普洱茶抑制SREBP和SCD降低秀丽线虫脂肪储存的研究
文献类型：学位论文
| 作者 | 丁毅弘 |
| 学位类别 | 硕士 |
| 答辩日期 | 2014-11 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 梁斌 |
| 关键词 | 普洱茶 秀丽隐杆线虫 脂肪储存 SREBP 硬脂酰去饱和酶（SCD） |
| 其他题名 | Pu-erh tea down-regulates SREBP and SCD to reduce fat storage in Caenorhaditis elegans |
| 学位专业 | 生物工程 |
| 中文摘要 | 随着经济的发展，人口饮食结构和生活方式的改变，肥胖症在全球的发生率呈快速上升的趋势。肥胖不仅影响到体态和美观，而且还和糖尿病、心脑血管疾病、高血压、非酒精性脂肪肝等重大代谢性疾病密切相关。普洱茶传统上一直被认为具有良好的减肥、降脂、防止心血管疾病、降血糖等保健功效，但是目前对普洱茶减肥降脂机理的研究却相对较少。秀丽隐杆线虫中许多脂代谢相关基因都很保守，且培养方便，易于检测，因此本文以秀丽线虫为研究对象，从普洱茶的减肥功效以及普洱茶可能作用的靶基因和信号通路等方面对普洱茶减肥机理进行了深入的探讨。 本文首先通过尼罗红固定染色的方法检测了不同浓度普洱茶水提液处理后线虫脂肪储存的变化，结果发现经过普洱茶水提液处理后，线虫脂滴变小，密度降低，脂肪含量下降。通过薄层层析和气相色谱对普洱茶处理后线虫的脂肪定量分析后发现，与对照组相比，普洱茶水提液(0.05g/ml)使线虫TAG/Total lipid比值明显下降(5%)。以上结果表明，普洱茶确实具有良好的减肥效果。于是我们进一步分析了普洱茶水提液处理后线虫脂肪酸含量的变化，结果发现普洱茶使线虫的硬脂酸(18:0)含量上升，油酸(18:1n9)的含量下降，这一结果暗示普洱茶可能是通过作用于线虫SCD来减肥的，同时鉴于SREBP是调控SCD的重要转录因子，因此，我们通过以下实验来验证普洱茶对线虫SCD和SREBP的作用。我们首先用普洱茶水提液处理FAT-6::GFP, FAT-7::GFP, SBP-1::GFP转基因线虫后观察其GFP信号表达的变化，其次通过实时定量PCR的方法检测普洱茶水提取液处理后线虫SCD及SREBP基因转录水平的变化，最后用尼罗红固定染色及薄层层析和气相色谱的方法对普洱茶水提取液处理后线虫SCD及SREBP功能缺失突变体脂肪储存的变化进行了检测，结果证实了普洱茶的确能在蛋白水平和mRNA水平上显著降低线虫SCD和SREBP的表达，同时普洱茶处理过后，线虫SCD及SREBP功能缺失突变体脂肪含量的变化并不显著，因此证明SREBP和SCD确实是普洱茶减肥的作用靶点。另外，我们发现普洱茶水提取物使线虫摄食速率下降，同时我们还对线虫饲喂带有GFP信号的大肠杆菌，然后观察普洱茶水提取液处理后该线虫体内GFP信号的变化，从而间接反映出普洱茶水提取液处理后线虫体内食物吸收，消化，排泄的情况，结果表明普洱茶能抑制线虫对食物的摄取。最后我们还对不同浓度普洱茶水提取液对线虫的生长发育，后代数目，寿命产生的影响也进行了评估。 |
| 英文摘要 | With economic development, changes in diet and lifestyle of the population, the incidence of obesity in the world showed an upward tendency. Obesity not only affects the posture and beauty, but also closely related to diabetes, cardiovascular disease, hypertension, non-alcoholic fatty liver disease and other major metabolic diseases. Pu-erh tea has traditionally been considered to have good health effects such as weight cutting, lipid-lowering, prevent cardiovascular disease, reducing blood sugar and so on, but the researches about the mechanism of Pu-erh tea weight-cutting effect are relatively scarce. Many of the lipid metabolism related genes are conservative in Caenorhabditis elegans, moreover Caenorhabditis elegans easy to be cultured and detected, so in this article we use Caenorhabditis elegans as a model to have a in-depth discuss in the mechanism of Pu-erh tea weight-cutting effect from the anti-obesity efficacy, possible target genes and signaling pathways of Pu-erh tea and so on. Firstly, by Nile Red staining of fixed worms, we detected the change of fat storage in different concentrations of Pu-erh tea water extract treated worms, the results show that after Pu-erh tea water extract treated, the worms’ fat content was decreased, and lipid droplet becomes smaller, the density was reduced. By thin layer chromatography and gas chromatography quantitative analysis the fat content of Pu-erh tea treated worms, we found that, compared with the control group, the Pu-erh tea water extract (0.05g/ml) make worms TAG / Total lipid ratio significantly decreased (5%) . These results suggest that tea indeed have a good weight-cutting effects. So we further analyzed the composition of fatty acid of worms after treatment of Pu-erh tea water extract, and found that Pu-erh tea increased the level of stearic acid (C18:0), while decreased the level of oleic acid (C18:1n9), this result suggests that Pu-erh tea is probably by acting on SCD to reduce fat, and in view of SREBP is an important transcription factor for regulate SCD, therefore, we used the following experiments to verify the role of Pu-erh tea act on SCD and SREBP. First we treated fat-6::GFP, fat-7::GFP, sbp-1::GFP transgenic worms with different concentrations of Pu-erh tea water extract and test the change of GFP signal in these transgenic worms, then we tested the transcription level of SCD and SREBP in different concentration of Pu-erh tea water extract treated worms by real-time quantitative PCR, at last we detected the fat content change in SCD and SREBP deficient worm mutant after different concentration of Pu-erh tea water extract treated by Nile Red staining of fixed worms and thin layer chromatography and gas chromatography quantification. the results confirmed that the tea can indeed significantly reduce worms’ SCD and SREBP expression both in the protein level and the mRNA level, while the fat content of SCD and SREBP deficient worm mutant didn’t obviously change after Pu-erh tea water extract treated. Thus the results demonstrating SREBP and SCD indeed are the target of Pu-erh tea to cut weight. In addition, we found that the Pu-erh tea water extract decreased the worm pumping rate, and we also fed E. coli transformed with GFP to the worms, and then test the changes of GFP signal distributed in the worm body after Pu-erh tea water extract treated, then food absorption, digestion, excretion of Pu-erh tea water extract treated worms were indirectly reflects through the change of GFP signal. The results show that tea can inhibit food intake among C.elegans. Finally, we also assessed the effects of different concentrations of tea water extract on growth、offspring and lifespan. |
| 语种 | 中文 |
| 源URL | [http://159.226.149.26:8080/handle/152453/10196]  |
| 专题 | 昆明动物研究所_脂类代谢与疾病 |
| 推荐引用方式 GB/T 7714 | 丁毅弘. 普洱茶抑制SREBP和SCD降低秀丽线虫脂肪储存的研究[D]. 北京. 中国科学院研究生院. 2014. |
入库方式： OAI收割
来源：昆明动物研究所
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