小鼠肠道菌群构成与肥胖易发性的研究
发布时间：2018-07-23 17:47
【摘要】：人体肠道中庞大的微生物群落与宿主互惠共生,在机体能量的获取、脂肪代谢的调控以及慢性炎症的发生等方面起重要作用,与肥胖和代谢综合征的发生发展密切相关。小鼠摄取高脂膳食可产生肥胖抵抗(DIO-R)和肥胖易感(DIO-P)两种表型,探究其肠道菌群有利于阐明肥胖产生机制,对提出干预肥胖的营养手段具有重要意义。将C57BL/6小鼠随机分为对照组(标准饲料饲养,NC)和高脂饲料组,7周后将高脂组分为肥胖抵抗组(DIO-R)和肥胖易感组(DIO-P)。在实验起始(0周)、肥胖发展的早期(7周)、中期(17周)和末期(27周),利用末端限制性片段长度多态性(T-RFLP)和定量PCR检测粪便菌群结构,并测定短链脂肪酸含量;在7、17和27周分别处死部分小鼠,分析血液脂多糖(LPS)和炎性因子水平,检测回肠上皮TLR4、GPR41、Fiaf、Zo-1、Occludin和Muc2基因的表达情况。结果显示:DIO-P组7周后体重显著高于另两组(P0.05),DIO-R组体重在中后期逐渐高于NC组(P0.05);DIO-P组肠道菌群与NC组和DIO-R组差异明显,多样性和稳定性下降,同时乳杆菌(Lactobacillus)、双歧杆菌(Bifidobacterium)、Akkermansia muciniphila丰度下降,而脱硫弧菌(Desulfovibrio)、金黄色葡萄球菌(Staphylococcus aureus)、阴沟肠杆菌(Enterobacter cloacae)丰度上升;DIO-R组这几种菌的含量在早期变化不明显,中后期逐渐趋向DIO-P组特征。试验起始时DIO-R组和DIO-P组小鼠肠道菌群构成存在显著差异,与肥胖发生具有显著相关性。与NC组相比,DIO-P组粪便乙酸、丁酸含量在7周后显著降低,而丙酸显著升高(P0.05),在前期与DIO-R组差异显著(P0.05),而后期无显著差异。DIO-P组血液LPS在7周后显著高于NC组(P0.05),在7周和17周时显著高于DIO-R组(P0.05)。回肠TLR4(LPS受体)和GPR41(短链脂肪酸的受体)基因表达在DIO-P组显著高于DIO-R组(P0.05),说明肠道可能出现炎性反应和短链脂肪酸代谢异常;Fiaf介导信号参与脂肪分解、抑制脂肪合成,DIO-P组该基因表达显著低于DIO-R组(P0.05)。Zo-1、Occludin和Muc2三个基因与肠粘膜屏障完整性有关,DIO-P组这三个基因的表达显著低于DIO-R组(P0.05),说明肥胖易感组的肠道屏障可能受损。ELISA分析显示DIO-P组小鼠血液促炎性细胞因子显著上升(P0.05),而抗炎性细胞因子含量显著下降(P0.05),说明发生了慢性系统性炎症。总之,本研究说明小鼠在高脂膳食下肥胖的易发性与肠道菌群有密切的联系,肠道菌群影响肥胖易感的机制可能与损伤肠黏膜屏障、诱发炎性反应和扰乱脂代谢有关。
[Abstract]:The large microbial community in the human intestine is symbiotic with the host, which plays an important role in the acquisition of body energy, the regulation of fat metabolism and the occurrence of chronic inflammation, which is closely related to the occurrence and development of obesity and metabolic syndrome. Two phenotypes of obesity resistance (DIO-R) and obesity susceptibility (DIO-P) can be produced in mice fed with high fat diet. It is important to explore the intestinal flora of mice to elucidate the mechanism of obesity and to propose nutritional intervention for obesity. C57BL/6 mice were randomly divided into control group (standard diet feeding NC) and high-fat diet group after 7 weeks. The high-fat group was divided into obese resistance group (DIO-R) and obesity susceptible group (DIO-P). At the beginning of the experiment (0 weeks), the early stage of obesity (7 weeks), the middle stage (17 weeks) and the end stage (27 weeks), the fecal microflora structure was detected by terminal restriction fragment length polymorphism (T-RFLP) and quantitative PCR, and the content of short chain fatty acids was measured. The blood lipopolysaccharide (LPS) and inflammatory cytokines were analyzed, and the expression of TLR4GPR41 Fiafo Zo-1Occludin and Muc2 genes in ileum epithelium were detected at 7 ~ (th) ~ (th) week and 27 ~ (th) week, respectively. The results showed that the body weight of the DIO-P group was significantly higher than that of the other two groups after 7 weeks (P0.05). The intestinal flora of the DIO-P group was significantly higher than that of the NC group (P0.05), the diversity and stability of the intestinal flora of the DIO-P group were significantly lower than those of the NC group and the DIO-R group, and the abundance of (Bifidobacterium) Akkermansia muciniphila in the DIO-R group was decreased. However, the (Enterobacter cloacae) abundance of (Desulfovibrio), Staphylococcus aureus (Staphylococcus aureus), increased in DIO-R group, and the content of these bacteria did not change obviously at the early stage, but gradually tended to the characteristics of DIO-P group in the middle and late stage. At the beginning of the experiment, there was a significant difference between DIO-R group and DIO-P group in the composition of intestinal flora, and there was a significant correlation between the composition of intestinal flora and obesity. Compared with NC group, the content of acetic acid and butyric acid in feces of DIO-P group decreased significantly after 7 weeks. But propionic acid significantly increased (P0.05), in the Prophase and DIO-R group significant difference (P0.05), but no significant difference in late stage. DIO-P group was significantly higher than NC group after 7 weeks (P0.05), at 7 weeks and 17 weeks significantly higher than DIO-R group (P0.05). The expression of TLR4 (LPS receptor) and GPR41 (short chain fatty acid receptor) gene in ileum was significantly higher in DIO-P group than in DIO-R group (P0.05). The expression of this gene in the DIO-P group was significantly lower than that in the DIO-R group (P0.05). Zo-1Occludin and Muc2 were associated with the integrity of intestinal mucosal barrier. The expression of these three genes in the DIO-P group was significantly lower than that in the DIO-R group (P0.05), indicating that the intestinal barrier might be damaged in the obese susceptible group. Elisa analysis showed that the blood pro-inflammatory cytokines in DIO-P group increased significantly (P0.05), while the content of anti-inflammatory cytokines decreased significantly (P0.05), indicating the occurrence of chronic systemic inflammation. In conclusion, this study shows that the susceptibility to obesity in mice under high fat diet is closely related to intestinal flora. The mechanism of intestinal flora affecting obesity susceptibility may be related to injury of intestinal mucosal barrier, inflammatory response and disturbance of lipid metabolism.
【学位授予单位】：江南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R589.2
本文编号：2140163
[Abstract]:The large microbial community in the human intestine is symbiotic with the host, which plays an important role in the acquisition of body energy, the regulation of fat metabolism and the occurrence of chronic inflammation, which is closely related to the occurrence and development of obesity and metabolic syndrome. Two phenotypes of obesity resistance (DIO-R) and obesity susceptibility (DIO-P) can be produced in mice fed with high fat diet. It is important to explore the intestinal flora of mice to elucidate the mechanism of obesity and to propose nutritional intervention for obesity. C57BL/6 mice were randomly divided into control group (standard diet feeding NC) and high-fat diet group after 7 weeks. The high-fat group was divided into obese resistance group (DIO-R) and obesity susceptible group (DIO-P). At the beginning of the experiment (0 weeks), the early stage of obesity (7 weeks), the middle stage (17 weeks) and the end stage (27 weeks), the fecal microflora structure was detected by terminal restriction fragment length polymorphism (T-RFLP) and quantitative PCR, and the content of short chain fatty acids was measured. The blood lipopolysaccharide (LPS) and inflammatory cytokines were analyzed, and the expression of TLR4GPR41 Fiafo Zo-1Occludin and Muc2 genes in ileum epithelium were detected at 7 ~ (th) ~ (th) week and 27 ~ (th) week, respectively. The results showed that the body weight of the DIO-P group was significantly higher than that of the other two groups after 7 weeks (P0.05). The intestinal flora of the DIO-P group was significantly higher than that of the NC group (P0.05), the diversity and stability of the intestinal flora of the DIO-P group were significantly lower than those of the NC group and the DIO-R group, and the abundance of (Bifidobacterium) Akkermansia muciniphila in the DIO-R group was decreased. However, the (Enterobacter cloacae) abundance of (Desulfovibrio), Staphylococcus aureus (Staphylococcus aureus), increased in DIO-R group, and the content of these bacteria did not change obviously at the early stage, but gradually tended to the characteristics of DIO-P group in the middle and late stage. At the beginning of the experiment, there was a significant difference between DIO-R group and DIO-P group in the composition of intestinal flora, and there was a significant correlation between the composition of intestinal flora and obesity. Compared with NC group, the content of acetic acid and butyric acid in feces of DIO-P group decreased significantly after 7 weeks. But propionic acid significantly increased (P0.05), in the Prophase and DIO-R group significant difference (P0.05), but no significant difference in late stage. DIO-P group was significantly higher than NC group after 7 weeks (P0.05), at 7 weeks and 17 weeks significantly higher than DIO-R group (P0.05). The expression of TLR4 (LPS receptor) and GPR41 (short chain fatty acid receptor) gene in ileum was significantly higher in DIO-P group than in DIO-R group (P0.05). The expression of this gene in the DIO-P group was significantly lower than that in the DIO-R group (P0.05). Zo-1Occludin and Muc2 were associated with the integrity of intestinal mucosal barrier. The expression of these three genes in the DIO-P group was significantly lower than that in the DIO-R group (P0.05), indicating that the intestinal barrier might be damaged in the obese susceptible group. Elisa analysis showed that the blood pro-inflammatory cytokines in DIO-P group increased significantly (P0.05), while the content of anti-inflammatory cytokines decreased significantly (P0.05), indicating the occurrence of chronic systemic inflammation. In conclusion, this study shows that the susceptibility to obesity in mice under high fat diet is closely related to intestinal flora. The mechanism of intestinal flora affecting obesity susceptibility may be related to injury of intestinal mucosal barrier, inflammatory response and disturbance of lipid metabolism.
【学位授予单位】：江南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R589.2
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