- 學位論文
缺鋅飼料對大鼠小腸食慾控制與全基因表現之影響
Zinc deficient diet affects intestinal gene expression including appetite-regulating genes in rat
摘要
鋅為生物體必需微量營養素，是構成轉錄因子之必要組成而調節基因表現。鋅缺乏時以食慾不振為最早出現的症狀，而已知腸道也會參與食慾的調控。因此本實驗針對缺鋅大鼠，利用高通量（high-throughput）DNA微陣列分析技術，以全面性分析缺鋅對小腸基因表現的影響，並嘗試發現缺鋅影響食慾的腸道因子。實驗動物為Wistar品系公鼠，分控制組、缺鋅組、中度缺鋅組及缺鋅對飼育組；中度缺鋅組於第19天改餵食缺鋅飼料（MD）及正常飼料（MPF）5天。各組於餵養24天後犧牲，然後收取小腸組織，萃取小腸黏膜total RNA，首先以pooled樣品進行DNA微陣列實驗分析，並以Q-PCR驗證有差異之特定基因。缺鋅組之體重、攝食量、飼料利用率、肝臟及血漿鋅含量都顯著低於控制組；對飼育組之飼料利用率、肝臟及血漿鋅濃度則高於缺鋅組。由微陣列分析結果顯示，與控制組比較，缺鋅組之基因表現升為2倍以上的基因有90個，表現降為50％以下之基因有84個，總有174個表現差異基因；對飼育組之表現升為2倍以上的基因有47個，表現降為50％以下之基因有29個，共76個表現差異基因。比較缺鋅與對飼育組可見，缺鋅改變之基因數較限食多，兩組只有20個基因表現是變化相同，其餘皆不同，表示缺鋅對基因的影響具有獨特性，並非透過限食作用。缺鋅組之metallothionein與ZnT-2基因表現減少，只有控制組的0.28倍與0.41倍，而neuromedin U則增加，為控制組的2.76倍；對飼育組則沒有這些變化。以Q-PCR進行大鼠個別驗證可見，缺鋅鼠之neuromedin U基因表現與缺鋅時攝食波動趨勢相關，已知neuromedin U在腦部有抑制食慾的作用，故推測neuromedin U與缺鋅對於食慾的影響有關。本研究首度發現鋅營養狀態可改變neuromedin U之基因表現，有助於探討缺鋅對食慾的影響。
並列摘要
Zinc is an indispensable metal nutrient and involved in regulation of gene expression since it is an essential component of the zinc finger structure of many transcription factors. One of the early zinc deficiency symptoms is depressed food intake, which can be regulated by both central nervous and peripheral neuroendocrine systems including the gastrointestinal tract. Therefore, we used a high-throughput, whole genome rat DNA microarray assay to analyze the gene expression profile of the small intestinal in zinc deficient rat, and tried to identify novel intestinal factor for appetite control by zinc. Twenty Wistar male rats were divided into four groups: a zinc-deficient group (ZnD, zinc < 2 ppm), a pair-fed group (PZnD, zinc = 50 ppm), a marginal zinc-deficient group (MZnD, zinc = 5 ppm), and a zinc-adequate group (ZnA, zinc = 50 ppm). All groups were fed with corresponding diet for 24 days except the MZnD, in which 3 rats were changed to the zinc deficient diet and two to the pair-fed diet for the last 5 five days. Total RNA of small intestinal mucosa was extracted from each rat, pooled RNA samples corresponding to ZnD, PZnD and ZnA were subjected to Agilent Rat V2 Oligo Microarray assay. Zn deficient rats had significantly lower body weight, food intake, feed efficiency and zinc content in liver and plasma. Between ZnD and ZnA groups, a total of 174 genes were differentially expressed, with 90 genes one-fold higher and 84 genes 50% lower in the ZnD group; whereas between PZnD and ZnA groups, a total of 76 genes were differentially expressed, with 47 genes one-fold higher and 29 genes 50% lower in the PZnD group. Between ZnD and PZnD groups, only 20 genes were similarly changed, indicating that zinc deficiency exerts a specific effect other than food restriction on intestinal gene expression. In addition to metallothionein and zinc transporter ZnT-2, neuromedin U expression (NmU) was specifically altered by zinc deficiency and increased to 2.76-fold. NmU expression of individual rat was measured with quantitative RT-PCR and showed an association with zinc status that NmU expression can be linked to the cyclical pattern of food intake in the ZnD group. In conclusion, we first report a novel relationship between zinc nutrition and intestinal neuromedin U expression, which may help to elucidate the appetite regulation modulated by zinc deficiency.
參考文獻
延伸閱讀
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