PPARγ基因修饰对非酒精性脂肪性肝纤维化的防治机制
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摘要
目的:非酒性脂肪性肝病(non-alcoholic fatty liver disease, NAFLD)是代谢综合征的重要危险因素之一,主要包括单纯性脂肪肝、脂肪性肝炎、脂肪性肝纤维化和肝硬化等一系列病变过程。非酒精性脂肪性肝炎(non-alcoholic steatohepatitis, NASH)是NAFLD病变过程的重要阶段,以肝细胞脂肪变性伴有坏死性炎症为主要特征,进一步进展可导致肝纤维化和肝硬化,近年研究证实NASH亦是肝细胞癌的致病因素之一。过氧化物增殖酶体激活物受体γ(peroxisome proliferator activated receptor gamma, PPARγ)为核激素受体超家族成员,是促脂肪细胞分化的细胞核转录因子之一,其经配体激活后调节体内糖、脂代谢,对细胞生长、分化、炎症及凋亡有重要影响。肝脏发生脂肪变时,胰岛素抵抗的程度随脂肪性肝炎及纤维化的发展逐渐加重,PPARγ的外源性调节可能有助于改善糖和脂肪代谢,延缓或阻止脂肪性肝炎、肝纤维化的发生及进展。本研究旨在探讨选择性的PPARγ基因导入治疗和PPARγ激动剂在非酒精性脂肪性肝炎、肝纤维化中的靶向性治疗效果,为临床有效防治非酒性脂肪性肝炎、肝纤维化提供新的途径和理论依据。
方法:采用高脂、胆碱-蛋氨酸缺乏饮食(high fat, methionine and choline deficient diet, MCD)建立小鼠非酒精性脂肪性肝纤维化模型,分别应用PPARγ拮抗剂、激动剂及/或腺病毒载体PPARγ基因导入(Ad-PPARγ)及β-半乳糖苷酶转基因腺病毒(Ad-LacZ)进行干预实验,并以胆碱-蛋氨酸充足饮食设立对照组。血清丙氨酸氨基转移酶(alanine aminotransferase, ALT)及甘油三酯(triglyceride, TG)采用日本Olympus AU 2700全自动生化分析仪酶法测定。肝组织切片采用HE染色观察肝脂肪变、炎症活动及纤维化程度;Masson染色观察肝脏纤维化程度。PPARγ、转化生长因子β1(transforming growth factor beta 1,TGFβ1)、α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)及间质金属蛋白酶-2(Matrix Metalloproteinase-2, MMP-2)、肿瘤坏死因子-α(Tumor necrosis factor-α, TNF-α)mRNA和蛋白表达分别采用RT-PCR、western blot及免疫组织化学检测。实验结果采用SPSS13.0单因素方差分析进行统计学处理,组间比较采用Student- Newman-Keuls分析。
结果:MCD饮食小鼠可形成典型脂肪性肝纤维化模型,PPARγ激动剂及/或腺病毒载体基因导入可显著改善肝细胞脂肪变性、坏死性炎症及纤维化程度,尤以PPARγ激动剂联合腺病毒载体PPARγ基因导入组为著。反之,PPARγ拮抗剂GW9662则加重肝细胞脂变、炎症坏死及肝纤维化程度。TGFβ1、α-SMA、MMP-2及TNF-αmRNA及蛋白表达随PPARγ表达增强及肝脏纤维化程度的减轻而改善(P<0.05)。
1实验动物一般情况对照组小鼠活泼、毛发有光泽,体重及肝重随造模时间逐渐增加,模型组及干预组小鼠体重及肝重则明显降低(P<0.05),各组间肝指数并无显著差异(P>0.05)(Table2)。
2实验小鼠血清生化学改变MCD组、Ad-LacZ组血清谷丙转氨酶(alanine aminotransferase, ALT)水平与对照组相比显著升高,罗格列酮组、Ad-PPARγ组及罗格列酮与Ad-PPARγ联合组ALT水平较模型组降低,尤以联合治疗组为著;PPARγ阻滞剂GW9662组ALT水平高于模型组。模型组及各干预组血清甘油三酯(triglyceride, TG)水平均显著高于对照组,P<0.05。
3实验小鼠肝组织病理学变化肝组织切片HE及Masson染色显示,MCD饮食喂养小鼠及Ad-LacZ基因导入小鼠可见重度肝细胞大泡性脂肪变性、炎细胞浸润、纤维化形成。罗格列酮组、Ad-PPARγ组及罗格列酮与Ad-PPARγ联合组肝细胞脂肪变、炎症及纤维化均明显减轻,尤以罗格列酮与Ad-PPARγ联合组肝损伤改善明显。GW9662组小鼠上述肝损伤较模型组加重(Fig.1,Fig.2)。
4 PPARγ基因调控对肝内促炎及促纤维化因子表达的作用MCD饮食喂养小鼠PPARγmRNA及蛋白的表达显著低于对照组(P<0.05);应用PPARγ激动剂罗格列酮及Ad- PPARγ导入后,小鼠肝组织PPARγmRNA及蛋白的表达显著增强,尤其联合应用罗格列酮及Ad-PPARγ导入组明显,伴随炎症因子TNF-α、促纤维化形成因子α-SMA,TGFβ1,MMP-2的表达下调。而PPARγ阻滞剂GW9662干预组呈现相反趋势。肝组织切片免疫组织化学染色显示:TNF-α、α-SMA,TGFβ1及MMP-2表达与western blot呈一致趋势,TNF-α主要表达于炎症活动区的肝窦壁细胞及炎细胞;TGFβ1、MMP2及α-SMA主要表达于炎症及纤维化明显区域,TGFβ1主要表达于星状细胞(hepatic stellate cell, HSC)及Kupffer细胞胞浆,MMP-2主要表达于纤维间隔内及汇管区的间质细胞胞浆,α-SMA主要表达于肝窦周隙HSCs胞浆,并与肝纤维化程度密切相关。
结论:PPARγ是调节肝脏脂质代谢、炎症及纤维化形成的关键因子,PPARγ靶向激活或基因导入可显著改善MCD饮食诱导的肝损伤,包括肝脂肪变、炎症和肝纤维化,提示PPARγ靶向基因调控可能为非酒精性脂肪性肝纤维化的治疗策略之一。
Objective: Non-alcoholic fatty liver disease (NAFLD) is a common liver disease. NAFLD and related metabolic disorder are increasing in recent years. As a hepatic manifestion of metabolic syndrome, NAFLD is closely associated with multiple factors such as obesity, hyperlipidemia, type 2 diabetes mullitus and hypertension. Nonalcoholic steatohepatitis (NASH), including hepatic steatosis, necrotic inflammation and fibrosis, is the key stage from simple steatosis to liver cirrhosis. NAFLD has actually been shown to be one of the etiologies in those patients with cryptogenic cirrhosis. Up to now, the mechanism of NASH was unclear. Peroxisome proliferator activated receptor gamma (PPARγ), one of nuclear receptor superfamily of transcription factors, is a key regulator of adipogenesis, inflammatory infiltration and fibrogenesis. This study aimed to elucidate the effect of targeted genetic modulation of PPARγon nutritional fibrotic steatohepatitis in mice.
Methods: Forty two healthy 7 weeks old male C57BL/6J mice were fed with methionine-choline supplemented diet for one week. Then they were randomly divided into seven groups: control group were fed with methionine-choline supplemented diet (MCD+) for eight weeks. Model group were fed a methionine-choline deficient (MCD) diet for eight weeks to induce fibrotic steatohepatitis. PPARγagonist rosiglitazone (50mg/kg/d) orally (n=6) and/or adenovirus-PPARγ(1×1010v.p, three time/week, p.i.), adenovirus-LacZ (1×1010vp,three time/week, p.i.), antagonist 2-chloro-5-nitrobenzaniliden (GW9662) (1mg/kg,three time/week, p.i.) were supplement simultaneous with MCD diet for 8 weeks, respectively. After 8 weeks, all rats were sacrificed after 12h of fasting. Blood samples were collected for biochemical assays. The liver was removed, and sampled for histological study and RNA extraction. Liver weight, body weight, liver index, serum activitis of alanine aminotransferase (ALT) and liver histology of mice of all groups were assayed. The grade of hepatic steatosis, inflammation and fibrosi were observed by Hematoxylin and eosin, and Masson trichromatism stained paraffin-embedded liver tissues isolated from mice. Effect of PPARγregulation was assessed by comparing severity of hepatic fibrosis in the liver sections, the mRNA and protein expression of hepatic inflammatory and fibrogenetic factors,α-SMA, TGFβ1, and MMP2.
Results:
1 The common behavior of mice: control mice were active, whose hair was bright and weight increased gradually. Body weight of model and intervention mice decreased remarkably compared to control group. The liver weight in model group and treatment group were notedly decreased compared with control group (both P<0.05). There was no significant difference in liver indexs among the seven groups (P>0.05). (Table2)
2 Measurement of serum ALT and triglyceride: mice given the MCD diet and adenovirus-LacZ group showed significantly higher serum ALT level and triglyceride content compared with control mice, indicating hepatic injury, and a significant reduction was noticed after rosiglitazone and/or adenovirus-PPARγtreatment, especially in combinated rosiglitazone and adenovirus-PPARγtreatment group. Conversely, the GW9662 group showed the highest ALT level. However, serum triglyceride and cholesterol level was on significant altered except the control group (Table3).
3 Liver histopathology showed that MCD fed mice and adenovirus-LacZ injected mice presented severe hepatic macrovesicular steatosis, inflammatory infiltratory and fibrosis. Severer liver injury was presented in giving GW9662 animals. Ad-PPARγor rosiglitazone administration could ameliorate hepatic steatosis, necrotic inflammation and progressive fibrosis. Further more, combination of rosiglitazone and Ad-PPARγshowed an additive effect on Ad-PPARγup-regulation and protection against liver injury.
4 Effect of PPARγgene modulation on expression of inflammatory and fibrotic factors: PPARγmRNA and protein expression were up-regulated by given rosiglitazone and adenovirus-PPARγ, further enhanced by combinated rosiglitazone plus adenovirus-PPARγ, which companied with blunting of the pro-inflammatory and pro-fibrosis factors TNF-α,α-SMA, TGFβ1 and MMP2 mRNA and protein expression. Contrarily, PPARγexpression was down-regulated by given GW9662 (P<0.05). By immunochemical staning, the expression of TNFαwas located in inflammatory area and endotheliocyte of hepatic sinus.α-SMA, TGFβ1, MMP2 was expressed in inflammatory and fibrogenesis area.
Conclusions: The present study provided a clear morphological and molecular biological evidence for the protective role of up-regulated PPARγin ameliorating heptic steatosis, inflammation and fibrosis in experimental nutritional steatohepatitis and fibrosis. Targeted genetic modulation of PPARγmight serve as a new approach for therapy of fibrotic steatohepatitis.
方法:采用高脂、胆碱-蛋氨酸缺乏饮食(high fat, methionine and choline deficient diet, MCD)建立小鼠非酒精性脂肪性肝纤维化模型,分别应用PPARγ拮抗剂、激动剂及/或腺病毒载体PPARγ基因导入(Ad-PPARγ)及β-半乳糖苷酶转基因腺病毒(Ad-LacZ)进行干预实验,并以胆碱-蛋氨酸充足饮食设立对照组。血清丙氨酸氨基转移酶(alanine aminotransferase, ALT)及甘油三酯(triglyceride, TG)采用日本Olympus AU 2700全自动生化分析仪酶法测定。肝组织切片采用HE染色观察肝脂肪变、炎症活动及纤维化程度;Masson染色观察肝脏纤维化程度。PPARγ、转化生长因子β1(transforming growth factor beta 1,TGFβ1)、α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)及间质金属蛋白酶-2(Matrix Metalloproteinase-2, MMP-2)、肿瘤坏死因子-α(Tumor necrosis factor-α, TNF-α)mRNA和蛋白表达分别采用RT-PCR、western blot及免疫组织化学检测。实验结果采用SPSS13.0单因素方差分析进行统计学处理,组间比较采用Student- Newman-Keuls分析。
结果:MCD饮食小鼠可形成典型脂肪性肝纤维化模型,PPARγ激动剂及/或腺病毒载体基因导入可显著改善肝细胞脂肪变性、坏死性炎症及纤维化程度,尤以PPARγ激动剂联合腺病毒载体PPARγ基因导入组为著。反之,PPARγ拮抗剂GW9662则加重肝细胞脂变、炎症坏死及肝纤维化程度。TGFβ1、α-SMA、MMP-2及TNF-αmRNA及蛋白表达随PPARγ表达增强及肝脏纤维化程度的减轻而改善(P<0.05)。
1实验动物一般情况对照组小鼠活泼、毛发有光泽,体重及肝重随造模时间逐渐增加,模型组及干预组小鼠体重及肝重则明显降低(P<0.05),各组间肝指数并无显著差异(P>0.05)(Table2)。
2实验小鼠血清生化学改变MCD组、Ad-LacZ组血清谷丙转氨酶(alanine aminotransferase, ALT)水平与对照组相比显著升高,罗格列酮组、Ad-PPARγ组及罗格列酮与Ad-PPARγ联合组ALT水平较模型组降低,尤以联合治疗组为著;PPARγ阻滞剂GW9662组ALT水平高于模型组。模型组及各干预组血清甘油三酯(triglyceride, TG)水平均显著高于对照组,P<0.05。
3实验小鼠肝组织病理学变化肝组织切片HE及Masson染色显示,MCD饮食喂养小鼠及Ad-LacZ基因导入小鼠可见重度肝细胞大泡性脂肪变性、炎细胞浸润、纤维化形成。罗格列酮组、Ad-PPARγ组及罗格列酮与Ad-PPARγ联合组肝细胞脂肪变、炎症及纤维化均明显减轻,尤以罗格列酮与Ad-PPARγ联合组肝损伤改善明显。GW9662组小鼠上述肝损伤较模型组加重(Fig.1,Fig.2)。
4 PPARγ基因调控对肝内促炎及促纤维化因子表达的作用MCD饮食喂养小鼠PPARγmRNA及蛋白的表达显著低于对照组(P<0.05);应用PPARγ激动剂罗格列酮及Ad- PPARγ导入后,小鼠肝组织PPARγmRNA及蛋白的表达显著增强,尤其联合应用罗格列酮及Ad-PPARγ导入组明显,伴随炎症因子TNF-α、促纤维化形成因子α-SMA,TGFβ1,MMP-2的表达下调。而PPARγ阻滞剂GW9662干预组呈现相反趋势。肝组织切片免疫组织化学染色显示:TNF-α、α-SMA,TGFβ1及MMP-2表达与western blot呈一致趋势,TNF-α主要表达于炎症活动区的肝窦壁细胞及炎细胞;TGFβ1、MMP2及α-SMA主要表达于炎症及纤维化明显区域,TGFβ1主要表达于星状细胞(hepatic stellate cell, HSC)及Kupffer细胞胞浆,MMP-2主要表达于纤维间隔内及汇管区的间质细胞胞浆,α-SMA主要表达于肝窦周隙HSCs胞浆,并与肝纤维化程度密切相关。
结论:PPARγ是调节肝脏脂质代谢、炎症及纤维化形成的关键因子,PPARγ靶向激活或基因导入可显著改善MCD饮食诱导的肝损伤,包括肝脂肪变、炎症和肝纤维化,提示PPARγ靶向基因调控可能为非酒精性脂肪性肝纤维化的治疗策略之一。
Objective: Non-alcoholic fatty liver disease (NAFLD) is a common liver disease. NAFLD and related metabolic disorder are increasing in recent years. As a hepatic manifestion of metabolic syndrome, NAFLD is closely associated with multiple factors such as obesity, hyperlipidemia, type 2 diabetes mullitus and hypertension. Nonalcoholic steatohepatitis (NASH), including hepatic steatosis, necrotic inflammation and fibrosis, is the key stage from simple steatosis to liver cirrhosis. NAFLD has actually been shown to be one of the etiologies in those patients with cryptogenic cirrhosis. Up to now, the mechanism of NASH was unclear. Peroxisome proliferator activated receptor gamma (PPARγ), one of nuclear receptor superfamily of transcription factors, is a key regulator of adipogenesis, inflammatory infiltration and fibrogenesis. This study aimed to elucidate the effect of targeted genetic modulation of PPARγon nutritional fibrotic steatohepatitis in mice.
Methods: Forty two healthy 7 weeks old male C57BL/6J mice were fed with methionine-choline supplemented diet for one week. Then they were randomly divided into seven groups: control group were fed with methionine-choline supplemented diet (MCD+) for eight weeks. Model group were fed a methionine-choline deficient (MCD) diet for eight weeks to induce fibrotic steatohepatitis. PPARγagonist rosiglitazone (50mg/kg/d) orally (n=6) and/or adenovirus-PPARγ(1×1010v.p, three time/week, p.i.), adenovirus-LacZ (1×1010vp,three time/week, p.i.), antagonist 2-chloro-5-nitrobenzaniliden (GW9662) (1mg/kg,three time/week, p.i.) were supplement simultaneous with MCD diet for 8 weeks, respectively. After 8 weeks, all rats were sacrificed after 12h of fasting. Blood samples were collected for biochemical assays. The liver was removed, and sampled for histological study and RNA extraction. Liver weight, body weight, liver index, serum activitis of alanine aminotransferase (ALT) and liver histology of mice of all groups were assayed. The grade of hepatic steatosis, inflammation and fibrosi were observed by Hematoxylin and eosin, and Masson trichromatism stained paraffin-embedded liver tissues isolated from mice. Effect of PPARγregulation was assessed by comparing severity of hepatic fibrosis in the liver sections, the mRNA and protein expression of hepatic inflammatory and fibrogenetic factors,α-SMA, TGFβ1, and MMP2.
Results:
1 The common behavior of mice: control mice were active, whose hair was bright and weight increased gradually. Body weight of model and intervention mice decreased remarkably compared to control group. The liver weight in model group and treatment group were notedly decreased compared with control group (both P<0.05). There was no significant difference in liver indexs among the seven groups (P>0.05). (Table2)
2 Measurement of serum ALT and triglyceride: mice given the MCD diet and adenovirus-LacZ group showed significantly higher serum ALT level and triglyceride content compared with control mice, indicating hepatic injury, and a significant reduction was noticed after rosiglitazone and/or adenovirus-PPARγtreatment, especially in combinated rosiglitazone and adenovirus-PPARγtreatment group. Conversely, the GW9662 group showed the highest ALT level. However, serum triglyceride and cholesterol level was on significant altered except the control group (Table3).
3 Liver histopathology showed that MCD fed mice and adenovirus-LacZ injected mice presented severe hepatic macrovesicular steatosis, inflammatory infiltratory and fibrosis. Severer liver injury was presented in giving GW9662 animals. Ad-PPARγor rosiglitazone administration could ameliorate hepatic steatosis, necrotic inflammation and progressive fibrosis. Further more, combination of rosiglitazone and Ad-PPARγshowed an additive effect on Ad-PPARγup-regulation and protection against liver injury.
4 Effect of PPARγgene modulation on expression of inflammatory and fibrotic factors: PPARγmRNA and protein expression were up-regulated by given rosiglitazone and adenovirus-PPARγ, further enhanced by combinated rosiglitazone plus adenovirus-PPARγ, which companied with blunting of the pro-inflammatory and pro-fibrosis factors TNF-α,α-SMA, TGFβ1 and MMP2 mRNA and protein expression. Contrarily, PPARγexpression was down-regulated by given GW9662 (P<0.05). By immunochemical staning, the expression of TNFαwas located in inflammatory area and endotheliocyte of hepatic sinus.α-SMA, TGFβ1, MMP2 was expressed in inflammatory and fibrogenesis area.
Conclusions: The present study provided a clear morphological and molecular biological evidence for the protective role of up-regulated PPARγin ameliorating heptic steatosis, inflammation and fibrosis in experimental nutritional steatohepatitis and fibrosis. Targeted genetic modulation of PPARγmight serve as a new approach for therapy of fibrotic steatohepatitis.
引文
1 Yue-Min Nan, Na Fu, Wen-Juan Wu, et al. Rosiglitazone prevents nutritional fibrosis and steatohepatitis in mice. Scandinavian Journal of Gastroenterology, 2009, 44(3): 358-365
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6 Gauthier MS, Couturier K, Latour JG, et al. Concurrent exercise prevents high-fat-diet-induced macrovesicular hepatic steatosis. J Appl Physiol, 2003, 94:2127-2134
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11王战建,张耀,苏杰英等.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.中国糖尿病杂志, 2008,16(9):529-532
12 Dela Pena A, I. Leclercq, J. Field, et al. NF-kappaB activation, rather than TNF, mediates hepatic inflammation in a murine dietary model of steatohepatitis. Gastroenterology, 2005, 129: 1663-1674
13 Gauthier MS, Couturier K, Latour JG, et al. Concurrent exercise prevents high-fat-diet-induced macrovesicular hepatic steatosis. J Appl Physiol, 2003, 94:2127-2134
14 Caldwell SH, Patrie JT, Brunt EM, et al. The effects of 48 weeks of rosiglitazone on hepatocyte mitochondria in human nonalcoholic steatohepatitis. Hepatology, 2007, 46: 1101-1107
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9 Wei S, Kulp SK, Chen CS. Energy restriction as an antitumor target of thiazolidinediones. The Journal of biological chemistry, 2010 Jan 21[Epub ahead of print]
10 Haraguchi G, Kosuge H, Maejima Y, et al. Pioglitazone reduces systematic inflammation and improves mortality in apolipoprotein E knockout mice with sepsis. Journal of intensive care medicine, 2008 Jul, 34(7): 1304-12
11 Sanyal AJ. AGA technical review on nonalcoholic fatty liver disease. Gastroenterology, 2002, 123: 1705-1725
12 Lee JY, Lee HR, et al. Association between serum uric acid and non-alcoholic fatty liver disease in Korean adults. Clin Chem Lab Med, 2009 [Epub ahead of print]
13 Paschos P, Paletas K. Non alcoholic fatty liver disease and metabolic syndrome. Hippokratia, 2009, 13,1: 9-19
14 He W, Barak Y, Hevener A, et al. Adipose-specifid peroxisome proliferator-activate receptorγknockout causes insulin resistance in fat and liber but not in muscle. Proc Natl Acad Sci USA, 2003, 100(26): 15712-15717
15 Begriche K, Igoudjil A, Pessayre D, et al. Mitochondrial dysfunction in NASH: causes, consequences and possible means to prevent it.Mitochondrion, 2006, 6: 1-28
16 Caldwell SH, Patrie JT, Brunt EM, et al. The effects of 48 weeks of rosiglitazone on hepatocyte mitochondria in human nonalcoholic steatohepatitis. Hepatology, 2007, 46: 1101-1107
17 Assy N, Grozovski M, Bersudsky I, et al. Effect of insulin-sensitizing agents in combination with ezetimibe, and valsartan in rats with non-alcoholic fatty liver disease. World J Gastroenterol, 2006, 12: 4369-4376
18 Bouskila M, Pajvani UB, Scherer PE. Adiponectin: a relevant player in PPARγ-agonist-mediated improvements in hepatic insulin sensitivity? Int J Obes Relat Metab Disord 29, 2005, (Suppl. 1): S17 -S23
19 Hauner H. The mode of action of thiazolidinediones. Diabetes Metab Res Rev2002, 18(Suppl. 2): S10-S15
20 Chinetti G, Fruchart JC, Staels B. Peroxisome proliferator-activated receptors (PPARs): nuclear receptors at the crossroads between lipid metabolism and inflammation. Inflamm, 2000, Res49: 497 -505
21 Moller DE, Berger JP. Role of PPARs in the regulation of obesity-related insulin sensitivity and inflammation. Int J Obes Relat Metab Disord27, 2003 (Suppl. 3): S17 -S21
22 Ricote M, Li AC, Willson TM, et al. The peroxisome proliferator-activated receptor-gamma is a negative regulator of macrophage activation. Nature, 1998, 391:79 -82
23 Patrice Dallaire, Kerstin Bellmann, et al. Obese mice lacking inducible nitric oxide synthase are sensitized to the metabolic actions of peroxisome proliferator-activated receptor-gamma agonism. Diabetes, 2008, 57(8): 1999-2011
24 Ogata H, Chinen T, Yoshida T, et al. Loss of SOCS3 in the liver promotes fibrosis by enhancing STAT3-ediated TGF-etal production. Oncogene, 2006, 25(17):2520 - 30
25 Marra F, Efsen E, Romanelli RG,e t al. Ligands of perosisome proliferator-activated receptor gamma modulate profibrogenic andproinflammatory actions in hepatic stellate cell. Gastroenterology, 2000, 116(2): 466-478.
26钟朝辉,郭晏同,周迈等.过氧化物酶题激活物活化的受体γ对肝星状细胞作用机制的研究.中华普通外科杂志, 2006, 21(11): 776-779
27 Yue-Min Nan, Na Fu, Wen-Juan Wu, et al. Rosiglitazone prevents nutritional fibrosis and steatohepatitis in mice. Scandinavian Journal of Gastroenterology, 2009, 44(3):358-365
28 Papai Z ,Feja CN, et al. p53 onconsuppressor as an idicator of overall surival and response to treatment in osteosarcomas. Pathol Ocol Res, 1997, 3(1)15-19
29 Park JK, Ki MR, Lee HR, et al. Vitamin C deficiency attenuates liver fibrosis by way of up-regulated peroxisome proliferator-activated receptor-gamma expression in senescence marker protein 30 knockout mice. Hepatology, 2009 Dec 9[Epub ahead of print]
30 MCKECHNIE NM,KING BC,FLETCHER E,et al.Fas-ligand is stored in secretory lysosomes of ocular barrier epithelia and released with microvesicles. Exp Eye Res, 2006, 83(2): 304-314
2中华医学会肝脏病学分会脂肪肝和酒精性肝病学组.非酒精性脂肪性肝病诊断标准.中华肝脏病杂志. 2005,6(36):364-365
3中华医学会传染病与寄生虫病学会分会、肝病学分会.病毒性肝炎防治方案.中华肝脏病学杂志, 2001, 40: 62~68
4 Brunt EM.Pathology of nonalcoholic steatohepatitis. Hepatol Res, 2005, 33:68–71
5 Dela Pena A, I. Leclercq, J. Field, et al. NF-kappaB activation, rather than TNF, mediates hepatic inflammation in a murine dietary model of steatohepatitis. Gastroenterology, 2005, 129: 1663-1674
6 Gauthier MS, Couturier K, Latour JG, et al. Concurrent exercise prevents high-fat-diet-induced macrovesicular hepatic steatosis. J Appl Physiol, 2003, 94:2127-2134
7 Caldwell SH, Patrie JT, Brunt EM, et al. The effects of 48 weeks of rosiglitazone on hepatocyte mitochondria in human nonalcoholic steatohepatitis. Hepatology, 2007, 46:1101-1107
8范建高,曾民德.脂肪性肝病.第一版,人民卫生出版社,2005:138
9 Feldstein A, Gores GJ. Steatohepatitis and apoptosis: therapeutic implications. Am J Gastroenterol, 2004, 99:1718-1719
10 Crespo J, Cayón A, Fernández-Gil P, et al. Gene expression of tumor necrosis factor alpha and TNF-receptors, p55 and p75, in nonalcoholic steatohepatitis patients. Hepatology,2001,34:1158-1163
11王战建,张耀,苏杰英等.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.中国糖尿病杂志, 2008,16(9):529-532
12 Dela Pena A, I. Leclercq, J. Field, et al. NF-kappaB activation, rather than TNF, mediates hepatic inflammation in a murine dietary model of steatohepatitis. Gastroenterology, 2005, 129: 1663-1674
13 Gauthier MS, Couturier K, Latour JG, et al. Concurrent exercise prevents high-fat-diet-induced macrovesicular hepatic steatosis. J Appl Physiol, 2003, 94:2127-2134
14 Caldwell SH, Patrie JT, Brunt EM, et al. The effects of 48 weeks of rosiglitazone on hepatocyte mitochondria in human nonalcoholic steatohepatitis. Hepatology, 2007, 46: 1101-1107
1 Browning JD, Szczepaniak LS, Dobbins R, et al. Prevalence of hepatic steatosis in an urban population in the United States: impact of ethnicity. Hepatology 2004, 40: 1387-1395
2 Weston SR, Leyden W, Murphy R et al. Racial and ethnic distribution of nonalcoholic fatty liver in persons with newly diagnosed chronic liver disease. Hepatology. 2005, 41: 372-379
3 Larsen TM, Toubro S, Astrup A. PPAR gamma agonists in the treatment of typeⅡdiabetes: is increased fatness commensurate with long-terms efficacy? Interm J Obes, 2003, 27: 147-161
4 Tzameli I, Fang H, Ollero M, et al. Regulated production of a peroxisomeproliferator-activated receptor-γligand during an early phase of adipocyte differentiation in 3T3-L1 adipocytes. Journal of Biological Chemistry, 2004, 279(34): 36093-36102
5 Khan SA, Heuvel JPV. J Nurti Biochem, 2003, 14: 554-567
6 M. Perreault, S. Will, D. Panza, et al. Modulation of nutrient sensing nuclear hormone receptors promotes weight loss through appetite suppression in mice. Diabets, Obesity and Metabolism, 2010 Mar,12(3): 234-45
7 Younossi ZM, Diehl AM, Ong JP. Nonalcoholic fatty liver disease: an agenda for clinical research. Hepatology, 2002, 35(4): 746-75
8 Xia Zhang, howatd A. PPAR and immune system-what do wo know? Int immunopharmacol, 2002, 2 (8): 1029-1044
9 Wei S, Kulp SK, Chen CS. Energy restriction as an antitumor target of thiazolidinediones. The Journal of biological chemistry, 2010 Jan 21[Epub ahead of print]
10 Haraguchi G, Kosuge H, Maejima Y, et al. Pioglitazone reduces systematic inflammation and improves mortality in apolipoprotein E knockout mice with sepsis. Journal of intensive care medicine, 2008 Jul, 34(7): 1304-12
11 Sanyal AJ. AGA technical review on nonalcoholic fatty liver disease. Gastroenterology, 2002, 123: 1705-1725
12 Lee JY, Lee HR, et al. Association between serum uric acid and non-alcoholic fatty liver disease in Korean adults. Clin Chem Lab Med, 2009 [Epub ahead of print]
13 Paschos P, Paletas K. Non alcoholic fatty liver disease and metabolic syndrome. Hippokratia, 2009, 13,1: 9-19
14 He W, Barak Y, Hevener A, et al. Adipose-specifid peroxisome proliferator-activate receptorγknockout causes insulin resistance in fat and liber but not in muscle. Proc Natl Acad Sci USA, 2003, 100(26): 15712-15717
15 Begriche K, Igoudjil A, Pessayre D, et al. Mitochondrial dysfunction in NASH: causes, consequences and possible means to prevent it.Mitochondrion, 2006, 6: 1-28
16 Caldwell SH, Patrie JT, Brunt EM, et al. The effects of 48 weeks of rosiglitazone on hepatocyte mitochondria in human nonalcoholic steatohepatitis. Hepatology, 2007, 46: 1101-1107
17 Assy N, Grozovski M, Bersudsky I, et al. Effect of insulin-sensitizing agents in combination with ezetimibe, and valsartan in rats with non-alcoholic fatty liver disease. World J Gastroenterol, 2006, 12: 4369-4376
18 Bouskila M, Pajvani UB, Scherer PE. Adiponectin: a relevant player in PPARγ-agonist-mediated improvements in hepatic insulin sensitivity? Int J Obes Relat Metab Disord 29, 2005, (Suppl. 1): S17 -S23
19 Hauner H. The mode of action of thiazolidinediones. Diabetes Metab Res Rev2002, 18(Suppl. 2): S10-S15
20 Chinetti G, Fruchart JC, Staels B. Peroxisome proliferator-activated receptors (PPARs): nuclear receptors at the crossroads between lipid metabolism and inflammation. Inflamm, 2000, Res49: 497 -505
21 Moller DE, Berger JP. Role of PPARs in the regulation of obesity-related insulin sensitivity and inflammation. Int J Obes Relat Metab Disord27, 2003 (Suppl. 3): S17 -S21
22 Ricote M, Li AC, Willson TM, et al. The peroxisome proliferator-activated receptor-gamma is a negative regulator of macrophage activation. Nature, 1998, 391:79 -82
23 Patrice Dallaire, Kerstin Bellmann, et al. Obese mice lacking inducible nitric oxide synthase are sensitized to the metabolic actions of peroxisome proliferator-activated receptor-gamma agonism. Diabetes, 2008, 57(8): 1999-2011
24 Ogata H, Chinen T, Yoshida T, et al. Loss of SOCS3 in the liver promotes fibrosis by enhancing STAT3-ediated TGF-etal production. Oncogene, 2006, 25(17):2520 - 30
25 Marra F, Efsen E, Romanelli RG,e t al. Ligands of perosisome proliferator-activated receptor gamma modulate profibrogenic andproinflammatory actions in hepatic stellate cell. Gastroenterology, 2000, 116(2): 466-478.
26钟朝辉,郭晏同,周迈等.过氧化物酶题激活物活化的受体γ对肝星状细胞作用机制的研究.中华普通外科杂志, 2006, 21(11): 776-779
27 Yue-Min Nan, Na Fu, Wen-Juan Wu, et al. Rosiglitazone prevents nutritional fibrosis and steatohepatitis in mice. Scandinavian Journal of Gastroenterology, 2009, 44(3):358-365
28 Papai Z ,Feja CN, et al. p53 onconsuppressor as an idicator of overall surival and response to treatment in osteosarcomas. Pathol Ocol Res, 1997, 3(1)15-19
29 Park JK, Ki MR, Lee HR, et al. Vitamin C deficiency attenuates liver fibrosis by way of up-regulated peroxisome proliferator-activated receptor-gamma expression in senescence marker protein 30 knockout mice. Hepatology, 2009 Dec 9[Epub ahead of print]
30 MCKECHNIE NM,KING BC,FLETCHER E,et al.Fas-ligand is stored in secretory lysosomes of ocular barrier epithelia and released with microvesicles. Exp Eye Res, 2006, 83(2): 304-314