生命早期营养过剩加重成年期饮食诱导的非酒精性脂肪肝炎

发布时间：2018-03-15 02:03

本文选题：生命早期营养过剩　切入点：胰岛素抵抗　出处：《重庆医科大学》2013年硕士论文　论文类型：学位论文

【摘要】：背景：生命早期营养过剩（Chronic postnatal overnutrition，CPO）与机体发生肥胖和成年后胰岛素抵抗（insulin resistance，IR）密切相关，而IR是非酒精性脂肪肝（Non-alcoholic fatty liver disease, NAFLD）始发的中心环节。NAFLD并非一种独立疾病，它是单纯脂肪变性，非酒精性脂肪性肝炎(Non-alcoholic steatohepatitis, NASH)，肝硬化和肝癌一系列进展性疾病的总称。目前对NAFLD发生发展的机制研究中，经典的“二次打击学说”认为IR是对肝脏的首次打击，但其具体的机制尚未明确。CPO与NAFLD发生及进展的直接关系也尚未见报道。因此，本课题拟通过建立稳定的动物模型探究生命早期营养过剩是否增加成年期胆碱蛋氨酸缺乏饮食（Methionine and choline deficientdiet，MCD）诱导NASH的易感性。方法：通过调整母鼠喂养子鼠的数量建立早期营养过剩模型，，喂养3只子鼠为早期营养过剩组（Chronic postnatal overnutrition, CPO）,喂养10只子鼠为对照组（Control，CTR）,在3周龄断奶后喂养标准饲料。在11周龄时，分别给予标准饮食（Standard chow diet, S）或MCD饮食持续喂养四周,据此将子鼠分为四组：CTR-S, CTR-MCD,CPO-S和CPO-MCD。3周龄、15周龄两个时间点处死子鼠。3周龄时，采用透射电镜技术观察肝细胞超微结构的改变；实时荧光定量PCR检测相关基因的表达；Western Blot检测肝脏中肉碱棕榈酰转移酶-1（Carnitine palmitoyltransferase-1， CPT-1）和细胞色素C（Cytochrome C, Cyt-C）蛋白表达。15周龄时，分别采用HE染色、油红O染色、F4/80免疫组织化学染色、天狼猩红染色及ɑ-SMA免疫组织化学染色观测肝脏组织形态、脂质沉积、炎症及纤维化程度；全自动生化分析仪检测小鼠血清生化指标的改变情况；实时荧光定量PCR检测相关基因表达；Western blot检测肝组织中固醇调节元件蛋白-1c（Sterol regulatory element-binding protein-1c, SREBP-1c）蛋白表达水平。结果：与CTR相比，CPO组从第2周龄开始体重增长更为明显并持续至成年期导致成年后IR。在3周龄时，CPO组肝脏组织中SREBP-1c，FAS和SCD-1基因表达明显上调；肝细胞内线粒体出现肿胀，并伴有CTP-1基因及其蛋白表达水平降低，细胞色素C蛋白表达量降低。MCD喂养可导致小鼠肝脏出现典型的NSAH样改变，尤其在MCD-CPO组最为明显，主要表现为：明显的脂质沉积，脂肪变性，大泡样改变，炎性细胞浸润；肝组织TNF-ɑ表达上调；血清ALT及AST水平显著升高。然而肝纤维化程度和TGF-β1mRNA表达各组间无统计学差异。MCD喂养后CPO小鼠肝组织中SREBP-1c, FAS,ACC-1, SCD, MTTP, FABP1, ABCA1和ABCA2基因表达明显下调。结论：生命早期营养过剩对机体代谢功能可能产生长期负面影响并促进成年期NASH进展。此现象可能与肝细胞脂质外排功能受损和炎症反应加重有关。生命早期营养过剩可能是NASH发展的重要风险因素之一。
[Abstract]:Background: Chronic postnatal overnutrition (postnatal) is closely associated with obesity and insulin resistance in adulthood, while IR is not an independent disease, which is the central link of non-alcoholic fatty liver disease (NAFLD). It is a general name for a series of progressive diseases such as simple steatosis, non-alcoholic steatohepatitis, NASH, liver cirrhosis and liver cancer. The classical "second strike theory" holds that IR is the first attack on the liver, but the specific mechanism of IR is not clear about the direct relationship between CPO and the occurrence and progression of NAFLD. The aim of this study was to establish a stable animal model to investigate whether early life excess increased the susceptibility to NASH induced by dietary methionine and choline deficiency MCD-induced by adult choline methionine deficiency. Methods: a model of early nutritional overnourishment was established by adjusting the number of offspring fed to female rats. Three of them were fed with Chronic postnatal overnutrition (CPO), and 10 of them were fed as control group, and fed with standard diet after weaning at 3 weeks of age. Standard chow diet (S) or MCD diet were given continuously for four weeks. The offspring were divided into four groups: CTR-S, CTR-MCDCPO-S and CPO-MCD.3 15-week-old. The ultrastructural changes of hepatocytes were observed by transmission electron microscopy (TEM). The expression of carnitine palmitoyltransferase-1 (CPT-1) and cytochrome C (Cyt-C) protein in liver were detected by real-time fluorescence quantitative PCR. The expression of carnitine palmitoyltransferase-1 (CPT-1) and cytochrome C (Cyt-C) in liver were detected by HE staining and oil red O staining respectively. The degree of liver tissue morphology, lipid deposition, inflammation and fibrosis were observed by Sirius red staining and SMA immunohistochemical staining, and the changes of serum biochemical indexes in mice were detected by automatic biochemical analyzer. The expression of sterol regulatory element-binding protein-1c (SREBP-1c) protein in liver tissue was detected by real-time fluorescence quantitative PCR and Western blot. Results: compared with CTR group, the weight gain was more obvious from the second week of age and continued until adulthood. At the age of 3 weeks, the expression of SREBP-1cFASA and SCD-1 genes in liver tissue was upregulated, and mitochondria swelling appeared in the hepatocytes. In addition, the expression of CTP-1 gene and its protein decreased, and the expression of cytochrome C protein decreased. MCD induced typical NSAH like changes in the liver of mice, especially in the group of MCD-CPO, which was characterized by obvious lipid deposition. Fatty degeneration, vesicular changes, inflammatory cell infiltration, up-regulation of TNF-T expression in liver tissue; The levels of serum ALT and AST were significantly increased. However, there was no significant difference in the degree of hepatic fibrosis and the expression of TGF- 尾 1 mRNA among the groups. The expression of SREBP-1c, FASACC-1, SCD-1, MTTP, FABP1, ABCA1 and ABCA2 genes in liver tissues of CPO mice fed with MCD was significantly down-regulated. Conclusion: excessive nutrition in early life may have long-term negative effects on metabolic function and promote the development of NASH in adulthood. This phenomenon may be related to impaired lipid efflux function of hepatocytes and aggravation of inflammatory reaction in early stage of life. Excess nutrition may be one of the important risk factors for the development of NASH.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R151.2

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