本文選題：生命早期營養(yǎng)過剩　切入點：胰島素抵抗　出處：《重慶醫(yī)科大學(xué)》2013年碩士論文　論文類型：學(xué)位論文

【摘要】：背景：生命早期營養(yǎng)過剩（Chronic postnatal overnutrition，CPO）與機體發(fā)生肥胖和成年后胰島素抵抗（insulin resistance，IR）密切相關(guān)，而IR是非酒精性脂肪肝（Non-alcoholic fatty liver disease, NAFLD）始發(fā)的中心環(huán)節(jié)。NAFLD并非一種獨立疾病，它是單純脂肪變性，非酒精性脂肪性肝炎(Non-alcoholic steatohepatitis, NASH)，肝硬化和肝癌一系列進展性疾病的總稱。目前對NAFLD發(fā)生發(fā)展的機制研究中，經(jīng)典的“二次打擊學(xué)說”認(rèn)為IR是對肝臟的首次打擊，但其具體的機制尚未明確。CPO與NAFLD發(fā)生及進展的直接關(guān)系也尚未見報道。因此，本課題擬通過建立穩(wěn)定的動物模型探究生命早期營養(yǎng)過剩是否增加成年期膽堿蛋氨酸缺乏飲食（Methionine and choline deficientdiet，MCD）誘導(dǎo)NASH的易感性。 方法：通過調(diào)整母鼠喂養(yǎng)子鼠的數(shù)量建立早期營養(yǎng)過剩模型，，喂養(yǎng)3只子鼠為早期營養(yǎng)過剩組（Chronic postnatal overnutrition, CPO）,喂養(yǎng)10只子鼠為對照組（Control，CTR）,在3周齡斷奶后喂養(yǎng)標(biāo)準(zhǔn)飼料。在11周齡時，分別給予標(biāo)準(zhǔn)飲食（Standard chow diet, S）或MCD飲食持續(xù)喂養(yǎng)四周,據(jù)此將子鼠分為四組：CTR-S, CTR-MCD,CPO-S和CPO-MCD。3周齡、15周齡兩個時間點處死子鼠。3周齡時，采用透射電鏡技術(shù)觀察肝細(xì)胞超微結(jié)構(gòu)的改變；實時熒光定量PCR檢測相關(guān)基因的表達(dá)；Western Blot檢測肝臟中肉堿棕櫚酰轉(zhuǎn)移酶-1（Carnitine palmitoyltransferase-1， CPT-1）和細(xì)胞色素C（Cytochrome C, Cyt-C）蛋白表達(dá)。15周齡時，分別采用HE染色、油紅O染色、F4/80免疫組織化學(xué)染色、天狼猩紅染色及ɑ-SMA免疫組織化學(xué)染色觀測肝臟組織形態(tài)、脂質(zhì)沉積、炎癥及纖維化程度；全自動生化分析儀檢測小鼠血清生化指標(biāo)的改變情況；實時熒光定量PCR檢測相關(guān)基因表達(dá)；Western blot檢測肝組織中固醇調(diào)節(jié)元件蛋白-1c（Sterol regulatory element-binding protein-1c, SREBP-1c）蛋白表達(dá)水平。 結(jié)果：與CTR相比，CPO組從第2周齡開始體重增長更為明顯并持續(xù)至成年期導(dǎo)致成年后IR。在3周齡時，CPO組肝臟組織中SREBP-1c，F(xiàn)AS和SCD-1基因表達(dá)明顯上調(diào)；肝細(xì)胞內(nèi)線粒體出現(xiàn)腫脹，并伴有CTP-1基因及其蛋白表達(dá)水平降低，細(xì)胞色素C蛋白表達(dá)量降低。MCD喂養(yǎng)可導(dǎo)致小鼠肝臟出現(xiàn)典型的NSAH樣改變，尤其在MCD-CPO組最為明顯，主要表現(xiàn)為：明顯的脂質(zhì)沉積，脂肪變性，大泡樣改變，炎性細(xì)胞浸潤；肝組織TNF-ɑ表達(dá)上調(diào)；血清ALT及AST水平顯著升高。然而肝纖維化程度和TGF-β1mRNA表達(dá)各組間無統(tǒng)計學(xué)差異。MCD喂養(yǎng)后CPO小鼠肝組織中SREBP-1c, FAS,ACC-1, SCD, MTTP, FABP1, ABCA1和ABCA2基因表達(dá)明顯下調(diào)。 結(jié)論：生命早期營養(yǎng)過剩對機體代謝功能可能產(chǎn)生長期負(fù)面影響并促進成年期NASH進展。此現(xiàn)象可能與肝細(xì)胞脂質(zhì)外排功能受損和炎癥反應(yīng)加重有關(guān)。生命早期營養(yǎng)過�？赡苁荖ASH發(fā)展的重要風(fēng)險因素之一。
[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.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R151.2

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