转录因子Nrf2对高脂饮食诱导小鼠肝脏胰岛素抵抗的影响和机制

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第四军医大学硕士学位论文

目录

缩略语表 (1)

中文摘要 (3)

英文摘要 (7)

前言 (12)

文献回顾 (13)

正文 (30)

实验一敲除Nrf2加重高脂饮食诱导小鼠肝脏氧化应激水平 (30)

引言 (30)

1 材料 (31)

2 方法 (31)

3 结果 (34)

4 讨论 (37)

实验二转录因子Nrf2对小鼠肝脏胰岛素抵抗的影响及其机制 (39)

引言 (39)

1 材料 (40)

2 方法 (40)

3 结果 (43)

4 讨论 (47)

小结 (50)

参考文献 (51)

个人简历和研究成果 (62)

致谢 (63)

第四军医大学硕士学位论文

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缩略语表

缩略词 英文全称

中文全称

NAFLD

Nonalcoholic fatty liver disease

非酒精性脂肪性肝病 IR Insulin resistance 胰岛素抵抗

NASH Nonalcoholic steatohepatitis 非酒精性脂肪性肝炎 IR Insulin receptor 胰岛素受体

Nrf2

NF-E2-related factor 2

转录因子NF-E2相关因子2 IRS Insulin receptor substrate 胰岛素受体底物 NF-κB

Nucleus factor-kappa B

核因子-kappa B IKK Inhibitor of nuclear factor kappa-B kinase 核因子kB 抑制物激酶 PI3K Phosphatidylinositol-3-Kinase 磷脂酰肌醇-3-激酶 Akt Protein Kinase B

蛋白激酶B GSK-3β Glycogen synthase kinase 3β 糖原合成激酶3β FoXO1

Forkhead box O1

叉头蛋白O3 4HNE 4-hydroxynonenal 四羟壬烯醛 ROS

Reactive oxygen species

活性氧 ARE Antioxidant-response element 抗氧化反应元件 MDA Malondialdehyde 丙二醛 GSH Glutathione 还原型谷胱甘肽 Nqo1 NAD(P)H:quinine oxidoreductase 1 醌氧化还原酶1 TNF-α

Tumor necrosis factor α

肿瘤坏死因子α IL-6 Interleukin-6 白细胞介素6 IL-10 Interleukin-10 白细胞介素10 TG Triglyceride 甘油三酯 FFA

Free fat acid

游离脂肪酸

第四军医大学硕士学位论文

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缩略词 英文全称 中文全称

DGAT2 Diacylglycerol acyltransferase2 二酰基甘油酰基转移酶2 AhR

A hydrocarbon receptor

芳香烃受体 MS Metabolic syndrome 代谢综合征 LPS Lipopolysaccharide 脂多糖 TLR Toll-like receptor Toll 样受体 KC Kupffer cell Kupffer 细胞 TGF-β Transforming growth factor β 转化生长因子β LXR Liver X receptor

肝脏X 受体 mTOR Mammalian Target Of Rapamycin 雷帕霉素靶蛋白

SREBP-1c Sterol regulatory element binding protein 1c 甾固醇调节元件结合蛋白1c ChREBP

Carbohydrate responsive element binding protein

碳水化合物反应元件结合蛋白

DNL

De novo lipogenesis

脂质新生 ER Endoplasmic reticulum 内质网应激 CYP2E1 Cytochrome P4502E1 细胞色素P4502E1

PPAR α

Peroxisome proliferator activatived receptors α

过氧化物酶体增殖物活化受体αUCP2 Uncoupling protein 2 解偶联蛋白2 ATP Adenosine Triphosphate 三磷酸腺苷 XO Xanthine Oxidase 黄嘌呤氧化酶 SOD Superoxide Dismutase 超氧化物歧化酶 GPx Glutathione peroxidase 谷胱甘肽过氧化物酶 Keap1 Kelch-like ECH-associating protein 1 Kelch 样ECH 联合蛋白1 Bach1

BTB and CNC homology 1

BTB-CNC 异体同源体1 LCFA-CoA Long chain fatty acid CoA

长链脂肪酸辅酶A

第四军医大学硕士学位论文

转录因子Nrf2对高脂饮食诱导小鼠肝脏胰岛素抵抗的

作用及机制

硕士研究生：倪 阵

导 师：闻勤生 教授

辅导教师：刘震雄 讲师

第四军医大学唐都医院消化内科，西安 710038

资助基金项目：国家自然科学基金（81270485;81170376）

中文摘要

关键词：NASH；Nrf2；胰岛素抵抗；氧化应激；NF-κB；

背景

非酒精性脂肪性肝病（nonalcoholic fatty liver disease, NAFLD）是一种组织学上表现为肝脏脂肪变性的临床病理综合征，但应排除酒精性、感染性、药物性和先天性代谢异常等继发因素，其疾病谱包括单纯性脂肪肝、非酒精性脂肪性肝炎（nonalcoholic steatohepatitis, NASH）及其进展而来的肝硬化和肝癌[1]。目前观点认为，单纯性脂肪肝属于良性病变，而NASH则可以进展至肝脏纤维化、肝硬化和肝癌[2]，但是NASH的发病机制至今尚未完全阐明。胰岛素抵抗在NASH患者中普遍存在，进一步发现肝脏胰岛素抵抗可以先于外周胰岛素抵抗出现[3]，因此，延缓或阻止肝脏胰岛素抵抗的发生和进展，则有可能成为治疗NASH的重要靶点。但是目前有关肝脏胰岛素抵抗的确切发病机制，仍有待于进一步研究。

肝脏胰岛素作用主要是通过IR/IRS/PI3K/Akt信号通路发挥作用。正常情况下，胰岛素可以通过抑制糖原异生和肝脏糖原分解，从而调节肝脏糖原输出，而在肝脏

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第四军医大学硕士学位论文

胰岛素抵抗状态下，此种抑制作用受损导致高血糖和代偿性高胰岛素血症[4]。大量研究证实，氧化应激是高脂饮食诱导肝脏胰岛素抵抗发生的关键机制。脂质过氧化产物和氧化应激导致的DNA损伤产物，如4HNE和8羟基脱氧鸟苷，与胰岛素抵抗严重程度呈现正相关[5]。另外，有证据提示脂质过氧化产物还可以激活炎症信号通路IKK/NF-κB并促进炎症因子TNFα和IL-6等的释放，间接诱导胰岛素抵抗。和健康人群相比，NASH患者ROS表达和炎症因子水平都明显增加[6]。事实上，脂质过氧化产物和炎症因子具有协同作用，它们可以通过促使胰岛素信号通路关键蛋白的磷酸化而触发胰岛素抵抗的发生。

转录因子NF-E2相关因子2（NF-E2-related factor 2，Nrf2）是细胞防御氧化应激反应的重要转录因子，它可以与抗氧化反应元件(antioxidant-response element, ARE)结合，上调抗氧化蛋白和Ⅱ相解毒酶的表达，增强细胞的抗氧化能力[7]。最近研究认为Nrf2是细胞抗氧化反应的中枢调节者，Nrf2 能够维持细胞内的氧化平衡，降低细胞对死亡信号的敏感性，在细胞抵御外源性或内源性的氧化应激的机制中起重要作用[8]。但目前关于Nrf2在肝脏胰岛素抵抗中的作用仍然不是很清楚。

因此本研究利用Nrf2基因敲除小鼠，通过高脂饮食诱导肝脏胰岛素抵抗，探讨其发生的相关具体机制。

目的

本实验采用高脂饮食制备小鼠肝脏胰岛素抵抗动物模型，观察氧化应激对肝脏细胞转录因子Nrf2核转位的影响，研究敲除Nrf2对肝脏胰岛素信号通路（IRS/Akt）和炎症信号通路（IKK/NF-κB）的影响，为以Nrf2为靶点进行肝脏胰岛素抵抗的防治提供新的实验基础和理论依据。

方法

1、Nrf2基因敲除（Nrf2-null）和野生型（WT）ICR雄性小鼠各10只，随机分为WT组（n=5）、WT高脂饮食组（WT+HFD）（n=5）、Nrf2-null组（n=5）和Nrf2-null 高脂饮食组（Nrf2-null+HFD）（n=5），喂养8周，每周记录体重变化，8周末，空腹12 h，腹腔注射戊巴比妥钠麻醉后，称取肝脏重量；生化法检测肝脏甘油三酯（triglyceride, TG）含量；分光光度法检测肝脏丙二醛（malondialdehyde, MDA）和

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第四军医大学硕士学位论文

谷胱甘肽水平（glutathione, GSH）；Western-blot法检测肝脏Nrf2、Nqo1等的表达变化；肝脏组织HE染色观察肝脏病理学改变。

2、Nrf2基因敲除（Nrf2-null）和野生型（WT）ICR雄性小鼠各10只，随机分为WT组（n=5）、WT高脂饮食组（WT+HFD）（n=5）、Nrf2-null组（n=5）和Nrf2-null 高脂饮食组（Nrf2-null+HFD）（n=5），喂养8周，每周记录体重变化，8周末空腹12 h行腹腔注射葡萄糖耐量实验；生化法检测空腹血清血糖；分光光度法检测肝脏丙二醛（malondialdehyde, MDA）和谷胱甘肽（glutathione, GSH）水平；ELISA和RT-PCR检测肝脏TNF-α和IL-6 mRNA和蛋白表达；Western blot法检测肝脏NF-κB、IRS1、Akt、GSK-3β、FoXO1等表达的变化。

结果

1．8周末，高脂饮食组小鼠体重显著高于普通饮食组（P<0.05），且Nrf2-null 高脂饮食小鼠体重高于WT高脂饮食小鼠，但二者差异无统计学意义（P>0.05）。生化结果显示，高脂饮食组小鼠肝脏重量和肝脏TG含量显著高于普通饮食组（P<0.05），且Nrf2-null高脂饮食小鼠显著高于WT高脂饮食小鼠（P<0.05）。肝脏组织学显示，和普通饮食组小鼠相比，WT高脂饮食小鼠肝脏组织学结构紊乱，肝细胞出现明显的脂肪沉积，表现为肝细胞大疱性脂肪变性；Nrf2-null小鼠肝脏肝细胞呈现气球样变性，伴小叶静脉周围及肝细胞周围大量炎症细胞浸润。

2．8周末，高脂饮食组小鼠肝脏MDA显著高于普通饮食组小鼠（P<0.05），GSH水平显著低于高脂饮食组小鼠（P<0.05），而且Nrf2-null高脂饮食组小鼠肝脏MDA水平显著高于WT高脂饮食组（P<0.05），GSH水平显著低于WT高脂饮食组小鼠（P<0.05）。Western Blot结果显示，Nrf2-null组小鼠肝脏无明显Nrf2总蛋白的表达，WT高脂饮食组小鼠肝脏Nrf2总蛋白表达和WT普通饮食组未见明显变化，而WT高脂饮食组小鼠肝脏Nrf2胞核蛋白表达显著高于WT普通饮食组；Nqo1表达在Nrf2-null小鼠中显著减少，而WT高脂饮食组小鼠肝脏Nqo1表达显著高于WT 普通饮食组小鼠。

3．8周末，高脂饮食组小鼠空腹血糖显著高于普通饮食组（P<0.05），Nrf2-null 高脂饮食组小鼠空腹血糖高于WT高脂饮食组小鼠，但差异无统计学意义（P>0.05）。腹腔注射葡萄糖耐量实验结果显示，高脂饮食组小鼠15min、30min、

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第四军医大学硕士学位论文

60min血糖和AUC显著高于普通饮食小鼠（P<0.05），且Nrf2-null高脂饮食组小鼠显著高于WT高脂饮食组小鼠（P<0.05）。Western-Blot结果显示，肝脏总IRS-1和Akt在各组间无明显差异，WT高脂饮食组小鼠肝脏p-IRS1（Tyr）、p-Akt（Ser307）、p-GSK-3β和p-FoXO1表达较WT组减少，且Nrf2-null+HFD组p-IRS1（Tyr）、p-Akt （Ser307）、p-GSK-3β和p-FoXO1表达较WT高脂饮食组显著减少。

4．8周末，ELISA和RT-PCR结果显示，高脂饮食组小鼠肝脏TNFα和IL-6显著高于普通饮食组小鼠，而Nrf2-null高脂饮食组小鼠肝脏TNFα和IL-6水平显著高于WT高脂饮食组。Western Blot结果显示，NF-κB总蛋白在各组表达无明显差异，而WT高脂饮食组小鼠肝脏NF-κB胞核蛋白表达明显高于普通饮食组，且Nrf2-null 高脂饮食组小鼠肝脏NF-κB胞核蛋白表达明显高于WT高脂饮食组。

结论

8周高脂饮食诱导小鼠出现明显氧化应激，证实在氧化应激条件下肝脏转录因子Nrf2发生核转位，发挥其抗氧化的作用。同时也证实，8周高脂饮食可以诱导小鼠肝脏NF-κB信号通路活化并导致胰岛素抵抗的发生，而敲除Nrf2后，由于NF-κB 信号通路活化和炎症因子释放加重，从而导致肝脏胰岛素抵抗程度进一步加重，为以Nrf2为靶点治疗和预防NASH提供新的理论基础和实验依据。

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第四军医大学硕士学位论文

Deletion of Nrf2 aggravate inhepatic insulin resistance in

mice fed with high fat diet

Candidate for master:Ni Zhen

Supervisor:Wen Qinsheng

Tutor:Liu Zhenxiong

Department of Gastrainterology,Tanddu hospital, Fourth Military Medical University,

Xi’an 710038, China

Sponsored Programs: NNSF(81270485;81170376)

Abstract

Key words：NASH; Nrf2; hepatic insulin resistance; oxidative stress; NF-κB；

Background

Non-alcoholic fatty liver disease (NAFLD), ranging from simple steatosis to steatohepatitis(NASH) which can progress to liver cirrhosis and hepatocellular cancer，is the most common form of chronic liver disease all over the world. But the exact pathological mechanism is still not so clear.

Because of its strong association with obesity, type 2 diabetes mellitus, hypertension and hypertriglyceridaemia, NAFLD is universally considered as the hepatic manifestation of metabolic syndrome. It is generally accepted that hepatic insulin resistance is a key pathophysiological hallmark in NAFLD and might play a role in the progression from simple fatty liver to NASH by promoting the development of steatosis and the subsequent cell injury and inflammation. So a better understanding on the underlying mechanisms how hepatic insulin resistance develops is pivotal to devise rational treatments that could

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第四军医大学硕士学位论文

prevent the progression of NAFLD to advanced forms.

One of the mechanisms proposed to account for the development of insulin resistance after the administration of a high fat diet is ROS upregulation in the liver. Markers of lipid peroxidation and oxidative DNA damage, such as hepatic content of both HNE and 8-hydroxydeoxygnanosine are correlated with the severity of insulin resistance. Indeed, ROS and lipid peroxidation products act together to trigger the development of insulin resistance through phosphorylation of proteins of the insulin signaling cascade.

In addition, recent evidence suggest that part of the role of oxidative stress on insulin resistance could contribute to secondary inflammation state. Furthermore, it is noteworty that these two events are intertwined and a vicious cycle can take place between them. One hand, ROS and products of lipid peroxidation can activate IKK/NF-kappaB signal pathway, which induce the synthesis of several proinflammatory cytokines including TNFalpha and IL-6, leading to insulin resistance via paracrine effects. On the other hand, those cytokines, especially TNFalpha and IL-6, can induce mitochoudrial dysfunction by promoting ROS and RNS production, which contributes significantly to the progression of insulin resistance and NASH. So it is reasonable to speculate that cutting off the loop will be an attractive therapeutic target to inhibit the development of insulin resistance and the progression of NAFLD to NASH.

Nrf2 is a key transcription factor that regulates many kinds of antioxidant genes and detoxifying enzymes against oxidative stress and electronic stress. Under normal condition, Nrf2 is sequestered in the cytosol by Kelch-like Ech-associated protein (Keap1). But when challenged with oxidative stress or electronic stress, Nrf2 dissociates with Keap1, then translocates into nucleus, and promotes the expression of a number of antioxidant genes such as GCLC,Nqo1 and Gsta1. It was showed that MCD or HFD could induce more aggravated oxidative stress injury and inflammation in Nrf2-null mice than in WT mice. On the contrary, such damages can be attenuated through elevating the level of Nrf2 by its inducer or genenic disruption of Keap1.Our previous study also showed that activation of Nrf2 by curcumin could significantly attenuate the oxidative stress injury in hepatocyte and hepatic stellate cells induced by GO.

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第四军医大学硕士学位论文

Objective

To investigate the influence of oxidation stress on nuclear translocation of Nrf2 in mouse hepatic insulin resistance model induced by high fat diet. Furthermore, we hypothesis that deletion of Nrf2 markedly accelerates the onset and progression of hepatic insulin resistance due to the more serious oxidative stress and inflammation. It will provide a new experimental and theoretical basis for the prevention and treatment of NASH.

Methods

1．Male 8 week-old WT and Nrf2-null mices on ICR background were randomly divided into four groups after adopt to the lab condition for 1 week and were free access to irradiated hight-fat diet(10% lard, 2% cholesterol, 0.5% bile salt and 87.5% base forage) or a control diet for 8 weeks(n=5). Body weight were measured weekly. At the end of the experiment,liver weight were measured. After that, the changes of pathology of liver were observed. Then the levels of liver TG, hepatic methane dicarboxylic aldehyde (MDA) and glutathione (GSH) were examined by spectrophotometric method. The expression of liver Nrf2 and Nqo1 were determined by western-blot.

2．Male 8 week-old WT and Nrf2-null mices on ICR background were randomly divided into four groups after adopt to the lab condition for 1 week and were free access to irradiated hight-fat diet(10% lard, 2% cholesterol, 0.5% bile salt and 87.5% base forage) or a control diet for 8 weeks(n=5). Body weight were measured weekly. At the end of the experiment, iPGTT was performed before sacrifice. After that, the changes of pathology of liver were observed. Then the levels of blood glucose, hepatic methane dicarboxylic aldehyde (MDA) and glutathione (GSH) were examined by spectrophotometric method. The levels hepatic TNFα and IL-6 were examined by ELISA and RT-PCR. The expression of liver NF-κB, IRS-1, Akt, p-GSK-3β and p-FoXO1 were determined by western-blot. Results

1．After feeding the diet for 8 weeks, weight gain in both WT and Nrf2-null mice increased significantly compared with weight gain after intake of control diet. And the

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第四军医大学硕士学位论文

tendency to be higher in Nrf2-null mice, although there were no significantly differences when compared to WT intake of HFD. Also the liver weight increased significantly both in WT and Nrf2-null mice after feeding HFD for 8 weeks in compared with control diet, but no significant differences were noted between WT and Nrf2-null mice. In line with this, WT and Nrf2-null on HFD showed a comparable increase in hepatic TG at 8 weeks. Moreover, the magnitudes were significantly higher in Nrf2-null mice than in WT mice.

2．After start the HF diet for 8 weeks, both WT and Nrf2-null mice showed more microvesicular lipid accumulation in livers compared with the normal diet groups. As expected, the Nrf2-null mice exhibited increases in in?ltration of in?ammatory cells in the livers, compared with the ?ndings in the livers of the WT mice intake of HFD 3．The level of liver MDA increased significantly both in WT and Nrf2-null mice after feeding HFD for 8 weeks in compared with control diet. Furthermore, the magnitudes were significantly greater in Nrf2-null mice than in WT mice. In contrast, feeding HFD for 8 weeks significantly reduced the concentration of liver GSH compared with intake of control diet and the tendency was to be more obervious in Nrf2-null mouse. Feeding HFD for 8 weeks significantly increased the expression of nuclear Nrf2 protein in WT mouse liver, wherase no Nrf2 expression were noted in Nrf2-null mouse liver. Furthermore, the basal protein levels of the typical Nrf2-regulated genes, Nqo1, were signi?cantly lower in the livers of Nrf2-null mice than in WT mice fed the control diet and were significantly induced by HFD feeding in liver of WT mouse.

4．8 weeks HFD fed resulted in a notable increase of FGB both in WT mice and Nrf2-null compared with control. Moreover, the increase in Nrf2-null mice were more significant than that in WT mices. Also, 8 weeks HFD fed leaded to markedly increase of the blood glucose of WT and Nrf2-null mices following i.p. glucose load at both baseline and after 15min, 60min and 90mmin. Additionally, the area under the curve (AUC), which is a measure of impaired glucose tolerance, also signi?cantly increased in iPGTT in WT and Nrf2-null mices. Further,the magnitude of the above increase were both greater in the Nrf2-null mice than those in the WT mice.

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第四军医大学硕士学位论文

5．After start the HF diet for 8 weeks, there were no significant changes of total IRS-1 and total Akt, but tyrosine phosphorylation of IRS-1 and the phosphorylation of Akt were signi?cantly reduced after the intake of HFD for 8 weeks. Also, the phosphorylation of GSK-3β and FoXO1 were significantly reduced. Moreover, the above decreases tended to be more significant in Nrf2-null mice indicating that HFD feeding induced more serious hepatic insulin resistance in nrf2-null mice.

6．The mRNA levels of TNFα and IL-6 were significantly higher in the livers of Nrf2-null mice than in those of WT mice at 8 weeks after the start of HFD feeding. Furthermore, the protein levels of TNFα and IL-6 were correlated with the mRNA levels. As expected, the nucleus expression of NF-κB was significantly upregulated both in WT and Nrf2-null mice liver at 8 weeks after the start of HFD feeding and the upregulation was more significant in Nrf2-null mice liver than in WT mice liver.

Conclusion

The study shows that oxidative stress induced by high fat diet can activate the nuclear translocation of Nrf2 to develop its biological effects. Meanwhile, deletion of Nrf2 significantly exacerbate intrahepatic insulin resistance through decreasing intracellar GSH level and aggravatting the degress of oxidative stress, promotting the translocation of NF-κB and the release of inflammatory factors. The findings imply that the translocation of Nrf2 could be a target for the prevention and treatment of NASH.

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第四军医大学硕士学位论文

前言

非酒精性脂肪性肝炎（NASH）是全球范围内最常见的慢性肝脏疾病之一，被认为是代谢综合征在肝脏的表现。胰岛素抵抗(IR)和NASH关系密切，并能通过促进肝脏脂肪变性、细胞损伤和炎症反应，最终导致非酒精性单纯性脂肪肝（NAFL）向非酒精性脂肪性肝炎（NASH）的发展，而氧化应激是诱导IR的重要危险因素，其可以直接或间接活化多种应激通路，进一步增加胰岛素受体底物-1（IRS-1）不连续的丝氨酸或苏氨酸磷酸化，从而抑制IRS-1酪氨酸磷酸化，最终导致IR的发生。核转录因子NF-E2相关因子2（Nrf2）是体内调节氧化应激的关键靶点，其可以通过促进多种抗氧化酶和II相解毒酶的表达，发挥重要的抗氧化作用，但Nrf2在肝脏IR 中的作用尚未见明确报道。

本实验以Nrf2基因敲除和WT小鼠为研究对象，8周高脂饮食诱导肝脏氧化应激，生化法检测肝脏MDA及GSH水平，Western Blot法观察肝脏Nrf2及Nqo1的表达，分光光度法检测血清葡萄糖水平，腹腔注射葡萄糖耐量实验观察胰岛素敏感性，ELISA和RT-PCR检测肝脏TNFα和IL-6表达，Western-blot检测肝脏IRS-1、Akt、p-IRS-1、p-Akt、p-GSK-3β、p-FoXO1和NF-κB表达变化，从而观察氧化应激对肝脏Nrf2表达的影响，以及敲除Nrf2对肝脏氧化应激、肝脏炎症水平及肝脏胰岛素抵抗的影响并初步探讨其机制，从而为以Nrf2为靶点治疗肝脏胰岛素抵抗提供理论基础。

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第四军医大学硕士学位论文

文献回顾

1 非酒精性脂肪性肝病

1.1 非酒精性脂肪性肝病的基本概念及流行病学

NAFLD是描述一个相关疾病综合征的涵盖性术语，其最早期阶段为肝脏脂肪变性，主要表现为甘油三酯（TG）以脂滴的形式在肝细胞胞浆内沉积，当肝脏TG 含量超过55 mg/g或者超过5%的肝细胞出现脂滴即可诊断为脂肪肝[9]。肝脏脂肪变性具有自限性，但是它可以进展至非酒精性脂肪性肝炎（NASH）。NASH不同于单纯性脂肪肝，表现为肝细胞损伤（肝细胞气球样变性和死亡）、炎症细胞浸润和胶原沉积（纤维化）。目前单纯性脂肪肝与NASH之间的关系仍有待进一步研究。NASH具有进展性，10%-29%的NASH患者在10年内可以发展至肝硬化，4%-27%的NASH相关肝硬化可以发展至肝癌[10]。NAFLD常与肥胖、2型糖尿病、高血脂、高血压等代谢综合征并存，并且严重程度也密切相关。因此，NAFLD被认为是代谢综合征（metabolic syndrome, MS）在肝脏的表现。

NAFLD流行病学和自然病程仍未完全阐明，目前只有少部分研究涉及，其在全球的发病率还不清楚，而且其在各个病理阶段（单纯脂肪肝、NASH、肝硬化）的流行情况仍然有待于进一步研究。在北美洲、欧洲和澳洲等发达国家和地区，其发病率约为30%，而在糖尿病患者、肥胖和病态肥胖患者中其发病率则分别达到50%、76%和100%[11]。在我国，最近研究显示上海和广东的发病率约为17%和15%，其中，90%的脂肪肝患者病因可以归因于代谢因素,广东地区的儿童和青少年NAFLD的发病率约为1.3%,香港和台湾地区的发病率为16%和12%,目前，NAFLD 已经成为大陆和台湾地区健康人群体检中，无症状肝脏转氨酶升高的主要原因[12]。

1.2 非酒精性脂肪性肝炎的发病机制

NASH 是NAFLD发病过程由单纯性脂肪肝发展至肝脏纤维化的一个重要的中间阶段，也是病情发展的一个关键转折点。但是到目前为止，NASH的发病机制尚不明确。

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