中文摘要 I
Abstract III
第一章 前言 1
第二章 文獻回顧 3
一、肥胖(obesity) 3
(一) 肥胖的定義 3
(二)肥胖盛行率 4
二、脂肪組織(Adipose tissue) 5
(一)棕色脂肪組織(Brown adipose tissue) 5
(二)白色脂肪組織(white adipose tissue) 7
(三)類棕色脂肪細胞(brown-like adipocyte) 8
三、脂肪組織的生熱(褐化)反應 9
(一)褐化過程 9
(二)影響因素 10
(三)生熱基因&類棕色脂肪細胞的標誌基因 11
四、PPARα & FGF21 13
(一)PPARα (Peroxisome proliferator-activated receptor alpha) 13
(二)FGF21 (Fibroblast growth factor 21) 14
第三章 材料與方法 17
一、實驗設計與假說 17
二、試驗飼料配製 20
三、動物飼養 22
四、檢體收集 23
五、血清脂質分析 24
六、血清FGF21分析 26
七、抽取RNA及cDNA的製備 28
八、Real time polymerase chain reaction(同步定量PCR ; qRT-PCR) 35
九、西方墨點法 (Western blotting) 40
第四章 結果 46
一、孕期給予clofibrate對母體及胚胎PPARα活化情形及FGF21生成之影響 46
(一) Clofibrate對懷孕母鼠體重和攝食之影響 46
(二) Clofibrate對懷孕母鼠及胚胎肝臟PPARα活化之影響 46
(三) 孕期攝取clofibrate對懷孕d18母鼠、胚胎肝臟及出生後第七天仔鼠之肝臟FGF21表現之影響 47
(四) Clofibrate對胚胎子宮內及出生後(斷乳前)血清FGF21濃度之影響 47
(五) Clofibrate對胚胎棕色脂肪組織生熱基因表現之影響 48
二、孕期給予clofibrate對子代成年後肥胖敏感性之影響 54
(一) Clofibrate對子代生長與攝食之影響 54
(二) Clofibrate對成年子代肥胖敏感性之影響 54
(三) Clofibrate對成年子代血脂之影響 55
(四) Clofibrate對成年子代肝臟PPARα活化效應之影響 55
(五) Clofibrate對成年子代白色脂肪組織生熱基因表現之影響 56
(六) Clofibrate對成年子代白色脂肪組織生熱蛋白之影響 56
第五章 討論 65
第六章 結論 71
第七章 參考文獻 72

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