摘要
目的:观察大鼠酒精性肝病组织病理形态学改变,探讨细胞凋亡与细胞色素P4502E1的表达以及和氧化应激的关系.方法:用酒精灌胃法制备酒精性肝病大鼠模型,模型组给予酒精8 g/kg,每天分2次灌胃连续8 wk,对照组给予等量的生理盐水灌胃.实验8 wk末,观察肝组织的病理形态学改变,用原位末端标记法(TUNEL)检测肝细胞凋亡,用免疫组化法检测肝组织中Caspase-3蛋白表达,用全自动生化仪检测ALT和AST的含量,用PCR法测定肝细胞色素P4502E1的基因表达,分别用硫代巴比妥酸法(TBA法)和黄嘌呤氧化酶法测定肝组织丙二醛(MDA)的含量和超氧化物歧化酶(SOD)的活力.结果:模型组凋亡的肝细胞明显增多,主要分布在中央静脉周围、点状和灶状坏死区;Caspase-3主要分布于中央静脉及肝细胞坏死灶周围细胞的胞质中.模型组肝细胞凋亡指数(AI)和Caspase-3蛋白表达强度明显高于对照组(AI:6.2%±1.7% vs 1.7%±0.8%;Caspase-3:66.7% vs 9.5%,P<0.05,P<0.01).CYP2E1表达:对照组c1基因频率为91.6%,c2基因频率为8.4%;模型组c1基因频率为53.4%,c2基因频率为46.6%,均有显著性差异(P<0.05).长期酒精摄入大鼠血清MDA含量增加(41.53±7.43μmol/L vs 15.72±2.06μmol/L,P<0.05),SOD活力下降(353.12±61.02 kU/L vs 636.82±138.60 kU/L,P<0.05),与酒精性肝病肝细胞凋亡程度有相关性(r=0.644,r=-0.511).结论:长期酒精摄入可引起大鼠酒精性肝病及及肝功能损伤,肝细胞凋亡明显增加.CYP2E1基因PstⅠ及RsaⅠRFLPs与酒精性肝病有关,其中c2基因可能与大鼠酒精性肝病的发生有关.MDA含量和SOD活力在酒精性肝病的肝细胞凋亡过程及脂质过氧化反应中发挥重要作用.
AIM： To observe the pathological changes and investigate the relationship between hepatocyte apoptosis, cytochrome P4502E1 （CYP2E1） and oxygen free radicals in alcoholic liver disease （ALD） in rats. METHODS： Ethanol （400 mL/L） was administered at 8 g/kg body weight by gavage twice daily for 8 weeks in the ALD model group, and the control group received isovolumic saline by gavage. By the end of the eighth week, the serum activity of alanine aminotransferase and aspartate aminotransferase were detected by automatic biochemistry analyzer. The pathological changes in the liver were observed by light microscopy of HE-stained sections; hepatocyte apoptosis was detected by the TdT-mediated d-UTP nick end labeling （TUNEL） method; and expression of caspase-3 in the liver was detected by immunohistochemistry. Expression of serum CYP2E1 was determined by polymerase chain reaction, and maleic dialdehyde （MDA） level and superoxide dismutase （SOD） activity in serum were detected by the thiobarbituric acid （TBA） method. RESULTS： TUNEL-positive cells were located around the central vein, spotty and focal necrosis area in the ALD group, and Caspase-3 was observed around the central vein and areas of hepatocellular necrosis. The apoptotic index （AI） and expression of caspase-3 in the ALD group were significantly higher than those in the control group （AI, 6.2% ± 1.7% vs 1.7% ± 0.8%; Caspase-3, 66.7% vs 9.5%, P 〈 0.05, P 〈 0.01）. The allelic frequency of cl and c2 subtypes of CYP2E1 gene in control group was 91.6% and 8.4%, respectively, and 53.4% and 46.6%, respectively, in the ALD group. There were significant differences between the two groups （both P 〈 0.05）. The serum MDA content showed a positive correlation with hepatocyte AI （r = 0.644）, and serum activity of SOD showed a negative correlation with AI （r = -0.511） in the ALD group. CONCLUSION： Chronic alcohol administration induced ALD and liver dysfunction, and increased hepatocyte apoptosis. Rsa I and Pst I restriction fragment length polymorphisms （RFLPs） were related to ALD in rats, and the c2 gene might be related to development of ALD. The MDA content and SOD activity play an important role in the process of hepatocyte apoptosis and lipid peroxidation.
出处
《世界华人消化杂志》
CAS
北大核心
2007年第20期2181-2185,共5页
World Chinese Journal of Digestology
关键词
酒精性肝病
细胞凋亡
Caspase-3
细胞色素P4502E1
氧化应激
原位末端标记法
免疫组化法
聚合酶链式反应
Alcoholic liver disease
Apoptosis
Cystein-dependent aspartate-specific programed-3
Cytochrome P4502E1
Oxidative stress
TdT-mediated d-UTP nick end labeling
Immunohistochemistry
Polymerase chain reaction
