本文選題：大鼠 + 營養(yǎng)過度�。� 參考：《上海交通大學(xué)》2015年博士論文

【摘要】：研究目的本研究通過生后早期營養(yǎng)干預(yù)致大鼠成年期胰島素抵抗的動物模型來探討生后早期營養(yǎng)環(huán)境與遠期代謝效應(yīng)的關(guān)聯(lián)及其部分內(nèi)在機制,闡明生后早期營養(yǎng)狀況對成年期胰島素抵抗的重要作用,為制定合理的生后早期喂養(yǎng)策略、預(yù)防和減少包括胰島素抵抗在內(nèi)的代謝綜合癥的發(fā)生提供依據(jù)。內(nèi)容本研究將分成三部分。第一部分為表型確立,建立生后早期營養(yǎng)過度、營養(yǎng)正常和營養(yǎng)不足的大鼠模型,比較各組大鼠胰島素抵抗差異。第二部分為機制探索,篩選與胰島素抵抗相關(guān)的信號通路并測定其中重要信號因子在胰島素效應(yīng)器官的表達和表觀遺傳修飾作用,同時對血清代謝組學(xué)進行檢測分析。第三部分為膳食刺激,比較生后早期不同營養(yǎng)狀況大鼠在高脂飲食暴露下發(fā)生成年期IR的風(fēng)險和重要信號因子表達的差異。方法新生SD大鼠,于生后第2天按每窩每只保姆鼠攜帶仔鼠數(shù)量的不同分別制備生后早期營養(yǎng)過度(3只/窩,SL組)、營養(yǎng)正常(10只/窩,NL組)和營養(yǎng)不足(20只/窩,LL組)大鼠模型。母鼠予常規(guī)飼料喂養(yǎng)。雄性大鼠為研究對象。仔鼠至日齡21天斷乳后繼續(xù)予常規(guī)飼料喂養(yǎng)。至6周齡時,每組再隨機分成兩個亞組,分別給予常規(guī)飼料和高脂飼料喂養(yǎng)至16周齡(成年期)止。動態(tài)觀察各組大鼠生理生化指標(biāo)、胰島素抵抗情況和胰島素外周效應(yīng)器官(骨骼肌、內(nèi)臟白色脂肪組織和肝臟)的形態(tài)學(xué)改變。運用轉(zhuǎn)錄組高通量測序技術(shù)篩選與胰島素抵抗相關(guān)的信號通路,采用實時定量PCR和western blotting方法檢測主要信號因子的表達,并使用重亞硫酸鹽測序法對重要差異信號因子DNA甲基化的程度進行分析。此外,氣相-質(zhì)譜、超高液相色譜-質(zhì)譜技術(shù)分別被用于大鼠血清脂肪酸譜和氨基酸譜的檢測。結(jié)果1. SL組大鼠在3周齡和16周齡時體重分別較NL組大鼠增加了37.5%和15.1%,LL組大鼠體重分別降低了34.9%和12.6%。肝臟、骨骼肌和內(nèi)臟白色脂肪(附睪脂肪、腎周脂肪)重量和組間差異與體重相-致,但經(jīng)體重糾正后只有SL組與NL組內(nèi)臟白色脂肪重量的組間差異得以保留。高脂喂養(yǎng)后仍然維持大鼠體重和內(nèi)臟白色脂肪組織重量的組間差異。2.16周齡SL組大鼠出現(xiàn)胰島素抵抗指數(shù)明顯上升和葡萄糖不耐受,同時伴有血清甘油三酯和游離脂肪酸水平升高,但空腹血糖水平未受影響。LL組大鼠胰島素抵抗指標(biāo)與NL組相比沒有差異。3. SL組大鼠3周齡時附睪脂肪細胞數(shù)目增加,16周齡時單個脂肪細胞面積增加。LL組大鼠單個附睪脂肪細胞面積均少于NL組大鼠。4.篩選出胰島素信號通路作為研究的目標(biāo)信號通路,其中主要關(guān)注的信號因子為胰島素受體(Insr)、胰島素受體底物(Irsl)、蛋白激酶B(Akt2)和葡萄糖轉(zhuǎn)運體4(Glut4)。16周齡SL組大鼠附睪脂肪Glut4和Irs1的mRNA表達下調(diào),Insr、Irs1、Akt2和Glut4的蛋白水平降低；腓腸肌Akt2和Glut4的mRNA表達下調(diào)、Insr和Glut4的蛋白水平下降。16周齡LL組大鼠附睪脂肪Glut4mRNA表達下調(diào)、腓腸肌Akt2mRNA表達上調(diào)和Irsl蛋白水平上升。5. 與NL組相比,SL組大鼠附睪脂肪Glut 4基因3周齡時為低甲基化高表達,16周齡時為高甲基化低表達。6.16周齡SL組大鼠血清長鏈飽和脂肪酸、單不飽和脂肪酸和花生四烯酸水平升高,同時伴有血清�；撬崴降慕档汀�7. SL組大鼠高脂喂養(yǎng)后胰島素抵抗指數(shù)和葡萄糖不耐受程度進一步上升,并出現(xiàn)明顯的高瘦素血癥和肝臟脂肪變性。附睪脂肪和腓腸肌胰島素信號通路信號因子表達進一步下調(diào)。8. LL組大鼠高脂喂養(yǎng)后出現(xiàn)葡萄糖不耐受程度增加、內(nèi)臟脂肪細胞面積明顯擴大和顯著的肝臟脂肪變性,同時伴有附睪脂肪和腓腸肌胰島素信號通路部分信號因子表達明顯下調(diào)。結(jié)論1. 生后早期營養(yǎng)過度可導(dǎo)致大鼠持續(xù)的體重過重和內(nèi)臟白色脂肪堆積,生后早期營養(yǎng)不足與之相反,這種變化不受飲食因素的影響。2. 生后早期營養(yǎng)過度導(dǎo)致大鼠成年期IR。大鼠骨骼肌和內(nèi)臟白色脂肪組織胰島素信號通路受損、內(nèi)臟白色脂肪組織Glut4基因甲基化程度改變、循環(huán)FFA水平增加、脂肪酸譜和氨基酸譜異�？赡軈⑴c了IR的發(fā)生和進展。3. 高脂飲食加劇了生后早期營養(yǎng)過度大鼠的IR和胰島素信號通路受損。4. 生后早期營養(yǎng)不足未對大鼠成年期IR產(chǎn)生影響,但高脂飲食可顯著增加生后早期營養(yǎng)不足大鼠發(fā)生IR的風(fēng)險。
[Abstract]:The purpose of this study is to explore the relationship between the early postnatal nutrition environment and the long-term metabolic effect and the intrinsic mechanism in order to elucidate the important role of early postnatal nutrition status to adult insulin resistance, and to formulate a reasonable early feeding strategy by using the animal model of adult insulin resistance induced by early postnatal nutrition intervention to explore the relationship between the early postnatal nutrition environment and the long-term metabolic effect. To prevent and reduce the occurrence of metabolic syndrome, including insulin resistance, this study will be divided into three parts. The first part is the establishment of phenotypes, the establishment of early postnatal overnutrition, normal nutrition and deficiency of the rat model, and the comparison of islet resistance differences in each group. The second part is the mechanism exploration, screening and analysis. Insulin resistance related signaling pathways and the determination of important signal factors in the expression of insulin effector organs and epigenetic modification, and the detection and analysis of serum metabolomics. The third part is dietary stimulation, and the risk of adult IR during the early stage of high fat diet exposure in rats with different nutritional status after birth is compared. Methods the difference in the expression of important signal factors. Method new SD rats were prepared on the second day after birth by each nanny rat of each nest to prepare early postnatal overnutrition (3 / nest, SL group), nutrition normal (10 / nest, NL group) and undernourishment (20 / LL) rats model. The female rats were fed with conventional feed. The male rats were fed. At the age of 21 days, the offspring continued to feed with conventional feed. At the age of 6 weeks, each group was randomly divided into two subgroups and fed with conventional and high fat feed to 16 weeks of age (adult). Dynamic observation of the physiological and biochemical indices of the rats, insulin resistance and the insulin peripheral effect organs (skeletal muscle, visceral white) Morphological changes in color adipose tissue and liver. The signal pathway related to insulin resistance was screened by high throughput sequencing technology in the transcriptional group. The expression of major signal factors was detected by real-time quantitative PCR and Western blotting methods, and the degree of DNA methylation of important differential signal factors was analyzed by heavy sulfite sequencing. In addition, GC-MS was used to detect the serum fatty acid spectrum and amino acid spectrum of rat serum. Results 1. SL rats were increased by 37.5% and 15.1% at the age of 3 and 16 weeks, respectively. The body weight of group LL decreased by 34.9% and 12.6%., and in the skeletal and visceral white fat respectively. The difference between the weight and the weight difference between the group SL and the NL group was retained after the weight correction. The difference between the weight of the body weight and the visceral white fat tissue after the high fat feeding was still maintained after the high fat feeding. The insulin resistance index in the group of SL rats at the age of.2.16 weeks was significantly higher. The level of triglyceride and free fatty acid in the serum was accompanied by elevated glucose intolerance and glucose intolerance, but the insulin resistance index of the.LL group was not significantly different from that of the NL group. The number of epididymal adipocytes in the group of.3. SL rats increased at the age of 3 weeks, and the area of the single adipocyte increased in the group of.LL rats at the age of 16 weeks. The area of the adipocyte was less than that of the NL group.4.. The insulin signaling pathway was selected as the target signal pathway for the study. The main signal factors concerned were insulin receptor (Insr), insulin receptor substrate (Irsl), protein kinase B (Akt2) and glucose transporter 4 (Glut4).16 weeks old rats in the epididymal fat Glut4 and Irs1 mRNA The protein levels of Insr, Irs1, Akt2 and Glut4 decreased, the expression of the mRNA expression of the gastrocnemius Akt2 and Glut4 was down, the protein levels of Insr and Glut4 decreased in the LL group of.16 weeks old rats, and the expression of the epididymal expression of the gastrocnemius muscle and the level of protein increased at the age of 3 weeks of age of the epididymal fat 4 of the rats. The high expression of low methylation was high methylation at the age of.6.16 week old SL group, the level of serum long chain saturated fatty acid, monounsaturated fatty acid and arachidic acid increased, and the level of serum taurine decreased in group.7. SL, and the index of insulin resistance and the degree of glucose intolerance increased further after high fat feeding in group.7. SL. The expression of signal factor of epididymal fat and gastrocnemius insulin signaling pathway further downregulated the increase of glucose intolerance after high fat feeding in the.8. LL group, significantly increased visceral adipocyte area and significant hepatic steatosis, accompanied by epididymal fat and gastrocnemius. The expression of partial signal factors in the signaling pathway of muscle insulin was obviously down-regulated. Conclusion early nutrition over 1. birth could lead to excessive weight loss and visceral white fat accumulation in rats. The early nutritional deficiency after birth was the opposite. This change was not influenced by dietary factors. The early period of.2. after birth was caused by the skeletal muscle of IR. rats in adult rats. The insulin signaling pathway in muscle and visceral white adipose tissue is damaged, the degree of methylation of Glut4 gene in visceral white adipose tissue changes, the level of circulating FFA increases, the abnormal fatty acid spectrum and amino acid spectrum may participate in the occurrence and progression of IR, and.3. high fat diet aggravates the IR and insulin signaling pathway of.4. in early postnatal rats. Early postnatal undernutrition did not affect IR in adult rats, but high fat diet significantly increased the risk of IR in the early postnatal undernourished rats.
【學(xué)位授予單位】：上海交通大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R153.1

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