【作者】 王玉峰；

【导师】 赵茹茜；

【作者基本信息】 南京农业大学 ， 基础兽医学， 2009， 硕士

【摘要】 FTO基因(fat mass and obesity-associated gene)是人类发现的第一个与非病理性肥胖相关的基因。人们对哺乳动物FTO基因的研究已经取得了一定的进展,但鸡FTO的研究目前仍属空白。本试验分析了鸡FTO基因的组织特异性表达以及日龄和品种的影响,并检测了FTO mRNA在鸡下丘脑和丘脑的分布以及下丘脑室旁核和腹内侧核FTO mRNA丰度表达的品种差异和进食的影响,为鸡FTO功能的进一步研究提供基础资料。1鸡FTO基因的组织特异性表达以及日龄和品种的影响本研究选择1周龄及19周龄的雄性罗斯(Ross)肉鸡和莱航(Leghorn)蛋鸡,采集下丘脑、端脑、小脑、肌肉、肝脏、内脏脂肪六种主要组织器官并采用实时定量荧光PCR方法检测FTO mRNA在不同组织器官中的表达。结果显示：FTO在所检测的六个组织器官中均有不同程度表达。其中,FTO在鸡的肝脏、下丘脑、内脏脂肪和小脑有较高丰度的表达,而肌肉和端脑表达丰度相对低。总的来说,周龄及品种对FTOmRNA的组织分布模式影响不大,但有些组织存在显著的周龄和品种差异。1周龄肉鸡肝脏中FTO的mRNA表达量极显著高于同周龄蛋鸡(P<0.01)；而内脏脂肪和小脑的FTO mRNA表达量则显著地低于蛋鸡(P<0.05)。19周龄蛋鸡肝脏和内脏脂肪中FTO的表达量均极显著高于1周龄蛋鸡(P<0.01)；19周龄肉鸡下丘脑FTO的mRNA表达水平显著高于1周龄肉鸡(P<0.05),小脑中FTO的mRNA表达量则极显著高于1周龄肉鸡(P<0.01)。2 FTO在鸡-下丘脑及丘脑的表达分布本研究采用原位杂交的方法检测FTO在鸡下丘脑和丘脑的表达分布情况。原位杂交结果显示：FTO基因在下丘脑调节能量平衡的室旁核(PVN, paraventricular nucleus)、腹内侧核(VMN, ventromedial nucleus)中有表达。FTO在丘脑与视觉、采食和生殖节律有关的豆状袢前核(ALA, ansae lenticularis anterior)、丘脑前背外侧核(DLA, nucleus dorsolateralis anterior)、枕中脑束(OM, Tractus occipitomesenphalicus)、外细胞层(SCE, stratum cellulare externum)、达克谢维奇(氏)核(ND, nucleus of Darkschewitsch)、中间核(nI, nucleus intramedialis)均有表达。3禁食及品种差异对FTO在室旁核和腹内侧核表达的影响选择1日龄雄性罗斯(Ross)肉鸡和莱航(Leghorn)蛋鸡,分别分为对照和24小时禁食组。限饲处理从第6日龄开始,7日龄时屠宰采集整脑,打孔(punch)收集下丘脑室旁核和腹内侧核,提取总RNA采用实时定量荧光PCR方法检测FTO mRNA在这些核团中的相对表达量。结果显示：肉鸡室旁核内FTO表达显著高于蛋鸡(P<0.05),而腹内侧核内FTO表达未表现品种差异。肉鸡下丘脑室旁核和腹内侧核中FTO基因表达均不受禁食的影响；而蛋鸡24小时禁食后,腹内侧核FTO基因表达量显著下降(P<0.05),室旁核FTO基因表达量没有显著变化。更多还原

【Abstract】 FTO gene (fat mass and obesity-associated gene) is the first gene we have found contributing to common forms of human obesity. Studies in mammalian have made certain progress, but chicken FTO study is still vacant. In present study, we tested FTO gene tissue-specific expresssion, especially chickens with different ages and strains. FTO mRNA distribution pattern in chicken hypothalamus and thalamus was examined, as well the PVN and VMN FTO expression comparison between different food intake status and breeds.1 FTO gene tissue specific expression in different ages and breeds of chickenHypothalamus, forebrain, cerebellum, muscle, liver and visceral fat were sampled from 1 week and 19 weeks old male Ross and Leghorn chicken. FTO gene expression in these tissue samples was tested with Real-time PCR. Our results showed that FTO gene was highly expressed in liver, hypothalamus, visceral fat and cerebellum. Although factors of age and strain did not affect the FTO expression pattern in different tissues very much, there are still some breed-specific and age-specific expressions in certain tissues.In breeds comparison, the FTO gene expression in 1 week old broiler liver is severely higher than 1 week layers (P<0.01); FTO expression in 1 week broiler visceral fat and cerebellum is significantly lower compared with 1 week layers (P<0.05). In ages comparison, FTO expression level in 19 weeks old layer liver and visceral fat were markedly higher than 1 week layers (P<0.01); FTO expression level in 19 weeks old broilers hypothalamus was higher than lweek layers(P<0.05), and in cerebellum was profoundly higher than 1 week layers (P<0.01).2 FTO mRNA distributions in hypothalamus and thalamusIn this study we examined the FTO mRNA distribution pattern in hypothalamus and thalamus with in situ hybridization (ISH). Our ISH result shows that FTO gene expressed in hypothalamic PVN (paraventricular nucleus) and VMN (ventromedial nucleus), and in thylamic DLA (nucleus dorsolateralis anterior), ALA (ansae lenticularis anterior), OM (Tractus occipitomesenphalicus), SCE (stratum cellulare externum), D (nucleus of Darkschewitsch), nl (nucleus intramedialis). These nucleuses relate to food intake, visual and reproductive rhythm function.3 Effects of fasting and breed on PVN and VMN FTO expression1 week old male Ross and Leghorn chicken were divided into broiler 24 hours fasting group, broiler control, layer 24 hours fasting group and layer control. Nucleus of PVN and VMN were collected from hypothalamus by punch, and FTO mRNA levels were determined by Real-time PCR. Our result shows FTO breed specific expression in chicken PVN, that FTO expression in 1 week old broiler PVN is significantly higher than 1 week layers (P<0.05); no breed specific expression was detected in chicken VMN.24 hours fasting didn’t affect both hypothalamic PVN and VMN FTO expression in 1 week male broilers. After the feed restriction, FTO expression level in VMN was significantly reduced (P<0.05), while no difference was found in PVN.更多还原