二甲双胍对高糖高脂饮食诱导的糖尿病大鼠脂肪肝的干预作用

目的 观察高糖、高脂饮食诱导的糖尿病大鼠肝脏病理改变,探讨二甲双胍的干预作用.方法 40只SD大鼠随机分为正常对照(NC)组、糖尿病对照(DM-C)组和DM+二甲双胍干预(DM-M)组.比较各组生化指标、肝重及肝脏指数的改变.HE染色观察大鼠肝脏形态学变化;免疫组化法检测肿瘤坏死因子α(TNF-α)在肝脏的表达.结果 高糖、高脂饲养4周后诱导大鼠出现胰岛素抵抗;8周成功建立T2DM大鼠模型.18周末 DM-C 组大鼠肝脏出现严重脂肪变性,TNF-α在肝脏的蛋白表达水平增加(P<0.01).经二甲双胍治疗后,DM-M组大鼠肝脏脂肪变性程度明显减轻,TNF-α的蛋白表达水平下降(P<0.05).结论 二甲双胍可降低高糖、高脂饮食诱导的糖尿病大鼠肝脏脂肪含量,降低TNF-α在肝组织的蛋白表达,对治疗2型糖尿病脂肪肝具有一定的作用.     

二甲双胍对高糖高脂饮食诱导的糖尿病大鼠脂肪肝治疗作用的研究

目的探讨二甲双胍对脂肪肝的治疗作用及在改善胰岛素抵抗、调节糖脂代谢中的作用机制。方法2003年2月至2003年7月河北医科大学第三医院通过高糖、高脂饮食诱导并结合小剂量链脲佐菌素注射的方法,建立2型糖尿病大鼠合并脂肪肝的动物模型,再给予二甲双胍625mg/(kg.d)灌胃进行干预治疗10周。18周末检测大鼠血糖、血脂、肝功、肝脏脂质定量、肝脏指数、胰岛素抵抗指数及肝组织学变化,肿瘤坏死因子-α(TNF-α)在肝组织的蛋白表达。结果二甲双胍可以显著降低血糖、血脂、肝脏脂质量,增加胰岛素敏感性,改善肝脏脂肪变程度,降低TNF-α在肝组织的蛋白表达。结论二甲双胍对2型糖尿病合并脂肪肝大鼠的肝脏脂肪变性具有一定的治疗意义。     

高脂饮食对大鼠脂肪肝胰岛素受体底物1、2表达的影响

目的探讨高脂饮食对大鼠脂肪肝胰岛素受体底物（IRS）1、IRS-2分子表达的影响。方法对8周高脂饲养（n=7）和正常饲养（n=7）大鼠的胰岛素敏感性、脂质代谢、肿瘤坏死因子（TNF）α以及肝脏IRS-1、IRS-2的mRNA和蛋白表达进行检测。结果与正常组比，高脂组大鼠呈现胰岛素抵抗，游离脂肪酸（FFA）和TNF-α增高；肝脏的IRS-1 mRNA和蛋白含量分别降低了28％和32％（P〈0．01），IRS-2mRNA和蛋白含量分别降低了30％和27％（P〈0．01）。结论高脂饮食导致大鼠肝脏IRS-1、IRS-2的mRNA和蛋白含量明显降低，这可能是高脂饮食引起脂肪变肝脏胰岛素抵抗的重要分子机制。     

高脂高糖诱导的非酒精性脂肪肝小鼠肝胰岛素底物受体-2的表达

目的探讨非酒精性脂肪肝小鼠肝脏胰岛素受体底物-2(insulin receptor substrate-2,IRS-2)的表达。方法于2002年1月至2003年3月在中国医科大学附属盛京医院内分泌科将30只6~8周龄的雌性SPF级C57BL6J小鼠随机分为3组饲养,其中正常饮食组10只、高脂高糖饮食8周组10只(实验过程中死亡5只)及高脂高糖食16周组10只。测定其血脂、血清胰岛素及空腹血糖、肝功能、肝脏重量、腹部瘦肉及瘦肉含量,肝脏脂质含量及IRS-2蛋白表达。并做肝脏病理检查。结果高脂高糖饮食16周的小鼠腹部脂肪量、血清甘油三酯、空腹血糖、碱性磷酸酶显著高于正常饮食小鼠(P<0.05~0.01);白蛋白及白蛋白球蛋白比值较正常饮食小鼠显著降低(P<0.05)。高脂高糖饮食小鼠肝脏有明显脂肪变性,16周的小鼠较8周的小鼠严重。高脂高糖饮食小鼠的肝脏总胆固醇和甘油三酯含量显著高于正常饮食小鼠,16周时达到正常饮食小鼠的2倍以上;IRS-2表达低于正常饮食小鼠,16周的小鼠高于8周的。结论高脂高糖饮食可使C57BL6J小鼠出现非酒精性脂肪肝,肝脏的IRS-2蛋白表达量低于正常饮食小鼠     

罗格列酮对高糖高脂饮食诱导糖尿病大鼠脂肪肝的治疗作用

目的探讨罗格列酮（RSG）对2型糖尿病（T2DM）大鼠合并脂肪肝的治疗作用。方法建立T2DM合并脂肪肝的大鼠模型，以RSG进行干预治疗。18周末留取肝组织及血清，检测比较各组血糖、血脂、胰岛素、谷丙转氨酶、TNF-α、瘦素等指标及肝脏病理变化和TNF-α在肝脏的表达。结果RSG治疗组大鼠肝脏结构较糖尿病对照组明显好转，TNF-α在肝脏的表达水平降低（P〈0.05），TG、TC、瘦素水平均明显下降（P〈0.05，P〈0.01，P〈0.05）。结论RSG对实验性糖尿病大鼠合并脂肪肝具有一定的治疗作用。     

吡格列酮治疗大鼠非酒精性脂肪性肝病的研究

目的观察胰岛素抵抗与非酒精性脂肪性肝病的关系,探讨吡格列酮对高脂喂养诱导大鼠脂肪肝的治疗作用。方法32只雄性SD大鼠分为3组：对照组(11只)喂饲普通饮食16周,模型组(10只)喂饲高脂饮食16周,药物组(11只)为高脂饮食喂饲8周后改为高脂饮食同时加用吡格列酮4 mg·kg~(-1)·d~(-1)灌胃8周。16周末处死大鼠,称量肝湿重及体重,测定丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、碱性磷酸酶(ALP)和血糖水平。用放射免疫法测定空腹胰岛素和肿瘤坏死因子(TNF)-α,计算胰岛素抵抗指数(HOMA-IR),并观察肝脏病理变化。结果16周末,模型组肝脏出现明显肝细胞脂肪变性和肝小叶炎症,肝指数、ALT、AST、ALP、胰岛素水平及HOMA-IR均高于对照组(P＜0．05)；药物组和模型组相比,脂肪变性和肝小叶炎症明显减轻,肝指数、ALT、ALP、TNF-α和HOMA- IR均降低(P＜0．05),胰岛素水平有降低趋势。结论高脂喂养脂肪肝大鼠随病变进展可能存在胰岛素抵抗。吡格列酮治疗可以改善或延缓大鼠脂肪肝病变程度。     

罗格列酮对2型糖尿病并发非酒精性脂肪肝大鼠肝脏解偶联蛋白-2表达的影响

目的观察罗格列酮对2型糖尿病（T2DM）并发非酒精性脂肪肝（NAFLD）大鼠肝脏解偶联蛋白-2（UCP-2）表达的影响,探讨其防治T2DM并发NAFLD的部分作用机制。方法高脂高糖饮食结合小剂量链脲佐菌素（STZ）腹腔注射建立T2DM并发NAFLD大鼠模型,将成模大鼠随机分为模型组、罗格列酮治疗组,每组各16只,并设立正常对照组。治疗组给予罗格列酮3 mg/（kg.d）灌胃治疗。于实验第16（治疗后8周）、20周（治疗后12周）末分批处死大鼠,检测肝功能、空腹血糖、血脂和血清胰岛素水平,光镜下观察大鼠肝脏组织学形态,分别以免疫组织化学法和逆转录-聚合酶链反应（RT-PCR）法检测肝组织UCP-2蛋白和UCP-2mRNA的表达情况。结果与正常组相比,模型组大鼠血清转氨酶、空腹血糖、血清胰岛素及血脂水平均明显升高,胰岛素敏感指数明显下降（P均〈0.01）,肝脏出现不同程度脂肪变性,肝组织UCP-2蛋白和UCP-2 mRNA表达增高,以20周末最为明显。在16周末（治疗后8周）和20周末（治疗后12周）,治疗组大鼠血清转氨酶、血糖、血清胰岛素及血脂水平均有改善,胰岛素敏感指数明显升高,肝细胞脂肪变性明显减轻,肝组织UCP-2蛋白和UCP-2 mRNA表达明显下降,与模型组比较差异均有统计学意义（P〈0.01）。结论 T2DM并发NAFLD大鼠肝组织UCP-2表达增强,罗格列酮可以调控T2DM并发NAFLD大鼠肝脏UCP-2 mRNA和UCP-2蛋白的适度表达,这可能是其治疗T2DM并发NAFLD的分子机制之一。     

酒精性脂肪肝大鼠肝组织骨桥蛋白表达的变化

目的探讨乙醇联合高脂饮食诱导脂肪肝大鼠肝组织骨桥蛋白（OPN）表达的变化。方法采用高脂饮食联合乙醇灌胃制备大鼠脂肪肝模型。在第6周末分别取血和肝组织,常规检测血生化指标和采用Westernblot法检测大鼠肝组织骨桥蛋白表达的改变。结果乙醇联合高脂饮食条件下大鼠血清ALT、AST、TG、TC水平升高（P〈0．05）,HDL-C降低（P〈0．05）;油红O染色结果显示乙醇联合高脂饮食组大鼠肝脏脂肪变明显;模型组大鼠肝组织OPN蛋白表达明显增强（P〈0．05）。结论骨桥蛋白表达增强在大鼠乙醇性脂肪肝的发病过程中起一定的作用。     

高脂饮食对实验性大鼠非酒精性脂肪肝UCP2表达的影响

目的:通过高脂饮食制作非酒精性脂肪肝(NAFLD)动物模型;观察高脂饮食对实验性大鼠NAFLD解偶联蛋白2(UCP2)的影响。方法:通过高脂饮食建立大鼠非酒精性脂肪性肝病模型。观察肝脏病理改变并检测肝脏UCP2表达情况。结果:与正常对照组相比,高脂饮食大鼠肝脏UCP2表达显著上升,肝组织广泛脂肪变性,形成单纯性脂肪肝。继续给予高脂饮食使肝脏UCP2表达水平进一步增加,肝脏发生进一步病理改变而形成脂肪性肝炎。结论:高脂饮食成功地复制了NAFLD动物模型;NAFLD时肝脏UCP2表达上调,可能是机体的一种适应性反应;但是UCP2过度表达,可能诱导或加剧肝脏病理改变。     

吡格列酮对大鼠非酒精性脂肪性肝炎干预的实验研究

目的　用吡格列酮对试验动物进行干预 ,探讨胰岛素抵抗与非酒精性脂肪性肝炎的关系。方法　雄性SD大鼠 5 0只随机分组。正常对照组喂饲普通饮食 ,高脂模型组喂饲高脂饮食 ,给药组分别在喂饲高脂饮食 4周后、9周后分别给予吡格列酮灌胃 ,并分别在 9周末及 14周末处死各组大鼠 ,测定血清转氨酶、血糖、血脂、胰岛素及FFA水平 ,观察肝组织学改变。结果　 9周后模型组大鼠呈现明显腹型肥胖 ,产生脂质代谢紊乱及胰岛素抵抗 ,肝脏出现肝细胞脂肪变性 ,给药组各项指标均有明显改善 ;14周后模型组大鼠血清转氨酶水平升高 ,肝脏出现炎细胞浸润 ,除肝组织炎症外 ,给药组其他指标均有不同程度的改善。结论　胰岛素抵抗在大鼠肝脏脂肪变的发生中起到关键作用 ,与脂肪肝发生炎症坏死关系甚密切 :吡格列酮可明显改善高脂饮食诱发大鼠肝脏的脂肪变性 ,对炎症变化作用不明显     

蜕皮甾酮对2型糖尿病大鼠肝细胞IR S-2蛋白表达的影响

目的：探讨蜕皮甾酮对2型糖尿病大鼠肝细胞胰岛素受体底物2（ IRS-2）蛋白表达的影响。方法选择42只大鼠,14只作为正常组,常规饲料喂养,余下28只先以高脂喂养加小剂量STZ注射诱导2型糖尿病大鼠模型,造模成功后,随机分为模型组和治疗组,分别给予高脂饲料喂养、高脂饲料喂养＋蜕皮甾酮灌胃治疗。5周后取大鼠肝组织标本,应用免疫组化和RT-PCR技术检测IRS-2蛋白和mRNA含量。结果正常组和治疗组肝细胞IRS-2蛋白及mRNA含量均明显高于模型组（ P＜0．05或＜0．01）。结论蜕皮甾酮可提高2型糖尿病大鼠肝细胞IRS-2蛋白和mRNA含量,这可能是蜕皮甾酮改善2型糖尿病大鼠胰岛素抵抗的机制之一。     

二甲双胍对胰岛素抵抗大鼠脂肪肝的影响

目的探讨二甲双胍对胰岛素抵抗大鼠脂肪肝的影响。方法雄性wistar大鼠31只，随机分为正常组（n=8）和实验组（n=23）。正常组普通饮食，实验组高脂饮食，8周末从实验组中随机抽取7只证实造模成功。实验组再分为二甲双胍组（n=8）和模型对照组（n=8），6周后，胰岛素敏感性用正常血糖高胰岛素钳夹技术稳态时的葡萄糖输注率来评价；氨基转移酶、甘油三酯（TG）、游离脂肪酸（FFA）等以生物化学方法测定；胰岛素和肿瘤坏死因子α（TNFα）分别用放射免疫法和酶联免疫吸附法测定；肝细胞脂肪变性和肝脏的炎症程度在光学显微镜下评估。结果与模型对照组相比，二甲双胍组大鼠胰岛素抵抗明显改善，肝指数、附睾脂肪比重、体重明显下降，氨基转移酶、TG、FFA、TNFα明显降低，肝细胞脂肪变性和肝脏炎症情况有不同程度的改善。结论二甲双胍能够明显改善高脂饲养诱发脂肪肝大鼠的胰岛素抵抗和肝脏组织学，它可能成为治疗脂肪肝的新药物。     

The effect of leptin, tumor necrosis factor-alpha (TNF-alpha), and nitric oxide (NO) production on insulin resistance in Otsuka Long-Evans fatty rats

Adipocytokines and nitric oxide (NO) play important roles in type 2 diabetes; however, the regulatory mechanism has not been fully clarified. To investigate the role of adipocytokines and NO production on insulin resistance in type 2 diabetes, the LETO rats and the OLETF rats were fed a control diet or a high-fat diet for 4 weeks. After 4 weeks the blood levels of leptin, tumor necrosis factor-alpha (TNF-alpha), and NO were measured. As an indicator of insulin resistance, the homeostasis model assessment for insulin resistance (HOMA-R) was applied. Food intake in high-fat diet group rats was lower than in control diet group rats. The high fat diet increased body weight (BW), but did not significantly affect the HOMA-R and blood pressure (BP). Leptin and TNF-alpha levels were significantly higher in the OLETF rats than in the LETO rats, while NO levels did not change between the two groups. The high-fat diet elevated blood leptin levels, but not TNF-alpha and NO levels. The HOMA-R in the OLETF rats was correlated with leptin, but not with BP, BW, TNF-alpha or NO. NO showed an inverse correlation with BP. In conclusion, leptin, TNF-alpha, and NO may each regulate insulin sensitivity through their own unique pathways. The elucidation of the regulatory mechanism of adipocytokines and NO may give a clue to clarify the pathophysiology of insulin resistance.     

Caloric restriction in obese pre-diabetic rats prevents beta-cell depletion,loss of beta-cell GLUT 2 and glucose incompetence

Summary Pre-diabetic male Zucker diabetic fatty rats (ZDF) become diabetic between 8 and 10 weeks of age. At that time their beta cells exhibit high basal insulin secretion, absent insulin response to glucose and loss of GLUT 2 glucose transporter. Beta-cell volume, which is increased at the onset of non-insulin-dependent diabetes, declines precipitously by age 18 weeks. To determine if expression of this diabetic phenotype was dependent upon the increased food intake of these rats, they were diet-matched to lean littermates for 12 weeks beginning at 6 weeks of age. Untreated control ZDF rats received an unrestricted diet for 3 months. All of the controls became hyperglycaemic by 8 weeks of age, whereas all diet-matched rats remained euglycaemic throughout the 3 months, despite the fact that at 18 weeks of age their mean body weight equaled that of obese rats on an unrestricted diet. In the former rats glucose-stimulated insulin secretion was absent at 12 weeks of age and GLUT-2-positive beta cells had fallen below 30%. The volume fraction of their beta cells was 2.6 times normal at this age but by 18 weeks of age it had declined by 75%. Diet restriction for 3 months prevented the loss of glucose-stimulated insulin secretion and the reduction of beta-cell GLUT-2 and beta-cell volume fraction. However, neither the elevated basal insulin secretion nor the exaggerated arginine-stimulated insulin secretion of the obese rats was reversed or prevented by caloric restriction. We conclude that in diabetic ZDF rats the glucose incompetence of beta cells and the reduction of beta-cell GLUT 2, which coincide with the onset of hyperglycaemia, and the subsequent loss of beta-cell volume, occur only when the caloric intake is excessive. The increased basal insulin secretion and exaggerated insulin response to arginine appear to be relatively independent of caloric intake.Abbreviations NIDDM Non-insulin-dependent diabetes mellitus - ZDF Zucker diabetic fatty rats - GSIS glucose-stimulated insulin secretion - GLUT-2 glucose transporter     

Analysis of gene expression profiles in insulin-sensitive tissues from pre-diabetic and diabetic Zucker diabetic fatty rats

Insulin resistance occurs early in the disease process, preceding the development of type 2 diabetes. Therefore, the identification of molecules that contribute to insulin resistance and leading up to type 2 diabetes is important to elucidate the molecular pathogenesis of the disease. To this end, we characterized gene expression profiles from insulin-sensitive tissues, including adipose tissue, skeletal muscle, and liver tissue of Zucker diabetic fatty (ZDF) rats, a well characterized type 2 diabetes animal model. Gene expression profiles from ZDF rats at 6 weeks (pre-diabetes), 12 weeks (diabetes), and 20 weeks (late-stage diabetes) were compared with age- and sex-matched Zucker lean control (ZLC) rats using 5000 cDNA chips. Differentially regulated genes demonstrating > 1.3-fold change at age were identified and categorized through hierarchical clustering analysis. Our results showed that while expression of lipolytic genes was elevated in adipose tissue of diabetic ZDF rats at 12 weeks of age, expression of lipogenic genes was decreased in liver but increased in skeletal muscle of 12 week old diabetic ZDF rats. These results suggest that impairment of hepatic lipogenesis accompanied with the reduced lipogenesis of adipose tissue may contribute to development of diabetes in ZDF rats by increasing lipogenesis in skeletal muscle. Moreover, expression of antioxidant defense genes was decreased in the liver of 12-week old diabetic ZDF rats as well as in the adipose tissue of ZDF rats both at 6 and 12 weeks of age. Cytochrome P450 (CYP) genes were also significantly reduced in 12 week old diabetic liver of ZDF rats. Genes involved in glucose utilization were downregulated in skeletal muscle of diabetic ZDF rats, and the hepatic gluconeogenic gene was upregulated in diabetic ZDF rats. Genes commonly expressed in all three tissue types were also observed. These profilings might provide better fundamental understanding of insulin resistance and development of type 2 diabetes.     

Effect of early insulin therapy on nuclear factor κB and cytokine gene expressions in the liver and skeletal muscle of high-fat diet,streptozotocin-treated diabetic rats

To clarify the effect of early insulin therapy on nuclear factor kappaB (NFkappaB) pathway and inflammatory cytokine responses in the liver and skeletal muscle in type 2 diabetes. High-fat diet and low dose streptozotocin (STZ) induced diabetic rats were given NPH insulin or gliclazide for 3 weeks initiated at the 3rd day after STZ injection as early treatment and NPH for 3 weeks at 1 month as late treatment. Intraperitoneal glucose tolerance test (IPGTT) was performed at 3rd day after the end of treatment. Early interventions caused a decrease in glucose-insulin index in IPGTT, promoted glucose transporter 4 (Glut4) gene and protein expressions in muscle and reduced phosphoenolpyruvate carboxykinase (PEPCK) protein levels in the liver. There was an increase in inhibitor kappaB (IkappaBalpha) protein and a decrease in NFkappaB p65 DNA binding activity. A decreased level in mRNAs encoding tumor necrosis factor (TNF)alpha in the liver and muscle and interleukin (IL)-1beta in the liver were observed. Our results suggested that early insulin treatment inhibits NFkappaB activity and inflammatory cytokine responses in the liver and skeletal muscle that were involved in the amelioration of insulin resistance in type 2 diabetic rats.     

Ursodeoxycholic acid improves insulin sensitivity and hepatic steatosis by inducing the excretion of hepatic lipids in high-fat diet-fed KK-Ay mice

Type 2 diabetes mellitus is frequently accompanied by fatty liver/nonalcoholic fatty liver disease. Hence, accumulation of lipids in the liver is considered to be one of the risk factors for insulin resistance and metabolic syndrome. Ursodeoxycholic acid (UDCA) is widely used for the treatment of liver dysfunction. We investigated the therapeutic effects of UDCA on type 2 diabetes mellitus exacerbating hepatic steatosis and the underlying mechanisms of its action using KK-A(y) mice fed a high-fat diet. KK-A(y) mice were prefed a high-fat diet; and 50, 150, and 450 mg/kg of UDCA was orally administered for 2 or 3 weeks. Administration of UDCA decreased fasting hyperglycemia and hyperinsulinemia. Hyperinsulinemic-euglycemic clamp analyses showed that UDCA improved hepatic (but not peripheral) insulin resistance. Hepatic triglyceride and cholesterol contents were significantly reduced by treatment with UDCA, although the genes involved in the synthesis of fatty acids and cholesterol, including fatty acid synthase and 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, were upregulated. Fecal levels of bile acids, neutral sterols, fatty acids, and phospholipids were significantly increased by UDCA treatment. The gene expression levels and protein phosphorylation levels of endoplasmic reticulum stress markers were not changed by UDCA treatment. These results indicate that UDCA ameliorates hyperglycemia and hyperinsulinemia by improving hepatic insulin resistance and steatosis in high-fat diet-fed KK-A(y) mice. Reduction of hepatic lipids might be due to their excretion in feces, followed by enhanced utilization of glucose for the synthesis of fatty acids and cholesterol. Ursodeoxycholic acid should be effective for the treatment of type 2 diabetes mellitus accompanying hepatic steatosis.     

脂联素与非酒精性脂肪肝病

脂联素(adiponectin,ADP)作为一种新发现的脂肪激素,主要是由脂肪细胞分泌的,在肥胖者、糖尿病及非酒精性脂肪性肝病(nonalcoholic fatty liver disease,NAFLD)患者中血清ADP水平低于正常.ADP有胰岛素增敏作用,与胰岛素抵抗相关;能够使肝脏肿瘤坏死因子(tumor necrosis factor-alpha,TNF-α)分泌下降,减少肝中脂肪堆积和炎症介质作用,对ADP的研究将为探讨NAFLD的发病机制及治疗方案提供新的线索.     

TNF-α与2型糖尿病合并非酒精性脂肪肝胰岛素抵抗及动脉粥样硬化的关系

目的探讨2型糖尿病（T2DM）合并非酒精性脂肪肝（NAFL）患者肿瘤坏死因子-α（TNF-α）与胰岛素抵抗（IR）及颈动脉内膜-中层厚度（CIMT）的相关性。方法将192例T2DM患者根据是否合并NAFL进行分组并与正常对照组比较。采用多因素相关分析和回归分析评价TNF-α对IR及CIMT的影响。结果与正常对照组及T2DM无NAFL组相比,T2DM合并NAFL患者TNF-α显著升高（P〈0．01）;多因素相关分析和回归分析显示,TNF-α与IR（r=0．525,P〈0．01）及CIMT（r=0．548,P〈0．01）呈显著正相关。结论T2DM合并NAFL患者TNF-α与IR及CIMT密切相关,参与了动脉粥样硬化的形成。